Drug delivery to brain tumors has been a controversialsubject Some believe the blood-brain barrier is notimportant while others believe it is the major obstacle intreatment and have devised innovative approaches to cir-cumvent it These approaches can be divided into two cat-egories those that attempt to increase drug delivery ofintravascularly administered drugs by manipulating eitherthe drugs or capillary permeability and those that attemptto increase drug delivery by local administration Severalstrategies have been developed to increase the fraction ofintravascular drug reaching the tumor including intra-arterial administration barrier disruption new ways ofpackaging drugs and most recently inhibiting drugefux from tumor When given intravascularly all drugshave a common drawback the body acts as a sink andeven in the best situations only a small fraction of admin-istered drug actually reaches the tumor A consequence isthat systemic toxicity is usually the dose-limiting factorWhen given locally such as into the cerebrospinal uid ordirectly into the tumor 100 of an administered dose isdelivered to the target site However local delivery is asso-ciated with variable and unpredictable spatial distributionand variation in drug concentration The major dose-lim-

iting factor of most local delivery methods will be neuro-toxicity The relative advantages and disadvantages ofthe different methods of circumventing the blood-brainbarrier are presented in this review and special attentionis given to convection-enhanced delivery which has par-ticular promise for the local delivery of large therapeuticagents such as monoclonal antibodies antisenseoligonucleotides or viral vectors Neuro-Oncology 2 45ndash59 2000 (Posted to Neuro-Oncology [serial online]Doc 99-30 December 14 1999 URL ltneuro-oncol-ogymcdukeedugt)

Introduction

The delivery of drugs to brain tumors has long been acontroversial problem In 1977 Vick et al wrote in aneditorial ldquoWe believe that there is compelling evidence tosuggest that the lsquoblood-brain barrierrsquo as it is generallyconceived is not one of the factors impeding the successof brain tumor chemotherapyrdquo They went on to say thatldquodosage route of administration tumor cell uptakemetabolic fate within tumor cells and the washout or sinkeffect of the extracellular space and CSF are the issues thatwill have to be studiedrdquo (Vick et al 1977) Some clini-cians have agreed with Vick et al (Donelli et al 1992Stewart 1994) however on the whole the belief that theBBB3 and BTB prevent drugs from reaching brain tumorsin sufcient concentrations to kill the tumor cells hasmotivated numerous attempts to increase the amount ofdrug that reaches the tumor Many innovative methodshave been used to try to increase drug delivery includingmost recently a method in which drugs are infuseddirectly into brain tumors a method referred to as con-vection-enhanced delivery (CED) (Bobo et al 1994Laske et al 1997a Lieberman et al 1995) This review

Neuro-Oncology n JANUARY 2000 45

Neuro-Oncology

The blood-brain and blood-tumor barriers A review of strategies for increasing drugdelivery1

Dennis R Groothuis2

Evanston Northwestern Healthcare Department of Neurology Northwestern University Medical SchoolDepartment of Neurobiology and Physiology and the Institute for Neuroscience Northwestern UniversityEvanston IL 60201

Received 2 July 1999 accepted 5 August 1999

1This work was supported by National Institutes of Health Grant RO1-NS12745 and the Richard M Lilieneld and Mark Moritz Brain TumorMemorial Funds

2Address correspondence and reprint requests to Dennis R GroothuisMD Evanston Hospital 2650 Ridge Ave Evanston IL 60201

3Abbreviations used are as follows AUC area under the curve BBBblood-brain barrier BTB blood-tumor barrier CED convection-enhanced delivery CSF cerebrospinal uid HBBBD hyperosmoticblood-brain barrier disruption ia intra-arterial K1 blood-to-tissuetransfer constant k2 tissue-to-blood efux constant k3 tissue metabolism constant

attempts to place the various methods for increasing drugdelivery into context There is no ideal solution to thisproblem each delivery method has advantages and disad-vantages Changes are also occurring in two other arenasthat will have profound effects on brain tumor therapyFirst the study of drug effects and mechanisms of actionwhich is called pharmacodynamics is evolving (Ali-Osman 1999) Second the development of new classes oftherapeutic agents such as monoclonal antibodies anti-sense oligonucleotides and receptor-linked toxins intro-duces new conceptual problems in drug delivery becausemany of these compounds are large which restricts evenmore severely entry into brain tumors from the vascularcompartment (Jain 1996 1998)

There have been numerous recent reviews aboutincreasing drug delivery across the normal BBB(Pardridge 1998 Rapoport 1996 Tamai and Tsuji1996) However there are important differences betweenthe normal BBB and the BTB In this review I will try tocompare the extent to which the normal BBB and thevariably abnormal BTB restrict the delivery of blood-borne therapeutic agents to brain tumors I will then dis-cuss the various methods that are being explored toincrease the delivery of intravascular drugs to braintumors Finally I will review alternative methods of drugdelivery that completely circumvent the vascular com-partment including CED

The BBB and BTB Barriers

The normal BBB is a formidable obstacle to the move-ment of most drugs from the blood into the brain This is

illustrated in Fig 1 in which the permeability-surfacearea product for a series of compounds is shown in liverlung muscle brain and experimental brain tumors Inliver sinusoidal capillaries exhibit almost no permselec-tivity (decreasing permeability with increasing molecularsize) and as a consequence drug delivery is little affectedby molecular size Muscle capillaries are continuous buthave increased numbers of pinocytotic vesicles and vari-ably competent interendothelial junctions and as a conse-quence exhibit permselectivity Normal brain capillariesare also continuous but have tight interendothelial junc-tions few pinocytotic vesicles and no fenestrations Theprinciple route by which drugs cross the BBB is by simplediffusion The result is an astounding 8-log difference inthe rate at which an immunoglobulin will cross a livercapillary and the rate at which one will cross a capillaryin the brain

In brain tumors permeability is a complex topicThere are at least two major variables involved The rstvariable concerns the tumor microvessel populationsthat is the BTB We have recently presented evidencethat there may be three distinct microvessel populationsin brain tumors (Schlageter et al 1999) The rst con-sists of continuous nonfenestrated capillaries like thoseof normal brain Examples of brain tumors with thiscapillary population include experimental ethylni-trosourea-induced gliomas in animals (Blasberg et al1983) and in humans grade 2 astrocytomas and manyoligodendrogliomas These tumors may not enhancewith the contrast agents used with CT or MRI The sec-ond microvessel population consists of continuous fen-estrated capillaries Tumors with these microvesselsexhibit increased permeability to small but not to large

Neuro-Oncology n JANUARY 200046

DR Groothuis Review of BBB and BTB

Fig 1 The relationship between molecular size and capillary permeability The relationship between the rate of entry (expressed as the perme-ability-surface area product with units of ml gndash1 minndash1) and molecular size is shown for ve compounds (urea sucrose inulin albumin and IgGarranged in increasing molecular size) in liver lung muscle and brain capillaries (Parker et al 1984 Taylor and Granger 1984) The curve forexperimental gliomas represents a compilation of data from RG-2 (Nakagawa et al 1987) and D-54 MG gliomas (Blasberg et al 1987) Thecapillaries of liver and lung do not exhibit permselectivity whereas those of muscle and brain do exhibit permselectivity that is a decrease in therate of transcapillary passage as a function of molecular size The glioma capillaries do not exhibit signicant permselectivity however the rateof transcapillary passage of albumin and IgG is lower than liver or lung capillaries because of a fewer number of gaps in the endothelial lining

molecules To identify this tumor population permeabil-ity studies with two different-sized markers are requiredWith MRI Ostrowitzki et al beautifully illustrated dif-ferential permeability of a 9L rat glioma to gadopentetate(molecular mass = 05 kDa) and albumin-(Gd-DTPA)30(molecular mass = 92 kDa) (Ostrowitzki et al 1998)Because studies in humans are generally not performedwith two different-sized permeability markers the preva-lence of this tumor population in humans is not knownThe third capillary population represented in Fig 1 con-tains interendothelial gaps that may measure as large as 1 microm the RG-2 rat glioma (Nakagawa et al 1987Schlageter et al 1999) and D-54 MG human gliomamodel (Blasberg et al 1987) are examples As shown inFig 1 these tumor models do not exhibit permselectivityfor large molecules However Fig 1 also points out thatthe permeability-surface area product of the gliomas is 2ndash3 log units less than that of the liver which is a conse-quence of a smaller number of interendothelial gaps inthe tumors than in the liver

The second major variable with regard to capillarypermeability involves the spatial distribution of the targetcapillaries Although brain tumor capillaries may haveincreased permeability like those of the RG-2 and D-54MG tumors permeability in brain surrounding tumorrapidly returns to normal brain values within a few mil-limeters of the tumor margin If as shown by Burger(1987) individual tumor cells may reside centimetersaway from the edge of a tumor spatial variability in cap-illary function will affect drug delivery to all braintumors

To understand the effects of changes in capillary per-meability on drug delivery it may be useful to introducesome general concepts Once injected intravascularly adrug mixes with the total body volume of blood or in thecase of water-soluble compounds that are not protein-bound with total body plasma Because a 70-kg humancontains 3 liters of plasma to achieve a starting plasmaconcentration of 1 unit of ldquoidealrdquo drug per ml 3000units of drug must be given As will be shown thisunpleasant fact will color all attempts to increase drugdelivery associated with intravascular administrationThe human body acts as an enormous sink in which themajority of intravascularly administered drug will be dis-tributed not to the brain tumor but to other body tis-sues Once mixed with total body plasma the drugdistributes throughout body tissues and is then elimi-nated The time course of the mixing distribution andelimination of the drug in plasma is generally describedby several different half-times which represent theprocess of tting the plasma concentration-time data to amultiexponential expression of the form

Cp(t) 5 Ae2 a t 1 Be2 b t 1 Ce 2 g t (Eq 1)

where A B and C represent the y-intercepts and a b and g represent the time constants with units of minndash1The time constants are related to the half-times by theexpression

t a12 5

ln2(Eq 2)

a

where the half-time has units of minutes The inte-grated value of plasma concentration over time usuallyreferred to as the area under the curve (AUCPL) repre-sents the amount of drug passing through the brain ortumor vessels that is the amount of drug to which thebrain or tumor vessels are exposed

The time course of drug concentration over time inbrain or brain tumor tissue is more complex As we havediscussed elsewhere (Blasberg and Groothuis 1986) theconcentration of ideal drug in tissue (Ci) (assuming pas-sive transport across the capillary no plasma or tissuebinding and first order metabolism and inactivationkinetics) is given by

Ci(T) 5 K1 e 0

TCp(t)e

2 (k2 1 k3)(T 2 t)dt (Eq 3)

where K1 is the blood-to-tissue transfer constant (withunits of ml gndash1 minndash1) k2 is the tissue efux constant (withunits of reciprocal time min-1) and k3 is a metabolism orinactivation constant also with units of reciprocal timeEquation 3 can be used to calculate the tissue drug con-centration over time and can be used to explore theimpact of different values of K1 k2 and k3 on tissue drugconcentrations Assuming that the drug is passively dis-tributed across the BBB and not removed by other meansthen K1 and k2 are related by the expression l = K1k2where l is the equilibrium distribution volume of thedrug in the tissue

It is very useful to have some expression of the ef-ciency of the drug delivery process if for no other reasonthan to compare one experimental methodology forincreasing drug delivery with another Pardridge has usedan expression called the pharmacokinetic rule to expressthe efciency of drug entry (Pardridge 1997)

IDg 5 PS 3 AUC (Eq 4)

in which ID represents the percent of injected dose ofdrug delivered per gram of tissue PS is the permeabilitysurface area product (which for most water-soluble com-pounds is equal to K1 with units of ml gndash1 minndash1) andAUC is the plasma area under the curve (for whichPardridge uses the units IDminmicrol) However thisexpression does not consider drug efflux from tissueandor metabolism In this review I will use two expres-sions to indicate the fractional efficiency of the drugdelivery process Both expressions contain a term in thenumerator that refers to the concentration-time productof drug in brain or tumor tissue (AUCB) The rst expres-sion called the local exposure fraction represents thefraction of drug removed from the blood to which thebrain or tumor is exposed that is the blood circulatinglocally within the tissue

concentration 2 time product of drugAUCB

5per g of brain or tumor tissue

(Eq 5)AUCPL concentration 2 time product of drugper ml of perfusing plasma

Equation 5 expresses the fraction of drug removedfrom the plasma to which the tissue was exposed and istherefore an expression of local delivery efciency How-

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 47

ever this is but a small fraction of the drug in the entirebody The second expression called the total exposurefraction incorporates the total body plasma volume

concentration 2time product of drug per g

AUCB5

of brain or tumor tissue(Eq 6)3000 3 AUCPL concentration 2

time product of drugper ml of perfusing plasma

The AUCPL can be obtained from Equation 1 andAUCB can be obtained from integrating Equation 3Equation 6 reminds us that the fraction of drug enteringa gram of brain or tumor is but one small fraction of drugcirculating in the entire body and that this route of deliv-ery is inherently inefcient

Equations 5 and 6 can be used to illustrate drug deliv-ery to normal brain and brain tumor in what may beviewed as ldquobest caserdquo and ldquoworst caserdquo scenarios Theworst case scenario will almost always be represented bywater-soluble drugs and the normal BBB that is the mostrestrictive situation The best case scenario will be repre-sented by lipid-soluble drugs and highly permeabletumors such as those represented in Fig 1 by the RG-2and D-54 MG gliomas In Table 1 the local and totalexposure fractions are presented for four drugs with differ-ent permeabilities methotrexate (K1 = 00014) 5-uo-rouracil (K1 = 00096) aziridinylbenzoquinone (K1 =0145) and 13-bis(2-chloroethyl)-1-nitrosourea (K1 = 06ml gndash1 minndash1) and for two permeability scenarios that ofnormal brain and that where permeability has increased01 ml gndash1 minndash1 over normal brain The data for the calcu-lations were contained in a previous publication (Blasbergand Groothuis 1986) the reader should be aware that thistype of modeling is dependent on assumptions made aboutthe different parameters Nonetheless Table 1 can be usedto illustrate several different points First in the worst casescenario with tumor microvessel function like the normalBBB and a highly water-soluble drug (methotrexate)

about 1 of the drug to which the tumor is exposed willenter the tumor With a lipid-soluble drug like aziridinyl-benzoquinone and normal BBB function the local expo-sure fraction increases to 07 in other words increasinglipid solubility does increase delivery to brain tissue Whentumor microvessel permeability increases by 01 ml gndash1

minndash1 the fraction of methotrexate to which the tumor isexposed increases to 18 A local increase in tumor per-meability results in a potentially signicant increase indrug delivery for a water-soluble drug However note thatalthough the fractional removal across normal BBB is highfor the lipid-soluble drugs aziridinylbenzoquinone and13-bis(2-chloroethyl)-1-nitrosourea the local exposurefraction does not increase signicantly when permeabilityis increased (the increase is only 1ndash2 percentage pointsTable 1) Increased drug delivery as a result of increases incapillary permeability will be more important for water-soluble drugs than for lipid-soluble drugs for which per-meability was not as great an issue in the rst place

It is important to remember that the rest of the bodyrepresents a large sink into which most of the drug is dis-tributed The fraction of drug entering tumor comparedwith that circulating through the entire body is minisculein every case lt1 of circulating drug will reach thetumor regardless of the drugrsquos permeability and regardlessof changes in capillary permeability (Table 1) Thereforeany method that is used to increase brain tumor perme-ability will still have to deal with the reality that most ofthe administered drug is distributed to the rest of the bodyand so far as treatment of the brain tumor is concernedwasted In most instances the limit on the total dose ofdrug that can be given is imposed by the tolerance of nor-mal tissues which is systemic toxicity in the case of drugsgiven systemically Because the maximum dose that can begiven to a patient is determined by factors external totreatment of the tumor (that is systemic toxicity) we mustaccept as a starting principle that the administered drugdose cannot be increased and we must search for alterna-tive methods to increase the fraction of drug that reachesthe tumor

Neuro-Oncology n JANUARY 200048

DR Groothuis Review of BBB and BTB

Table 1 Fractional delivery of four different chemotherapeutic agents across the BBB and BTB

Permeability of normal brain Permeability increase = 01 ml gndash1 minndash1

Drug Local exposure fraction Total exposure fraction Local exposure fraction Total exposure fraction

MTX 0017a 000006a 018 000006

5-FU 011 000004 025 000008

AZQ 070 00002 072b 00002b

BCNU 054 00002 055b 00002b

Abbreviations MTX methotrexate 5-FU 5-uorouracil AZQ azridinylbenzoquinone BCNU 13-bis(2-chloroethyl)-1-nitrosourea

Four drugs with different rates of brain entry brain efux and plasma half-lives are shown For each drug the values represent the fraction of drug extracted by the tumor The localexposure fraction represents the fraction of drug removed by 1 g of tumor from the plasma to which it was exposed (Equation 5) The total exposure fraction represents the fraction of

drug removed by 1 g of tumor from the entire circulating volume of drug in plasma (Equation 6) aThe ldquoworst caserdquo scenario occurs when water-soluble drugs are crossing the normal BBB bThe ldquobest caserdquo scenario occurs when lipid-soluble drugs cross capillaries in brain tumors with increased permeability However the two columns labeled total exposure fractionemphasize that the amount of drug removed by 1 g of tumor is very small compared with the total drug circulating in the body and indicates the magnitude of the sink effect of the

rest of the body in brain tumor chemotherapy

Increasing Permeability of the BBB and BTBto Drugs Given Intravascularly

Table 2 summarizes many of the methods that are cur-rently being used to manipulate the permeability of theBBB and BTB as well as my own interpretation of theiradvantages and disadvantages In light of the previousdiscussion all of these methods share a major disadvan-tage the bulk of administered drug will be lost to the restof the body in a sink effect and the maximum adminis-tered dose will almost always be determined by systemictoxicity

Chemical Modication of Existing Drugs

Chemical modication of existing drugs may refer to sev-eral different approaches First an existing drug may bemodified to make it fit a receptor in the BBB Thisapproach has been used with melphalan in which nitro-gen mustard (mechlorethamine) was linked to phenylala-nine (Groothuis et al 1992) Chemical modication mayrefer to linking an active protein (for example growthfactor) or a peptide to an antibody against a BBB recep-tor (Pardridge 1995) More commonly chemical modi-cation refers to the process of making an existing drugmore lipid soluble with the intent of increasing BBB per-meability This approach has been used extensively withantiretroviral nucleoside analogs for treating AIDS buthas not been used much with brain tumor chemothera-peutic drugs The use of increasingly accurate means topredict physicochemical properties offers encouragementthat chemical modication will nd wider use in modify-ing brain tumor drugs (van de Waterbeemd et al 1998)

Prodrug Therapy

This refers to a special class of chemical modication inwhich a drug is chemically modied to increase its capil-

lary permeability Once in the brain the prodrug under-goes an enzymatic reaction that returns the drug to itsactive state and with reduced BBB permeability (Shermanet al 1991 Yoshikawa et al 1999) These approacheswhich have not yet been used for brain tumor chemother-apeutic drugs offer an advantage by increasing K1 inEquation 3 while k2 and k3 remain unchanged Thiswould increase the AUCB while AUCPL remainsunchanged Although this increases the fraction of drugentering the tissue it does not change total body expo-sure and unfortunately introduces the need to explorethe pharmacokinetics and pharmacodynamics of eachnew compound

Intra-Arterial Administration

This method of administration refers to the ia injectionof a therapeutic agent without the concomitant use of abarrier-modifying approach The principle advantage ofthis method is that during the course of injecting a druginto an artery the tissue perfused by that artery receives ahigher plasma concentration during the first passagethrough the circulation (AUCPL is increased) In a seriesof papers Fenstermacher and coworkers theoreticallyevaluated the efficacy of intra-arterial delivery to thebrain and brain tumors (Cowles and Fenstermacher1974 Eckman et al 1974 Fenstermacher and Cowles1977 Fenstermacher and Gazendam 1981) They evalu-ated the contributions of several variables and concludedthat there were significant delivery advantages to iaadministration in a setting where the rate of tumor bloodow is very low or where the rate of systemic transfor-mation or excretion is very high or in a unique situationwhere having crossed the BBB or BTB the drug binds tothe tissue Once the drug passes through the tumormicrovasculature it enters the systemic circulation andthe pharmacokinetics are the same as for an iv adminis-tration Thus the ideal drug for ia administration is one

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 49

Table 2 Comparison of different methods of increasing delivery of intravascularly administered compounds to brain tumors

Method of increasing delivery Principle advantages Principle disadvantages Limiting factor(s)

Chemical modication of Requirements are now Each new compound must be BBB BTB permeabilityexisting drugs understood for different evaluated individually systemic toxicity

delivery routes

Prodrug Utilizes inherent biochemical Difference between BBB and BTB BBB BTB permeabilitypathways in brain and tumor is not known systemic toxicity

Intra-arterial First pass increase in AUC Invasive small increase in AUCBT BBB BTB permeability systemic toxicity

Hyperosmotic disruption First pass increase in AUC and Invasive differential effect on BBB BTB permeability increased permeability brain and tumor short-lived systemic toxicity

Chemical modication First pass increase in AUC and Invasive no effect on brain BBB BTB permeabilityincreased permeability short-lived

Receptor-mediated transport Increased permeability Low capacity system BBB BTB permeability

Inhibiting drug efux Decreased efux may be used None known None knownwith existing drugs

Avant-garde methods Not yet known Not yet known Not yet known

Abbreviations BBB blood-brain barrier BTB blood-tumor barrier AUC area under the curve

This table lists the different approaches that are available to increase the amount of drug in a brain tumor in cases where the drug is given either by iv or ia injection The advantages

and disadvantages are an expression of the authorrsquos opinion about the most prominent feature of each method Details about each method may be found in the text

that rapidly crosses the BBB or BTB and is either boundto tissue elements or locally metabolized in the process ofexerting its anticancer effect There are practical prob-lems associated with ia administration however includ-ing the need for the tumor to reside within the arterialdistribution being infused and hemodynamic streamingof the administered drug Perhaps the most obvious evi-dence that increased tissue drug concentrations can beachieved with ia administration is the increased inci-dence of local toxicity (Arafat et al 1999) Neverthelessbecause most drugs being used to treat brain tumors donot have the ideal qualities to take advantage of ia deliv-ery the results of clinical trials of ia chemotherapy inbrain tumors show minimal if any improvement in sur-vival (Dropcho et al 1998 Gundersen et al 1998Hirano et al 1998)

Hyperosmolar Blood-Brain Barrier Disruption(HBBBD)

This method involves the infusion of a hyperosmolarsolution (usually 14 M mannitol) into a cerebral arterygenerally followed by the intra-arterial administration ofa drug The principle features of this method areincreased capillary permeability (caused by the hyperos-molar solution) followed by a rst pass advantage fromia infusion that is K1 is transiently increased as isAUCPL This method was originally proposed byRapoport (Rapoport and Thompson 1973) and has beenused extensively by Neuwelt and colleagues (Kroll andNeuwelt 1998 Neuwelt and Dahlborg 1989 Neuweltet al 1980 Neuwelt et al 1983) Controversy about theuse of this method arose because of differential suscepti-bility of the normal BBB as compared with the abnormalBTB Whereas all studies demonstrated increased perme-ability of the normal BBB some studies in animal tumorsfailed to demonstrate any increase in tumor permeability(Nakagawa et al 1984) others showed transient perme-ability increases (Groothuis et al 1990) and somereported signicant permeability increases (Neuwelt etal 1984) Most recently Zunkeler et al used PET tostudy the time course of BBB function in 13 patients withmalignant astrocytomas after HBBBD (Zunkeler et al1996) They conrmed the differential effect of HBBBDpermeability was increased 1000 in brain and 60 intumors The half-time of the osmotic effect was 81 minin brain and 42 min in tumor They modeled the effectsof HBBBD on methotrexate and concluded that concen-trations above 1 microM the minimal concentration requiredfor an effect from methotrexate would not be enhancedin tumor and would be enhanced only 10 in brainThus HBBBD in combination with ia administrationwould be most useful for drugs that once having crossedthe BBB bind to brain or tumor tissue and do not efuxback across the BBB In addition the differential effect ofHBBBD on normal brain compared with that of tumormust be considered when evaluating neurotoxicity

Chemical Modication of the BBB and BTB

The latest entry into the therapeutic arena is the use ofchemical agents to disrupt the BBB (Black 1995) The

agents are mostly derivatives of normal vasoactive com-pounds and include bradykinin (Inamura and Black1994) interleukin-2 (Gutman et al 1996) leukotrieneC4 (Black and Chio 1992) and others (de Vries et al1996) The principle feature of these drugs is that theyare given ia and followed by an ia injection of a thera-peutic agent RMP-7 is a commercially developedbradykinin analog that has reached clinical trials (Ford etal 1998) and acts by increasing permeability at the inter-cellular junction (Sanovick et al 1995) Studies withPET in 9 malignant gliomas have shown that permeabil-ity to 68Ga-EDTA was increased 46 plusmn 42 in tumorswithout a signicant increase in tumor-free brain (Blacket al 1997) Another report suggests that RMP-7 canincrease delivery across the normal BBB (Emerich et al1998) RMP-7 is generally administered over a 15-minperiod the timing of administration of the chemothera-peutic drug relative to the RMP-7 has varied eg as abolus after RMP-7 or alternating chemotherapeutic drugadministration with RMP-7 BBB function is restoredrapidly after the RMP-7 infusion ends continuing theinfusion for over 20 min results in tachyphylaxis withloss of responsiveness (Bartus 1999) As with ia andHBBBD administration the advantage with chemicalmodication of the BBB is limited to a fractional increasein the circulating drug to which the tumor is exposedand still represents a vanishingly small fraction of thetotal body drug exposure Like those previous tech-niques chemical modication of the BBB will be ideal fora drug with dened properties as discussed before How-ever this ideal drug has yet to be developed

Receptor-Mediated Transport

This is a very broad area because any receptor-mediatedtransport system in the BBB can be selected as a targetThe considerations about drug delivery in a facilitatedtransport system are different from those involving sim-ple diffusion First a facilitated transport system can becharacterized by the Michaelis-Menten constants Km(the concentration at which the reaction velocity is halfmaximal) and Vmax (the limiting velocity as the concen-tration approaches innity) An important factor in thefacilitated transport of drugs is the plasma concentrationof the native substrate for the receptor For examplephenylalanine crosses the BBB via the large neutral aminoacid transport (LNAA) system melphalan is achemotherapeutic drug with a phenylalanine backboneand could therefore be transported by the LNAA systemHowever the normal plasma levels of phenylalaninecompete with melphalan to such a degree that the rate ofmelphalan transport into experimental gliomas is notincreased (Groothuis et al 1992) Lowering plasmaphenylalanine levels increased the rate of melphalanentry into brain tumors but only by a modest 8 Mostof the available transport receptors are low afnity lowcapacity systems such as those for the nucleosides vari-ous peptides transferrin and insulin (Ermisch et al1993 Pardridge et al 1995 Friden et al 1996 Tamai etal 1997) Although these transport systems have beenused to increase brain delivery of oligonucleotides(Boado et al 1998 Normand-Sdiqui and Akhtar 1998)

Neuro-Oncology n JANUARY 200050

DR Groothuis Review of BBB and BTB

nerve growth factor (Friden et al 1993) and vasoactiveintestinal peptide (Bickel et al 1993) none of thesetransport systems has been explored to any extent inbrain tumors

Inhibiting Drug Efux

Throughout most of the history of the BBB movementacross the capillary for most compounds without a trans-port system has been assumed to occur by simple diffu-sion the process is passive and the rate that a compoundcrosses the barrier is the same in both directions so thatthe process is symmetrical Recent studies have begun tochallenge this idea In particular P-glycoprotein repre-sents an ATP-dependent efflux pathway that confersresistance to cancer cells by allowing them to move avariety of chemotherapeutic drugs out of the cell againsta concentration gradient (Schinkel 1999 Tsuji andTamai 1997) P-glycoprotein appears to be localized pri-marily to the luminal capillary membrane and the num-ber of drugs that are transported by this system may bequite large (Schinkel 1999) Immunohistochemically P-glycoprotein has been demonstrated in malignantglioma tumor cells (Leweke et al 1998 von Bossanyi etal 1997) Leweke et al commented that tumor endothe-lial cells stained in 20 of 21 cases and von Bossanyi et alstated that tumor blood vessels were positive in 60 oftumors PET has been used to demonstrate the reversal ofP-glycoprotein pump function in mdr1a knock-out micewith cyclosporin A (Hendrikse et al 1998) Since manychemotherapeutic drugs are transported by P-glycopro-tein this represents a logical and exciting target in braintumor chemotherapy especially because inhibition of P-glycoprotein function could occur at both the BTB andin tumor cells Inhibiting P-glycoprotein would increaseboth extracellular and intracellular drug concentration(by decreasing k2 in Equation 3) without increasing theamount of drug administered This is an avenue of modi-fying brain tumor therapy that must be explored

Avant-Garde Methods of Delivery

Included in this category are the newest methods forintravascular delivery including the use of biodegradablenanoparticles (Schroeder et al 1998 Song et al 1997)liposomes (Allen and Moase 1996 Sakamoto and Ido1993) monoglyceride-based systems (Chang et al1994) and magnetic microspheres (Chang and Bodmeier1997) Many of these novel approaches are evaluatingdifferent methods of giving drug-laden molecules thatwill preferentially accumulate in a tissue usually com-bined with selective methods of perfusion such as iadelivery For example Pulfer and Gallo delivered 1ndash2 micromia magnetic aminodextran microspheres to RG-2 ratbrain tumors and used a magnetic eld to retain the par-ticles within the tumor They showed that this methodresulted in signicant accumulation of the particles as aresult of surface charge and magnetic eld (Chang andBodmeier 1997) These methods are too new to beapplied to human brain tumors but are representative ofthe diverse and novel directions that investigators areusing to increase brain tumor delivery

Summary of Intravascular Drug Delivery

A tremendous amount of progress has been made in thepast few years in our understanding of drug delivery tobrain tumors This progress has not yet translated intoimpressive increases in survival of patients with braintumors There are major areas that remain to be exploredand developed in the arena of intravascularly adminis-tered drugs For example because ia administration(with or without HBBBD or chemical modification)offers a unique rst pass advantage a search for drugsthat have high lipid solubility and bind to brain tumortargets is likely to be protable Second increasing tissuedrug concentration by the use of efux inhibitors is obvi-ous there will be no increase in total body exposurewhile potentially achieving signicant increases in tumordrug concentration However the ultimate limiting factorfor all intravascularly administered drugs is that of thepotential toxicity from total body exposure It seemsunlikely that this important and unfortunately negativevariable can be overcome

Injection or Infusion of Drugs Directly intothe Brain or Its Cavities

Table 3 summarizes the different approaches that arebeing used to administer drugs directly into the brainthus bypassing the BBB All of these routes and methodsof administration drastically change the pharmacokineticlandscape Instead of concern about crossing the formi-dable BBB or BTB the drugs are administered directlyinto the CSF or brain extracellular space Instead of need-ing to measure the inux rate it is necessary to under-stand drug movement within the brain and to understandefux mechanisms In marked contrast to intravascularadministration 100 of the administered dose can bedelivered using these methods of administration Theproblem then becomes one of understanding the forcesthat control the movement of the drug within brain andtumor tissue One or more of four forces will affect drugmovement depending upon the particular mode of drugadministration bulk flow of CSF bulk flow of braininterstitial uid bulk ow due to the infusion of a solu-tion into the brain or CSF and diffusion Drug movementwill also be variably affected by the amount of whitematter in the target area as a result of the change fromthe highly tortuous environment of a gray matter struc-ture (isotropic) to the highly oriented ber pathways ofwhite matter (anisotropic)

Intrathecal and Intraventricular Drug Administration

In this review intrathecal drug administration refers tothe administration of drug into the subarachnoid spaceusually into the lumbar subarachnoid space whereasintraventricular administration refers to injection or infu-sion into the lateral ventricle They are discussed togetherbecause of the large number of features they have in com-mon The principle feature of drug administration bythese routes is dominated by the bulk uid ow of the

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 51

CSF The major source of CSF is from the choroidplexuses of the lateral and 4th ventricles as much asanother 30 may originate from the brain extracellularuid The direction of the ow is from the lateral ventri-cles into the 3rd and then 4th ventricles out through theforamina of Luschke and Magendie into the basal cis-terns around the convexities and out of the subarach-noid space through the arachnoid granulations In ratswhen 14C-sucrose was injected into the lateral ventriclethe sucrose distributed rapidly within 5 min it was foundin the basal cisterns and by 1 h the sucrose had largely leftthe subarachnoid space (Ghersi-Egea et al 1996) Whenthe injected agent is metabolically active it may enterselected cells (Rubertone et al 1993) In humans the nor-mal volume of CSF produced is between 400 and 500 mlper day Since the CSF spaces within the brain are about150 ml in volume there is a considerable bulk ow withinthis system although the velocity varies tremendouslyfrom site to site When isotopes are injected into the lum-bar subarachnoid space they travel up the spinal sub-arachnoid space and are found in the basal cisterns by 3 hBy 24 h they are found over the cerebral convexities CSFow from the lateral ventricles is directional isotopesinjected into the lumbar CSF do not normally enter thelateral ventricles As a result of these bulk ow pathwaysmaterials injected into the CSF will attain initial concen-trations that are directly proportional to the concentra-tion in the infusate This route of administration is idealfor situations in which the target is within the subarach-noid space (for example carcinomatous meningitis) or inwhich the target is close to the CSF brain interface Carci-nomatous meningitis has long been treated with intrathe-cal infusions including the more recent use of monoclonalantibodies (Brown et al 1996) However the capacity ofdrugs to enter the brain extracellular space from the CSFis limited Numerous studies have shown that entry intothe brain is diffusional and that concentrations declineexponentially from the brain surface (Blasberg 1977Blasberg et al 1977 Groothuis and Levy 1997 Patlak

and Fenstermacher 1975) This route of administration isboth impractical and inefcient for intraparenchymalbrain tumors

Intratumoral and Intracavitary Injection

These are discussed together and briey because they areboth limiting cases of convection-enhanced deliverywhich is discussed below These both represent localforms of drug administration in which the distributionwill be determined initially by the rate and duration ofthe injection and secondly by tissue bulk ow pathwaysand diffusion The history of intratumoral injection wasdiscussed by Tomita (1991) who recognized the poten-tial usefulness of this approach as well as the importanceof neurotoxicity as the limiting factor Intratumoral andintracavitary injections are being widely explored asmeans of delivering large agents such as viral vectorsgrowth factors and cells (Bigner et al 1998 Farkkila etal 1994 Hsiao et al 1997 Puri et al 1996 Ram et al1997 Yang et al 1997) All of the attributes discussed inthe section on CED will also apply to these forms ofdrug delivery

Microdialysis

Microdialysis is a method that employs the passive diffu-sion of a drug across a semipermeable membrane (deLange et al 1997 de Lange et al 1999 Parsons andJustice 1994) It can be used to both sample and deliverdrugs to the surrounding tissue and has been used tosample extracellular drug concentrations in experimentalbrain tumors (Devineni et al 1996 Nakashima et al1997) The principle features of microdialysis when usedas a drug delivery methodology are that the tissue con-centrations are a function of the drug concentration inthe dialysate (which places control of the concentrationin hands of the user) and that distribution of drug awayfrom the dialysis catheter occurs by diffusion (Dykstra et

Neuro-Oncology n JANUARY 200052

DR Groothuis Review of BBB and BTB

Table 3 Comparison of different methods for local drug delivery to brain tumors

Method of circumventing the BBB Principle advantages Principle disadvantages Limiting factor(s)

Intrathecal Easy access ideal for therapy of Not useful for parenchymal Bulk ow rate of CSFmeningeal disease disease

Intraventricular administration Easy access ideal for therapy of Not useful for parenchymal Bulk ow rate of CSFmeningeal disease disease

Intratumoral injection User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Intracavitary injection User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Microdialysis User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Biodegradable polymers User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Convection-enhanced delivery User control over administered Invasive Unpredictable distribution dose 100 reaches target neurotoxicity

Abbreviations BBB blood-brain barrier CSF cerebrospinal uid

This table lists the different approaches that are available to deliver therapeutic drugs and agents by local administration The advantages and disadvantages are an expression of theauthorrsquos opinion about the most prominent feature of each method Details about each method may be found in the text

al 1992) However the normal BBB is disrupted imme-diately around the catheter and stays disrupted for sometime after catheter insertion making drug efux in thisvicinity an additional problem when trying to understandlocal delivery kinetics (Groothuis et al 1998 Morgan etal 1996 Westergren et al 1995) Because the principlefactors in local tissue concentrations are diffusion andorlocal brain bulk flow patterns the volume of brainreached by microdialysis will be relatively small anddrug concentrations will in general decline exponen-tially away from the dialysis cannula This method willbe applicable when the volume of tissue that needs to bereached is small The effect of normal brain bulk owpathways on this delivery method remains to be studied

Biodegradable Polymers

This represents a local delivery system in whichbiodegradable polymers are loaded with drug andimplanted into brain or tumor tissue resulting in timed-release of drug (Cao and Shoichet 1999 Menei et al1997) The principle features of this system are that mul-tiple polymer pellets or wafers can be implanted whichgives control over the spatial distribution and that drugdistribution away from the source occurs by diffusionwith a variable contribution from the inherent bulk owpathways of the brain (Fung et al 1998) This approachhas been used in patients with malignant gliomas whereits safety and efcacy have been demonstrated (Brem etal 1995 Olivi and Brem 1994) As with microdialysisthe principle limiting feature of this method of drug deliv-ery is that drug distribution occurs mainly by diffusionin which drug concentrations fall exponentially awayfrom the source and are generally limited to a few mil-limeters of tissue penetration (Fung et al 1998)

Convection-Enhanced Delivery

This is a relatively new method of drug delivery to braintumors The principle feature of this delivery method isthat a drug solution is infused directly into the brain Theconcentration of the drug is independent of the infusionrate and duration and the distribution of the drug is afunction of the hydrostatic pressure of the infusion aswell as diffusion into surrounding tissue From a deliveryperspective the only limiting factor to the drug concen-tration that can be delivered is the solubility of the drugCED has been discussed from a theoretical perspective(Morrison et al 1994) it has been explored in severalanimal models (Bobo et al 1994 Groothuis et al 1999Laske et al 1997a Lieberman et al 1995 Viola et al1995) and it has been used for delivery of a targetedtoxin in human gliomas (Laske et al 1997b) The distri-bution of the infused drug is highly dependent uponwhether the drug is infused into a homogeneous graymatter structure or a structure containing ber pathwaysIn the case of the infusion of a drug-containing solutioninto a homogeneous brain structure the infusion initiallyproduces a spherical volume in which the concentrationof the drug is directly proportional to that of the infusate(Fig 2b) After the infusion reaches equilibrium at whichpoint the amount of drug leaving across brain capillaries

is equal to the amount being infused the drug begins todiffuse away from the edge of the sphere at concentra-tions that decline exponentially in accordance with themathematics of diffusion (Fig 2b) If the infusionencounters an organized white matter pathway the rateof movement is either increased or decreased (dependingupon the direction of the inherent bulk ow within thatpathway) (Fig 2c) Morrison stated that when the infu-sion is being made into a homogeneous isotropicmedium the radius of the central spherical componentmdashthe convective componentmdashcan be related to the infusionrate and the efux constant of the drug being infused

rp 5 3 2qv (4p K) (Eq 7)

where qv is the infusion rate and K represents thesteady-state efux rate constant Fig 3 shows a plot of theradius of the convective component as a function of infu-sion rate and efux constant From reviewing Fig 3

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 53

Fig 2 The distribution of 14C-sucrose in rat brain by convection-enhanced delivery The diagram in the upper left (A) shows the targetlocation (center of the caudate nucleus) as well as the organization ofthe brain at the target level After a 1-h 05-microlmin infusion the iso-tope is found in a spherical shape with even distribution across (B) By8 h the isotope is moving into the external capsule (C) A similarappearance is found after a constant infusion over 7 days the centralconvective component is clearly visible surrounded by decreasingconcentration from the edge (D) Note the isotopersquos rapid lateralmovement down the external capsule (probably representing a nor-mal brain bulk uid pathway) but not into the corpus callosum Infu-sion into rat brain for 7 days produced no neuropathologic changesother than those associated with catheter insertion

which extends from infusion rates provided by osmoticmini-pumps (1 microlh) to high infusion rates (10 microlmin) it is

apparent that the volume of affected brain remains quitesmall for most infusion rates Similarly as the efux con-stant decreases indicating slower rates of crossing theBBB the convection volume increases Fig 3 illustratesthat CED is best suited for delivery of large compounds(with low efux rates) and delivery at moderately highinfusion rates if the intent is to reach large brain volumes

The shape of the volume of distribution at steady statewill be highly dependent on the location in the brain thatis being infused Fig 4 shows a CT scan of a dog brainreceiving an infusion of iodinated contrast into whitematter The iodinated contrast rapidly distributes in thewhite matter of the ipsilateral hemisphere with rapidlydecreasing concentrations in the cortex on the affectedside Note that in both Figs 2 and 4 the infused materialdoes not cross the midline through the corpus callosumThis suggests that the direction of the normal bulk owpathway within the corpus callosum is away from themidline thus opposing the CED infusion component Incontrast to intravascularly administered drugs whereincreased capillary permeability results in increased tissueconcentrations (Table 1) the opposite effect will occur inCED infusions Increased efux across permeable tumorcapillaries will reduce both the extracellular drug concen-tration and the volume of distribution In Fig 5 highconcentrations of 14C-sucrose were achieved by a CEDinfusion into an RG-2 glioma but at a distance of 24mm from the infusion site tissue concentrationsdecreased by several logarithmic units Although studieshave not been done in humans it is easy to anticipatethat the volume of distribution from a CED infusion willfollow similar principles Fig 6 shows several levels of anMRI scan through a malignant glioma Each set of scansshows a gadolinium-enhanced tumor margin (which canbe considered naturersquos equivalent to a CED infusion) anda T2-weighted image (which corresponds to the extracel-lular water originating from the tumor source) The

Neuro-Oncology n JANUARY 200054

DR Groothuis Review of BBB and BTB

Fig 3 Relationship between radius of convective CED componentinfusion rate and efux rate The radius of the inection pointbetween the convective and diffusional components of CED infusionsis shown for infusion into a homogeneous brain structure (Equation7) The values on the vertical axis represent the radius inside whichthe local concentrations remain uniform and outside of which drugconcentrations decline exponentially Changes for each 5-cm incre-ment are represented by different shades of gray The infusion ratewas chosen from 1 acute 10ndash6 to 1 acute 10ndash2 ml per minndash1 The lower valuesare those seen with infusions from osmotic mini-pumps while thehigher values represent the upper limit of CED infusion rates in dogsThe efux rate was chosen to vary from 1 acute 10ndash1 to 1 acute 10ndash7 minndash1The volume of the convective component increases as a function ofboth increasing infusion rate and decreasing efux rate

Fig 4 Illustration of CED infusion in a dog The gure on the left shows the CT scan at the start of a 3 microl minndash1 infusion of Isovue (iopamidol)into the hemispheric white matter of a dog while that on the right shows the same coronal level on the same dog after 10 days of infusion Thescan on the left contains an air bubble in the lateral ventricle the cannula tip contains a metal marker (the high density circle) and a small amountof Isovue surrounding the catheter tip In the scan on the right the Isovue has distributed at a high concentration throughout the white matterof the ipsilateral hemisphere with a rapid decline in concentration at gray-white interfaces Note that the distribution of Isovue is limited to theipsilateral hemisphere No neuropathologic changes were observed as a result of the infusion

shape and distribution of the edema are highly irregularand very dependent upon the brain structures encoun-tered Note that the edema uid moves into the internalcapsule but not into the thalamus or cortex Notice alsothat the edema uid does not cross the posterior limb ofthe corpus callosum although it comes into contact withit at several levels

A dominant feature of CED is that distribution of thedrug is determined by the infusion parameters and thetype of tissue being infused whereas the concentration ofdrug in the infusate is unrelated to these parameters Themaximum concentration of the drug in the infusate willbe determined by neurotoxicity within the central con-vective component of the infusion with variable concen-trations in the anisotropic brain surrounding the centralcomponent and concentrations 50 to 100 times lower inthe diffusional component furthermost from the infusioncatheter The ability to manipulate drug concentration inbrain and tumor tissue with such ease and to maintainthese manipulations for long periods of time (Groothuiset al 1999) brings a need for the development ofentirely new concepts For example the limits withregard to the composition of the infusate need to beexplored such as pH osmolarity and ionic compositionFurthermore how does one reliably study neurotoxicityAre infusions into animal models going to be reliable pre-dictors of toxicity in humans Will differential neurotox-icity become an issue It is possible that neuronsastrocytes and oligodendrogliocytes will display differ-ent levels of susceptibility to many drugs Will we seedelayed neurotoxicity such as that seen with radiation Itwill probably be best for many of the variables associatedwith neurotoxicity to be explored in animals before sub-jecting humans to CED infusions

Summary of Drug Delivery that Circumvents the BBB

There has been an explosion of methodology involvinglocal tissue drug delivery during the past 10 years Someof these techniques such as CED fulll the promise tocircumvent the BBB entirely In fullling this promisethese methods introduce new problems such as the rela-tionship between the spatial distribution of drugs andneurotoxicity that need to be understood before thesetechniques are widely used However for viral vectorsmonoclonal antibodies antisense oligonucleotides andother therapeutic agents that will be unable to cross theBBB in therapeutically effective amounts CED may pro-vide the necessary delivery tool

The State of Drug Delivery to Brain Tumorsndash1999

If we return to the statement of Vick et al at the begin-ning of this review we can now state with certainty thatthe BBB and BTB are major factors limiting the access ofmany therapeutic agents to brain tumors and these bar-riers will become even more signicant with the develop-ment of new molecular biological therapies that willinvolve large molecules viruses and even cells It is alsoremarkable how prescient those authors were in identify-ing the other parts of the drug delivery process that mustnow be understood and how little progress we havemade in elucidating ldquotumor cell uptake metabolic fatewithin tumor cells and the washout or sink effect of theextracellular space and CSFrdquo (Vick et al 1977) Formost of the past 20 years we have focused so much on

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 55

Fig 5 Infusion of 14C-sucrose into an RG-2 rat glioma An infusion cannula was placed into the caudate nucleus of a Fisher-344 rat (targetshown in Fig 2A) and RG-2 cells were injected Ten days later 14C-sucrose was infused at a rate of 10 microlh for 7 days Autoradiographic imageswere made (right images) and the sections were then stained with hematoxylin-eosin (left images) The top two images show that the 14C-sucrose distributes in high concentration throughout the RG-2 tumor extending into the edematous external capsule (E) However thelower two sections taken from 24 mm behind the infusion site still contain tumor and show that the concentration of 14C-sucrose is 100000times lower These sections illustrate the problems associated with controlling drug delivery by convection-enhanceddelivery

ways to increase delivery across the BBB that we havefailed to address these other issues which now becomethe forefront in drug delivery methods such as CED Thearsenal from which to choose a drug delivery method istruly impressive We must now study the other factorsthat limit effective brain tumor therapy Perhaps the mostpressing of these includes the need to develop effective

laboratory tools for identifying the relationship betweenthe concentration-time product of therapeutic agents andtumor cell kill rather than relying on empirical testing inhuman subjects It is most likely that the new century willsee the development and application of tools for individ-ualizing chemotherapy for brain tumors and other solidhuman tumors

Ali-Osman F (1999) Human glioma cultures and in vitro analysis of therapeu-

tic response In Berger MS and Wilson CB (Eds)The Gliomas

Philadelphia WB Saunders Co pp 134ndash142

Allen TM and Moase EH (1996) Therapeutic opportunities for targeted

liposomal drug delivery Adv Drug Deliv Rev 21 117ndash133

Arafat T Hentschel P Madajewicz S Manzione J Chowhan N Davis

R Roche P Af I Roque C Meek A and Magdy S (1999) Toxicities

related to intraarterial infusion of cisplatin and etoposide in patients with

brain tumors J Neurooncol 42 73ndash77

Bartus RT (1999) The blood-brain barrier as a target for phamacological

modulation Curr Opin Drug Discov Dev 2 152ndash167

Bickel U Yoshikawa T Landaw EM Faull KF and Pardridge WM

(1993) Pharmacologic effects in vivo in brain by vector-mediated peptide

delivery Proc Natl Acad Sci USA 90 2618ndash2622

Bigner DD Brown MT Friedman AH Coleman RE Akabani G Fried-

man HS Thorstad WL McLendon RE Bigner SH Zhao XG

Pegram CN Wikstrand CJ Herndon JE II Vick NA Paleologos N

Cokgor I Provenzale JM and Zalutsky MR (1998) Iodine-131-labeled

antitenascin monoclonal antibody 81C6 treatment of patients with recur-

rent malignant gliomas Phase I trial results J Clin Oncol 16 2202ndash2212

Black KL (1995) Biochemical opening of the blood-brain barrier Adv Drug

Deliv Rev 15 37ndash52

Neuro-Oncology n JANUARY 200056

DR Groothuis Review of BBB and BTB

Fig 6 Distribution of vasogenic edema in a malignant glioma There are four pairs of scans extending through a tumor in the left temporal lobeextending from the lowest (A) to the highest level (D) Each pair of scans contains a gadolinium-enhanced scan (left) and a T2-weighted image(right) The gadolinium-enhanced scans show the area of abnormally permeable BBB The T2-weighted images show the distribution of edema(water) emanating from the tumor The tumor may be considered a natural equivalent of a convection-enhanced delivery infusion The resultingedema spreads extensively through white matter in the ipsilateral hemisphere reaching forward in the internal capsule (levels C and D) but notpenetrating gray matter structures such as the thalamus (levels C and D) or cortex (levels AndashD) Note that the edema contacts the posterior limbof the corpus callosum in levels C and D but does not extend into the corpus callosum suggesting that there is a physiological barrier to the bulkmovement of uid in that direction

References

attempts to place the various methods for increasing drugdelivery into context There is no ideal solution to thisproblem each delivery method has advantages and disad-vantages Changes are also occurring in two other arenasthat will have profound effects on brain tumor therapyFirst the study of drug effects and mechanisms of actionwhich is called pharmacodynamics is evolving (Ali-Osman 1999) Second the development of new classes oftherapeutic agents such as monoclonal antibodies anti-sense oligonucleotides and receptor-linked toxins intro-duces new conceptual problems in drug delivery becausemany of these compounds are large which restricts evenmore severely entry into brain tumors from the vascularcompartment (Jain 1996 1998)

There have been numerous recent reviews aboutincreasing drug delivery across the normal BBB(Pardridge 1998 Rapoport 1996 Tamai and Tsuji1996) However there are important differences betweenthe normal BBB and the BTB In this review I will try tocompare the extent to which the normal BBB and thevariably abnormal BTB restrict the delivery of blood-borne therapeutic agents to brain tumors I will then dis-cuss the various methods that are being explored toincrease the delivery of intravascular drugs to braintumors Finally I will review alternative methods of drugdelivery that completely circumvent the vascular com-partment including CED

The BBB and BTB Barriers

The normal BBB is a formidable obstacle to the move-ment of most drugs from the blood into the brain This is

illustrated in Fig 1 in which the permeability-surfacearea product for a series of compounds is shown in liverlung muscle brain and experimental brain tumors Inliver sinusoidal capillaries exhibit almost no permselec-tivity (decreasing permeability with increasing molecularsize) and as a consequence drug delivery is little affectedby molecular size Muscle capillaries are continuous buthave increased numbers of pinocytotic vesicles and vari-ably competent interendothelial junctions and as a conse-quence exhibit permselectivity Normal brain capillariesare also continuous but have tight interendothelial junc-tions few pinocytotic vesicles and no fenestrations Theprinciple route by which drugs cross the BBB is by simplediffusion The result is an astounding 8-log difference inthe rate at which an immunoglobulin will cross a livercapillary and the rate at which one will cross a capillaryin the brain

In brain tumors permeability is a complex topicThere are at least two major variables involved The rstvariable concerns the tumor microvessel populationsthat is the BTB We have recently presented evidencethat there may be three distinct microvessel populationsin brain tumors (Schlageter et al 1999) The rst con-sists of continuous nonfenestrated capillaries like thoseof normal brain Examples of brain tumors with thiscapillary population include experimental ethylni-trosourea-induced gliomas in animals (Blasberg et al1983) and in humans grade 2 astrocytomas and manyoligodendrogliomas These tumors may not enhancewith the contrast agents used with CT or MRI The sec-ond microvessel population consists of continuous fen-estrated capillaries Tumors with these microvesselsexhibit increased permeability to small but not to large

Neuro-Oncology n JANUARY 200046

DR Groothuis Review of BBB and BTB

Fig 1 The relationship between molecular size and capillary permeability The relationship between the rate of entry (expressed as the perme-ability-surface area product with units of ml gndash1 minndash1) and molecular size is shown for ve compounds (urea sucrose inulin albumin and IgGarranged in increasing molecular size) in liver lung muscle and brain capillaries (Parker et al 1984 Taylor and Granger 1984) The curve forexperimental gliomas represents a compilation of data from RG-2 (Nakagawa et al 1987) and D-54 MG gliomas (Blasberg et al 1987) Thecapillaries of liver and lung do not exhibit permselectivity whereas those of muscle and brain do exhibit permselectivity that is a decrease in therate of transcapillary passage as a function of molecular size The glioma capillaries do not exhibit signicant permselectivity however the rateof transcapillary passage of albumin and IgG is lower than liver or lung capillaries because of a fewer number of gaps in the endothelial lining

molecules To identify this tumor population permeabil-ity studies with two different-sized markers are requiredWith MRI Ostrowitzki et al beautifully illustrated dif-ferential permeability of a 9L rat glioma to gadopentetate(molecular mass = 05 kDa) and albumin-(Gd-DTPA)30(molecular mass = 92 kDa) (Ostrowitzki et al 1998)Because studies in humans are generally not performedwith two different-sized permeability markers the preva-lence of this tumor population in humans is not knownThe third capillary population represented in Fig 1 con-tains interendothelial gaps that may measure as large as 1 microm the RG-2 rat glioma (Nakagawa et al 1987Schlageter et al 1999) and D-54 MG human gliomamodel (Blasberg et al 1987) are examples As shown inFig 1 these tumor models do not exhibit permselectivityfor large molecules However Fig 1 also points out thatthe permeability-surface area product of the gliomas is 2ndash3 log units less than that of the liver which is a conse-quence of a smaller number of interendothelial gaps inthe tumors than in the liver

The second major variable with regard to capillarypermeability involves the spatial distribution of the targetcapillaries Although brain tumor capillaries may haveincreased permeability like those of the RG-2 and D-54MG tumors permeability in brain surrounding tumorrapidly returns to normal brain values within a few mil-limeters of the tumor margin If as shown by Burger(1987) individual tumor cells may reside centimetersaway from the edge of a tumor spatial variability in cap-illary function will affect drug delivery to all braintumors

To understand the effects of changes in capillary per-meability on drug delivery it may be useful to introducesome general concepts Once injected intravascularly adrug mixes with the total body volume of blood or in thecase of water-soluble compounds that are not protein-bound with total body plasma Because a 70-kg humancontains 3 liters of plasma to achieve a starting plasmaconcentration of 1 unit of ldquoidealrdquo drug per ml 3000units of drug must be given As will be shown thisunpleasant fact will color all attempts to increase drugdelivery associated with intravascular administrationThe human body acts as an enormous sink in which themajority of intravascularly administered drug will be dis-tributed not to the brain tumor but to other body tis-sues Once mixed with total body plasma the drugdistributes throughout body tissues and is then elimi-nated The time course of the mixing distribution andelimination of the drug in plasma is generally describedby several different half-times which represent theprocess of tting the plasma concentration-time data to amultiexponential expression of the form

Cp(t) 5 Ae2 a t 1 Be2 b t 1 Ce 2 g t (Eq 1)

where A B and C represent the y-intercepts and a b and g represent the time constants with units of minndash1The time constants are related to the half-times by theexpression

t a12 5

ln2(Eq 2)

a

where the half-time has units of minutes The inte-grated value of plasma concentration over time usuallyreferred to as the area under the curve (AUCPL) repre-sents the amount of drug passing through the brain ortumor vessels that is the amount of drug to which thebrain or tumor vessels are exposed

The time course of drug concentration over time inbrain or brain tumor tissue is more complex As we havediscussed elsewhere (Blasberg and Groothuis 1986) theconcentration of ideal drug in tissue (Ci) (assuming pas-sive transport across the capillary no plasma or tissuebinding and first order metabolism and inactivationkinetics) is given by

Ci(T) 5 K1 e 0

TCp(t)e

2 (k2 1 k3)(T 2 t)dt (Eq 3)

where K1 is the blood-to-tissue transfer constant (withunits of ml gndash1 minndash1) k2 is the tissue efux constant (withunits of reciprocal time min-1) and k3 is a metabolism orinactivation constant also with units of reciprocal timeEquation 3 can be used to calculate the tissue drug con-centration over time and can be used to explore theimpact of different values of K1 k2 and k3 on tissue drugconcentrations Assuming that the drug is passively dis-tributed across the BBB and not removed by other meansthen K1 and k2 are related by the expression l = K1k2where l is the equilibrium distribution volume of thedrug in the tissue

It is very useful to have some expression of the ef-ciency of the drug delivery process if for no other reasonthan to compare one experimental methodology forincreasing drug delivery with another Pardridge has usedan expression called the pharmacokinetic rule to expressthe efciency of drug entry (Pardridge 1997)

IDg 5 PS 3 AUC (Eq 4)

in which ID represents the percent of injected dose ofdrug delivered per gram of tissue PS is the permeabilitysurface area product (which for most water-soluble com-pounds is equal to K1 with units of ml gndash1 minndash1) andAUC is the plasma area under the curve (for whichPardridge uses the units IDminmicrol) However thisexpression does not consider drug efflux from tissueandor metabolism In this review I will use two expres-sions to indicate the fractional efficiency of the drugdelivery process Both expressions contain a term in thenumerator that refers to the concentration-time productof drug in brain or tumor tissue (AUCB) The rst expres-sion called the local exposure fraction represents thefraction of drug removed from the blood to which thebrain or tumor is exposed that is the blood circulatinglocally within the tissue

concentration 2 time product of drugAUCB

5per g of brain or tumor tissue

(Eq 5)AUCPL concentration 2 time product of drugper ml of perfusing plasma

Equation 5 expresses the fraction of drug removedfrom the plasma to which the tissue was exposed and istherefore an expression of local delivery efciency How-

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 47

ever this is but a small fraction of the drug in the entirebody The second expression called the total exposurefraction incorporates the total body plasma volume

concentration 2time product of drug per g

AUCB5

of brain or tumor tissue(Eq 6)3000 3 AUCPL concentration 2

time product of drugper ml of perfusing plasma

The AUCPL can be obtained from Equation 1 andAUCB can be obtained from integrating Equation 3Equation 6 reminds us that the fraction of drug enteringa gram of brain or tumor is but one small fraction of drugcirculating in the entire body and that this route of deliv-ery is inherently inefcient

Equations 5 and 6 can be used to illustrate drug deliv-ery to normal brain and brain tumor in what may beviewed as ldquobest caserdquo and ldquoworst caserdquo scenarios Theworst case scenario will almost always be represented bywater-soluble drugs and the normal BBB that is the mostrestrictive situation The best case scenario will be repre-sented by lipid-soluble drugs and highly permeabletumors such as those represented in Fig 1 by the RG-2and D-54 MG gliomas In Table 1 the local and totalexposure fractions are presented for four drugs with differ-ent permeabilities methotrexate (K1 = 00014) 5-uo-rouracil (K1 = 00096) aziridinylbenzoquinone (K1 =0145) and 13-bis(2-chloroethyl)-1-nitrosourea (K1 = 06ml gndash1 minndash1) and for two permeability scenarios that ofnormal brain and that where permeability has increased01 ml gndash1 minndash1 over normal brain The data for the calcu-lations were contained in a previous publication (Blasbergand Groothuis 1986) the reader should be aware that thistype of modeling is dependent on assumptions made aboutthe different parameters Nonetheless Table 1 can be usedto illustrate several different points First in the worst casescenario with tumor microvessel function like the normalBBB and a highly water-soluble drug (methotrexate)

about 1 of the drug to which the tumor is exposed willenter the tumor With a lipid-soluble drug like aziridinyl-benzoquinone and normal BBB function the local expo-sure fraction increases to 07 in other words increasinglipid solubility does increase delivery to brain tissue Whentumor microvessel permeability increases by 01 ml gndash1

minndash1 the fraction of methotrexate to which the tumor isexposed increases to 18 A local increase in tumor per-meability results in a potentially signicant increase indrug delivery for a water-soluble drug However note thatalthough the fractional removal across normal BBB is highfor the lipid-soluble drugs aziridinylbenzoquinone and13-bis(2-chloroethyl)-1-nitrosourea the local exposurefraction does not increase signicantly when permeabilityis increased (the increase is only 1ndash2 percentage pointsTable 1) Increased drug delivery as a result of increases incapillary permeability will be more important for water-soluble drugs than for lipid-soluble drugs for which per-meability was not as great an issue in the rst place

It is important to remember that the rest of the bodyrepresents a large sink into which most of the drug is dis-tributed The fraction of drug entering tumor comparedwith that circulating through the entire body is minisculein every case lt1 of circulating drug will reach thetumor regardless of the drugrsquos permeability and regardlessof changes in capillary permeability (Table 1) Thereforeany method that is used to increase brain tumor perme-ability will still have to deal with the reality that most ofthe administered drug is distributed to the rest of the bodyand so far as treatment of the brain tumor is concernedwasted In most instances the limit on the total dose ofdrug that can be given is imposed by the tolerance of nor-mal tissues which is systemic toxicity in the case of drugsgiven systemically Because the maximum dose that can begiven to a patient is determined by factors external totreatment of the tumor (that is systemic toxicity) we mustaccept as a starting principle that the administered drugdose cannot be increased and we must search for alterna-tive methods to increase the fraction of drug that reachesthe tumor

Neuro-Oncology n JANUARY 200048

DR Groothuis Review of BBB and BTB

Table 1 Fractional delivery of four different chemotherapeutic agents across the BBB and BTB

Permeability of normal brain Permeability increase = 01 ml gndash1 minndash1

Drug Local exposure fraction Total exposure fraction Local exposure fraction Total exposure fraction

MTX 0017a 000006a 018 000006

5-FU 011 000004 025 000008

AZQ 070 00002 072b 00002b

BCNU 054 00002 055b 00002b

Abbreviations MTX methotrexate 5-FU 5-uorouracil AZQ azridinylbenzoquinone BCNU 13-bis(2-chloroethyl)-1-nitrosourea

Four drugs with different rates of brain entry brain efux and plasma half-lives are shown For each drug the values represent the fraction of drug extracted by the tumor The localexposure fraction represents the fraction of drug removed by 1 g of tumor from the plasma to which it was exposed (Equation 5) The total exposure fraction represents the fraction of

drug removed by 1 g of tumor from the entire circulating volume of drug in plasma (Equation 6) aThe ldquoworst caserdquo scenario occurs when water-soluble drugs are crossing the normal BBB bThe ldquobest caserdquo scenario occurs when lipid-soluble drugs cross capillaries in brain tumors with increased permeability However the two columns labeled total exposure fractionemphasize that the amount of drug removed by 1 g of tumor is very small compared with the total drug circulating in the body and indicates the magnitude of the sink effect of the

rest of the body in brain tumor chemotherapy

Increasing Permeability of the BBB and BTBto Drugs Given Intravascularly

Table 2 summarizes many of the methods that are cur-rently being used to manipulate the permeability of theBBB and BTB as well as my own interpretation of theiradvantages and disadvantages In light of the previousdiscussion all of these methods share a major disadvan-tage the bulk of administered drug will be lost to the restof the body in a sink effect and the maximum adminis-tered dose will almost always be determined by systemictoxicity

Chemical Modication of Existing Drugs

Chemical modication of existing drugs may refer to sev-eral different approaches First an existing drug may bemodified to make it fit a receptor in the BBB Thisapproach has been used with melphalan in which nitro-gen mustard (mechlorethamine) was linked to phenylala-nine (Groothuis et al 1992) Chemical modication mayrefer to linking an active protein (for example growthfactor) or a peptide to an antibody against a BBB recep-tor (Pardridge 1995) More commonly chemical modi-cation refers to the process of making an existing drugmore lipid soluble with the intent of increasing BBB per-meability This approach has been used extensively withantiretroviral nucleoside analogs for treating AIDS buthas not been used much with brain tumor chemothera-peutic drugs The use of increasingly accurate means topredict physicochemical properties offers encouragementthat chemical modication will nd wider use in modify-ing brain tumor drugs (van de Waterbeemd et al 1998)

Prodrug Therapy

This refers to a special class of chemical modication inwhich a drug is chemically modied to increase its capil-

lary permeability Once in the brain the prodrug under-goes an enzymatic reaction that returns the drug to itsactive state and with reduced BBB permeability (Shermanet al 1991 Yoshikawa et al 1999) These approacheswhich have not yet been used for brain tumor chemother-apeutic drugs offer an advantage by increasing K1 inEquation 3 while k2 and k3 remain unchanged Thiswould increase the AUCB while AUCPL remainsunchanged Although this increases the fraction of drugentering the tissue it does not change total body expo-sure and unfortunately introduces the need to explorethe pharmacokinetics and pharmacodynamics of eachnew compound

Intra-Arterial Administration

This method of administration refers to the ia injectionof a therapeutic agent without the concomitant use of abarrier-modifying approach The principle advantage ofthis method is that during the course of injecting a druginto an artery the tissue perfused by that artery receives ahigher plasma concentration during the first passagethrough the circulation (AUCPL is increased) In a seriesof papers Fenstermacher and coworkers theoreticallyevaluated the efficacy of intra-arterial delivery to thebrain and brain tumors (Cowles and Fenstermacher1974 Eckman et al 1974 Fenstermacher and Cowles1977 Fenstermacher and Gazendam 1981) They evalu-ated the contributions of several variables and concludedthat there were significant delivery advantages to iaadministration in a setting where the rate of tumor bloodow is very low or where the rate of systemic transfor-mation or excretion is very high or in a unique situationwhere having crossed the BBB or BTB the drug binds tothe tissue Once the drug passes through the tumormicrovasculature it enters the systemic circulation andthe pharmacokinetics are the same as for an iv adminis-tration Thus the ideal drug for ia administration is one

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 49

Table 2 Comparison of different methods of increasing delivery of intravascularly administered compounds to brain tumors

Method of increasing delivery Principle advantages Principle disadvantages Limiting factor(s)

Chemical modication of Requirements are now Each new compound must be BBB BTB permeabilityexisting drugs understood for different evaluated individually systemic toxicity

delivery routes

Prodrug Utilizes inherent biochemical Difference between BBB and BTB BBB BTB permeabilitypathways in brain and tumor is not known systemic toxicity

Intra-arterial First pass increase in AUC Invasive small increase in AUCBT BBB BTB permeability systemic toxicity

Hyperosmotic disruption First pass increase in AUC and Invasive differential effect on BBB BTB permeability increased permeability brain and tumor short-lived systemic toxicity

Chemical modication First pass increase in AUC and Invasive no effect on brain BBB BTB permeabilityincreased permeability short-lived

Receptor-mediated transport Increased permeability Low capacity system BBB BTB permeability

Inhibiting drug efux Decreased efux may be used None known None knownwith existing drugs

Avant-garde methods Not yet known Not yet known Not yet known

Abbreviations BBB blood-brain barrier BTB blood-tumor barrier AUC area under the curve

This table lists the different approaches that are available to increase the amount of drug in a brain tumor in cases where the drug is given either by iv or ia injection The advantages

and disadvantages are an expression of the authorrsquos opinion about the most prominent feature of each method Details about each method may be found in the text

that rapidly crosses the BBB or BTB and is either boundto tissue elements or locally metabolized in the process ofexerting its anticancer effect There are practical prob-lems associated with ia administration however includ-ing the need for the tumor to reside within the arterialdistribution being infused and hemodynamic streamingof the administered drug Perhaps the most obvious evi-dence that increased tissue drug concentrations can beachieved with ia administration is the increased inci-dence of local toxicity (Arafat et al 1999) Neverthelessbecause most drugs being used to treat brain tumors donot have the ideal qualities to take advantage of ia deliv-ery the results of clinical trials of ia chemotherapy inbrain tumors show minimal if any improvement in sur-vival (Dropcho et al 1998 Gundersen et al 1998Hirano et al 1998)

Hyperosmolar Blood-Brain Barrier Disruption(HBBBD)

This method involves the infusion of a hyperosmolarsolution (usually 14 M mannitol) into a cerebral arterygenerally followed by the intra-arterial administration ofa drug The principle features of this method areincreased capillary permeability (caused by the hyperos-molar solution) followed by a rst pass advantage fromia infusion that is K1 is transiently increased as isAUCPL This method was originally proposed byRapoport (Rapoport and Thompson 1973) and has beenused extensively by Neuwelt and colleagues (Kroll andNeuwelt 1998 Neuwelt and Dahlborg 1989 Neuweltet al 1980 Neuwelt et al 1983) Controversy about theuse of this method arose because of differential suscepti-bility of the normal BBB as compared with the abnormalBTB Whereas all studies demonstrated increased perme-ability of the normal BBB some studies in animal tumorsfailed to demonstrate any increase in tumor permeability(Nakagawa et al 1984) others showed transient perme-ability increases (Groothuis et al 1990) and somereported signicant permeability increases (Neuwelt etal 1984) Most recently Zunkeler et al used PET tostudy the time course of BBB function in 13 patients withmalignant astrocytomas after HBBBD (Zunkeler et al1996) They conrmed the differential effect of HBBBDpermeability was increased 1000 in brain and 60 intumors The half-time of the osmotic effect was 81 minin brain and 42 min in tumor They modeled the effectsof HBBBD on methotrexate and concluded that concen-trations above 1 microM the minimal concentration requiredfor an effect from methotrexate would not be enhancedin tumor and would be enhanced only 10 in brainThus HBBBD in combination with ia administrationwould be most useful for drugs that once having crossedthe BBB bind to brain or tumor tissue and do not efuxback across the BBB In addition the differential effect ofHBBBD on normal brain compared with that of tumormust be considered when evaluating neurotoxicity

Chemical Modication of the BBB and BTB

The latest entry into the therapeutic arena is the use ofchemical agents to disrupt the BBB (Black 1995) The

agents are mostly derivatives of normal vasoactive com-pounds and include bradykinin (Inamura and Black1994) interleukin-2 (Gutman et al 1996) leukotrieneC4 (Black and Chio 1992) and others (de Vries et al1996) The principle feature of these drugs is that theyare given ia and followed by an ia injection of a thera-peutic agent RMP-7 is a commercially developedbradykinin analog that has reached clinical trials (Ford etal 1998) and acts by increasing permeability at the inter-cellular junction (Sanovick et al 1995) Studies withPET in 9 malignant gliomas have shown that permeabil-ity to 68Ga-EDTA was increased 46 plusmn 42 in tumorswithout a signicant increase in tumor-free brain (Blacket al 1997) Another report suggests that RMP-7 canincrease delivery across the normal BBB (Emerich et al1998) RMP-7 is generally administered over a 15-minperiod the timing of administration of the chemothera-peutic drug relative to the RMP-7 has varied eg as abolus after RMP-7 or alternating chemotherapeutic drugadministration with RMP-7 BBB function is restoredrapidly after the RMP-7 infusion ends continuing theinfusion for over 20 min results in tachyphylaxis withloss of responsiveness (Bartus 1999) As with ia andHBBBD administration the advantage with chemicalmodication of the BBB is limited to a fractional increasein the circulating drug to which the tumor is exposedand still represents a vanishingly small fraction of thetotal body drug exposure Like those previous tech-niques chemical modication of the BBB will be ideal fora drug with dened properties as discussed before How-ever this ideal drug has yet to be developed

Receptor-Mediated Transport

This is a very broad area because any receptor-mediatedtransport system in the BBB can be selected as a targetThe considerations about drug delivery in a facilitatedtransport system are different from those involving sim-ple diffusion First a facilitated transport system can becharacterized by the Michaelis-Menten constants Km(the concentration at which the reaction velocity is halfmaximal) and Vmax (the limiting velocity as the concen-tration approaches innity) An important factor in thefacilitated transport of drugs is the plasma concentrationof the native substrate for the receptor For examplephenylalanine crosses the BBB via the large neutral aminoacid transport (LNAA) system melphalan is achemotherapeutic drug with a phenylalanine backboneand could therefore be transported by the LNAA systemHowever the normal plasma levels of phenylalaninecompete with melphalan to such a degree that the rate ofmelphalan transport into experimental gliomas is notincreased (Groothuis et al 1992) Lowering plasmaphenylalanine levels increased the rate of melphalanentry into brain tumors but only by a modest 8 Mostof the available transport receptors are low afnity lowcapacity systems such as those for the nucleosides vari-ous peptides transferrin and insulin (Ermisch et al1993 Pardridge et al 1995 Friden et al 1996 Tamai etal 1997) Although these transport systems have beenused to increase brain delivery of oligonucleotides(Boado et al 1998 Normand-Sdiqui and Akhtar 1998)

Neuro-Oncology n JANUARY 200050

DR Groothuis Review of BBB and BTB

nerve growth factor (Friden et al 1993) and vasoactiveintestinal peptide (Bickel et al 1993) none of thesetransport systems has been explored to any extent inbrain tumors

Inhibiting Drug Efux

Throughout most of the history of the BBB movementacross the capillary for most compounds without a trans-port system has been assumed to occur by simple diffu-sion the process is passive and the rate that a compoundcrosses the barrier is the same in both directions so thatthe process is symmetrical Recent studies have begun tochallenge this idea In particular P-glycoprotein repre-sents an ATP-dependent efflux pathway that confersresistance to cancer cells by allowing them to move avariety of chemotherapeutic drugs out of the cell againsta concentration gradient (Schinkel 1999 Tsuji andTamai 1997) P-glycoprotein appears to be localized pri-marily to the luminal capillary membrane and the num-ber of drugs that are transported by this system may bequite large (Schinkel 1999) Immunohistochemically P-glycoprotein has been demonstrated in malignantglioma tumor cells (Leweke et al 1998 von Bossanyi etal 1997) Leweke et al commented that tumor endothe-lial cells stained in 20 of 21 cases and von Bossanyi et alstated that tumor blood vessels were positive in 60 oftumors PET has been used to demonstrate the reversal ofP-glycoprotein pump function in mdr1a knock-out micewith cyclosporin A (Hendrikse et al 1998) Since manychemotherapeutic drugs are transported by P-glycopro-tein this represents a logical and exciting target in braintumor chemotherapy especially because inhibition of P-glycoprotein function could occur at both the BTB andin tumor cells Inhibiting P-glycoprotein would increaseboth extracellular and intracellular drug concentration(by decreasing k2 in Equation 3) without increasing theamount of drug administered This is an avenue of modi-fying brain tumor therapy that must be explored

Avant-Garde Methods of Delivery

Included in this category are the newest methods forintravascular delivery including the use of biodegradablenanoparticles (Schroeder et al 1998 Song et al 1997)liposomes (Allen and Moase 1996 Sakamoto and Ido1993) monoglyceride-based systems (Chang et al1994) and magnetic microspheres (Chang and Bodmeier1997) Many of these novel approaches are evaluatingdifferent methods of giving drug-laden molecules thatwill preferentially accumulate in a tissue usually com-bined with selective methods of perfusion such as iadelivery For example Pulfer and Gallo delivered 1ndash2 micromia magnetic aminodextran microspheres to RG-2 ratbrain tumors and used a magnetic eld to retain the par-ticles within the tumor They showed that this methodresulted in signicant accumulation of the particles as aresult of surface charge and magnetic eld (Chang andBodmeier 1997) These methods are too new to beapplied to human brain tumors but are representative ofthe diverse and novel directions that investigators areusing to increase brain tumor delivery

Summary of Intravascular Drug Delivery

A tremendous amount of progress has been made in thepast few years in our understanding of drug delivery tobrain tumors This progress has not yet translated intoimpressive increases in survival of patients with braintumors There are major areas that remain to be exploredand developed in the arena of intravascularly adminis-tered drugs For example because ia administration(with or without HBBBD or chemical modification)offers a unique rst pass advantage a search for drugsthat have high lipid solubility and bind to brain tumortargets is likely to be protable Second increasing tissuedrug concentration by the use of efux inhibitors is obvi-ous there will be no increase in total body exposurewhile potentially achieving signicant increases in tumordrug concentration However the ultimate limiting factorfor all intravascularly administered drugs is that of thepotential toxicity from total body exposure It seemsunlikely that this important and unfortunately negativevariable can be overcome

Injection or Infusion of Drugs Directly intothe Brain or Its Cavities

Table 3 summarizes the different approaches that arebeing used to administer drugs directly into the brainthus bypassing the BBB All of these routes and methodsof administration drastically change the pharmacokineticlandscape Instead of concern about crossing the formi-dable BBB or BTB the drugs are administered directlyinto the CSF or brain extracellular space Instead of need-ing to measure the inux rate it is necessary to under-stand drug movement within the brain and to understandefux mechanisms In marked contrast to intravascularadministration 100 of the administered dose can bedelivered using these methods of administration Theproblem then becomes one of understanding the forcesthat control the movement of the drug within brain andtumor tissue One or more of four forces will affect drugmovement depending upon the particular mode of drugadministration bulk flow of CSF bulk flow of braininterstitial uid bulk ow due to the infusion of a solu-tion into the brain or CSF and diffusion Drug movementwill also be variably affected by the amount of whitematter in the target area as a result of the change fromthe highly tortuous environment of a gray matter struc-ture (isotropic) to the highly oriented ber pathways ofwhite matter (anisotropic)

Intrathecal and Intraventricular Drug Administration

In this review intrathecal drug administration refers tothe administration of drug into the subarachnoid spaceusually into the lumbar subarachnoid space whereasintraventricular administration refers to injection or infu-sion into the lateral ventricle They are discussed togetherbecause of the large number of features they have in com-mon The principle feature of drug administration bythese routes is dominated by the bulk uid ow of the

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 51

CSF The major source of CSF is from the choroidplexuses of the lateral and 4th ventricles as much asanother 30 may originate from the brain extracellularuid The direction of the ow is from the lateral ventri-cles into the 3rd and then 4th ventricles out through theforamina of Luschke and Magendie into the basal cis-terns around the convexities and out of the subarach-noid space through the arachnoid granulations In ratswhen 14C-sucrose was injected into the lateral ventriclethe sucrose distributed rapidly within 5 min it was foundin the basal cisterns and by 1 h the sucrose had largely leftthe subarachnoid space (Ghersi-Egea et al 1996) Whenthe injected agent is metabolically active it may enterselected cells (Rubertone et al 1993) In humans the nor-mal volume of CSF produced is between 400 and 500 mlper day Since the CSF spaces within the brain are about150 ml in volume there is a considerable bulk ow withinthis system although the velocity varies tremendouslyfrom site to site When isotopes are injected into the lum-bar subarachnoid space they travel up the spinal sub-arachnoid space and are found in the basal cisterns by 3 hBy 24 h they are found over the cerebral convexities CSFow from the lateral ventricles is directional isotopesinjected into the lumbar CSF do not normally enter thelateral ventricles As a result of these bulk ow pathwaysmaterials injected into the CSF will attain initial concen-trations that are directly proportional to the concentra-tion in the infusate This route of administration is idealfor situations in which the target is within the subarach-noid space (for example carcinomatous meningitis) or inwhich the target is close to the CSF brain interface Carci-nomatous meningitis has long been treated with intrathe-cal infusions including the more recent use of monoclonalantibodies (Brown et al 1996) However the capacity ofdrugs to enter the brain extracellular space from the CSFis limited Numerous studies have shown that entry intothe brain is diffusional and that concentrations declineexponentially from the brain surface (Blasberg 1977Blasberg et al 1977 Groothuis and Levy 1997 Patlak

and Fenstermacher 1975) This route of administration isboth impractical and inefcient for intraparenchymalbrain tumors

Intratumoral and Intracavitary Injection

These are discussed together and briey because they areboth limiting cases of convection-enhanced deliverywhich is discussed below These both represent localforms of drug administration in which the distributionwill be determined initially by the rate and duration ofthe injection and secondly by tissue bulk ow pathwaysand diffusion The history of intratumoral injection wasdiscussed by Tomita (1991) who recognized the poten-tial usefulness of this approach as well as the importanceof neurotoxicity as the limiting factor Intratumoral andintracavitary injections are being widely explored asmeans of delivering large agents such as viral vectorsgrowth factors and cells (Bigner et al 1998 Farkkila etal 1994 Hsiao et al 1997 Puri et al 1996 Ram et al1997 Yang et al 1997) All of the attributes discussed inthe section on CED will also apply to these forms ofdrug delivery

Microdialysis

Microdialysis is a method that employs the passive diffu-sion of a drug across a semipermeable membrane (deLange et al 1997 de Lange et al 1999 Parsons andJustice 1994) It can be used to both sample and deliverdrugs to the surrounding tissue and has been used tosample extracellular drug concentrations in experimentalbrain tumors (Devineni et al 1996 Nakashima et al1997) The principle features of microdialysis when usedas a drug delivery methodology are that the tissue con-centrations are a function of the drug concentration inthe dialysate (which places control of the concentrationin hands of the user) and that distribution of drug awayfrom the dialysis catheter occurs by diffusion (Dykstra et

Neuro-Oncology n JANUARY 200052

DR Groothuis Review of BBB and BTB

Table 3 Comparison of different methods for local drug delivery to brain tumors

Method of circumventing the BBB Principle advantages Principle disadvantages Limiting factor(s)

Intrathecal Easy access ideal for therapy of Not useful for parenchymal Bulk ow rate of CSFmeningeal disease disease

Intraventricular administration Easy access ideal for therapy of Not useful for parenchymal Bulk ow rate of CSFmeningeal disease disease

Intratumoral injection User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Intracavitary injection User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Microdialysis User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Biodegradable polymers User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Convection-enhanced delivery User control over administered Invasive Unpredictable distribution dose 100 reaches target neurotoxicity

Abbreviations BBB blood-brain barrier CSF cerebrospinal uid

This table lists the different approaches that are available to deliver therapeutic drugs and agents by local administration The advantages and disadvantages are an expression of theauthorrsquos opinion about the most prominent feature of each method Details about each method may be found in the text

al 1992) However the normal BBB is disrupted imme-diately around the catheter and stays disrupted for sometime after catheter insertion making drug efux in thisvicinity an additional problem when trying to understandlocal delivery kinetics (Groothuis et al 1998 Morgan etal 1996 Westergren et al 1995) Because the principlefactors in local tissue concentrations are diffusion andorlocal brain bulk flow patterns the volume of brainreached by microdialysis will be relatively small anddrug concentrations will in general decline exponen-tially away from the dialysis cannula This method willbe applicable when the volume of tissue that needs to bereached is small The effect of normal brain bulk owpathways on this delivery method remains to be studied

Biodegradable Polymers

This represents a local delivery system in whichbiodegradable polymers are loaded with drug andimplanted into brain or tumor tissue resulting in timed-release of drug (Cao and Shoichet 1999 Menei et al1997) The principle features of this system are that mul-tiple polymer pellets or wafers can be implanted whichgives control over the spatial distribution and that drugdistribution away from the source occurs by diffusionwith a variable contribution from the inherent bulk owpathways of the brain (Fung et al 1998) This approachhas been used in patients with malignant gliomas whereits safety and efcacy have been demonstrated (Brem etal 1995 Olivi and Brem 1994) As with microdialysisthe principle limiting feature of this method of drug deliv-ery is that drug distribution occurs mainly by diffusionin which drug concentrations fall exponentially awayfrom the source and are generally limited to a few mil-limeters of tissue penetration (Fung et al 1998)

Convection-Enhanced Delivery

This is a relatively new method of drug delivery to braintumors The principle feature of this delivery method isthat a drug solution is infused directly into the brain Theconcentration of the drug is independent of the infusionrate and duration and the distribution of the drug is afunction of the hydrostatic pressure of the infusion aswell as diffusion into surrounding tissue From a deliveryperspective the only limiting factor to the drug concen-tration that can be delivered is the solubility of the drugCED has been discussed from a theoretical perspective(Morrison et al 1994) it has been explored in severalanimal models (Bobo et al 1994 Groothuis et al 1999Laske et al 1997a Lieberman et al 1995 Viola et al1995) and it has been used for delivery of a targetedtoxin in human gliomas (Laske et al 1997b) The distri-bution of the infused drug is highly dependent uponwhether the drug is infused into a homogeneous graymatter structure or a structure containing ber pathwaysIn the case of the infusion of a drug-containing solutioninto a homogeneous brain structure the infusion initiallyproduces a spherical volume in which the concentrationof the drug is directly proportional to that of the infusate(Fig 2b) After the infusion reaches equilibrium at whichpoint the amount of drug leaving across brain capillaries

is equal to the amount being infused the drug begins todiffuse away from the edge of the sphere at concentra-tions that decline exponentially in accordance with themathematics of diffusion (Fig 2b) If the infusionencounters an organized white matter pathway the rateof movement is either increased or decreased (dependingupon the direction of the inherent bulk ow within thatpathway) (Fig 2c) Morrison stated that when the infu-sion is being made into a homogeneous isotropicmedium the radius of the central spherical componentmdashthe convective componentmdashcan be related to the infusionrate and the efux constant of the drug being infused

rp 5 3 2qv (4p K) (Eq 7)

where qv is the infusion rate and K represents thesteady-state efux rate constant Fig 3 shows a plot of theradius of the convective component as a function of infu-sion rate and efux constant From reviewing Fig 3

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 53

Fig 2 The distribution of 14C-sucrose in rat brain by convection-enhanced delivery The diagram in the upper left (A) shows the targetlocation (center of the caudate nucleus) as well as the organization ofthe brain at the target level After a 1-h 05-microlmin infusion the iso-tope is found in a spherical shape with even distribution across (B) By8 h the isotope is moving into the external capsule (C) A similarappearance is found after a constant infusion over 7 days the centralconvective component is clearly visible surrounded by decreasingconcentration from the edge (D) Note the isotopersquos rapid lateralmovement down the external capsule (probably representing a nor-mal brain bulk uid pathway) but not into the corpus callosum Infu-sion into rat brain for 7 days produced no neuropathologic changesother than those associated with catheter insertion

which extends from infusion rates provided by osmoticmini-pumps (1 microlh) to high infusion rates (10 microlmin) it is

apparent that the volume of affected brain remains quitesmall for most infusion rates Similarly as the efux con-stant decreases indicating slower rates of crossing theBBB the convection volume increases Fig 3 illustratesthat CED is best suited for delivery of large compounds(with low efux rates) and delivery at moderately highinfusion rates if the intent is to reach large brain volumes

The shape of the volume of distribution at steady statewill be highly dependent on the location in the brain thatis being infused Fig 4 shows a CT scan of a dog brainreceiving an infusion of iodinated contrast into whitematter The iodinated contrast rapidly distributes in thewhite matter of the ipsilateral hemisphere with rapidlydecreasing concentrations in the cortex on the affectedside Note that in both Figs 2 and 4 the infused materialdoes not cross the midline through the corpus callosumThis suggests that the direction of the normal bulk owpathway within the corpus callosum is away from themidline thus opposing the CED infusion component Incontrast to intravascularly administered drugs whereincreased capillary permeability results in increased tissueconcentrations (Table 1) the opposite effect will occur inCED infusions Increased efux across permeable tumorcapillaries will reduce both the extracellular drug concen-tration and the volume of distribution In Fig 5 highconcentrations of 14C-sucrose were achieved by a CEDinfusion into an RG-2 glioma but at a distance of 24mm from the infusion site tissue concentrationsdecreased by several logarithmic units Although studieshave not been done in humans it is easy to anticipatethat the volume of distribution from a CED infusion willfollow similar principles Fig 6 shows several levels of anMRI scan through a malignant glioma Each set of scansshows a gadolinium-enhanced tumor margin (which canbe considered naturersquos equivalent to a CED infusion) anda T2-weighted image (which corresponds to the extracel-lular water originating from the tumor source) The

Neuro-Oncology n JANUARY 200054

DR Groothuis Review of BBB and BTB

Fig 3 Relationship between radius of convective CED componentinfusion rate and efux rate The radius of the inection pointbetween the convective and diffusional components of CED infusionsis shown for infusion into a homogeneous brain structure (Equation7) The values on the vertical axis represent the radius inside whichthe local concentrations remain uniform and outside of which drugconcentrations decline exponentially Changes for each 5-cm incre-ment are represented by different shades of gray The infusion ratewas chosen from 1 acute 10ndash6 to 1 acute 10ndash2 ml per minndash1 The lower valuesare those seen with infusions from osmotic mini-pumps while thehigher values represent the upper limit of CED infusion rates in dogsThe efux rate was chosen to vary from 1 acute 10ndash1 to 1 acute 10ndash7 minndash1The volume of the convective component increases as a function ofboth increasing infusion rate and decreasing efux rate

Fig 4 Illustration of CED infusion in a dog The gure on the left shows the CT scan at the start of a 3 microl minndash1 infusion of Isovue (iopamidol)into the hemispheric white matter of a dog while that on the right shows the same coronal level on the same dog after 10 days of infusion Thescan on the left contains an air bubble in the lateral ventricle the cannula tip contains a metal marker (the high density circle) and a small amountof Isovue surrounding the catheter tip In the scan on the right the Isovue has distributed at a high concentration throughout the white matterof the ipsilateral hemisphere with a rapid decline in concentration at gray-white interfaces Note that the distribution of Isovue is limited to theipsilateral hemisphere No neuropathologic changes were observed as a result of the infusion

shape and distribution of the edema are highly irregularand very dependent upon the brain structures encoun-tered Note that the edema uid moves into the internalcapsule but not into the thalamus or cortex Notice alsothat the edema uid does not cross the posterior limb ofthe corpus callosum although it comes into contact withit at several levels

A dominant feature of CED is that distribution of thedrug is determined by the infusion parameters and thetype of tissue being infused whereas the concentration ofdrug in the infusate is unrelated to these parameters Themaximum concentration of the drug in the infusate willbe determined by neurotoxicity within the central con-vective component of the infusion with variable concen-trations in the anisotropic brain surrounding the centralcomponent and concentrations 50 to 100 times lower inthe diffusional component furthermost from the infusioncatheter The ability to manipulate drug concentration inbrain and tumor tissue with such ease and to maintainthese manipulations for long periods of time (Groothuiset al 1999) brings a need for the development ofentirely new concepts For example the limits withregard to the composition of the infusate need to beexplored such as pH osmolarity and ionic compositionFurthermore how does one reliably study neurotoxicityAre infusions into animal models going to be reliable pre-dictors of toxicity in humans Will differential neurotox-icity become an issue It is possible that neuronsastrocytes and oligodendrogliocytes will display differ-ent levels of susceptibility to many drugs Will we seedelayed neurotoxicity such as that seen with radiation Itwill probably be best for many of the variables associatedwith neurotoxicity to be explored in animals before sub-jecting humans to CED infusions

Summary of Drug Delivery that Circumvents the BBB

There has been an explosion of methodology involvinglocal tissue drug delivery during the past 10 years Someof these techniques such as CED fulll the promise tocircumvent the BBB entirely In fullling this promisethese methods introduce new problems such as the rela-tionship between the spatial distribution of drugs andneurotoxicity that need to be understood before thesetechniques are widely used However for viral vectorsmonoclonal antibodies antisense oligonucleotides andother therapeutic agents that will be unable to cross theBBB in therapeutically effective amounts CED may pro-vide the necessary delivery tool

The State of Drug Delivery to Brain Tumorsndash1999

If we return to the statement of Vick et al at the begin-ning of this review we can now state with certainty thatthe BBB and BTB are major factors limiting the access ofmany therapeutic agents to brain tumors and these bar-riers will become even more signicant with the develop-ment of new molecular biological therapies that willinvolve large molecules viruses and even cells It is alsoremarkable how prescient those authors were in identify-ing the other parts of the drug delivery process that mustnow be understood and how little progress we havemade in elucidating ldquotumor cell uptake metabolic fatewithin tumor cells and the washout or sink effect of theextracellular space and CSFrdquo (Vick et al 1977) Formost of the past 20 years we have focused so much on

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 55

Fig 5 Infusion of 14C-sucrose into an RG-2 rat glioma An infusion cannula was placed into the caudate nucleus of a Fisher-344 rat (targetshown in Fig 2A) and RG-2 cells were injected Ten days later 14C-sucrose was infused at a rate of 10 microlh for 7 days Autoradiographic imageswere made (right images) and the sections were then stained with hematoxylin-eosin (left images) The top two images show that the 14C-sucrose distributes in high concentration throughout the RG-2 tumor extending into the edematous external capsule (E) However thelower two sections taken from 24 mm behind the infusion site still contain tumor and show that the concentration of 14C-sucrose is 100000times lower These sections illustrate the problems associated with controlling drug delivery by convection-enhanceddelivery

ways to increase delivery across the BBB that we havefailed to address these other issues which now becomethe forefront in drug delivery methods such as CED Thearsenal from which to choose a drug delivery method istruly impressive We must now study the other factorsthat limit effective brain tumor therapy Perhaps the mostpressing of these includes the need to develop effective

laboratory tools for identifying the relationship betweenthe concentration-time product of therapeutic agents andtumor cell kill rather than relying on empirical testing inhuman subjects It is most likely that the new century willsee the development and application of tools for individ-ualizing chemotherapy for brain tumors and other solidhuman tumors

Ali-Osman F (1999) Human glioma cultures and in vitro analysis of therapeu-

tic response In Berger MS and Wilson CB (Eds)The Gliomas

Philadelphia WB Saunders Co pp 134ndash142

Allen TM and Moase EH (1996) Therapeutic opportunities for targeted

liposomal drug delivery Adv Drug Deliv Rev 21 117ndash133

Arafat T Hentschel P Madajewicz S Manzione J Chowhan N Davis

R Roche P Af I Roque C Meek A and Magdy S (1999) Toxicities

related to intraarterial infusion of cisplatin and etoposide in patients with

brain tumors J Neurooncol 42 73ndash77

Bartus RT (1999) The blood-brain barrier as a target for phamacological

modulation Curr Opin Drug Discov Dev 2 152ndash167

Bickel U Yoshikawa T Landaw EM Faull KF and Pardridge WM

(1993) Pharmacologic effects in vivo in brain by vector-mediated peptide

delivery Proc Natl Acad Sci USA 90 2618ndash2622

Bigner DD Brown MT Friedman AH Coleman RE Akabani G Fried-

man HS Thorstad WL McLendon RE Bigner SH Zhao XG

Pegram CN Wikstrand CJ Herndon JE II Vick NA Paleologos N

Cokgor I Provenzale JM and Zalutsky MR (1998) Iodine-131-labeled

antitenascin monoclonal antibody 81C6 treatment of patients with recur-

rent malignant gliomas Phase I trial results J Clin Oncol 16 2202ndash2212

Black KL (1995) Biochemical opening of the blood-brain barrier Adv Drug

Deliv Rev 15 37ndash52

Neuro-Oncology n JANUARY 200056

DR Groothuis Review of BBB and BTB

Fig 6 Distribution of vasogenic edema in a malignant glioma There are four pairs of scans extending through a tumor in the left temporal lobeextending from the lowest (A) to the highest level (D) Each pair of scans contains a gadolinium-enhanced scan (left) and a T2-weighted image(right) The gadolinium-enhanced scans show the area of abnormally permeable BBB The T2-weighted images show the distribution of edema(water) emanating from the tumor The tumor may be considered a natural equivalent of a convection-enhanced delivery infusion The resultingedema spreads extensively through white matter in the ipsilateral hemisphere reaching forward in the internal capsule (levels C and D) but notpenetrating gray matter structures such as the thalamus (levels C and D) or cortex (levels AndashD) Note that the edema contacts the posterior limbof the corpus callosum in levels C and D but does not extend into the corpus callosum suggesting that there is a physiological barrier to the bulkmovement of uid in that direction

References

molecules To identify this tumor population permeabil-ity studies with two different-sized markers are requiredWith MRI Ostrowitzki et al beautifully illustrated dif-ferential permeability of a 9L rat glioma to gadopentetate(molecular mass = 05 kDa) and albumin-(Gd-DTPA)30(molecular mass = 92 kDa) (Ostrowitzki et al 1998)Because studies in humans are generally not performedwith two different-sized permeability markers the preva-lence of this tumor population in humans is not knownThe third capillary population represented in Fig 1 con-tains interendothelial gaps that may measure as large as 1 microm the RG-2 rat glioma (Nakagawa et al 1987Schlageter et al 1999) and D-54 MG human gliomamodel (Blasberg et al 1987) are examples As shown inFig 1 these tumor models do not exhibit permselectivityfor large molecules However Fig 1 also points out thatthe permeability-surface area product of the gliomas is 2ndash3 log units less than that of the liver which is a conse-quence of a smaller number of interendothelial gaps inthe tumors than in the liver

The second major variable with regard to capillarypermeability involves the spatial distribution of the targetcapillaries Although brain tumor capillaries may haveincreased permeability like those of the RG-2 and D-54MG tumors permeability in brain surrounding tumorrapidly returns to normal brain values within a few mil-limeters of the tumor margin If as shown by Burger(1987) individual tumor cells may reside centimetersaway from the edge of a tumor spatial variability in cap-illary function will affect drug delivery to all braintumors

To understand the effects of changes in capillary per-meability on drug delivery it may be useful to introducesome general concepts Once injected intravascularly adrug mixes with the total body volume of blood or in thecase of water-soluble compounds that are not protein-bound with total body plasma Because a 70-kg humancontains 3 liters of plasma to achieve a starting plasmaconcentration of 1 unit of ldquoidealrdquo drug per ml 3000units of drug must be given As will be shown thisunpleasant fact will color all attempts to increase drugdelivery associated with intravascular administrationThe human body acts as an enormous sink in which themajority of intravascularly administered drug will be dis-tributed not to the brain tumor but to other body tis-sues Once mixed with total body plasma the drugdistributes throughout body tissues and is then elimi-nated The time course of the mixing distribution andelimination of the drug in plasma is generally describedby several different half-times which represent theprocess of tting the plasma concentration-time data to amultiexponential expression of the form

Cp(t) 5 Ae2 a t 1 Be2 b t 1 Ce 2 g t (Eq 1)

where A B and C represent the y-intercepts and a b and g represent the time constants with units of minndash1The time constants are related to the half-times by theexpression

t a12 5

ln2(Eq 2)

a

where the half-time has units of minutes The inte-grated value of plasma concentration over time usuallyreferred to as the area under the curve (AUCPL) repre-sents the amount of drug passing through the brain ortumor vessels that is the amount of drug to which thebrain or tumor vessels are exposed

The time course of drug concentration over time inbrain or brain tumor tissue is more complex As we havediscussed elsewhere (Blasberg and Groothuis 1986) theconcentration of ideal drug in tissue (Ci) (assuming pas-sive transport across the capillary no plasma or tissuebinding and first order metabolism and inactivationkinetics) is given by

Ci(T) 5 K1 e 0

TCp(t)e

2 (k2 1 k3)(T 2 t)dt (Eq 3)

where K1 is the blood-to-tissue transfer constant (withunits of ml gndash1 minndash1) k2 is the tissue efux constant (withunits of reciprocal time min-1) and k3 is a metabolism orinactivation constant also with units of reciprocal timeEquation 3 can be used to calculate the tissue drug con-centration over time and can be used to explore theimpact of different values of K1 k2 and k3 on tissue drugconcentrations Assuming that the drug is passively dis-tributed across the BBB and not removed by other meansthen K1 and k2 are related by the expression l = K1k2where l is the equilibrium distribution volume of thedrug in the tissue

It is very useful to have some expression of the ef-ciency of the drug delivery process if for no other reasonthan to compare one experimental methodology forincreasing drug delivery with another Pardridge has usedan expression called the pharmacokinetic rule to expressthe efciency of drug entry (Pardridge 1997)

IDg 5 PS 3 AUC (Eq 4)

in which ID represents the percent of injected dose ofdrug delivered per gram of tissue PS is the permeabilitysurface area product (which for most water-soluble com-pounds is equal to K1 with units of ml gndash1 minndash1) andAUC is the plasma area under the curve (for whichPardridge uses the units IDminmicrol) However thisexpression does not consider drug efflux from tissueandor metabolism In this review I will use two expres-sions to indicate the fractional efficiency of the drugdelivery process Both expressions contain a term in thenumerator that refers to the concentration-time productof drug in brain or tumor tissue (AUCB) The rst expres-sion called the local exposure fraction represents thefraction of drug removed from the blood to which thebrain or tumor is exposed that is the blood circulatinglocally within the tissue

concentration 2 time product of drugAUCB

5per g of brain or tumor tissue

(Eq 5)AUCPL concentration 2 time product of drugper ml of perfusing plasma

Equation 5 expresses the fraction of drug removedfrom the plasma to which the tissue was exposed and istherefore an expression of local delivery efciency How-

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 47

ever this is but a small fraction of the drug in the entirebody The second expression called the total exposurefraction incorporates the total body plasma volume

concentration 2time product of drug per g

AUCB5

of brain or tumor tissue(Eq 6)3000 3 AUCPL concentration 2

time product of drugper ml of perfusing plasma

The AUCPL can be obtained from Equation 1 andAUCB can be obtained from integrating Equation 3Equation 6 reminds us that the fraction of drug enteringa gram of brain or tumor is but one small fraction of drugcirculating in the entire body and that this route of deliv-ery is inherently inefcient

Equations 5 and 6 can be used to illustrate drug deliv-ery to normal brain and brain tumor in what may beviewed as ldquobest caserdquo and ldquoworst caserdquo scenarios Theworst case scenario will almost always be represented bywater-soluble drugs and the normal BBB that is the mostrestrictive situation The best case scenario will be repre-sented by lipid-soluble drugs and highly permeabletumors such as those represented in Fig 1 by the RG-2and D-54 MG gliomas In Table 1 the local and totalexposure fractions are presented for four drugs with differ-ent permeabilities methotrexate (K1 = 00014) 5-uo-rouracil (K1 = 00096) aziridinylbenzoquinone (K1 =0145) and 13-bis(2-chloroethyl)-1-nitrosourea (K1 = 06ml gndash1 minndash1) and for two permeability scenarios that ofnormal brain and that where permeability has increased01 ml gndash1 minndash1 over normal brain The data for the calcu-lations were contained in a previous publication (Blasbergand Groothuis 1986) the reader should be aware that thistype of modeling is dependent on assumptions made aboutthe different parameters Nonetheless Table 1 can be usedto illustrate several different points First in the worst casescenario with tumor microvessel function like the normalBBB and a highly water-soluble drug (methotrexate)

about 1 of the drug to which the tumor is exposed willenter the tumor With a lipid-soluble drug like aziridinyl-benzoquinone and normal BBB function the local expo-sure fraction increases to 07 in other words increasinglipid solubility does increase delivery to brain tissue Whentumor microvessel permeability increases by 01 ml gndash1

minndash1 the fraction of methotrexate to which the tumor isexposed increases to 18 A local increase in tumor per-meability results in a potentially signicant increase indrug delivery for a water-soluble drug However note thatalthough the fractional removal across normal BBB is highfor the lipid-soluble drugs aziridinylbenzoquinone and13-bis(2-chloroethyl)-1-nitrosourea the local exposurefraction does not increase signicantly when permeabilityis increased (the increase is only 1ndash2 percentage pointsTable 1) Increased drug delivery as a result of increases incapillary permeability will be more important for water-soluble drugs than for lipid-soluble drugs for which per-meability was not as great an issue in the rst place

It is important to remember that the rest of the bodyrepresents a large sink into which most of the drug is dis-tributed The fraction of drug entering tumor comparedwith that circulating through the entire body is minisculein every case lt1 of circulating drug will reach thetumor regardless of the drugrsquos permeability and regardlessof changes in capillary permeability (Table 1) Thereforeany method that is used to increase brain tumor perme-ability will still have to deal with the reality that most ofthe administered drug is distributed to the rest of the bodyand so far as treatment of the brain tumor is concernedwasted In most instances the limit on the total dose ofdrug that can be given is imposed by the tolerance of nor-mal tissues which is systemic toxicity in the case of drugsgiven systemically Because the maximum dose that can begiven to a patient is determined by factors external totreatment of the tumor (that is systemic toxicity) we mustaccept as a starting principle that the administered drugdose cannot be increased and we must search for alterna-tive methods to increase the fraction of drug that reachesthe tumor

Neuro-Oncology n JANUARY 200048

DR Groothuis Review of BBB and BTB

Table 1 Fractional delivery of four different chemotherapeutic agents across the BBB and BTB

Permeability of normal brain Permeability increase = 01 ml gndash1 minndash1

Drug Local exposure fraction Total exposure fraction Local exposure fraction Total exposure fraction

MTX 0017a 000006a 018 000006

5-FU 011 000004 025 000008

AZQ 070 00002 072b 00002b

BCNU 054 00002 055b 00002b

Abbreviations MTX methotrexate 5-FU 5-uorouracil AZQ azridinylbenzoquinone BCNU 13-bis(2-chloroethyl)-1-nitrosourea

Four drugs with different rates of brain entry brain efux and plasma half-lives are shown For each drug the values represent the fraction of drug extracted by the tumor The localexposure fraction represents the fraction of drug removed by 1 g of tumor from the plasma to which it was exposed (Equation 5) The total exposure fraction represents the fraction of

drug removed by 1 g of tumor from the entire circulating volume of drug in plasma (Equation 6) aThe ldquoworst caserdquo scenario occurs when water-soluble drugs are crossing the normal BBB bThe ldquobest caserdquo scenario occurs when lipid-soluble drugs cross capillaries in brain tumors with increased permeability However the two columns labeled total exposure fractionemphasize that the amount of drug removed by 1 g of tumor is very small compared with the total drug circulating in the body and indicates the magnitude of the sink effect of the

rest of the body in brain tumor chemotherapy

Increasing Permeability of the BBB and BTBto Drugs Given Intravascularly

Table 2 summarizes many of the methods that are cur-rently being used to manipulate the permeability of theBBB and BTB as well as my own interpretation of theiradvantages and disadvantages In light of the previousdiscussion all of these methods share a major disadvan-tage the bulk of administered drug will be lost to the restof the body in a sink effect and the maximum adminis-tered dose will almost always be determined by systemictoxicity

Chemical Modication of Existing Drugs

Chemical modication of existing drugs may refer to sev-eral different approaches First an existing drug may bemodified to make it fit a receptor in the BBB Thisapproach has been used with melphalan in which nitro-gen mustard (mechlorethamine) was linked to phenylala-nine (Groothuis et al 1992) Chemical modication mayrefer to linking an active protein (for example growthfactor) or a peptide to an antibody against a BBB recep-tor (Pardridge 1995) More commonly chemical modi-cation refers to the process of making an existing drugmore lipid soluble with the intent of increasing BBB per-meability This approach has been used extensively withantiretroviral nucleoside analogs for treating AIDS buthas not been used much with brain tumor chemothera-peutic drugs The use of increasingly accurate means topredict physicochemical properties offers encouragementthat chemical modication will nd wider use in modify-ing brain tumor drugs (van de Waterbeemd et al 1998)

Prodrug Therapy

This refers to a special class of chemical modication inwhich a drug is chemically modied to increase its capil-

lary permeability Once in the brain the prodrug under-goes an enzymatic reaction that returns the drug to itsactive state and with reduced BBB permeability (Shermanet al 1991 Yoshikawa et al 1999) These approacheswhich have not yet been used for brain tumor chemother-apeutic drugs offer an advantage by increasing K1 inEquation 3 while k2 and k3 remain unchanged Thiswould increase the AUCB while AUCPL remainsunchanged Although this increases the fraction of drugentering the tissue it does not change total body expo-sure and unfortunately introduces the need to explorethe pharmacokinetics and pharmacodynamics of eachnew compound

Intra-Arterial Administration

This method of administration refers to the ia injectionof a therapeutic agent without the concomitant use of abarrier-modifying approach The principle advantage ofthis method is that during the course of injecting a druginto an artery the tissue perfused by that artery receives ahigher plasma concentration during the first passagethrough the circulation (AUCPL is increased) In a seriesof papers Fenstermacher and coworkers theoreticallyevaluated the efficacy of intra-arterial delivery to thebrain and brain tumors (Cowles and Fenstermacher1974 Eckman et al 1974 Fenstermacher and Cowles1977 Fenstermacher and Gazendam 1981) They evalu-ated the contributions of several variables and concludedthat there were significant delivery advantages to iaadministration in a setting where the rate of tumor bloodow is very low or where the rate of systemic transfor-mation or excretion is very high or in a unique situationwhere having crossed the BBB or BTB the drug binds tothe tissue Once the drug passes through the tumormicrovasculature it enters the systemic circulation andthe pharmacokinetics are the same as for an iv adminis-tration Thus the ideal drug for ia administration is one

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 49

Table 2 Comparison of different methods of increasing delivery of intravascularly administered compounds to brain tumors

Method of increasing delivery Principle advantages Principle disadvantages Limiting factor(s)

Chemical modication of Requirements are now Each new compound must be BBB BTB permeabilityexisting drugs understood for different evaluated individually systemic toxicity

delivery routes

Prodrug Utilizes inherent biochemical Difference between BBB and BTB BBB BTB permeabilitypathways in brain and tumor is not known systemic toxicity

Intra-arterial First pass increase in AUC Invasive small increase in AUCBT BBB BTB permeability systemic toxicity

Hyperosmotic disruption First pass increase in AUC and Invasive differential effect on BBB BTB permeability increased permeability brain and tumor short-lived systemic toxicity

Chemical modication First pass increase in AUC and Invasive no effect on brain BBB BTB permeabilityincreased permeability short-lived

Receptor-mediated transport Increased permeability Low capacity system BBB BTB permeability

Inhibiting drug efux Decreased efux may be used None known None knownwith existing drugs

Avant-garde methods Not yet known Not yet known Not yet known

Abbreviations BBB blood-brain barrier BTB blood-tumor barrier AUC area under the curve

This table lists the different approaches that are available to increase the amount of drug in a brain tumor in cases where the drug is given either by iv or ia injection The advantages

and disadvantages are an expression of the authorrsquos opinion about the most prominent feature of each method Details about each method may be found in the text

that rapidly crosses the BBB or BTB and is either boundto tissue elements or locally metabolized in the process ofexerting its anticancer effect There are practical prob-lems associated with ia administration however includ-ing the need for the tumor to reside within the arterialdistribution being infused and hemodynamic streamingof the administered drug Perhaps the most obvious evi-dence that increased tissue drug concentrations can beachieved with ia administration is the increased inci-dence of local toxicity (Arafat et al 1999) Neverthelessbecause most drugs being used to treat brain tumors donot have the ideal qualities to take advantage of ia deliv-ery the results of clinical trials of ia chemotherapy inbrain tumors show minimal if any improvement in sur-vival (Dropcho et al 1998 Gundersen et al 1998Hirano et al 1998)

Hyperosmolar Blood-Brain Barrier Disruption(HBBBD)

This method involves the infusion of a hyperosmolarsolution (usually 14 M mannitol) into a cerebral arterygenerally followed by the intra-arterial administration ofa drug The principle features of this method areincreased capillary permeability (caused by the hyperos-molar solution) followed by a rst pass advantage fromia infusion that is K1 is transiently increased as isAUCPL This method was originally proposed byRapoport (Rapoport and Thompson 1973) and has beenused extensively by Neuwelt and colleagues (Kroll andNeuwelt 1998 Neuwelt and Dahlborg 1989 Neuweltet al 1980 Neuwelt et al 1983) Controversy about theuse of this method arose because of differential suscepti-bility of the normal BBB as compared with the abnormalBTB Whereas all studies demonstrated increased perme-ability of the normal BBB some studies in animal tumorsfailed to demonstrate any increase in tumor permeability(Nakagawa et al 1984) others showed transient perme-ability increases (Groothuis et al 1990) and somereported signicant permeability increases (Neuwelt etal 1984) Most recently Zunkeler et al used PET tostudy the time course of BBB function in 13 patients withmalignant astrocytomas after HBBBD (Zunkeler et al1996) They conrmed the differential effect of HBBBDpermeability was increased 1000 in brain and 60 intumors The half-time of the osmotic effect was 81 minin brain and 42 min in tumor They modeled the effectsof HBBBD on methotrexate and concluded that concen-trations above 1 microM the minimal concentration requiredfor an effect from methotrexate would not be enhancedin tumor and would be enhanced only 10 in brainThus HBBBD in combination with ia administrationwould be most useful for drugs that once having crossedthe BBB bind to brain or tumor tissue and do not efuxback across the BBB In addition the differential effect ofHBBBD on normal brain compared with that of tumormust be considered when evaluating neurotoxicity

Chemical Modication of the BBB and BTB

The latest entry into the therapeutic arena is the use ofchemical agents to disrupt the BBB (Black 1995) The

agents are mostly derivatives of normal vasoactive com-pounds and include bradykinin (Inamura and Black1994) interleukin-2 (Gutman et al 1996) leukotrieneC4 (Black and Chio 1992) and others (de Vries et al1996) The principle feature of these drugs is that theyare given ia and followed by an ia injection of a thera-peutic agent RMP-7 is a commercially developedbradykinin analog that has reached clinical trials (Ford etal 1998) and acts by increasing permeability at the inter-cellular junction (Sanovick et al 1995) Studies withPET in 9 malignant gliomas have shown that permeabil-ity to 68Ga-EDTA was increased 46 plusmn 42 in tumorswithout a signicant increase in tumor-free brain (Blacket al 1997) Another report suggests that RMP-7 canincrease delivery across the normal BBB (Emerich et al1998) RMP-7 is generally administered over a 15-minperiod the timing of administration of the chemothera-peutic drug relative to the RMP-7 has varied eg as abolus after RMP-7 or alternating chemotherapeutic drugadministration with RMP-7 BBB function is restoredrapidly after the RMP-7 infusion ends continuing theinfusion for over 20 min results in tachyphylaxis withloss of responsiveness (Bartus 1999) As with ia andHBBBD administration the advantage with chemicalmodication of the BBB is limited to a fractional increasein the circulating drug to which the tumor is exposedand still represents a vanishingly small fraction of thetotal body drug exposure Like those previous tech-niques chemical modication of the BBB will be ideal fora drug with dened properties as discussed before How-ever this ideal drug has yet to be developed

Receptor-Mediated Transport

This is a very broad area because any receptor-mediatedtransport system in the BBB can be selected as a targetThe considerations about drug delivery in a facilitatedtransport system are different from those involving sim-ple diffusion First a facilitated transport system can becharacterized by the Michaelis-Menten constants Km(the concentration at which the reaction velocity is halfmaximal) and Vmax (the limiting velocity as the concen-tration approaches innity) An important factor in thefacilitated transport of drugs is the plasma concentrationof the native substrate for the receptor For examplephenylalanine crosses the BBB via the large neutral aminoacid transport (LNAA) system melphalan is achemotherapeutic drug with a phenylalanine backboneand could therefore be transported by the LNAA systemHowever the normal plasma levels of phenylalaninecompete with melphalan to such a degree that the rate ofmelphalan transport into experimental gliomas is notincreased (Groothuis et al 1992) Lowering plasmaphenylalanine levels increased the rate of melphalanentry into brain tumors but only by a modest 8 Mostof the available transport receptors are low afnity lowcapacity systems such as those for the nucleosides vari-ous peptides transferrin and insulin (Ermisch et al1993 Pardridge et al 1995 Friden et al 1996 Tamai etal 1997) Although these transport systems have beenused to increase brain delivery of oligonucleotides(Boado et al 1998 Normand-Sdiqui and Akhtar 1998)

Neuro-Oncology n JANUARY 200050

DR Groothuis Review of BBB and BTB

nerve growth factor (Friden et al 1993) and vasoactiveintestinal peptide (Bickel et al 1993) none of thesetransport systems has been explored to any extent inbrain tumors

Inhibiting Drug Efux

Throughout most of the history of the BBB movementacross the capillary for most compounds without a trans-port system has been assumed to occur by simple diffu-sion the process is passive and the rate that a compoundcrosses the barrier is the same in both directions so thatthe process is symmetrical Recent studies have begun tochallenge this idea In particular P-glycoprotein repre-sents an ATP-dependent efflux pathway that confersresistance to cancer cells by allowing them to move avariety of chemotherapeutic drugs out of the cell againsta concentration gradient (Schinkel 1999 Tsuji andTamai 1997) P-glycoprotein appears to be localized pri-marily to the luminal capillary membrane and the num-ber of drugs that are transported by this system may bequite large (Schinkel 1999) Immunohistochemically P-glycoprotein has been demonstrated in malignantglioma tumor cells (Leweke et al 1998 von Bossanyi etal 1997) Leweke et al commented that tumor endothe-lial cells stained in 20 of 21 cases and von Bossanyi et alstated that tumor blood vessels were positive in 60 oftumors PET has been used to demonstrate the reversal ofP-glycoprotein pump function in mdr1a knock-out micewith cyclosporin A (Hendrikse et al 1998) Since manychemotherapeutic drugs are transported by P-glycopro-tein this represents a logical and exciting target in braintumor chemotherapy especially because inhibition of P-glycoprotein function could occur at both the BTB andin tumor cells Inhibiting P-glycoprotein would increaseboth extracellular and intracellular drug concentration(by decreasing k2 in Equation 3) without increasing theamount of drug administered This is an avenue of modi-fying brain tumor therapy that must be explored

Avant-Garde Methods of Delivery

Included in this category are the newest methods forintravascular delivery including the use of biodegradablenanoparticles (Schroeder et al 1998 Song et al 1997)liposomes (Allen and Moase 1996 Sakamoto and Ido1993) monoglyceride-based systems (Chang et al1994) and magnetic microspheres (Chang and Bodmeier1997) Many of these novel approaches are evaluatingdifferent methods of giving drug-laden molecules thatwill preferentially accumulate in a tissue usually com-bined with selective methods of perfusion such as iadelivery For example Pulfer and Gallo delivered 1ndash2 micromia magnetic aminodextran microspheres to RG-2 ratbrain tumors and used a magnetic eld to retain the par-ticles within the tumor They showed that this methodresulted in signicant accumulation of the particles as aresult of surface charge and magnetic eld (Chang andBodmeier 1997) These methods are too new to beapplied to human brain tumors but are representative ofthe diverse and novel directions that investigators areusing to increase brain tumor delivery

Summary of Intravascular Drug Delivery

A tremendous amount of progress has been made in thepast few years in our understanding of drug delivery tobrain tumors This progress has not yet translated intoimpressive increases in survival of patients with braintumors There are major areas that remain to be exploredand developed in the arena of intravascularly adminis-tered drugs For example because ia administration(with or without HBBBD or chemical modification)offers a unique rst pass advantage a search for drugsthat have high lipid solubility and bind to brain tumortargets is likely to be protable Second increasing tissuedrug concentration by the use of efux inhibitors is obvi-ous there will be no increase in total body exposurewhile potentially achieving signicant increases in tumordrug concentration However the ultimate limiting factorfor all intravascularly administered drugs is that of thepotential toxicity from total body exposure It seemsunlikely that this important and unfortunately negativevariable can be overcome

Injection or Infusion of Drugs Directly intothe Brain or Its Cavities

Table 3 summarizes the different approaches that arebeing used to administer drugs directly into the brainthus bypassing the BBB All of these routes and methodsof administration drastically change the pharmacokineticlandscape Instead of concern about crossing the formi-dable BBB or BTB the drugs are administered directlyinto the CSF or brain extracellular space Instead of need-ing to measure the inux rate it is necessary to under-stand drug movement within the brain and to understandefux mechanisms In marked contrast to intravascularadministration 100 of the administered dose can bedelivered using these methods of administration Theproblem then becomes one of understanding the forcesthat control the movement of the drug within brain andtumor tissue One or more of four forces will affect drugmovement depending upon the particular mode of drugadministration bulk flow of CSF bulk flow of braininterstitial uid bulk ow due to the infusion of a solu-tion into the brain or CSF and diffusion Drug movementwill also be variably affected by the amount of whitematter in the target area as a result of the change fromthe highly tortuous environment of a gray matter struc-ture (isotropic) to the highly oriented ber pathways ofwhite matter (anisotropic)

Intrathecal and Intraventricular Drug Administration

In this review intrathecal drug administration refers tothe administration of drug into the subarachnoid spaceusually into the lumbar subarachnoid space whereasintraventricular administration refers to injection or infu-sion into the lateral ventricle They are discussed togetherbecause of the large number of features they have in com-mon The principle feature of drug administration bythese routes is dominated by the bulk uid ow of the

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 51

CSF The major source of CSF is from the choroidplexuses of the lateral and 4th ventricles as much asanother 30 may originate from the brain extracellularuid The direction of the ow is from the lateral ventri-cles into the 3rd and then 4th ventricles out through theforamina of Luschke and Magendie into the basal cis-terns around the convexities and out of the subarach-noid space through the arachnoid granulations In ratswhen 14C-sucrose was injected into the lateral ventriclethe sucrose distributed rapidly within 5 min it was foundin the basal cisterns and by 1 h the sucrose had largely leftthe subarachnoid space (Ghersi-Egea et al 1996) Whenthe injected agent is metabolically active it may enterselected cells (Rubertone et al 1993) In humans the nor-mal volume of CSF produced is between 400 and 500 mlper day Since the CSF spaces within the brain are about150 ml in volume there is a considerable bulk ow withinthis system although the velocity varies tremendouslyfrom site to site When isotopes are injected into the lum-bar subarachnoid space they travel up the spinal sub-arachnoid space and are found in the basal cisterns by 3 hBy 24 h they are found over the cerebral convexities CSFow from the lateral ventricles is directional isotopesinjected into the lumbar CSF do not normally enter thelateral ventricles As a result of these bulk ow pathwaysmaterials injected into the CSF will attain initial concen-trations that are directly proportional to the concentra-tion in the infusate This route of administration is idealfor situations in which the target is within the subarach-noid space (for example carcinomatous meningitis) or inwhich the target is close to the CSF brain interface Carci-nomatous meningitis has long been treated with intrathe-cal infusions including the more recent use of monoclonalantibodies (Brown et al 1996) However the capacity ofdrugs to enter the brain extracellular space from the CSFis limited Numerous studies have shown that entry intothe brain is diffusional and that concentrations declineexponentially from the brain surface (Blasberg 1977Blasberg et al 1977 Groothuis and Levy 1997 Patlak

and Fenstermacher 1975) This route of administration isboth impractical and inefcient for intraparenchymalbrain tumors

Intratumoral and Intracavitary Injection

These are discussed together and briey because they areboth limiting cases of convection-enhanced deliverywhich is discussed below These both represent localforms of drug administration in which the distributionwill be determined initially by the rate and duration ofthe injection and secondly by tissue bulk ow pathwaysand diffusion The history of intratumoral injection wasdiscussed by Tomita (1991) who recognized the poten-tial usefulness of this approach as well as the importanceof neurotoxicity as the limiting factor Intratumoral andintracavitary injections are being widely explored asmeans of delivering large agents such as viral vectorsgrowth factors and cells (Bigner et al 1998 Farkkila etal 1994 Hsiao et al 1997 Puri et al 1996 Ram et al1997 Yang et al 1997) All of the attributes discussed inthe section on CED will also apply to these forms ofdrug delivery

Microdialysis

Microdialysis is a method that employs the passive diffu-sion of a drug across a semipermeable membrane (deLange et al 1997 de Lange et al 1999 Parsons andJustice 1994) It can be used to both sample and deliverdrugs to the surrounding tissue and has been used tosample extracellular drug concentrations in experimentalbrain tumors (Devineni et al 1996 Nakashima et al1997) The principle features of microdialysis when usedas a drug delivery methodology are that the tissue con-centrations are a function of the drug concentration inthe dialysate (which places control of the concentrationin hands of the user) and that distribution of drug awayfrom the dialysis catheter occurs by diffusion (Dykstra et

Neuro-Oncology n JANUARY 200052

DR Groothuis Review of BBB and BTB

Table 3 Comparison of different methods for local drug delivery to brain tumors

Method of circumventing the BBB Principle advantages Principle disadvantages Limiting factor(s)

Intrathecal Easy access ideal for therapy of Not useful for parenchymal Bulk ow rate of CSFmeningeal disease disease

Intraventricular administration Easy access ideal for therapy of Not useful for parenchymal Bulk ow rate of CSFmeningeal disease disease

Intratumoral injection User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Intracavitary injection User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Microdialysis User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Biodegradable polymers User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Convection-enhanced delivery User control over administered Invasive Unpredictable distribution dose 100 reaches target neurotoxicity

Abbreviations BBB blood-brain barrier CSF cerebrospinal uid

This table lists the different approaches that are available to deliver therapeutic drugs and agents by local administration The advantages and disadvantages are an expression of theauthorrsquos opinion about the most prominent feature of each method Details about each method may be found in the text

al 1992) However the normal BBB is disrupted imme-diately around the catheter and stays disrupted for sometime after catheter insertion making drug efux in thisvicinity an additional problem when trying to understandlocal delivery kinetics (Groothuis et al 1998 Morgan etal 1996 Westergren et al 1995) Because the principlefactors in local tissue concentrations are diffusion andorlocal brain bulk flow patterns the volume of brainreached by microdialysis will be relatively small anddrug concentrations will in general decline exponen-tially away from the dialysis cannula This method willbe applicable when the volume of tissue that needs to bereached is small The effect of normal brain bulk owpathways on this delivery method remains to be studied

Biodegradable Polymers

This represents a local delivery system in whichbiodegradable polymers are loaded with drug andimplanted into brain or tumor tissue resulting in timed-release of drug (Cao and Shoichet 1999 Menei et al1997) The principle features of this system are that mul-tiple polymer pellets or wafers can be implanted whichgives control over the spatial distribution and that drugdistribution away from the source occurs by diffusionwith a variable contribution from the inherent bulk owpathways of the brain (Fung et al 1998) This approachhas been used in patients with malignant gliomas whereits safety and efcacy have been demonstrated (Brem etal 1995 Olivi and Brem 1994) As with microdialysisthe principle limiting feature of this method of drug deliv-ery is that drug distribution occurs mainly by diffusionin which drug concentrations fall exponentially awayfrom the source and are generally limited to a few mil-limeters of tissue penetration (Fung et al 1998)

Convection-Enhanced Delivery

This is a relatively new method of drug delivery to braintumors The principle feature of this delivery method isthat a drug solution is infused directly into the brain Theconcentration of the drug is independent of the infusionrate and duration and the distribution of the drug is afunction of the hydrostatic pressure of the infusion aswell as diffusion into surrounding tissue From a deliveryperspective the only limiting factor to the drug concen-tration that can be delivered is the solubility of the drugCED has been discussed from a theoretical perspective(Morrison et al 1994) it has been explored in severalanimal models (Bobo et al 1994 Groothuis et al 1999Laske et al 1997a Lieberman et al 1995 Viola et al1995) and it has been used for delivery of a targetedtoxin in human gliomas (Laske et al 1997b) The distri-bution of the infused drug is highly dependent uponwhether the drug is infused into a homogeneous graymatter structure or a structure containing ber pathwaysIn the case of the infusion of a drug-containing solutioninto a homogeneous brain structure the infusion initiallyproduces a spherical volume in which the concentrationof the drug is directly proportional to that of the infusate(Fig 2b) After the infusion reaches equilibrium at whichpoint the amount of drug leaving across brain capillaries

is equal to the amount being infused the drug begins todiffuse away from the edge of the sphere at concentra-tions that decline exponentially in accordance with themathematics of diffusion (Fig 2b) If the infusionencounters an organized white matter pathway the rateof movement is either increased or decreased (dependingupon the direction of the inherent bulk ow within thatpathway) (Fig 2c) Morrison stated that when the infu-sion is being made into a homogeneous isotropicmedium the radius of the central spherical componentmdashthe convective componentmdashcan be related to the infusionrate and the efux constant of the drug being infused

rp 5 3 2qv (4p K) (Eq 7)

where qv is the infusion rate and K represents thesteady-state efux rate constant Fig 3 shows a plot of theradius of the convective component as a function of infu-sion rate and efux constant From reviewing Fig 3

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 53

Fig 2 The distribution of 14C-sucrose in rat brain by convection-enhanced delivery The diagram in the upper left (A) shows the targetlocation (center of the caudate nucleus) as well as the organization ofthe brain at the target level After a 1-h 05-microlmin infusion the iso-tope is found in a spherical shape with even distribution across (B) By8 h the isotope is moving into the external capsule (C) A similarappearance is found after a constant infusion over 7 days the centralconvective component is clearly visible surrounded by decreasingconcentration from the edge (D) Note the isotopersquos rapid lateralmovement down the external capsule (probably representing a nor-mal brain bulk uid pathway) but not into the corpus callosum Infu-sion into rat brain for 7 days produced no neuropathologic changesother than those associated with catheter insertion

which extends from infusion rates provided by osmoticmini-pumps (1 microlh) to high infusion rates (10 microlmin) it is

apparent that the volume of affected brain remains quitesmall for most infusion rates Similarly as the efux con-stant decreases indicating slower rates of crossing theBBB the convection volume increases Fig 3 illustratesthat CED is best suited for delivery of large compounds(with low efux rates) and delivery at moderately highinfusion rates if the intent is to reach large brain volumes

The shape of the volume of distribution at steady statewill be highly dependent on the location in the brain thatis being infused Fig 4 shows a CT scan of a dog brainreceiving an infusion of iodinated contrast into whitematter The iodinated contrast rapidly distributes in thewhite matter of the ipsilateral hemisphere with rapidlydecreasing concentrations in the cortex on the affectedside Note that in both Figs 2 and 4 the infused materialdoes not cross the midline through the corpus callosumThis suggests that the direction of the normal bulk owpathway within the corpus callosum is away from themidline thus opposing the CED infusion component Incontrast to intravascularly administered drugs whereincreased capillary permeability results in increased tissueconcentrations (Table 1) the opposite effect will occur inCED infusions Increased efux across permeable tumorcapillaries will reduce both the extracellular drug concen-tration and the volume of distribution In Fig 5 highconcentrations of 14C-sucrose were achieved by a CEDinfusion into an RG-2 glioma but at a distance of 24mm from the infusion site tissue concentrationsdecreased by several logarithmic units Although studieshave not been done in humans it is easy to anticipatethat the volume of distribution from a CED infusion willfollow similar principles Fig 6 shows several levels of anMRI scan through a malignant glioma Each set of scansshows a gadolinium-enhanced tumor margin (which canbe considered naturersquos equivalent to a CED infusion) anda T2-weighted image (which corresponds to the extracel-lular water originating from the tumor source) The

Neuro-Oncology n JANUARY 200054

DR Groothuis Review of BBB and BTB

Fig 3 Relationship between radius of convective CED componentinfusion rate and efux rate The radius of the inection pointbetween the convective and diffusional components of CED infusionsis shown for infusion into a homogeneous brain structure (Equation7) The values on the vertical axis represent the radius inside whichthe local concentrations remain uniform and outside of which drugconcentrations decline exponentially Changes for each 5-cm incre-ment are represented by different shades of gray The infusion ratewas chosen from 1 acute 10ndash6 to 1 acute 10ndash2 ml per minndash1 The lower valuesare those seen with infusions from osmotic mini-pumps while thehigher values represent the upper limit of CED infusion rates in dogsThe efux rate was chosen to vary from 1 acute 10ndash1 to 1 acute 10ndash7 minndash1The volume of the convective component increases as a function ofboth increasing infusion rate and decreasing efux rate

Fig 4 Illustration of CED infusion in a dog The gure on the left shows the CT scan at the start of a 3 microl minndash1 infusion of Isovue (iopamidol)into the hemispheric white matter of a dog while that on the right shows the same coronal level on the same dog after 10 days of infusion Thescan on the left contains an air bubble in the lateral ventricle the cannula tip contains a metal marker (the high density circle) and a small amountof Isovue surrounding the catheter tip In the scan on the right the Isovue has distributed at a high concentration throughout the white matterof the ipsilateral hemisphere with a rapid decline in concentration at gray-white interfaces Note that the distribution of Isovue is limited to theipsilateral hemisphere No neuropathologic changes were observed as a result of the infusion

shape and distribution of the edema are highly irregularand very dependent upon the brain structures encoun-tered Note that the edema uid moves into the internalcapsule but not into the thalamus or cortex Notice alsothat the edema uid does not cross the posterior limb ofthe corpus callosum although it comes into contact withit at several levels

A dominant feature of CED is that distribution of thedrug is determined by the infusion parameters and thetype of tissue being infused whereas the concentration ofdrug in the infusate is unrelated to these parameters Themaximum concentration of the drug in the infusate willbe determined by neurotoxicity within the central con-vective component of the infusion with variable concen-trations in the anisotropic brain surrounding the centralcomponent and concentrations 50 to 100 times lower inthe diffusional component furthermost from the infusioncatheter The ability to manipulate drug concentration inbrain and tumor tissue with such ease and to maintainthese manipulations for long periods of time (Groothuiset al 1999) brings a need for the development ofentirely new concepts For example the limits withregard to the composition of the infusate need to beexplored such as pH osmolarity and ionic compositionFurthermore how does one reliably study neurotoxicityAre infusions into animal models going to be reliable pre-dictors of toxicity in humans Will differential neurotox-icity become an issue It is possible that neuronsastrocytes and oligodendrogliocytes will display differ-ent levels of susceptibility to many drugs Will we seedelayed neurotoxicity such as that seen with radiation Itwill probably be best for many of the variables associatedwith neurotoxicity to be explored in animals before sub-jecting humans to CED infusions

Summary of Drug Delivery that Circumvents the BBB

There has been an explosion of methodology involvinglocal tissue drug delivery during the past 10 years Someof these techniques such as CED fulll the promise tocircumvent the BBB entirely In fullling this promisethese methods introduce new problems such as the rela-tionship between the spatial distribution of drugs andneurotoxicity that need to be understood before thesetechniques are widely used However for viral vectorsmonoclonal antibodies antisense oligonucleotides andother therapeutic agents that will be unable to cross theBBB in therapeutically effective amounts CED may pro-vide the necessary delivery tool

The State of Drug Delivery to Brain Tumorsndash1999

If we return to the statement of Vick et al at the begin-ning of this review we can now state with certainty thatthe BBB and BTB are major factors limiting the access ofmany therapeutic agents to brain tumors and these bar-riers will become even more signicant with the develop-ment of new molecular biological therapies that willinvolve large molecules viruses and even cells It is alsoremarkable how prescient those authors were in identify-ing the other parts of the drug delivery process that mustnow be understood and how little progress we havemade in elucidating ldquotumor cell uptake metabolic fatewithin tumor cells and the washout or sink effect of theextracellular space and CSFrdquo (Vick et al 1977) Formost of the past 20 years we have focused so much on

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 55

Fig 5 Infusion of 14C-sucrose into an RG-2 rat glioma An infusion cannula was placed into the caudate nucleus of a Fisher-344 rat (targetshown in Fig 2A) and RG-2 cells were injected Ten days later 14C-sucrose was infused at a rate of 10 microlh for 7 days Autoradiographic imageswere made (right images) and the sections were then stained with hematoxylin-eosin (left images) The top two images show that the 14C-sucrose distributes in high concentration throughout the RG-2 tumor extending into the edematous external capsule (E) However thelower two sections taken from 24 mm behind the infusion site still contain tumor and show that the concentration of 14C-sucrose is 100000times lower These sections illustrate the problems associated with controlling drug delivery by convection-enhanceddelivery

ways to increase delivery across the BBB that we havefailed to address these other issues which now becomethe forefront in drug delivery methods such as CED Thearsenal from which to choose a drug delivery method istruly impressive We must now study the other factorsthat limit effective brain tumor therapy Perhaps the mostpressing of these includes the need to develop effective

laboratory tools for identifying the relationship betweenthe concentration-time product of therapeutic agents andtumor cell kill rather than relying on empirical testing inhuman subjects It is most likely that the new century willsee the development and application of tools for individ-ualizing chemotherapy for brain tumors and other solidhuman tumors

Ali-Osman F (1999) Human glioma cultures and in vitro analysis of therapeu-

tic response In Berger MS and Wilson CB (Eds)The Gliomas

Philadelphia WB Saunders Co pp 134ndash142

Allen TM and Moase EH (1996) Therapeutic opportunities for targeted

liposomal drug delivery Adv Drug Deliv Rev 21 117ndash133

Arafat T Hentschel P Madajewicz S Manzione J Chowhan N Davis

R Roche P Af I Roque C Meek A and Magdy S (1999) Toxicities

related to intraarterial infusion of cisplatin and etoposide in patients with

brain tumors J Neurooncol 42 73ndash77

Bartus RT (1999) The blood-brain barrier as a target for phamacological

modulation Curr Opin Drug Discov Dev 2 152ndash167

Bickel U Yoshikawa T Landaw EM Faull KF and Pardridge WM

(1993) Pharmacologic effects in vivo in brain by vector-mediated peptide

delivery Proc Natl Acad Sci USA 90 2618ndash2622

Bigner DD Brown MT Friedman AH Coleman RE Akabani G Fried-

man HS Thorstad WL McLendon RE Bigner SH Zhao XG

Pegram CN Wikstrand CJ Herndon JE II Vick NA Paleologos N

Cokgor I Provenzale JM and Zalutsky MR (1998) Iodine-131-labeled

antitenascin monoclonal antibody 81C6 treatment of patients with recur-

rent malignant gliomas Phase I trial results J Clin Oncol 16 2202ndash2212

Black KL (1995) Biochemical opening of the blood-brain barrier Adv Drug

Deliv Rev 15 37ndash52

Neuro-Oncology n JANUARY 200056

DR Groothuis Review of BBB and BTB

Fig 6 Distribution of vasogenic edema in a malignant glioma There are four pairs of scans extending through a tumor in the left temporal lobeextending from the lowest (A) to the highest level (D) Each pair of scans contains a gadolinium-enhanced scan (left) and a T2-weighted image(right) The gadolinium-enhanced scans show the area of abnormally permeable BBB The T2-weighted images show the distribution of edema(water) emanating from the tumor The tumor may be considered a natural equivalent of a convection-enhanced delivery infusion The resultingedema spreads extensively through white matter in the ipsilateral hemisphere reaching forward in the internal capsule (levels C and D) but notpenetrating gray matter structures such as the thalamus (levels C and D) or cortex (levels AndashD) Note that the edema contacts the posterior limbof the corpus callosum in levels C and D but does not extend into the corpus callosum suggesting that there is a physiological barrier to the bulkmovement of uid in that direction

References

ever this is but a small fraction of the drug in the entirebody The second expression called the total exposurefraction incorporates the total body plasma volume

concentration 2time product of drug per g

AUCB5

of brain or tumor tissue(Eq 6)3000 3 AUCPL concentration 2

time product of drugper ml of perfusing plasma

The AUCPL can be obtained from Equation 1 andAUCB can be obtained from integrating Equation 3Equation 6 reminds us that the fraction of drug enteringa gram of brain or tumor is but one small fraction of drugcirculating in the entire body and that this route of deliv-ery is inherently inefcient

Equations 5 and 6 can be used to illustrate drug deliv-ery to normal brain and brain tumor in what may beviewed as ldquobest caserdquo and ldquoworst caserdquo scenarios Theworst case scenario will almost always be represented bywater-soluble drugs and the normal BBB that is the mostrestrictive situation The best case scenario will be repre-sented by lipid-soluble drugs and highly permeabletumors such as those represented in Fig 1 by the RG-2and D-54 MG gliomas In Table 1 the local and totalexposure fractions are presented for four drugs with differ-ent permeabilities methotrexate (K1 = 00014) 5-uo-rouracil (K1 = 00096) aziridinylbenzoquinone (K1 =0145) and 13-bis(2-chloroethyl)-1-nitrosourea (K1 = 06ml gndash1 minndash1) and for two permeability scenarios that ofnormal brain and that where permeability has increased01 ml gndash1 minndash1 over normal brain The data for the calcu-lations were contained in a previous publication (Blasbergand Groothuis 1986) the reader should be aware that thistype of modeling is dependent on assumptions made aboutthe different parameters Nonetheless Table 1 can be usedto illustrate several different points First in the worst casescenario with tumor microvessel function like the normalBBB and a highly water-soluble drug (methotrexate)

about 1 of the drug to which the tumor is exposed willenter the tumor With a lipid-soluble drug like aziridinyl-benzoquinone and normal BBB function the local expo-sure fraction increases to 07 in other words increasinglipid solubility does increase delivery to brain tissue Whentumor microvessel permeability increases by 01 ml gndash1

minndash1 the fraction of methotrexate to which the tumor isexposed increases to 18 A local increase in tumor per-meability results in a potentially signicant increase indrug delivery for a water-soluble drug However note thatalthough the fractional removal across normal BBB is highfor the lipid-soluble drugs aziridinylbenzoquinone and13-bis(2-chloroethyl)-1-nitrosourea the local exposurefraction does not increase signicantly when permeabilityis increased (the increase is only 1ndash2 percentage pointsTable 1) Increased drug delivery as a result of increases incapillary permeability will be more important for water-soluble drugs than for lipid-soluble drugs for which per-meability was not as great an issue in the rst place

It is important to remember that the rest of the bodyrepresents a large sink into which most of the drug is dis-tributed The fraction of drug entering tumor comparedwith that circulating through the entire body is minisculein every case lt1 of circulating drug will reach thetumor regardless of the drugrsquos permeability and regardlessof changes in capillary permeability (Table 1) Thereforeany method that is used to increase brain tumor perme-ability will still have to deal with the reality that most ofthe administered drug is distributed to the rest of the bodyand so far as treatment of the brain tumor is concernedwasted In most instances the limit on the total dose ofdrug that can be given is imposed by the tolerance of nor-mal tissues which is systemic toxicity in the case of drugsgiven systemically Because the maximum dose that can begiven to a patient is determined by factors external totreatment of the tumor (that is systemic toxicity) we mustaccept as a starting principle that the administered drugdose cannot be increased and we must search for alterna-tive methods to increase the fraction of drug that reachesthe tumor

Neuro-Oncology n JANUARY 200048

DR Groothuis Review of BBB and BTB

Table 1 Fractional delivery of four different chemotherapeutic agents across the BBB and BTB

Permeability of normal brain Permeability increase = 01 ml gndash1 minndash1

Drug Local exposure fraction Total exposure fraction Local exposure fraction Total exposure fraction

MTX 0017a 000006a 018 000006

5-FU 011 000004 025 000008

AZQ 070 00002 072b 00002b

BCNU 054 00002 055b 00002b

Abbreviations MTX methotrexate 5-FU 5-uorouracil AZQ azridinylbenzoquinone BCNU 13-bis(2-chloroethyl)-1-nitrosourea

Four drugs with different rates of brain entry brain efux and plasma half-lives are shown For each drug the values represent the fraction of drug extracted by the tumor The localexposure fraction represents the fraction of drug removed by 1 g of tumor from the plasma to which it was exposed (Equation 5) The total exposure fraction represents the fraction of

drug removed by 1 g of tumor from the entire circulating volume of drug in plasma (Equation 6) aThe ldquoworst caserdquo scenario occurs when water-soluble drugs are crossing the normal BBB bThe ldquobest caserdquo scenario occurs when lipid-soluble drugs cross capillaries in brain tumors with increased permeability However the two columns labeled total exposure fractionemphasize that the amount of drug removed by 1 g of tumor is very small compared with the total drug circulating in the body and indicates the magnitude of the sink effect of the

rest of the body in brain tumor chemotherapy

Increasing Permeability of the BBB and BTBto Drugs Given Intravascularly

Table 2 summarizes many of the methods that are cur-rently being used to manipulate the permeability of theBBB and BTB as well as my own interpretation of theiradvantages and disadvantages In light of the previousdiscussion all of these methods share a major disadvan-tage the bulk of administered drug will be lost to the restof the body in a sink effect and the maximum adminis-tered dose will almost always be determined by systemictoxicity

Chemical Modication of Existing Drugs

Chemical modication of existing drugs may refer to sev-eral different approaches First an existing drug may bemodified to make it fit a receptor in the BBB Thisapproach has been used with melphalan in which nitro-gen mustard (mechlorethamine) was linked to phenylala-nine (Groothuis et al 1992) Chemical modication mayrefer to linking an active protein (for example growthfactor) or a peptide to an antibody against a BBB recep-tor (Pardridge 1995) More commonly chemical modi-cation refers to the process of making an existing drugmore lipid soluble with the intent of increasing BBB per-meability This approach has been used extensively withantiretroviral nucleoside analogs for treating AIDS buthas not been used much with brain tumor chemothera-peutic drugs The use of increasingly accurate means topredict physicochemical properties offers encouragementthat chemical modication will nd wider use in modify-ing brain tumor drugs (van de Waterbeemd et al 1998)

Prodrug Therapy

This refers to a special class of chemical modication inwhich a drug is chemically modied to increase its capil-

lary permeability Once in the brain the prodrug under-goes an enzymatic reaction that returns the drug to itsactive state and with reduced BBB permeability (Shermanet al 1991 Yoshikawa et al 1999) These approacheswhich have not yet been used for brain tumor chemother-apeutic drugs offer an advantage by increasing K1 inEquation 3 while k2 and k3 remain unchanged Thiswould increase the AUCB while AUCPL remainsunchanged Although this increases the fraction of drugentering the tissue it does not change total body expo-sure and unfortunately introduces the need to explorethe pharmacokinetics and pharmacodynamics of eachnew compound

Intra-Arterial Administration

This method of administration refers to the ia injectionof a therapeutic agent without the concomitant use of abarrier-modifying approach The principle advantage ofthis method is that during the course of injecting a druginto an artery the tissue perfused by that artery receives ahigher plasma concentration during the first passagethrough the circulation (AUCPL is increased) In a seriesof papers Fenstermacher and coworkers theoreticallyevaluated the efficacy of intra-arterial delivery to thebrain and brain tumors (Cowles and Fenstermacher1974 Eckman et al 1974 Fenstermacher and Cowles1977 Fenstermacher and Gazendam 1981) They evalu-ated the contributions of several variables and concludedthat there were significant delivery advantages to iaadministration in a setting where the rate of tumor bloodow is very low or where the rate of systemic transfor-mation or excretion is very high or in a unique situationwhere having crossed the BBB or BTB the drug binds tothe tissue Once the drug passes through the tumormicrovasculature it enters the systemic circulation andthe pharmacokinetics are the same as for an iv adminis-tration Thus the ideal drug for ia administration is one

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 49

Table 2 Comparison of different methods of increasing delivery of intravascularly administered compounds to brain tumors

Method of increasing delivery Principle advantages Principle disadvantages Limiting factor(s)

Chemical modication of Requirements are now Each new compound must be BBB BTB permeabilityexisting drugs understood for different evaluated individually systemic toxicity

delivery routes

Prodrug Utilizes inherent biochemical Difference between BBB and BTB BBB BTB permeabilitypathways in brain and tumor is not known systemic toxicity

Intra-arterial First pass increase in AUC Invasive small increase in AUCBT BBB BTB permeability systemic toxicity

Hyperosmotic disruption First pass increase in AUC and Invasive differential effect on BBB BTB permeability increased permeability brain and tumor short-lived systemic toxicity

Chemical modication First pass increase in AUC and Invasive no effect on brain BBB BTB permeabilityincreased permeability short-lived

Receptor-mediated transport Increased permeability Low capacity system BBB BTB permeability

Inhibiting drug efux Decreased efux may be used None known None knownwith existing drugs

Avant-garde methods Not yet known Not yet known Not yet known

Abbreviations BBB blood-brain barrier BTB blood-tumor barrier AUC area under the curve

This table lists the different approaches that are available to increase the amount of drug in a brain tumor in cases where the drug is given either by iv or ia injection The advantages

and disadvantages are an expression of the authorrsquos opinion about the most prominent feature of each method Details about each method may be found in the text

that rapidly crosses the BBB or BTB and is either boundto tissue elements or locally metabolized in the process ofexerting its anticancer effect There are practical prob-lems associated with ia administration however includ-ing the need for the tumor to reside within the arterialdistribution being infused and hemodynamic streamingof the administered drug Perhaps the most obvious evi-dence that increased tissue drug concentrations can beachieved with ia administration is the increased inci-dence of local toxicity (Arafat et al 1999) Neverthelessbecause most drugs being used to treat brain tumors donot have the ideal qualities to take advantage of ia deliv-ery the results of clinical trials of ia chemotherapy inbrain tumors show minimal if any improvement in sur-vival (Dropcho et al 1998 Gundersen et al 1998Hirano et al 1998)

Hyperosmolar Blood-Brain Barrier Disruption(HBBBD)

This method involves the infusion of a hyperosmolarsolution (usually 14 M mannitol) into a cerebral arterygenerally followed by the intra-arterial administration ofa drug The principle features of this method areincreased capillary permeability (caused by the hyperos-molar solution) followed by a rst pass advantage fromia infusion that is K1 is transiently increased as isAUCPL This method was originally proposed byRapoport (Rapoport and Thompson 1973) and has beenused extensively by Neuwelt and colleagues (Kroll andNeuwelt 1998 Neuwelt and Dahlborg 1989 Neuweltet al 1980 Neuwelt et al 1983) Controversy about theuse of this method arose because of differential suscepti-bility of the normal BBB as compared with the abnormalBTB Whereas all studies demonstrated increased perme-ability of the normal BBB some studies in animal tumorsfailed to demonstrate any increase in tumor permeability(Nakagawa et al 1984) others showed transient perme-ability increases (Groothuis et al 1990) and somereported signicant permeability increases (Neuwelt etal 1984) Most recently Zunkeler et al used PET tostudy the time course of BBB function in 13 patients withmalignant astrocytomas after HBBBD (Zunkeler et al1996) They conrmed the differential effect of HBBBDpermeability was increased 1000 in brain and 60 intumors The half-time of the osmotic effect was 81 minin brain and 42 min in tumor They modeled the effectsof HBBBD on methotrexate and concluded that concen-trations above 1 microM the minimal concentration requiredfor an effect from methotrexate would not be enhancedin tumor and would be enhanced only 10 in brainThus HBBBD in combination with ia administrationwould be most useful for drugs that once having crossedthe BBB bind to brain or tumor tissue and do not efuxback across the BBB In addition the differential effect ofHBBBD on normal brain compared with that of tumormust be considered when evaluating neurotoxicity

Chemical Modication of the BBB and BTB

The latest entry into the therapeutic arena is the use ofchemical agents to disrupt the BBB (Black 1995) The

agents are mostly derivatives of normal vasoactive com-pounds and include bradykinin (Inamura and Black1994) interleukin-2 (Gutman et al 1996) leukotrieneC4 (Black and Chio 1992) and others (de Vries et al1996) The principle feature of these drugs is that theyare given ia and followed by an ia injection of a thera-peutic agent RMP-7 is a commercially developedbradykinin analog that has reached clinical trials (Ford etal 1998) and acts by increasing permeability at the inter-cellular junction (Sanovick et al 1995) Studies withPET in 9 malignant gliomas have shown that permeabil-ity to 68Ga-EDTA was increased 46 plusmn 42 in tumorswithout a signicant increase in tumor-free brain (Blacket al 1997) Another report suggests that RMP-7 canincrease delivery across the normal BBB (Emerich et al1998) RMP-7 is generally administered over a 15-minperiod the timing of administration of the chemothera-peutic drug relative to the RMP-7 has varied eg as abolus after RMP-7 or alternating chemotherapeutic drugadministration with RMP-7 BBB function is restoredrapidly after the RMP-7 infusion ends continuing theinfusion for over 20 min results in tachyphylaxis withloss of responsiveness (Bartus 1999) As with ia andHBBBD administration the advantage with chemicalmodication of the BBB is limited to a fractional increasein the circulating drug to which the tumor is exposedand still represents a vanishingly small fraction of thetotal body drug exposure Like those previous tech-niques chemical modication of the BBB will be ideal fora drug with dened properties as discussed before How-ever this ideal drug has yet to be developed

Receptor-Mediated Transport

This is a very broad area because any receptor-mediatedtransport system in the BBB can be selected as a targetThe considerations about drug delivery in a facilitatedtransport system are different from those involving sim-ple diffusion First a facilitated transport system can becharacterized by the Michaelis-Menten constants Km(the concentration at which the reaction velocity is halfmaximal) and Vmax (the limiting velocity as the concen-tration approaches innity) An important factor in thefacilitated transport of drugs is the plasma concentrationof the native substrate for the receptor For examplephenylalanine crosses the BBB via the large neutral aminoacid transport (LNAA) system melphalan is achemotherapeutic drug with a phenylalanine backboneand could therefore be transported by the LNAA systemHowever the normal plasma levels of phenylalaninecompete with melphalan to such a degree that the rate ofmelphalan transport into experimental gliomas is notincreased (Groothuis et al 1992) Lowering plasmaphenylalanine levels increased the rate of melphalanentry into brain tumors but only by a modest 8 Mostof the available transport receptors are low afnity lowcapacity systems such as those for the nucleosides vari-ous peptides transferrin and insulin (Ermisch et al1993 Pardridge et al 1995 Friden et al 1996 Tamai etal 1997) Although these transport systems have beenused to increase brain delivery of oligonucleotides(Boado et al 1998 Normand-Sdiqui and Akhtar 1998)

Neuro-Oncology n JANUARY 200050

DR Groothuis Review of BBB and BTB

nerve growth factor (Friden et al 1993) and vasoactiveintestinal peptide (Bickel et al 1993) none of thesetransport systems has been explored to any extent inbrain tumors

Inhibiting Drug Efux

Throughout most of the history of the BBB movementacross the capillary for most compounds without a trans-port system has been assumed to occur by simple diffu-sion the process is passive and the rate that a compoundcrosses the barrier is the same in both directions so thatthe process is symmetrical Recent studies have begun tochallenge this idea In particular P-glycoprotein repre-sents an ATP-dependent efflux pathway that confersresistance to cancer cells by allowing them to move avariety of chemotherapeutic drugs out of the cell againsta concentration gradient (Schinkel 1999 Tsuji andTamai 1997) P-glycoprotein appears to be localized pri-marily to the luminal capillary membrane and the num-ber of drugs that are transported by this system may bequite large (Schinkel 1999) Immunohistochemically P-glycoprotein has been demonstrated in malignantglioma tumor cells (Leweke et al 1998 von Bossanyi etal 1997) Leweke et al commented that tumor endothe-lial cells stained in 20 of 21 cases and von Bossanyi et alstated that tumor blood vessels were positive in 60 oftumors PET has been used to demonstrate the reversal ofP-glycoprotein pump function in mdr1a knock-out micewith cyclosporin A (Hendrikse et al 1998) Since manychemotherapeutic drugs are transported by P-glycopro-tein this represents a logical and exciting target in braintumor chemotherapy especially because inhibition of P-glycoprotein function could occur at both the BTB andin tumor cells Inhibiting P-glycoprotein would increaseboth extracellular and intracellular drug concentration(by decreasing k2 in Equation 3) without increasing theamount of drug administered This is an avenue of modi-fying brain tumor therapy that must be explored

Avant-Garde Methods of Delivery

Included in this category are the newest methods forintravascular delivery including the use of biodegradablenanoparticles (Schroeder et al 1998 Song et al 1997)liposomes (Allen and Moase 1996 Sakamoto and Ido1993) monoglyceride-based systems (Chang et al1994) and magnetic microspheres (Chang and Bodmeier1997) Many of these novel approaches are evaluatingdifferent methods of giving drug-laden molecules thatwill preferentially accumulate in a tissue usually com-bined with selective methods of perfusion such as iadelivery For example Pulfer and Gallo delivered 1ndash2 micromia magnetic aminodextran microspheres to RG-2 ratbrain tumors and used a magnetic eld to retain the par-ticles within the tumor They showed that this methodresulted in signicant accumulation of the particles as aresult of surface charge and magnetic eld (Chang andBodmeier 1997) These methods are too new to beapplied to human brain tumors but are representative ofthe diverse and novel directions that investigators areusing to increase brain tumor delivery

Summary of Intravascular Drug Delivery

A tremendous amount of progress has been made in thepast few years in our understanding of drug delivery tobrain tumors This progress has not yet translated intoimpressive increases in survival of patients with braintumors There are major areas that remain to be exploredand developed in the arena of intravascularly adminis-tered drugs For example because ia administration(with or without HBBBD or chemical modification)offers a unique rst pass advantage a search for drugsthat have high lipid solubility and bind to brain tumortargets is likely to be protable Second increasing tissuedrug concentration by the use of efux inhibitors is obvi-ous there will be no increase in total body exposurewhile potentially achieving signicant increases in tumordrug concentration However the ultimate limiting factorfor all intravascularly administered drugs is that of thepotential toxicity from total body exposure It seemsunlikely that this important and unfortunately negativevariable can be overcome

Injection or Infusion of Drugs Directly intothe Brain or Its Cavities

Table 3 summarizes the different approaches that arebeing used to administer drugs directly into the brainthus bypassing the BBB All of these routes and methodsof administration drastically change the pharmacokineticlandscape Instead of concern about crossing the formi-dable BBB or BTB the drugs are administered directlyinto the CSF or brain extracellular space Instead of need-ing to measure the inux rate it is necessary to under-stand drug movement within the brain and to understandefux mechanisms In marked contrast to intravascularadministration 100 of the administered dose can bedelivered using these methods of administration Theproblem then becomes one of understanding the forcesthat control the movement of the drug within brain andtumor tissue One or more of four forces will affect drugmovement depending upon the particular mode of drugadministration bulk flow of CSF bulk flow of braininterstitial uid bulk ow due to the infusion of a solu-tion into the brain or CSF and diffusion Drug movementwill also be variably affected by the amount of whitematter in the target area as a result of the change fromthe highly tortuous environment of a gray matter struc-ture (isotropic) to the highly oriented ber pathways ofwhite matter (anisotropic)

Intrathecal and Intraventricular Drug Administration

In this review intrathecal drug administration refers tothe administration of drug into the subarachnoid spaceusually into the lumbar subarachnoid space whereasintraventricular administration refers to injection or infu-sion into the lateral ventricle They are discussed togetherbecause of the large number of features they have in com-mon The principle feature of drug administration bythese routes is dominated by the bulk uid ow of the

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 51

CSF The major source of CSF is from the choroidplexuses of the lateral and 4th ventricles as much asanother 30 may originate from the brain extracellularuid The direction of the ow is from the lateral ventri-cles into the 3rd and then 4th ventricles out through theforamina of Luschke and Magendie into the basal cis-terns around the convexities and out of the subarach-noid space through the arachnoid granulations In ratswhen 14C-sucrose was injected into the lateral ventriclethe sucrose distributed rapidly within 5 min it was foundin the basal cisterns and by 1 h the sucrose had largely leftthe subarachnoid space (Ghersi-Egea et al 1996) Whenthe injected agent is metabolically active it may enterselected cells (Rubertone et al 1993) In humans the nor-mal volume of CSF produced is between 400 and 500 mlper day Since the CSF spaces within the brain are about150 ml in volume there is a considerable bulk ow withinthis system although the velocity varies tremendouslyfrom site to site When isotopes are injected into the lum-bar subarachnoid space they travel up the spinal sub-arachnoid space and are found in the basal cisterns by 3 hBy 24 h they are found over the cerebral convexities CSFow from the lateral ventricles is directional isotopesinjected into the lumbar CSF do not normally enter thelateral ventricles As a result of these bulk ow pathwaysmaterials injected into the CSF will attain initial concen-trations that are directly proportional to the concentra-tion in the infusate This route of administration is idealfor situations in which the target is within the subarach-noid space (for example carcinomatous meningitis) or inwhich the target is close to the CSF brain interface Carci-nomatous meningitis has long been treated with intrathe-cal infusions including the more recent use of monoclonalantibodies (Brown et al 1996) However the capacity ofdrugs to enter the brain extracellular space from the CSFis limited Numerous studies have shown that entry intothe brain is diffusional and that concentrations declineexponentially from the brain surface (Blasberg 1977Blasberg et al 1977 Groothuis and Levy 1997 Patlak

and Fenstermacher 1975) This route of administration isboth impractical and inefcient for intraparenchymalbrain tumors

Intratumoral and Intracavitary Injection

These are discussed together and briey because they areboth limiting cases of convection-enhanced deliverywhich is discussed below These both represent localforms of drug administration in which the distributionwill be determined initially by the rate and duration ofthe injection and secondly by tissue bulk ow pathwaysand diffusion The history of intratumoral injection wasdiscussed by Tomita (1991) who recognized the poten-tial usefulness of this approach as well as the importanceof neurotoxicity as the limiting factor Intratumoral andintracavitary injections are being widely explored asmeans of delivering large agents such as viral vectorsgrowth factors and cells (Bigner et al 1998 Farkkila etal 1994 Hsiao et al 1997 Puri et al 1996 Ram et al1997 Yang et al 1997) All of the attributes discussed inthe section on CED will also apply to these forms ofdrug delivery

Microdialysis

Microdialysis is a method that employs the passive diffu-sion of a drug across a semipermeable membrane (deLange et al 1997 de Lange et al 1999 Parsons andJustice 1994) It can be used to both sample and deliverdrugs to the surrounding tissue and has been used tosample extracellular drug concentrations in experimentalbrain tumors (Devineni et al 1996 Nakashima et al1997) The principle features of microdialysis when usedas a drug delivery methodology are that the tissue con-centrations are a function of the drug concentration inthe dialysate (which places control of the concentrationin hands of the user) and that distribution of drug awayfrom the dialysis catheter occurs by diffusion (Dykstra et

Neuro-Oncology n JANUARY 200052

DR Groothuis Review of BBB and BTB

Table 3 Comparison of different methods for local drug delivery to brain tumors

Method of circumventing the BBB Principle advantages Principle disadvantages Limiting factor(s)

Intrathecal Easy access ideal for therapy of Not useful for parenchymal Bulk ow rate of CSFmeningeal disease disease

Intraventricular administration Easy access ideal for therapy of Not useful for parenchymal Bulk ow rate of CSFmeningeal disease disease

Intratumoral injection User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Intracavitary injection User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Microdialysis User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Biodegradable polymers User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Convection-enhanced delivery User control over administered Invasive Unpredictable distribution dose 100 reaches target neurotoxicity

Abbreviations BBB blood-brain barrier CSF cerebrospinal uid

This table lists the different approaches that are available to deliver therapeutic drugs and agents by local administration The advantages and disadvantages are an expression of theauthorrsquos opinion about the most prominent feature of each method Details about each method may be found in the text

al 1992) However the normal BBB is disrupted imme-diately around the catheter and stays disrupted for sometime after catheter insertion making drug efux in thisvicinity an additional problem when trying to understandlocal delivery kinetics (Groothuis et al 1998 Morgan etal 1996 Westergren et al 1995) Because the principlefactors in local tissue concentrations are diffusion andorlocal brain bulk flow patterns the volume of brainreached by microdialysis will be relatively small anddrug concentrations will in general decline exponen-tially away from the dialysis cannula This method willbe applicable when the volume of tissue that needs to bereached is small The effect of normal brain bulk owpathways on this delivery method remains to be studied

Biodegradable Polymers

This represents a local delivery system in whichbiodegradable polymers are loaded with drug andimplanted into brain or tumor tissue resulting in timed-release of drug (Cao and Shoichet 1999 Menei et al1997) The principle features of this system are that mul-tiple polymer pellets or wafers can be implanted whichgives control over the spatial distribution and that drugdistribution away from the source occurs by diffusionwith a variable contribution from the inherent bulk owpathways of the brain (Fung et al 1998) This approachhas been used in patients with malignant gliomas whereits safety and efcacy have been demonstrated (Brem etal 1995 Olivi and Brem 1994) As with microdialysisthe principle limiting feature of this method of drug deliv-ery is that drug distribution occurs mainly by diffusionin which drug concentrations fall exponentially awayfrom the source and are generally limited to a few mil-limeters of tissue penetration (Fung et al 1998)

Convection-Enhanced Delivery

This is a relatively new method of drug delivery to braintumors The principle feature of this delivery method isthat a drug solution is infused directly into the brain Theconcentration of the drug is independent of the infusionrate and duration and the distribution of the drug is afunction of the hydrostatic pressure of the infusion aswell as diffusion into surrounding tissue From a deliveryperspective the only limiting factor to the drug concen-tration that can be delivered is the solubility of the drugCED has been discussed from a theoretical perspective(Morrison et al 1994) it has been explored in severalanimal models (Bobo et al 1994 Groothuis et al 1999Laske et al 1997a Lieberman et al 1995 Viola et al1995) and it has been used for delivery of a targetedtoxin in human gliomas (Laske et al 1997b) The distri-bution of the infused drug is highly dependent uponwhether the drug is infused into a homogeneous graymatter structure or a structure containing ber pathwaysIn the case of the infusion of a drug-containing solutioninto a homogeneous brain structure the infusion initiallyproduces a spherical volume in which the concentrationof the drug is directly proportional to that of the infusate(Fig 2b) After the infusion reaches equilibrium at whichpoint the amount of drug leaving across brain capillaries

is equal to the amount being infused the drug begins todiffuse away from the edge of the sphere at concentra-tions that decline exponentially in accordance with themathematics of diffusion (Fig 2b) If the infusionencounters an organized white matter pathway the rateof movement is either increased or decreased (dependingupon the direction of the inherent bulk ow within thatpathway) (Fig 2c) Morrison stated that when the infu-sion is being made into a homogeneous isotropicmedium the radius of the central spherical componentmdashthe convective componentmdashcan be related to the infusionrate and the efux constant of the drug being infused

rp 5 3 2qv (4p K) (Eq 7)

where qv is the infusion rate and K represents thesteady-state efux rate constant Fig 3 shows a plot of theradius of the convective component as a function of infu-sion rate and efux constant From reviewing Fig 3

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 53

Fig 2 The distribution of 14C-sucrose in rat brain by convection-enhanced delivery The diagram in the upper left (A) shows the targetlocation (center of the caudate nucleus) as well as the organization ofthe brain at the target level After a 1-h 05-microlmin infusion the iso-tope is found in a spherical shape with even distribution across (B) By8 h the isotope is moving into the external capsule (C) A similarappearance is found after a constant infusion over 7 days the centralconvective component is clearly visible surrounded by decreasingconcentration from the edge (D) Note the isotopersquos rapid lateralmovement down the external capsule (probably representing a nor-mal brain bulk uid pathway) but not into the corpus callosum Infu-sion into rat brain for 7 days produced no neuropathologic changesother than those associated with catheter insertion

which extends from infusion rates provided by osmoticmini-pumps (1 microlh) to high infusion rates (10 microlmin) it is

apparent that the volume of affected brain remains quitesmall for most infusion rates Similarly as the efux con-stant decreases indicating slower rates of crossing theBBB the convection volume increases Fig 3 illustratesthat CED is best suited for delivery of large compounds(with low efux rates) and delivery at moderately highinfusion rates if the intent is to reach large brain volumes

The shape of the volume of distribution at steady statewill be highly dependent on the location in the brain thatis being infused Fig 4 shows a CT scan of a dog brainreceiving an infusion of iodinated contrast into whitematter The iodinated contrast rapidly distributes in thewhite matter of the ipsilateral hemisphere with rapidlydecreasing concentrations in the cortex on the affectedside Note that in both Figs 2 and 4 the infused materialdoes not cross the midline through the corpus callosumThis suggests that the direction of the normal bulk owpathway within the corpus callosum is away from themidline thus opposing the CED infusion component Incontrast to intravascularly administered drugs whereincreased capillary permeability results in increased tissueconcentrations (Table 1) the opposite effect will occur inCED infusions Increased efux across permeable tumorcapillaries will reduce both the extracellular drug concen-tration and the volume of distribution In Fig 5 highconcentrations of 14C-sucrose were achieved by a CEDinfusion into an RG-2 glioma but at a distance of 24mm from the infusion site tissue concentrationsdecreased by several logarithmic units Although studieshave not been done in humans it is easy to anticipatethat the volume of distribution from a CED infusion willfollow similar principles Fig 6 shows several levels of anMRI scan through a malignant glioma Each set of scansshows a gadolinium-enhanced tumor margin (which canbe considered naturersquos equivalent to a CED infusion) anda T2-weighted image (which corresponds to the extracel-lular water originating from the tumor source) The

Neuro-Oncology n JANUARY 200054

DR Groothuis Review of BBB and BTB

Fig 3 Relationship between radius of convective CED componentinfusion rate and efux rate The radius of the inection pointbetween the convective and diffusional components of CED infusionsis shown for infusion into a homogeneous brain structure (Equation7) The values on the vertical axis represent the radius inside whichthe local concentrations remain uniform and outside of which drugconcentrations decline exponentially Changes for each 5-cm incre-ment are represented by different shades of gray The infusion ratewas chosen from 1 acute 10ndash6 to 1 acute 10ndash2 ml per minndash1 The lower valuesare those seen with infusions from osmotic mini-pumps while thehigher values represent the upper limit of CED infusion rates in dogsThe efux rate was chosen to vary from 1 acute 10ndash1 to 1 acute 10ndash7 minndash1The volume of the convective component increases as a function ofboth increasing infusion rate and decreasing efux rate

Fig 4 Illustration of CED infusion in a dog The gure on the left shows the CT scan at the start of a 3 microl minndash1 infusion of Isovue (iopamidol)into the hemispheric white matter of a dog while that on the right shows the same coronal level on the same dog after 10 days of infusion Thescan on the left contains an air bubble in the lateral ventricle the cannula tip contains a metal marker (the high density circle) and a small amountof Isovue surrounding the catheter tip In the scan on the right the Isovue has distributed at a high concentration throughout the white matterof the ipsilateral hemisphere with a rapid decline in concentration at gray-white interfaces Note that the distribution of Isovue is limited to theipsilateral hemisphere No neuropathologic changes were observed as a result of the infusion

shape and distribution of the edema are highly irregularand very dependent upon the brain structures encoun-tered Note that the edema uid moves into the internalcapsule but not into the thalamus or cortex Notice alsothat the edema uid does not cross the posterior limb ofthe corpus callosum although it comes into contact withit at several levels

A dominant feature of CED is that distribution of thedrug is determined by the infusion parameters and thetype of tissue being infused whereas the concentration ofdrug in the infusate is unrelated to these parameters Themaximum concentration of the drug in the infusate willbe determined by neurotoxicity within the central con-vective component of the infusion with variable concen-trations in the anisotropic brain surrounding the centralcomponent and concentrations 50 to 100 times lower inthe diffusional component furthermost from the infusioncatheter The ability to manipulate drug concentration inbrain and tumor tissue with such ease and to maintainthese manipulations for long periods of time (Groothuiset al 1999) brings a need for the development ofentirely new concepts For example the limits withregard to the composition of the infusate need to beexplored such as pH osmolarity and ionic compositionFurthermore how does one reliably study neurotoxicityAre infusions into animal models going to be reliable pre-dictors of toxicity in humans Will differential neurotox-icity become an issue It is possible that neuronsastrocytes and oligodendrogliocytes will display differ-ent levels of susceptibility to many drugs Will we seedelayed neurotoxicity such as that seen with radiation Itwill probably be best for many of the variables associatedwith neurotoxicity to be explored in animals before sub-jecting humans to CED infusions

Summary of Drug Delivery that Circumvents the BBB

There has been an explosion of methodology involvinglocal tissue drug delivery during the past 10 years Someof these techniques such as CED fulll the promise tocircumvent the BBB entirely In fullling this promisethese methods introduce new problems such as the rela-tionship between the spatial distribution of drugs andneurotoxicity that need to be understood before thesetechniques are widely used However for viral vectorsmonoclonal antibodies antisense oligonucleotides andother therapeutic agents that will be unable to cross theBBB in therapeutically effective amounts CED may pro-vide the necessary delivery tool

The State of Drug Delivery to Brain Tumorsndash1999

If we return to the statement of Vick et al at the begin-ning of this review we can now state with certainty thatthe BBB and BTB are major factors limiting the access ofmany therapeutic agents to brain tumors and these bar-riers will become even more signicant with the develop-ment of new molecular biological therapies that willinvolve large molecules viruses and even cells It is alsoremarkable how prescient those authors were in identify-ing the other parts of the drug delivery process that mustnow be understood and how little progress we havemade in elucidating ldquotumor cell uptake metabolic fatewithin tumor cells and the washout or sink effect of theextracellular space and CSFrdquo (Vick et al 1977) Formost of the past 20 years we have focused so much on

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 55

Fig 5 Infusion of 14C-sucrose into an RG-2 rat glioma An infusion cannula was placed into the caudate nucleus of a Fisher-344 rat (targetshown in Fig 2A) and RG-2 cells were injected Ten days later 14C-sucrose was infused at a rate of 10 microlh for 7 days Autoradiographic imageswere made (right images) and the sections were then stained with hematoxylin-eosin (left images) The top two images show that the 14C-sucrose distributes in high concentration throughout the RG-2 tumor extending into the edematous external capsule (E) However thelower two sections taken from 24 mm behind the infusion site still contain tumor and show that the concentration of 14C-sucrose is 100000times lower These sections illustrate the problems associated with controlling drug delivery by convection-enhanceddelivery

ways to increase delivery across the BBB that we havefailed to address these other issues which now becomethe forefront in drug delivery methods such as CED Thearsenal from which to choose a drug delivery method istruly impressive We must now study the other factorsthat limit effective brain tumor therapy Perhaps the mostpressing of these includes the need to develop effective

laboratory tools for identifying the relationship betweenthe concentration-time product of therapeutic agents andtumor cell kill rather than relying on empirical testing inhuman subjects It is most likely that the new century willsee the development and application of tools for individ-ualizing chemotherapy for brain tumors and other solidhuman tumors

Ali-Osman F (1999) Human glioma cultures and in vitro analysis of therapeu-

tic response In Berger MS and Wilson CB (Eds)The Gliomas

Philadelphia WB Saunders Co pp 134ndash142

Allen TM and Moase EH (1996) Therapeutic opportunities for targeted

liposomal drug delivery Adv Drug Deliv Rev 21 117ndash133

Arafat T Hentschel P Madajewicz S Manzione J Chowhan N Davis

R Roche P Af I Roque C Meek A and Magdy S (1999) Toxicities

related to intraarterial infusion of cisplatin and etoposide in patients with

brain tumors J Neurooncol 42 73ndash77

Bartus RT (1999) The blood-brain barrier as a target for phamacological

modulation Curr Opin Drug Discov Dev 2 152ndash167

Bickel U Yoshikawa T Landaw EM Faull KF and Pardridge WM

(1993) Pharmacologic effects in vivo in brain by vector-mediated peptide

delivery Proc Natl Acad Sci USA 90 2618ndash2622

Bigner DD Brown MT Friedman AH Coleman RE Akabani G Fried-

man HS Thorstad WL McLendon RE Bigner SH Zhao XG

Pegram CN Wikstrand CJ Herndon JE II Vick NA Paleologos N

Cokgor I Provenzale JM and Zalutsky MR (1998) Iodine-131-labeled

antitenascin monoclonal antibody 81C6 treatment of patients with recur-

rent malignant gliomas Phase I trial results J Clin Oncol 16 2202ndash2212

Black KL (1995) Biochemical opening of the blood-brain barrier Adv Drug

Deliv Rev 15 37ndash52

Neuro-Oncology n JANUARY 200056

DR Groothuis Review of BBB and BTB

Fig 6 Distribution of vasogenic edema in a malignant glioma There are four pairs of scans extending through a tumor in the left temporal lobeextending from the lowest (A) to the highest level (D) Each pair of scans contains a gadolinium-enhanced scan (left) and a T2-weighted image(right) The gadolinium-enhanced scans show the area of abnormally permeable BBB The T2-weighted images show the distribution of edema(water) emanating from the tumor The tumor may be considered a natural equivalent of a convection-enhanced delivery infusion The resultingedema spreads extensively through white matter in the ipsilateral hemisphere reaching forward in the internal capsule (levels C and D) but notpenetrating gray matter structures such as the thalamus (levels C and D) or cortex (levels AndashD) Note that the edema contacts the posterior limbof the corpus callosum in levels C and D but does not extend into the corpus callosum suggesting that there is a physiological barrier to the bulkmovement of uid in that direction

References

Increasing Permeability of the BBB and BTBto Drugs Given Intravascularly

Table 2 summarizes many of the methods that are cur-rently being used to manipulate the permeability of theBBB and BTB as well as my own interpretation of theiradvantages and disadvantages In light of the previousdiscussion all of these methods share a major disadvan-tage the bulk of administered drug will be lost to the restof the body in a sink effect and the maximum adminis-tered dose will almost always be determined by systemictoxicity

Chemical Modication of Existing Drugs

Chemical modication of existing drugs may refer to sev-eral different approaches First an existing drug may bemodified to make it fit a receptor in the BBB Thisapproach has been used with melphalan in which nitro-gen mustard (mechlorethamine) was linked to phenylala-nine (Groothuis et al 1992) Chemical modication mayrefer to linking an active protein (for example growthfactor) or a peptide to an antibody against a BBB recep-tor (Pardridge 1995) More commonly chemical modi-cation refers to the process of making an existing drugmore lipid soluble with the intent of increasing BBB per-meability This approach has been used extensively withantiretroviral nucleoside analogs for treating AIDS buthas not been used much with brain tumor chemothera-peutic drugs The use of increasingly accurate means topredict physicochemical properties offers encouragementthat chemical modication will nd wider use in modify-ing brain tumor drugs (van de Waterbeemd et al 1998)

Prodrug Therapy

This refers to a special class of chemical modication inwhich a drug is chemically modied to increase its capil-

lary permeability Once in the brain the prodrug under-goes an enzymatic reaction that returns the drug to itsactive state and with reduced BBB permeability (Shermanet al 1991 Yoshikawa et al 1999) These approacheswhich have not yet been used for brain tumor chemother-apeutic drugs offer an advantage by increasing K1 inEquation 3 while k2 and k3 remain unchanged Thiswould increase the AUCB while AUCPL remainsunchanged Although this increases the fraction of drugentering the tissue it does not change total body expo-sure and unfortunately introduces the need to explorethe pharmacokinetics and pharmacodynamics of eachnew compound

Intra-Arterial Administration

This method of administration refers to the ia injectionof a therapeutic agent without the concomitant use of abarrier-modifying approach The principle advantage ofthis method is that during the course of injecting a druginto an artery the tissue perfused by that artery receives ahigher plasma concentration during the first passagethrough the circulation (AUCPL is increased) In a seriesof papers Fenstermacher and coworkers theoreticallyevaluated the efficacy of intra-arterial delivery to thebrain and brain tumors (Cowles and Fenstermacher1974 Eckman et al 1974 Fenstermacher and Cowles1977 Fenstermacher and Gazendam 1981) They evalu-ated the contributions of several variables and concludedthat there were significant delivery advantages to iaadministration in a setting where the rate of tumor bloodow is very low or where the rate of systemic transfor-mation or excretion is very high or in a unique situationwhere having crossed the BBB or BTB the drug binds tothe tissue Once the drug passes through the tumormicrovasculature it enters the systemic circulation andthe pharmacokinetics are the same as for an iv adminis-tration Thus the ideal drug for ia administration is one

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 49

Table 2 Comparison of different methods of increasing delivery of intravascularly administered compounds to brain tumors

Method of increasing delivery Principle advantages Principle disadvantages Limiting factor(s)

Chemical modication of Requirements are now Each new compound must be BBB BTB permeabilityexisting drugs understood for different evaluated individually systemic toxicity

delivery routes

Prodrug Utilizes inherent biochemical Difference between BBB and BTB BBB BTB permeabilitypathways in brain and tumor is not known systemic toxicity

Intra-arterial First pass increase in AUC Invasive small increase in AUCBT BBB BTB permeability systemic toxicity

Hyperosmotic disruption First pass increase in AUC and Invasive differential effect on BBB BTB permeability increased permeability brain and tumor short-lived systemic toxicity

Chemical modication First pass increase in AUC and Invasive no effect on brain BBB BTB permeabilityincreased permeability short-lived

Receptor-mediated transport Increased permeability Low capacity system BBB BTB permeability

Inhibiting drug efux Decreased efux may be used None known None knownwith existing drugs

Avant-garde methods Not yet known Not yet known Not yet known

Abbreviations BBB blood-brain barrier BTB blood-tumor barrier AUC area under the curve

This table lists the different approaches that are available to increase the amount of drug in a brain tumor in cases where the drug is given either by iv or ia injection The advantages

and disadvantages are an expression of the authorrsquos opinion about the most prominent feature of each method Details about each method may be found in the text

that rapidly crosses the BBB or BTB and is either boundto tissue elements or locally metabolized in the process ofexerting its anticancer effect There are practical prob-lems associated with ia administration however includ-ing the need for the tumor to reside within the arterialdistribution being infused and hemodynamic streamingof the administered drug Perhaps the most obvious evi-dence that increased tissue drug concentrations can beachieved with ia administration is the increased inci-dence of local toxicity (Arafat et al 1999) Neverthelessbecause most drugs being used to treat brain tumors donot have the ideal qualities to take advantage of ia deliv-ery the results of clinical trials of ia chemotherapy inbrain tumors show minimal if any improvement in sur-vival (Dropcho et al 1998 Gundersen et al 1998Hirano et al 1998)

Hyperosmolar Blood-Brain Barrier Disruption(HBBBD)

This method involves the infusion of a hyperosmolarsolution (usually 14 M mannitol) into a cerebral arterygenerally followed by the intra-arterial administration ofa drug The principle features of this method areincreased capillary permeability (caused by the hyperos-molar solution) followed by a rst pass advantage fromia infusion that is K1 is transiently increased as isAUCPL This method was originally proposed byRapoport (Rapoport and Thompson 1973) and has beenused extensively by Neuwelt and colleagues (Kroll andNeuwelt 1998 Neuwelt and Dahlborg 1989 Neuweltet al 1980 Neuwelt et al 1983) Controversy about theuse of this method arose because of differential suscepti-bility of the normal BBB as compared with the abnormalBTB Whereas all studies demonstrated increased perme-ability of the normal BBB some studies in animal tumorsfailed to demonstrate any increase in tumor permeability(Nakagawa et al 1984) others showed transient perme-ability increases (Groothuis et al 1990) and somereported signicant permeability increases (Neuwelt etal 1984) Most recently Zunkeler et al used PET tostudy the time course of BBB function in 13 patients withmalignant astrocytomas after HBBBD (Zunkeler et al1996) They conrmed the differential effect of HBBBDpermeability was increased 1000 in brain and 60 intumors The half-time of the osmotic effect was 81 minin brain and 42 min in tumor They modeled the effectsof HBBBD on methotrexate and concluded that concen-trations above 1 microM the minimal concentration requiredfor an effect from methotrexate would not be enhancedin tumor and would be enhanced only 10 in brainThus HBBBD in combination with ia administrationwould be most useful for drugs that once having crossedthe BBB bind to brain or tumor tissue and do not efuxback across the BBB In addition the differential effect ofHBBBD on normal brain compared with that of tumormust be considered when evaluating neurotoxicity

Chemical Modication of the BBB and BTB

The latest entry into the therapeutic arena is the use ofchemical agents to disrupt the BBB (Black 1995) The

agents are mostly derivatives of normal vasoactive com-pounds and include bradykinin (Inamura and Black1994) interleukin-2 (Gutman et al 1996) leukotrieneC4 (Black and Chio 1992) and others (de Vries et al1996) The principle feature of these drugs is that theyare given ia and followed by an ia injection of a thera-peutic agent RMP-7 is a commercially developedbradykinin analog that has reached clinical trials (Ford etal 1998) and acts by increasing permeability at the inter-cellular junction (Sanovick et al 1995) Studies withPET in 9 malignant gliomas have shown that permeabil-ity to 68Ga-EDTA was increased 46 plusmn 42 in tumorswithout a signicant increase in tumor-free brain (Blacket al 1997) Another report suggests that RMP-7 canincrease delivery across the normal BBB (Emerich et al1998) RMP-7 is generally administered over a 15-minperiod the timing of administration of the chemothera-peutic drug relative to the RMP-7 has varied eg as abolus after RMP-7 or alternating chemotherapeutic drugadministration with RMP-7 BBB function is restoredrapidly after the RMP-7 infusion ends continuing theinfusion for over 20 min results in tachyphylaxis withloss of responsiveness (Bartus 1999) As with ia andHBBBD administration the advantage with chemicalmodication of the BBB is limited to a fractional increasein the circulating drug to which the tumor is exposedand still represents a vanishingly small fraction of thetotal body drug exposure Like those previous tech-niques chemical modication of the BBB will be ideal fora drug with dened properties as discussed before How-ever this ideal drug has yet to be developed

Receptor-Mediated Transport

This is a very broad area because any receptor-mediatedtransport system in the BBB can be selected as a targetThe considerations about drug delivery in a facilitatedtransport system are different from those involving sim-ple diffusion First a facilitated transport system can becharacterized by the Michaelis-Menten constants Km(the concentration at which the reaction velocity is halfmaximal) and Vmax (the limiting velocity as the concen-tration approaches innity) An important factor in thefacilitated transport of drugs is the plasma concentrationof the native substrate for the receptor For examplephenylalanine crosses the BBB via the large neutral aminoacid transport (LNAA) system melphalan is achemotherapeutic drug with a phenylalanine backboneand could therefore be transported by the LNAA systemHowever the normal plasma levels of phenylalaninecompete with melphalan to such a degree that the rate ofmelphalan transport into experimental gliomas is notincreased (Groothuis et al 1992) Lowering plasmaphenylalanine levels increased the rate of melphalanentry into brain tumors but only by a modest 8 Mostof the available transport receptors are low afnity lowcapacity systems such as those for the nucleosides vari-ous peptides transferrin and insulin (Ermisch et al1993 Pardridge et al 1995 Friden et al 1996 Tamai etal 1997) Although these transport systems have beenused to increase brain delivery of oligonucleotides(Boado et al 1998 Normand-Sdiqui and Akhtar 1998)

Neuro-Oncology n JANUARY 200050

DR Groothuis Review of BBB and BTB

nerve growth factor (Friden et al 1993) and vasoactiveintestinal peptide (Bickel et al 1993) none of thesetransport systems has been explored to any extent inbrain tumors

Inhibiting Drug Efux

Throughout most of the history of the BBB movementacross the capillary for most compounds without a trans-port system has been assumed to occur by simple diffu-sion the process is passive and the rate that a compoundcrosses the barrier is the same in both directions so thatthe process is symmetrical Recent studies have begun tochallenge this idea In particular P-glycoprotein repre-sents an ATP-dependent efflux pathway that confersresistance to cancer cells by allowing them to move avariety of chemotherapeutic drugs out of the cell againsta concentration gradient (Schinkel 1999 Tsuji andTamai 1997) P-glycoprotein appears to be localized pri-marily to the luminal capillary membrane and the num-ber of drugs that are transported by this system may bequite large (Schinkel 1999) Immunohistochemically P-glycoprotein has been demonstrated in malignantglioma tumor cells (Leweke et al 1998 von Bossanyi etal 1997) Leweke et al commented that tumor endothe-lial cells stained in 20 of 21 cases and von Bossanyi et alstated that tumor blood vessels were positive in 60 oftumors PET has been used to demonstrate the reversal ofP-glycoprotein pump function in mdr1a knock-out micewith cyclosporin A (Hendrikse et al 1998) Since manychemotherapeutic drugs are transported by P-glycopro-tein this represents a logical and exciting target in braintumor chemotherapy especially because inhibition of P-glycoprotein function could occur at both the BTB andin tumor cells Inhibiting P-glycoprotein would increaseboth extracellular and intracellular drug concentration(by decreasing k2 in Equation 3) without increasing theamount of drug administered This is an avenue of modi-fying brain tumor therapy that must be explored

Avant-Garde Methods of Delivery

Included in this category are the newest methods forintravascular delivery including the use of biodegradablenanoparticles (Schroeder et al 1998 Song et al 1997)liposomes (Allen and Moase 1996 Sakamoto and Ido1993) monoglyceride-based systems (Chang et al1994) and magnetic microspheres (Chang and Bodmeier1997) Many of these novel approaches are evaluatingdifferent methods of giving drug-laden molecules thatwill preferentially accumulate in a tissue usually com-bined with selective methods of perfusion such as iadelivery For example Pulfer and Gallo delivered 1ndash2 micromia magnetic aminodextran microspheres to RG-2 ratbrain tumors and used a magnetic eld to retain the par-ticles within the tumor They showed that this methodresulted in signicant accumulation of the particles as aresult of surface charge and magnetic eld (Chang andBodmeier 1997) These methods are too new to beapplied to human brain tumors but are representative ofthe diverse and novel directions that investigators areusing to increase brain tumor delivery

Summary of Intravascular Drug Delivery

A tremendous amount of progress has been made in thepast few years in our understanding of drug delivery tobrain tumors This progress has not yet translated intoimpressive increases in survival of patients with braintumors There are major areas that remain to be exploredand developed in the arena of intravascularly adminis-tered drugs For example because ia administration(with or without HBBBD or chemical modification)offers a unique rst pass advantage a search for drugsthat have high lipid solubility and bind to brain tumortargets is likely to be protable Second increasing tissuedrug concentration by the use of efux inhibitors is obvi-ous there will be no increase in total body exposurewhile potentially achieving signicant increases in tumordrug concentration However the ultimate limiting factorfor all intravascularly administered drugs is that of thepotential toxicity from total body exposure It seemsunlikely that this important and unfortunately negativevariable can be overcome

Injection or Infusion of Drugs Directly intothe Brain or Its Cavities

Table 3 summarizes the different approaches that arebeing used to administer drugs directly into the brainthus bypassing the BBB All of these routes and methodsof administration drastically change the pharmacokineticlandscape Instead of concern about crossing the formi-dable BBB or BTB the drugs are administered directlyinto the CSF or brain extracellular space Instead of need-ing to measure the inux rate it is necessary to under-stand drug movement within the brain and to understandefux mechanisms In marked contrast to intravascularadministration 100 of the administered dose can bedelivered using these methods of administration Theproblem then becomes one of understanding the forcesthat control the movement of the drug within brain andtumor tissue One or more of four forces will affect drugmovement depending upon the particular mode of drugadministration bulk flow of CSF bulk flow of braininterstitial uid bulk ow due to the infusion of a solu-tion into the brain or CSF and diffusion Drug movementwill also be variably affected by the amount of whitematter in the target area as a result of the change fromthe highly tortuous environment of a gray matter struc-ture (isotropic) to the highly oriented ber pathways ofwhite matter (anisotropic)

Intrathecal and Intraventricular Drug Administration

In this review intrathecal drug administration refers tothe administration of drug into the subarachnoid spaceusually into the lumbar subarachnoid space whereasintraventricular administration refers to injection or infu-sion into the lateral ventricle They are discussed togetherbecause of the large number of features they have in com-mon The principle feature of drug administration bythese routes is dominated by the bulk uid ow of the

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 51

CSF The major source of CSF is from the choroidplexuses of the lateral and 4th ventricles as much asanother 30 may originate from the brain extracellularuid The direction of the ow is from the lateral ventri-cles into the 3rd and then 4th ventricles out through theforamina of Luschke and Magendie into the basal cis-terns around the convexities and out of the subarach-noid space through the arachnoid granulations In ratswhen 14C-sucrose was injected into the lateral ventriclethe sucrose distributed rapidly within 5 min it was foundin the basal cisterns and by 1 h the sucrose had largely leftthe subarachnoid space (Ghersi-Egea et al 1996) Whenthe injected agent is metabolically active it may enterselected cells (Rubertone et al 1993) In humans the nor-mal volume of CSF produced is between 400 and 500 mlper day Since the CSF spaces within the brain are about150 ml in volume there is a considerable bulk ow withinthis system although the velocity varies tremendouslyfrom site to site When isotopes are injected into the lum-bar subarachnoid space they travel up the spinal sub-arachnoid space and are found in the basal cisterns by 3 hBy 24 h they are found over the cerebral convexities CSFow from the lateral ventricles is directional isotopesinjected into the lumbar CSF do not normally enter thelateral ventricles As a result of these bulk ow pathwaysmaterials injected into the CSF will attain initial concen-trations that are directly proportional to the concentra-tion in the infusate This route of administration is idealfor situations in which the target is within the subarach-noid space (for example carcinomatous meningitis) or inwhich the target is close to the CSF brain interface Carci-nomatous meningitis has long been treated with intrathe-cal infusions including the more recent use of monoclonalantibodies (Brown et al 1996) However the capacity ofdrugs to enter the brain extracellular space from the CSFis limited Numerous studies have shown that entry intothe brain is diffusional and that concentrations declineexponentially from the brain surface (Blasberg 1977Blasberg et al 1977 Groothuis and Levy 1997 Patlak

and Fenstermacher 1975) This route of administration isboth impractical and inefcient for intraparenchymalbrain tumors

Intratumoral and Intracavitary Injection

These are discussed together and briey because they areboth limiting cases of convection-enhanced deliverywhich is discussed below These both represent localforms of drug administration in which the distributionwill be determined initially by the rate and duration ofthe injection and secondly by tissue bulk ow pathwaysand diffusion The history of intratumoral injection wasdiscussed by Tomita (1991) who recognized the poten-tial usefulness of this approach as well as the importanceof neurotoxicity as the limiting factor Intratumoral andintracavitary injections are being widely explored asmeans of delivering large agents such as viral vectorsgrowth factors and cells (Bigner et al 1998 Farkkila etal 1994 Hsiao et al 1997 Puri et al 1996 Ram et al1997 Yang et al 1997) All of the attributes discussed inthe section on CED will also apply to these forms ofdrug delivery

Microdialysis

Microdialysis is a method that employs the passive diffu-sion of a drug across a semipermeable membrane (deLange et al 1997 de Lange et al 1999 Parsons andJustice 1994) It can be used to both sample and deliverdrugs to the surrounding tissue and has been used tosample extracellular drug concentrations in experimentalbrain tumors (Devineni et al 1996 Nakashima et al1997) The principle features of microdialysis when usedas a drug delivery methodology are that the tissue con-centrations are a function of the drug concentration inthe dialysate (which places control of the concentrationin hands of the user) and that distribution of drug awayfrom the dialysis catheter occurs by diffusion (Dykstra et

Neuro-Oncology n JANUARY 200052

DR Groothuis Review of BBB and BTB

Table 3 Comparison of different methods for local drug delivery to brain tumors

Method of circumventing the BBB Principle advantages Principle disadvantages Limiting factor(s)

Intrathecal Easy access ideal for therapy of Not useful for parenchymal Bulk ow rate of CSFmeningeal disease disease

Intraventricular administration Easy access ideal for therapy of Not useful for parenchymal Bulk ow rate of CSFmeningeal disease disease

Intratumoral injection User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Intracavitary injection User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Microdialysis User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Biodegradable polymers User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Convection-enhanced delivery User control over administered Invasive Unpredictable distribution dose 100 reaches target neurotoxicity

Abbreviations BBB blood-brain barrier CSF cerebrospinal uid

This table lists the different approaches that are available to deliver therapeutic drugs and agents by local administration The advantages and disadvantages are an expression of theauthorrsquos opinion about the most prominent feature of each method Details about each method may be found in the text

al 1992) However the normal BBB is disrupted imme-diately around the catheter and stays disrupted for sometime after catheter insertion making drug efux in thisvicinity an additional problem when trying to understandlocal delivery kinetics (Groothuis et al 1998 Morgan etal 1996 Westergren et al 1995) Because the principlefactors in local tissue concentrations are diffusion andorlocal brain bulk flow patterns the volume of brainreached by microdialysis will be relatively small anddrug concentrations will in general decline exponen-tially away from the dialysis cannula This method willbe applicable when the volume of tissue that needs to bereached is small The effect of normal brain bulk owpathways on this delivery method remains to be studied

Biodegradable Polymers

This represents a local delivery system in whichbiodegradable polymers are loaded with drug andimplanted into brain or tumor tissue resulting in timed-release of drug (Cao and Shoichet 1999 Menei et al1997) The principle features of this system are that mul-tiple polymer pellets or wafers can be implanted whichgives control over the spatial distribution and that drugdistribution away from the source occurs by diffusionwith a variable contribution from the inherent bulk owpathways of the brain (Fung et al 1998) This approachhas been used in patients with malignant gliomas whereits safety and efcacy have been demonstrated (Brem etal 1995 Olivi and Brem 1994) As with microdialysisthe principle limiting feature of this method of drug deliv-ery is that drug distribution occurs mainly by diffusionin which drug concentrations fall exponentially awayfrom the source and are generally limited to a few mil-limeters of tissue penetration (Fung et al 1998)

Convection-Enhanced Delivery

This is a relatively new method of drug delivery to braintumors The principle feature of this delivery method isthat a drug solution is infused directly into the brain Theconcentration of the drug is independent of the infusionrate and duration and the distribution of the drug is afunction of the hydrostatic pressure of the infusion aswell as diffusion into surrounding tissue From a deliveryperspective the only limiting factor to the drug concen-tration that can be delivered is the solubility of the drugCED has been discussed from a theoretical perspective(Morrison et al 1994) it has been explored in severalanimal models (Bobo et al 1994 Groothuis et al 1999Laske et al 1997a Lieberman et al 1995 Viola et al1995) and it has been used for delivery of a targetedtoxin in human gliomas (Laske et al 1997b) The distri-bution of the infused drug is highly dependent uponwhether the drug is infused into a homogeneous graymatter structure or a structure containing ber pathwaysIn the case of the infusion of a drug-containing solutioninto a homogeneous brain structure the infusion initiallyproduces a spherical volume in which the concentrationof the drug is directly proportional to that of the infusate(Fig 2b) After the infusion reaches equilibrium at whichpoint the amount of drug leaving across brain capillaries

is equal to the amount being infused the drug begins todiffuse away from the edge of the sphere at concentra-tions that decline exponentially in accordance with themathematics of diffusion (Fig 2b) If the infusionencounters an organized white matter pathway the rateof movement is either increased or decreased (dependingupon the direction of the inherent bulk ow within thatpathway) (Fig 2c) Morrison stated that when the infu-sion is being made into a homogeneous isotropicmedium the radius of the central spherical componentmdashthe convective componentmdashcan be related to the infusionrate and the efux constant of the drug being infused

rp 5 3 2qv (4p K) (Eq 7)

where qv is the infusion rate and K represents thesteady-state efux rate constant Fig 3 shows a plot of theradius of the convective component as a function of infu-sion rate and efux constant From reviewing Fig 3

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 53

Fig 2 The distribution of 14C-sucrose in rat brain by convection-enhanced delivery The diagram in the upper left (A) shows the targetlocation (center of the caudate nucleus) as well as the organization ofthe brain at the target level After a 1-h 05-microlmin infusion the iso-tope is found in a spherical shape with even distribution across (B) By8 h the isotope is moving into the external capsule (C) A similarappearance is found after a constant infusion over 7 days the centralconvective component is clearly visible surrounded by decreasingconcentration from the edge (D) Note the isotopersquos rapid lateralmovement down the external capsule (probably representing a nor-mal brain bulk uid pathway) but not into the corpus callosum Infu-sion into rat brain for 7 days produced no neuropathologic changesother than those associated with catheter insertion

which extends from infusion rates provided by osmoticmini-pumps (1 microlh) to high infusion rates (10 microlmin) it is

apparent that the volume of affected brain remains quitesmall for most infusion rates Similarly as the efux con-stant decreases indicating slower rates of crossing theBBB the convection volume increases Fig 3 illustratesthat CED is best suited for delivery of large compounds(with low efux rates) and delivery at moderately highinfusion rates if the intent is to reach large brain volumes

The shape of the volume of distribution at steady statewill be highly dependent on the location in the brain thatis being infused Fig 4 shows a CT scan of a dog brainreceiving an infusion of iodinated contrast into whitematter The iodinated contrast rapidly distributes in thewhite matter of the ipsilateral hemisphere with rapidlydecreasing concentrations in the cortex on the affectedside Note that in both Figs 2 and 4 the infused materialdoes not cross the midline through the corpus callosumThis suggests that the direction of the normal bulk owpathway within the corpus callosum is away from themidline thus opposing the CED infusion component Incontrast to intravascularly administered drugs whereincreased capillary permeability results in increased tissueconcentrations (Table 1) the opposite effect will occur inCED infusions Increased efux across permeable tumorcapillaries will reduce both the extracellular drug concen-tration and the volume of distribution In Fig 5 highconcentrations of 14C-sucrose were achieved by a CEDinfusion into an RG-2 glioma but at a distance of 24mm from the infusion site tissue concentrationsdecreased by several logarithmic units Although studieshave not been done in humans it is easy to anticipatethat the volume of distribution from a CED infusion willfollow similar principles Fig 6 shows several levels of anMRI scan through a malignant glioma Each set of scansshows a gadolinium-enhanced tumor margin (which canbe considered naturersquos equivalent to a CED infusion) anda T2-weighted image (which corresponds to the extracel-lular water originating from the tumor source) The

Neuro-Oncology n JANUARY 200054

DR Groothuis Review of BBB and BTB

Fig 3 Relationship between radius of convective CED componentinfusion rate and efux rate The radius of the inection pointbetween the convective and diffusional components of CED infusionsis shown for infusion into a homogeneous brain structure (Equation7) The values on the vertical axis represent the radius inside whichthe local concentrations remain uniform and outside of which drugconcentrations decline exponentially Changes for each 5-cm incre-ment are represented by different shades of gray The infusion ratewas chosen from 1 acute 10ndash6 to 1 acute 10ndash2 ml per minndash1 The lower valuesare those seen with infusions from osmotic mini-pumps while thehigher values represent the upper limit of CED infusion rates in dogsThe efux rate was chosen to vary from 1 acute 10ndash1 to 1 acute 10ndash7 minndash1The volume of the convective component increases as a function ofboth increasing infusion rate and decreasing efux rate

Fig 4 Illustration of CED infusion in a dog The gure on the left shows the CT scan at the start of a 3 microl minndash1 infusion of Isovue (iopamidol)into the hemispheric white matter of a dog while that on the right shows the same coronal level on the same dog after 10 days of infusion Thescan on the left contains an air bubble in the lateral ventricle the cannula tip contains a metal marker (the high density circle) and a small amountof Isovue surrounding the catheter tip In the scan on the right the Isovue has distributed at a high concentration throughout the white matterof the ipsilateral hemisphere with a rapid decline in concentration at gray-white interfaces Note that the distribution of Isovue is limited to theipsilateral hemisphere No neuropathologic changes were observed as a result of the infusion

shape and distribution of the edema are highly irregularand very dependent upon the brain structures encoun-tered Note that the edema uid moves into the internalcapsule but not into the thalamus or cortex Notice alsothat the edema uid does not cross the posterior limb ofthe corpus callosum although it comes into contact withit at several levels

A dominant feature of CED is that distribution of thedrug is determined by the infusion parameters and thetype of tissue being infused whereas the concentration ofdrug in the infusate is unrelated to these parameters Themaximum concentration of the drug in the infusate willbe determined by neurotoxicity within the central con-vective component of the infusion with variable concen-trations in the anisotropic brain surrounding the centralcomponent and concentrations 50 to 100 times lower inthe diffusional component furthermost from the infusioncatheter The ability to manipulate drug concentration inbrain and tumor tissue with such ease and to maintainthese manipulations for long periods of time (Groothuiset al 1999) brings a need for the development ofentirely new concepts For example the limits withregard to the composition of the infusate need to beexplored such as pH osmolarity and ionic compositionFurthermore how does one reliably study neurotoxicityAre infusions into animal models going to be reliable pre-dictors of toxicity in humans Will differential neurotox-icity become an issue It is possible that neuronsastrocytes and oligodendrogliocytes will display differ-ent levels of susceptibility to many drugs Will we seedelayed neurotoxicity such as that seen with radiation Itwill probably be best for many of the variables associatedwith neurotoxicity to be explored in animals before sub-jecting humans to CED infusions

Summary of Drug Delivery that Circumvents the BBB

There has been an explosion of methodology involvinglocal tissue drug delivery during the past 10 years Someof these techniques such as CED fulll the promise tocircumvent the BBB entirely In fullling this promisethese methods introduce new problems such as the rela-tionship between the spatial distribution of drugs andneurotoxicity that need to be understood before thesetechniques are widely used However for viral vectorsmonoclonal antibodies antisense oligonucleotides andother therapeutic agents that will be unable to cross theBBB in therapeutically effective amounts CED may pro-vide the necessary delivery tool

The State of Drug Delivery to Brain Tumorsndash1999

If we return to the statement of Vick et al at the begin-ning of this review we can now state with certainty thatthe BBB and BTB are major factors limiting the access ofmany therapeutic agents to brain tumors and these bar-riers will become even more signicant with the develop-ment of new molecular biological therapies that willinvolve large molecules viruses and even cells It is alsoremarkable how prescient those authors were in identify-ing the other parts of the drug delivery process that mustnow be understood and how little progress we havemade in elucidating ldquotumor cell uptake metabolic fatewithin tumor cells and the washout or sink effect of theextracellular space and CSFrdquo (Vick et al 1977) Formost of the past 20 years we have focused so much on

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 55

Fig 5 Infusion of 14C-sucrose into an RG-2 rat glioma An infusion cannula was placed into the caudate nucleus of a Fisher-344 rat (targetshown in Fig 2A) and RG-2 cells were injected Ten days later 14C-sucrose was infused at a rate of 10 microlh for 7 days Autoradiographic imageswere made (right images) and the sections were then stained with hematoxylin-eosin (left images) The top two images show that the 14C-sucrose distributes in high concentration throughout the RG-2 tumor extending into the edematous external capsule (E) However thelower two sections taken from 24 mm behind the infusion site still contain tumor and show that the concentration of 14C-sucrose is 100000times lower These sections illustrate the problems associated with controlling drug delivery by convection-enhanceddelivery

ways to increase delivery across the BBB that we havefailed to address these other issues which now becomethe forefront in drug delivery methods such as CED Thearsenal from which to choose a drug delivery method istruly impressive We must now study the other factorsthat limit effective brain tumor therapy Perhaps the mostpressing of these includes the need to develop effective

laboratory tools for identifying the relationship betweenthe concentration-time product of therapeutic agents andtumor cell kill rather than relying on empirical testing inhuman subjects It is most likely that the new century willsee the development and application of tools for individ-ualizing chemotherapy for brain tumors and other solidhuman tumors

Ali-Osman F (1999) Human glioma cultures and in vitro analysis of therapeu-

tic response In Berger MS and Wilson CB (Eds)The Gliomas

Philadelphia WB Saunders Co pp 134ndash142

Allen TM and Moase EH (1996) Therapeutic opportunities for targeted

liposomal drug delivery Adv Drug Deliv Rev 21 117ndash133

Arafat T Hentschel P Madajewicz S Manzione J Chowhan N Davis

R Roche P Af I Roque C Meek A and Magdy S (1999) Toxicities

related to intraarterial infusion of cisplatin and etoposide in patients with

brain tumors J Neurooncol 42 73ndash77

Bartus RT (1999) The blood-brain barrier as a target for phamacological

modulation Curr Opin Drug Discov Dev 2 152ndash167

Bickel U Yoshikawa T Landaw EM Faull KF and Pardridge WM

(1993) Pharmacologic effects in vivo in brain by vector-mediated peptide

delivery Proc Natl Acad Sci USA 90 2618ndash2622

Bigner DD Brown MT Friedman AH Coleman RE Akabani G Fried-

man HS Thorstad WL McLendon RE Bigner SH Zhao XG

Pegram CN Wikstrand CJ Herndon JE II Vick NA Paleologos N

Cokgor I Provenzale JM and Zalutsky MR (1998) Iodine-131-labeled

antitenascin monoclonal antibody 81C6 treatment of patients with recur-

rent malignant gliomas Phase I trial results J Clin Oncol 16 2202ndash2212

Black KL (1995) Biochemical opening of the blood-brain barrier Adv Drug

Deliv Rev 15 37ndash52

Neuro-Oncology n JANUARY 200056

DR Groothuis Review of BBB and BTB

Fig 6 Distribution of vasogenic edema in a malignant glioma There are four pairs of scans extending through a tumor in the left temporal lobeextending from the lowest (A) to the highest level (D) Each pair of scans contains a gadolinium-enhanced scan (left) and a T2-weighted image(right) The gadolinium-enhanced scans show the area of abnormally permeable BBB The T2-weighted images show the distribution of edema(water) emanating from the tumor The tumor may be considered a natural equivalent of a convection-enhanced delivery infusion The resultingedema spreads extensively through white matter in the ipsilateral hemisphere reaching forward in the internal capsule (levels C and D) but notpenetrating gray matter structures such as the thalamus (levels C and D) or cortex (levels AndashD) Note that the edema contacts the posterior limbof the corpus callosum in levels C and D but does not extend into the corpus callosum suggesting that there is a physiological barrier to the bulkmovement of uid in that direction

References

that rapidly crosses the BBB or BTB and is either boundto tissue elements or locally metabolized in the process ofexerting its anticancer effect There are practical prob-lems associated with ia administration however includ-ing the need for the tumor to reside within the arterialdistribution being infused and hemodynamic streamingof the administered drug Perhaps the most obvious evi-dence that increased tissue drug concentrations can beachieved with ia administration is the increased inci-dence of local toxicity (Arafat et al 1999) Neverthelessbecause most drugs being used to treat brain tumors donot have the ideal qualities to take advantage of ia deliv-ery the results of clinical trials of ia chemotherapy inbrain tumors show minimal if any improvement in sur-vival (Dropcho et al 1998 Gundersen et al 1998Hirano et al 1998)

Hyperosmolar Blood-Brain Barrier Disruption(HBBBD)

This method involves the infusion of a hyperosmolarsolution (usually 14 M mannitol) into a cerebral arterygenerally followed by the intra-arterial administration ofa drug The principle features of this method areincreased capillary permeability (caused by the hyperos-molar solution) followed by a rst pass advantage fromia infusion that is K1 is transiently increased as isAUCPL This method was originally proposed byRapoport (Rapoport and Thompson 1973) and has beenused extensively by Neuwelt and colleagues (Kroll andNeuwelt 1998 Neuwelt and Dahlborg 1989 Neuweltet al 1980 Neuwelt et al 1983) Controversy about theuse of this method arose because of differential suscepti-bility of the normal BBB as compared with the abnormalBTB Whereas all studies demonstrated increased perme-ability of the normal BBB some studies in animal tumorsfailed to demonstrate any increase in tumor permeability(Nakagawa et al 1984) others showed transient perme-ability increases (Groothuis et al 1990) and somereported signicant permeability increases (Neuwelt etal 1984) Most recently Zunkeler et al used PET tostudy the time course of BBB function in 13 patients withmalignant astrocytomas after HBBBD (Zunkeler et al1996) They conrmed the differential effect of HBBBDpermeability was increased 1000 in brain and 60 intumors The half-time of the osmotic effect was 81 minin brain and 42 min in tumor They modeled the effectsof HBBBD on methotrexate and concluded that concen-trations above 1 microM the minimal concentration requiredfor an effect from methotrexate would not be enhancedin tumor and would be enhanced only 10 in brainThus HBBBD in combination with ia administrationwould be most useful for drugs that once having crossedthe BBB bind to brain or tumor tissue and do not efuxback across the BBB In addition the differential effect ofHBBBD on normal brain compared with that of tumormust be considered when evaluating neurotoxicity

Chemical Modication of the BBB and BTB

The latest entry into the therapeutic arena is the use ofchemical agents to disrupt the BBB (Black 1995) The

agents are mostly derivatives of normal vasoactive com-pounds and include bradykinin (Inamura and Black1994) interleukin-2 (Gutman et al 1996) leukotrieneC4 (Black and Chio 1992) and others (de Vries et al1996) The principle feature of these drugs is that theyare given ia and followed by an ia injection of a thera-peutic agent RMP-7 is a commercially developedbradykinin analog that has reached clinical trials (Ford etal 1998) and acts by increasing permeability at the inter-cellular junction (Sanovick et al 1995) Studies withPET in 9 malignant gliomas have shown that permeabil-ity to 68Ga-EDTA was increased 46 plusmn 42 in tumorswithout a signicant increase in tumor-free brain (Blacket al 1997) Another report suggests that RMP-7 canincrease delivery across the normal BBB (Emerich et al1998) RMP-7 is generally administered over a 15-minperiod the timing of administration of the chemothera-peutic drug relative to the RMP-7 has varied eg as abolus after RMP-7 or alternating chemotherapeutic drugadministration with RMP-7 BBB function is restoredrapidly after the RMP-7 infusion ends continuing theinfusion for over 20 min results in tachyphylaxis withloss of responsiveness (Bartus 1999) As with ia andHBBBD administration the advantage with chemicalmodication of the BBB is limited to a fractional increasein the circulating drug to which the tumor is exposedand still represents a vanishingly small fraction of thetotal body drug exposure Like those previous tech-niques chemical modication of the BBB will be ideal fora drug with dened properties as discussed before How-ever this ideal drug has yet to be developed

Receptor-Mediated Transport

This is a very broad area because any receptor-mediatedtransport system in the BBB can be selected as a targetThe considerations about drug delivery in a facilitatedtransport system are different from those involving sim-ple diffusion First a facilitated transport system can becharacterized by the Michaelis-Menten constants Km(the concentration at which the reaction velocity is halfmaximal) and Vmax (the limiting velocity as the concen-tration approaches innity) An important factor in thefacilitated transport of drugs is the plasma concentrationof the native substrate for the receptor For examplephenylalanine crosses the BBB via the large neutral aminoacid transport (LNAA) system melphalan is achemotherapeutic drug with a phenylalanine backboneand could therefore be transported by the LNAA systemHowever the normal plasma levels of phenylalaninecompete with melphalan to such a degree that the rate ofmelphalan transport into experimental gliomas is notincreased (Groothuis et al 1992) Lowering plasmaphenylalanine levels increased the rate of melphalanentry into brain tumors but only by a modest 8 Mostof the available transport receptors are low afnity lowcapacity systems such as those for the nucleosides vari-ous peptides transferrin and insulin (Ermisch et al1993 Pardridge et al 1995 Friden et al 1996 Tamai etal 1997) Although these transport systems have beenused to increase brain delivery of oligonucleotides(Boado et al 1998 Normand-Sdiqui and Akhtar 1998)

Neuro-Oncology n JANUARY 200050

DR Groothuis Review of BBB and BTB

nerve growth factor (Friden et al 1993) and vasoactiveintestinal peptide (Bickel et al 1993) none of thesetransport systems has been explored to any extent inbrain tumors

Inhibiting Drug Efux

Throughout most of the history of the BBB movementacross the capillary for most compounds without a trans-port system has been assumed to occur by simple diffu-sion the process is passive and the rate that a compoundcrosses the barrier is the same in both directions so thatthe process is symmetrical Recent studies have begun tochallenge this idea In particular P-glycoprotein repre-sents an ATP-dependent efflux pathway that confersresistance to cancer cells by allowing them to move avariety of chemotherapeutic drugs out of the cell againsta concentration gradient (Schinkel 1999 Tsuji andTamai 1997) P-glycoprotein appears to be localized pri-marily to the luminal capillary membrane and the num-ber of drugs that are transported by this system may bequite large (Schinkel 1999) Immunohistochemically P-glycoprotein has been demonstrated in malignantglioma tumor cells (Leweke et al 1998 von Bossanyi etal 1997) Leweke et al commented that tumor endothe-lial cells stained in 20 of 21 cases and von Bossanyi et alstated that tumor blood vessels were positive in 60 oftumors PET has been used to demonstrate the reversal ofP-glycoprotein pump function in mdr1a knock-out micewith cyclosporin A (Hendrikse et al 1998) Since manychemotherapeutic drugs are transported by P-glycopro-tein this represents a logical and exciting target in braintumor chemotherapy especially because inhibition of P-glycoprotein function could occur at both the BTB andin tumor cells Inhibiting P-glycoprotein would increaseboth extracellular and intracellular drug concentration(by decreasing k2 in Equation 3) without increasing theamount of drug administered This is an avenue of modi-fying brain tumor therapy that must be explored

Avant-Garde Methods of Delivery

Included in this category are the newest methods forintravascular delivery including the use of biodegradablenanoparticles (Schroeder et al 1998 Song et al 1997)liposomes (Allen and Moase 1996 Sakamoto and Ido1993) monoglyceride-based systems (Chang et al1994) and magnetic microspheres (Chang and Bodmeier1997) Many of these novel approaches are evaluatingdifferent methods of giving drug-laden molecules thatwill preferentially accumulate in a tissue usually com-bined with selective methods of perfusion such as iadelivery For example Pulfer and Gallo delivered 1ndash2 micromia magnetic aminodextran microspheres to RG-2 ratbrain tumors and used a magnetic eld to retain the par-ticles within the tumor They showed that this methodresulted in signicant accumulation of the particles as aresult of surface charge and magnetic eld (Chang andBodmeier 1997) These methods are too new to beapplied to human brain tumors but are representative ofthe diverse and novel directions that investigators areusing to increase brain tumor delivery

Summary of Intravascular Drug Delivery

A tremendous amount of progress has been made in thepast few years in our understanding of drug delivery tobrain tumors This progress has not yet translated intoimpressive increases in survival of patients with braintumors There are major areas that remain to be exploredand developed in the arena of intravascularly adminis-tered drugs For example because ia administration(with or without HBBBD or chemical modification)offers a unique rst pass advantage a search for drugsthat have high lipid solubility and bind to brain tumortargets is likely to be protable Second increasing tissuedrug concentration by the use of efux inhibitors is obvi-ous there will be no increase in total body exposurewhile potentially achieving signicant increases in tumordrug concentration However the ultimate limiting factorfor all intravascularly administered drugs is that of thepotential toxicity from total body exposure It seemsunlikely that this important and unfortunately negativevariable can be overcome

Injection or Infusion of Drugs Directly intothe Brain or Its Cavities

Table 3 summarizes the different approaches that arebeing used to administer drugs directly into the brainthus bypassing the BBB All of these routes and methodsof administration drastically change the pharmacokineticlandscape Instead of concern about crossing the formi-dable BBB or BTB the drugs are administered directlyinto the CSF or brain extracellular space Instead of need-ing to measure the inux rate it is necessary to under-stand drug movement within the brain and to understandefux mechanisms In marked contrast to intravascularadministration 100 of the administered dose can bedelivered using these methods of administration Theproblem then becomes one of understanding the forcesthat control the movement of the drug within brain andtumor tissue One or more of four forces will affect drugmovement depending upon the particular mode of drugadministration bulk flow of CSF bulk flow of braininterstitial uid bulk ow due to the infusion of a solu-tion into the brain or CSF and diffusion Drug movementwill also be variably affected by the amount of whitematter in the target area as a result of the change fromthe highly tortuous environment of a gray matter struc-ture (isotropic) to the highly oriented ber pathways ofwhite matter (anisotropic)

Intrathecal and Intraventricular Drug Administration

In this review intrathecal drug administration refers tothe administration of drug into the subarachnoid spaceusually into the lumbar subarachnoid space whereasintraventricular administration refers to injection or infu-sion into the lateral ventricle They are discussed togetherbecause of the large number of features they have in com-mon The principle feature of drug administration bythese routes is dominated by the bulk uid ow of the

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 51

CSF The major source of CSF is from the choroidplexuses of the lateral and 4th ventricles as much asanother 30 may originate from the brain extracellularuid The direction of the ow is from the lateral ventri-cles into the 3rd and then 4th ventricles out through theforamina of Luschke and Magendie into the basal cis-terns around the convexities and out of the subarach-noid space through the arachnoid granulations In ratswhen 14C-sucrose was injected into the lateral ventriclethe sucrose distributed rapidly within 5 min it was foundin the basal cisterns and by 1 h the sucrose had largely leftthe subarachnoid space (Ghersi-Egea et al 1996) Whenthe injected agent is metabolically active it may enterselected cells (Rubertone et al 1993) In humans the nor-mal volume of CSF produced is between 400 and 500 mlper day Since the CSF spaces within the brain are about150 ml in volume there is a considerable bulk ow withinthis system although the velocity varies tremendouslyfrom site to site When isotopes are injected into the lum-bar subarachnoid space they travel up the spinal sub-arachnoid space and are found in the basal cisterns by 3 hBy 24 h they are found over the cerebral convexities CSFow from the lateral ventricles is directional isotopesinjected into the lumbar CSF do not normally enter thelateral ventricles As a result of these bulk ow pathwaysmaterials injected into the CSF will attain initial concen-trations that are directly proportional to the concentra-tion in the infusate This route of administration is idealfor situations in which the target is within the subarach-noid space (for example carcinomatous meningitis) or inwhich the target is close to the CSF brain interface Carci-nomatous meningitis has long been treated with intrathe-cal infusions including the more recent use of monoclonalantibodies (Brown et al 1996) However the capacity ofdrugs to enter the brain extracellular space from the CSFis limited Numerous studies have shown that entry intothe brain is diffusional and that concentrations declineexponentially from the brain surface (Blasberg 1977Blasberg et al 1977 Groothuis and Levy 1997 Patlak

and Fenstermacher 1975) This route of administration isboth impractical and inefcient for intraparenchymalbrain tumors

Intratumoral and Intracavitary Injection

These are discussed together and briey because they areboth limiting cases of convection-enhanced deliverywhich is discussed below These both represent localforms of drug administration in which the distributionwill be determined initially by the rate and duration ofthe injection and secondly by tissue bulk ow pathwaysand diffusion The history of intratumoral injection wasdiscussed by Tomita (1991) who recognized the poten-tial usefulness of this approach as well as the importanceof neurotoxicity as the limiting factor Intratumoral andintracavitary injections are being widely explored asmeans of delivering large agents such as viral vectorsgrowth factors and cells (Bigner et al 1998 Farkkila etal 1994 Hsiao et al 1997 Puri et al 1996 Ram et al1997 Yang et al 1997) All of the attributes discussed inthe section on CED will also apply to these forms ofdrug delivery

Microdialysis

Microdialysis is a method that employs the passive diffu-sion of a drug across a semipermeable membrane (deLange et al 1997 de Lange et al 1999 Parsons andJustice 1994) It can be used to both sample and deliverdrugs to the surrounding tissue and has been used tosample extracellular drug concentrations in experimentalbrain tumors (Devineni et al 1996 Nakashima et al1997) The principle features of microdialysis when usedas a drug delivery methodology are that the tissue con-centrations are a function of the drug concentration inthe dialysate (which places control of the concentrationin hands of the user) and that distribution of drug awayfrom the dialysis catheter occurs by diffusion (Dykstra et

Neuro-Oncology n JANUARY 200052

DR Groothuis Review of BBB and BTB

Table 3 Comparison of different methods for local drug delivery to brain tumors

Method of circumventing the BBB Principle advantages Principle disadvantages Limiting factor(s)

Intrathecal Easy access ideal for therapy of Not useful for parenchymal Bulk ow rate of CSFmeningeal disease disease

Intraventricular administration Easy access ideal for therapy of Not useful for parenchymal Bulk ow rate of CSFmeningeal disease disease

Intratumoral injection User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Intracavitary injection User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Microdialysis User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Biodegradable polymers User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Convection-enhanced delivery User control over administered Invasive Unpredictable distribution dose 100 reaches target neurotoxicity

Abbreviations BBB blood-brain barrier CSF cerebrospinal uid

This table lists the different approaches that are available to deliver therapeutic drugs and agents by local administration The advantages and disadvantages are an expression of theauthorrsquos opinion about the most prominent feature of each method Details about each method may be found in the text

al 1992) However the normal BBB is disrupted imme-diately around the catheter and stays disrupted for sometime after catheter insertion making drug efux in thisvicinity an additional problem when trying to understandlocal delivery kinetics (Groothuis et al 1998 Morgan etal 1996 Westergren et al 1995) Because the principlefactors in local tissue concentrations are diffusion andorlocal brain bulk flow patterns the volume of brainreached by microdialysis will be relatively small anddrug concentrations will in general decline exponen-tially away from the dialysis cannula This method willbe applicable when the volume of tissue that needs to bereached is small The effect of normal brain bulk owpathways on this delivery method remains to be studied

Biodegradable Polymers

This represents a local delivery system in whichbiodegradable polymers are loaded with drug andimplanted into brain or tumor tissue resulting in timed-release of drug (Cao and Shoichet 1999 Menei et al1997) The principle features of this system are that mul-tiple polymer pellets or wafers can be implanted whichgives control over the spatial distribution and that drugdistribution away from the source occurs by diffusionwith a variable contribution from the inherent bulk owpathways of the brain (Fung et al 1998) This approachhas been used in patients with malignant gliomas whereits safety and efcacy have been demonstrated (Brem etal 1995 Olivi and Brem 1994) As with microdialysisthe principle limiting feature of this method of drug deliv-ery is that drug distribution occurs mainly by diffusionin which drug concentrations fall exponentially awayfrom the source and are generally limited to a few mil-limeters of tissue penetration (Fung et al 1998)

Convection-Enhanced Delivery

This is a relatively new method of drug delivery to braintumors The principle feature of this delivery method isthat a drug solution is infused directly into the brain Theconcentration of the drug is independent of the infusionrate and duration and the distribution of the drug is afunction of the hydrostatic pressure of the infusion aswell as diffusion into surrounding tissue From a deliveryperspective the only limiting factor to the drug concen-tration that can be delivered is the solubility of the drugCED has been discussed from a theoretical perspective(Morrison et al 1994) it has been explored in severalanimal models (Bobo et al 1994 Groothuis et al 1999Laske et al 1997a Lieberman et al 1995 Viola et al1995) and it has been used for delivery of a targetedtoxin in human gliomas (Laske et al 1997b) The distri-bution of the infused drug is highly dependent uponwhether the drug is infused into a homogeneous graymatter structure or a structure containing ber pathwaysIn the case of the infusion of a drug-containing solutioninto a homogeneous brain structure the infusion initiallyproduces a spherical volume in which the concentrationof the drug is directly proportional to that of the infusate(Fig 2b) After the infusion reaches equilibrium at whichpoint the amount of drug leaving across brain capillaries

is equal to the amount being infused the drug begins todiffuse away from the edge of the sphere at concentra-tions that decline exponentially in accordance with themathematics of diffusion (Fig 2b) If the infusionencounters an organized white matter pathway the rateof movement is either increased or decreased (dependingupon the direction of the inherent bulk ow within thatpathway) (Fig 2c) Morrison stated that when the infu-sion is being made into a homogeneous isotropicmedium the radius of the central spherical componentmdashthe convective componentmdashcan be related to the infusionrate and the efux constant of the drug being infused

rp 5 3 2qv (4p K) (Eq 7)

where qv is the infusion rate and K represents thesteady-state efux rate constant Fig 3 shows a plot of theradius of the convective component as a function of infu-sion rate and efux constant From reviewing Fig 3

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 53

Fig 2 The distribution of 14C-sucrose in rat brain by convection-enhanced delivery The diagram in the upper left (A) shows the targetlocation (center of the caudate nucleus) as well as the organization ofthe brain at the target level After a 1-h 05-microlmin infusion the iso-tope is found in a spherical shape with even distribution across (B) By8 h the isotope is moving into the external capsule (C) A similarappearance is found after a constant infusion over 7 days the centralconvective component is clearly visible surrounded by decreasingconcentration from the edge (D) Note the isotopersquos rapid lateralmovement down the external capsule (probably representing a nor-mal brain bulk uid pathway) but not into the corpus callosum Infu-sion into rat brain for 7 days produced no neuropathologic changesother than those associated with catheter insertion

which extends from infusion rates provided by osmoticmini-pumps (1 microlh) to high infusion rates (10 microlmin) it is

apparent that the volume of affected brain remains quitesmall for most infusion rates Similarly as the efux con-stant decreases indicating slower rates of crossing theBBB the convection volume increases Fig 3 illustratesthat CED is best suited for delivery of large compounds(with low efux rates) and delivery at moderately highinfusion rates if the intent is to reach large brain volumes

The shape of the volume of distribution at steady statewill be highly dependent on the location in the brain thatis being infused Fig 4 shows a CT scan of a dog brainreceiving an infusion of iodinated contrast into whitematter The iodinated contrast rapidly distributes in thewhite matter of the ipsilateral hemisphere with rapidlydecreasing concentrations in the cortex on the affectedside Note that in both Figs 2 and 4 the infused materialdoes not cross the midline through the corpus callosumThis suggests that the direction of the normal bulk owpathway within the corpus callosum is away from themidline thus opposing the CED infusion component Incontrast to intravascularly administered drugs whereincreased capillary permeability results in increased tissueconcentrations (Table 1) the opposite effect will occur inCED infusions Increased efux across permeable tumorcapillaries will reduce both the extracellular drug concen-tration and the volume of distribution In Fig 5 highconcentrations of 14C-sucrose were achieved by a CEDinfusion into an RG-2 glioma but at a distance of 24mm from the infusion site tissue concentrationsdecreased by several logarithmic units Although studieshave not been done in humans it is easy to anticipatethat the volume of distribution from a CED infusion willfollow similar principles Fig 6 shows several levels of anMRI scan through a malignant glioma Each set of scansshows a gadolinium-enhanced tumor margin (which canbe considered naturersquos equivalent to a CED infusion) anda T2-weighted image (which corresponds to the extracel-lular water originating from the tumor source) The

Neuro-Oncology n JANUARY 200054

DR Groothuis Review of BBB and BTB

Fig 3 Relationship between radius of convective CED componentinfusion rate and efux rate The radius of the inection pointbetween the convective and diffusional components of CED infusionsis shown for infusion into a homogeneous brain structure (Equation7) The values on the vertical axis represent the radius inside whichthe local concentrations remain uniform and outside of which drugconcentrations decline exponentially Changes for each 5-cm incre-ment are represented by different shades of gray The infusion ratewas chosen from 1 acute 10ndash6 to 1 acute 10ndash2 ml per minndash1 The lower valuesare those seen with infusions from osmotic mini-pumps while thehigher values represent the upper limit of CED infusion rates in dogsThe efux rate was chosen to vary from 1 acute 10ndash1 to 1 acute 10ndash7 minndash1The volume of the convective component increases as a function ofboth increasing infusion rate and decreasing efux rate

Fig 4 Illustration of CED infusion in a dog The gure on the left shows the CT scan at the start of a 3 microl minndash1 infusion of Isovue (iopamidol)into the hemispheric white matter of a dog while that on the right shows the same coronal level on the same dog after 10 days of infusion Thescan on the left contains an air bubble in the lateral ventricle the cannula tip contains a metal marker (the high density circle) and a small amountof Isovue surrounding the catheter tip In the scan on the right the Isovue has distributed at a high concentration throughout the white matterof the ipsilateral hemisphere with a rapid decline in concentration at gray-white interfaces Note that the distribution of Isovue is limited to theipsilateral hemisphere No neuropathologic changes were observed as a result of the infusion

shape and distribution of the edema are highly irregularand very dependent upon the brain structures encoun-tered Note that the edema uid moves into the internalcapsule but not into the thalamus or cortex Notice alsothat the edema uid does not cross the posterior limb ofthe corpus callosum although it comes into contact withit at several levels

A dominant feature of CED is that distribution of thedrug is determined by the infusion parameters and thetype of tissue being infused whereas the concentration ofdrug in the infusate is unrelated to these parameters Themaximum concentration of the drug in the infusate willbe determined by neurotoxicity within the central con-vective component of the infusion with variable concen-trations in the anisotropic brain surrounding the centralcomponent and concentrations 50 to 100 times lower inthe diffusional component furthermost from the infusioncatheter The ability to manipulate drug concentration inbrain and tumor tissue with such ease and to maintainthese manipulations for long periods of time (Groothuiset al 1999) brings a need for the development ofentirely new concepts For example the limits withregard to the composition of the infusate need to beexplored such as pH osmolarity and ionic compositionFurthermore how does one reliably study neurotoxicityAre infusions into animal models going to be reliable pre-dictors of toxicity in humans Will differential neurotox-icity become an issue It is possible that neuronsastrocytes and oligodendrogliocytes will display differ-ent levels of susceptibility to many drugs Will we seedelayed neurotoxicity such as that seen with radiation Itwill probably be best for many of the variables associatedwith neurotoxicity to be explored in animals before sub-jecting humans to CED infusions

Summary of Drug Delivery that Circumvents the BBB

There has been an explosion of methodology involvinglocal tissue drug delivery during the past 10 years Someof these techniques such as CED fulll the promise tocircumvent the BBB entirely In fullling this promisethese methods introduce new problems such as the rela-tionship between the spatial distribution of drugs andneurotoxicity that need to be understood before thesetechniques are widely used However for viral vectorsmonoclonal antibodies antisense oligonucleotides andother therapeutic agents that will be unable to cross theBBB in therapeutically effective amounts CED may pro-vide the necessary delivery tool

The State of Drug Delivery to Brain Tumorsndash1999

If we return to the statement of Vick et al at the begin-ning of this review we can now state with certainty thatthe BBB and BTB are major factors limiting the access ofmany therapeutic agents to brain tumors and these bar-riers will become even more signicant with the develop-ment of new molecular biological therapies that willinvolve large molecules viruses and even cells It is alsoremarkable how prescient those authors were in identify-ing the other parts of the drug delivery process that mustnow be understood and how little progress we havemade in elucidating ldquotumor cell uptake metabolic fatewithin tumor cells and the washout or sink effect of theextracellular space and CSFrdquo (Vick et al 1977) Formost of the past 20 years we have focused so much on

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 55

Fig 5 Infusion of 14C-sucrose into an RG-2 rat glioma An infusion cannula was placed into the caudate nucleus of a Fisher-344 rat (targetshown in Fig 2A) and RG-2 cells were injected Ten days later 14C-sucrose was infused at a rate of 10 microlh for 7 days Autoradiographic imageswere made (right images) and the sections were then stained with hematoxylin-eosin (left images) The top two images show that the 14C-sucrose distributes in high concentration throughout the RG-2 tumor extending into the edematous external capsule (E) However thelower two sections taken from 24 mm behind the infusion site still contain tumor and show that the concentration of 14C-sucrose is 100000times lower These sections illustrate the problems associated with controlling drug delivery by convection-enhanceddelivery

ways to increase delivery across the BBB that we havefailed to address these other issues which now becomethe forefront in drug delivery methods such as CED Thearsenal from which to choose a drug delivery method istruly impressive We must now study the other factorsthat limit effective brain tumor therapy Perhaps the mostpressing of these includes the need to develop effective

laboratory tools for identifying the relationship betweenthe concentration-time product of therapeutic agents andtumor cell kill rather than relying on empirical testing inhuman subjects It is most likely that the new century willsee the development and application of tools for individ-ualizing chemotherapy for brain tumors and other solidhuman tumors

Ali-Osman F (1999) Human glioma cultures and in vitro analysis of therapeu-

tic response In Berger MS and Wilson CB (Eds)The Gliomas

Philadelphia WB Saunders Co pp 134ndash142

Allen TM and Moase EH (1996) Therapeutic opportunities for targeted

liposomal drug delivery Adv Drug Deliv Rev 21 117ndash133

Arafat T Hentschel P Madajewicz S Manzione J Chowhan N Davis

R Roche P Af I Roque C Meek A and Magdy S (1999) Toxicities

related to intraarterial infusion of cisplatin and etoposide in patients with

brain tumors J Neurooncol 42 73ndash77

Bartus RT (1999) The blood-brain barrier as a target for phamacological

modulation Curr Opin Drug Discov Dev 2 152ndash167

Bickel U Yoshikawa T Landaw EM Faull KF and Pardridge WM

(1993) Pharmacologic effects in vivo in brain by vector-mediated peptide

delivery Proc Natl Acad Sci USA 90 2618ndash2622

Bigner DD Brown MT Friedman AH Coleman RE Akabani G Fried-

man HS Thorstad WL McLendon RE Bigner SH Zhao XG

Pegram CN Wikstrand CJ Herndon JE II Vick NA Paleologos N

Cokgor I Provenzale JM and Zalutsky MR (1998) Iodine-131-labeled

antitenascin monoclonal antibody 81C6 treatment of patients with recur-

rent malignant gliomas Phase I trial results J Clin Oncol 16 2202ndash2212

Black KL (1995) Biochemical opening of the blood-brain barrier Adv Drug

Deliv Rev 15 37ndash52

Neuro-Oncology n JANUARY 200056

DR Groothuis Review of BBB and BTB

Fig 6 Distribution of vasogenic edema in a malignant glioma There are four pairs of scans extending through a tumor in the left temporal lobeextending from the lowest (A) to the highest level (D) Each pair of scans contains a gadolinium-enhanced scan (left) and a T2-weighted image(right) The gadolinium-enhanced scans show the area of abnormally permeable BBB The T2-weighted images show the distribution of edema(water) emanating from the tumor The tumor may be considered a natural equivalent of a convection-enhanced delivery infusion The resultingedema spreads extensively through white matter in the ipsilateral hemisphere reaching forward in the internal capsule (levels C and D) but notpenetrating gray matter structures such as the thalamus (levels C and D) or cortex (levels AndashD) Note that the edema contacts the posterior limbof the corpus callosum in levels C and D but does not extend into the corpus callosum suggesting that there is a physiological barrier to the bulkmovement of uid in that direction

References

nerve growth factor (Friden et al 1993) and vasoactiveintestinal peptide (Bickel et al 1993) none of thesetransport systems has been explored to any extent inbrain tumors

Inhibiting Drug Efux

Throughout most of the history of the BBB movementacross the capillary for most compounds without a trans-port system has been assumed to occur by simple diffu-sion the process is passive and the rate that a compoundcrosses the barrier is the same in both directions so thatthe process is symmetrical Recent studies have begun tochallenge this idea In particular P-glycoprotein repre-sents an ATP-dependent efflux pathway that confersresistance to cancer cells by allowing them to move avariety of chemotherapeutic drugs out of the cell againsta concentration gradient (Schinkel 1999 Tsuji andTamai 1997) P-glycoprotein appears to be localized pri-marily to the luminal capillary membrane and the num-ber of drugs that are transported by this system may bequite large (Schinkel 1999) Immunohistochemically P-glycoprotein has been demonstrated in malignantglioma tumor cells (Leweke et al 1998 von Bossanyi etal 1997) Leweke et al commented that tumor endothe-lial cells stained in 20 of 21 cases and von Bossanyi et alstated that tumor blood vessels were positive in 60 oftumors PET has been used to demonstrate the reversal ofP-glycoprotein pump function in mdr1a knock-out micewith cyclosporin A (Hendrikse et al 1998) Since manychemotherapeutic drugs are transported by P-glycopro-tein this represents a logical and exciting target in braintumor chemotherapy especially because inhibition of P-glycoprotein function could occur at both the BTB andin tumor cells Inhibiting P-glycoprotein would increaseboth extracellular and intracellular drug concentration(by decreasing k2 in Equation 3) without increasing theamount of drug administered This is an avenue of modi-fying brain tumor therapy that must be explored

Avant-Garde Methods of Delivery

Included in this category are the newest methods forintravascular delivery including the use of biodegradablenanoparticles (Schroeder et al 1998 Song et al 1997)liposomes (Allen and Moase 1996 Sakamoto and Ido1993) monoglyceride-based systems (Chang et al1994) and magnetic microspheres (Chang and Bodmeier1997) Many of these novel approaches are evaluatingdifferent methods of giving drug-laden molecules thatwill preferentially accumulate in a tissue usually com-bined with selective methods of perfusion such as iadelivery For example Pulfer and Gallo delivered 1ndash2 micromia magnetic aminodextran microspheres to RG-2 ratbrain tumors and used a magnetic eld to retain the par-ticles within the tumor They showed that this methodresulted in signicant accumulation of the particles as aresult of surface charge and magnetic eld (Chang andBodmeier 1997) These methods are too new to beapplied to human brain tumors but are representative ofthe diverse and novel directions that investigators areusing to increase brain tumor delivery

Summary of Intravascular Drug Delivery

A tremendous amount of progress has been made in thepast few years in our understanding of drug delivery tobrain tumors This progress has not yet translated intoimpressive increases in survival of patients with braintumors There are major areas that remain to be exploredand developed in the arena of intravascularly adminis-tered drugs For example because ia administration(with or without HBBBD or chemical modification)offers a unique rst pass advantage a search for drugsthat have high lipid solubility and bind to brain tumortargets is likely to be protable Second increasing tissuedrug concentration by the use of efux inhibitors is obvi-ous there will be no increase in total body exposurewhile potentially achieving signicant increases in tumordrug concentration However the ultimate limiting factorfor all intravascularly administered drugs is that of thepotential toxicity from total body exposure It seemsunlikely that this important and unfortunately negativevariable can be overcome

Injection or Infusion of Drugs Directly intothe Brain or Its Cavities

Table 3 summarizes the different approaches that arebeing used to administer drugs directly into the brainthus bypassing the BBB All of these routes and methodsof administration drastically change the pharmacokineticlandscape Instead of concern about crossing the formi-dable BBB or BTB the drugs are administered directlyinto the CSF or brain extracellular space Instead of need-ing to measure the inux rate it is necessary to under-stand drug movement within the brain and to understandefux mechanisms In marked contrast to intravascularadministration 100 of the administered dose can bedelivered using these methods of administration Theproblem then becomes one of understanding the forcesthat control the movement of the drug within brain andtumor tissue One or more of four forces will affect drugmovement depending upon the particular mode of drugadministration bulk flow of CSF bulk flow of braininterstitial uid bulk ow due to the infusion of a solu-tion into the brain or CSF and diffusion Drug movementwill also be variably affected by the amount of whitematter in the target area as a result of the change fromthe highly tortuous environment of a gray matter struc-ture (isotropic) to the highly oriented ber pathways ofwhite matter (anisotropic)

Intrathecal and Intraventricular Drug Administration

In this review intrathecal drug administration refers tothe administration of drug into the subarachnoid spaceusually into the lumbar subarachnoid space whereasintraventricular administration refers to injection or infu-sion into the lateral ventricle They are discussed togetherbecause of the large number of features they have in com-mon The principle feature of drug administration bythese routes is dominated by the bulk uid ow of the

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 51

CSF The major source of CSF is from the choroidplexuses of the lateral and 4th ventricles as much asanother 30 may originate from the brain extracellularuid The direction of the ow is from the lateral ventri-cles into the 3rd and then 4th ventricles out through theforamina of Luschke and Magendie into the basal cis-terns around the convexities and out of the subarach-noid space through the arachnoid granulations In ratswhen 14C-sucrose was injected into the lateral ventriclethe sucrose distributed rapidly within 5 min it was foundin the basal cisterns and by 1 h the sucrose had largely leftthe subarachnoid space (Ghersi-Egea et al 1996) Whenthe injected agent is metabolically active it may enterselected cells (Rubertone et al 1993) In humans the nor-mal volume of CSF produced is between 400 and 500 mlper day Since the CSF spaces within the brain are about150 ml in volume there is a considerable bulk ow withinthis system although the velocity varies tremendouslyfrom site to site When isotopes are injected into the lum-bar subarachnoid space they travel up the spinal sub-arachnoid space and are found in the basal cisterns by 3 hBy 24 h they are found over the cerebral convexities CSFow from the lateral ventricles is directional isotopesinjected into the lumbar CSF do not normally enter thelateral ventricles As a result of these bulk ow pathwaysmaterials injected into the CSF will attain initial concen-trations that are directly proportional to the concentra-tion in the infusate This route of administration is idealfor situations in which the target is within the subarach-noid space (for example carcinomatous meningitis) or inwhich the target is close to the CSF brain interface Carci-nomatous meningitis has long been treated with intrathe-cal infusions including the more recent use of monoclonalantibodies (Brown et al 1996) However the capacity ofdrugs to enter the brain extracellular space from the CSFis limited Numerous studies have shown that entry intothe brain is diffusional and that concentrations declineexponentially from the brain surface (Blasberg 1977Blasberg et al 1977 Groothuis and Levy 1997 Patlak

and Fenstermacher 1975) This route of administration isboth impractical and inefcient for intraparenchymalbrain tumors

Intratumoral and Intracavitary Injection

These are discussed together and briey because they areboth limiting cases of convection-enhanced deliverywhich is discussed below These both represent localforms of drug administration in which the distributionwill be determined initially by the rate and duration ofthe injection and secondly by tissue bulk ow pathwaysand diffusion The history of intratumoral injection wasdiscussed by Tomita (1991) who recognized the poten-tial usefulness of this approach as well as the importanceof neurotoxicity as the limiting factor Intratumoral andintracavitary injections are being widely explored asmeans of delivering large agents such as viral vectorsgrowth factors and cells (Bigner et al 1998 Farkkila etal 1994 Hsiao et al 1997 Puri et al 1996 Ram et al1997 Yang et al 1997) All of the attributes discussed inthe section on CED will also apply to these forms ofdrug delivery

Microdialysis

Microdialysis is a method that employs the passive diffu-sion of a drug across a semipermeable membrane (deLange et al 1997 de Lange et al 1999 Parsons andJustice 1994) It can be used to both sample and deliverdrugs to the surrounding tissue and has been used tosample extracellular drug concentrations in experimentalbrain tumors (Devineni et al 1996 Nakashima et al1997) The principle features of microdialysis when usedas a drug delivery methodology are that the tissue con-centrations are a function of the drug concentration inthe dialysate (which places control of the concentrationin hands of the user) and that distribution of drug awayfrom the dialysis catheter occurs by diffusion (Dykstra et

Neuro-Oncology n JANUARY 200052

DR Groothuis Review of BBB and BTB

Table 3 Comparison of different methods for local drug delivery to brain tumors

Method of circumventing the BBB Principle advantages Principle disadvantages Limiting factor(s)

Intrathecal Easy access ideal for therapy of Not useful for parenchymal Bulk ow rate of CSFmeningeal disease disease

Intraventricular administration Easy access ideal for therapy of Not useful for parenchymal Bulk ow rate of CSFmeningeal disease disease

Intratumoral injection User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Intracavitary injection User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Microdialysis User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Biodegradable polymers User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Convection-enhanced delivery User control over administered Invasive Unpredictable distribution dose 100 reaches target neurotoxicity

Abbreviations BBB blood-brain barrier CSF cerebrospinal uid

This table lists the different approaches that are available to deliver therapeutic drugs and agents by local administration The advantages and disadvantages are an expression of theauthorrsquos opinion about the most prominent feature of each method Details about each method may be found in the text

al 1992) However the normal BBB is disrupted imme-diately around the catheter and stays disrupted for sometime after catheter insertion making drug efux in thisvicinity an additional problem when trying to understandlocal delivery kinetics (Groothuis et al 1998 Morgan etal 1996 Westergren et al 1995) Because the principlefactors in local tissue concentrations are diffusion andorlocal brain bulk flow patterns the volume of brainreached by microdialysis will be relatively small anddrug concentrations will in general decline exponen-tially away from the dialysis cannula This method willbe applicable when the volume of tissue that needs to bereached is small The effect of normal brain bulk owpathways on this delivery method remains to be studied

Biodegradable Polymers

This represents a local delivery system in whichbiodegradable polymers are loaded with drug andimplanted into brain or tumor tissue resulting in timed-release of drug (Cao and Shoichet 1999 Menei et al1997) The principle features of this system are that mul-tiple polymer pellets or wafers can be implanted whichgives control over the spatial distribution and that drugdistribution away from the source occurs by diffusionwith a variable contribution from the inherent bulk owpathways of the brain (Fung et al 1998) This approachhas been used in patients with malignant gliomas whereits safety and efcacy have been demonstrated (Brem etal 1995 Olivi and Brem 1994) As with microdialysisthe principle limiting feature of this method of drug deliv-ery is that drug distribution occurs mainly by diffusionin which drug concentrations fall exponentially awayfrom the source and are generally limited to a few mil-limeters of tissue penetration (Fung et al 1998)

Convection-Enhanced Delivery

This is a relatively new method of drug delivery to braintumors The principle feature of this delivery method isthat a drug solution is infused directly into the brain Theconcentration of the drug is independent of the infusionrate and duration and the distribution of the drug is afunction of the hydrostatic pressure of the infusion aswell as diffusion into surrounding tissue From a deliveryperspective the only limiting factor to the drug concen-tration that can be delivered is the solubility of the drugCED has been discussed from a theoretical perspective(Morrison et al 1994) it has been explored in severalanimal models (Bobo et al 1994 Groothuis et al 1999Laske et al 1997a Lieberman et al 1995 Viola et al1995) and it has been used for delivery of a targetedtoxin in human gliomas (Laske et al 1997b) The distri-bution of the infused drug is highly dependent uponwhether the drug is infused into a homogeneous graymatter structure or a structure containing ber pathwaysIn the case of the infusion of a drug-containing solutioninto a homogeneous brain structure the infusion initiallyproduces a spherical volume in which the concentrationof the drug is directly proportional to that of the infusate(Fig 2b) After the infusion reaches equilibrium at whichpoint the amount of drug leaving across brain capillaries

is equal to the amount being infused the drug begins todiffuse away from the edge of the sphere at concentra-tions that decline exponentially in accordance with themathematics of diffusion (Fig 2b) If the infusionencounters an organized white matter pathway the rateof movement is either increased or decreased (dependingupon the direction of the inherent bulk ow within thatpathway) (Fig 2c) Morrison stated that when the infu-sion is being made into a homogeneous isotropicmedium the radius of the central spherical componentmdashthe convective componentmdashcan be related to the infusionrate and the efux constant of the drug being infused

rp 5 3 2qv (4p K) (Eq 7)

where qv is the infusion rate and K represents thesteady-state efux rate constant Fig 3 shows a plot of theradius of the convective component as a function of infu-sion rate and efux constant From reviewing Fig 3

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 53

Fig 2 The distribution of 14C-sucrose in rat brain by convection-enhanced delivery The diagram in the upper left (A) shows the targetlocation (center of the caudate nucleus) as well as the organization ofthe brain at the target level After a 1-h 05-microlmin infusion the iso-tope is found in a spherical shape with even distribution across (B) By8 h the isotope is moving into the external capsule (C) A similarappearance is found after a constant infusion over 7 days the centralconvective component is clearly visible surrounded by decreasingconcentration from the edge (D) Note the isotopersquos rapid lateralmovement down the external capsule (probably representing a nor-mal brain bulk uid pathway) but not into the corpus callosum Infu-sion into rat brain for 7 days produced no neuropathologic changesother than those associated with catheter insertion

which extends from infusion rates provided by osmoticmini-pumps (1 microlh) to high infusion rates (10 microlmin) it is

apparent that the volume of affected brain remains quitesmall for most infusion rates Similarly as the efux con-stant decreases indicating slower rates of crossing theBBB the convection volume increases Fig 3 illustratesthat CED is best suited for delivery of large compounds(with low efux rates) and delivery at moderately highinfusion rates if the intent is to reach large brain volumes

The shape of the volume of distribution at steady statewill be highly dependent on the location in the brain thatis being infused Fig 4 shows a CT scan of a dog brainreceiving an infusion of iodinated contrast into whitematter The iodinated contrast rapidly distributes in thewhite matter of the ipsilateral hemisphere with rapidlydecreasing concentrations in the cortex on the affectedside Note that in both Figs 2 and 4 the infused materialdoes not cross the midline through the corpus callosumThis suggests that the direction of the normal bulk owpathway within the corpus callosum is away from themidline thus opposing the CED infusion component Incontrast to intravascularly administered drugs whereincreased capillary permeability results in increased tissueconcentrations (Table 1) the opposite effect will occur inCED infusions Increased efux across permeable tumorcapillaries will reduce both the extracellular drug concen-tration and the volume of distribution In Fig 5 highconcentrations of 14C-sucrose were achieved by a CEDinfusion into an RG-2 glioma but at a distance of 24mm from the infusion site tissue concentrationsdecreased by several logarithmic units Although studieshave not been done in humans it is easy to anticipatethat the volume of distribution from a CED infusion willfollow similar principles Fig 6 shows several levels of anMRI scan through a malignant glioma Each set of scansshows a gadolinium-enhanced tumor margin (which canbe considered naturersquos equivalent to a CED infusion) anda T2-weighted image (which corresponds to the extracel-lular water originating from the tumor source) The

Neuro-Oncology n JANUARY 200054

DR Groothuis Review of BBB and BTB

Fig 3 Relationship between radius of convective CED componentinfusion rate and efux rate The radius of the inection pointbetween the convective and diffusional components of CED infusionsis shown for infusion into a homogeneous brain structure (Equation7) The values on the vertical axis represent the radius inside whichthe local concentrations remain uniform and outside of which drugconcentrations decline exponentially Changes for each 5-cm incre-ment are represented by different shades of gray The infusion ratewas chosen from 1 acute 10ndash6 to 1 acute 10ndash2 ml per minndash1 The lower valuesare those seen with infusions from osmotic mini-pumps while thehigher values represent the upper limit of CED infusion rates in dogsThe efux rate was chosen to vary from 1 acute 10ndash1 to 1 acute 10ndash7 minndash1The volume of the convective component increases as a function ofboth increasing infusion rate and decreasing efux rate

Fig 4 Illustration of CED infusion in a dog The gure on the left shows the CT scan at the start of a 3 microl minndash1 infusion of Isovue (iopamidol)into the hemispheric white matter of a dog while that on the right shows the same coronal level on the same dog after 10 days of infusion Thescan on the left contains an air bubble in the lateral ventricle the cannula tip contains a metal marker (the high density circle) and a small amountof Isovue surrounding the catheter tip In the scan on the right the Isovue has distributed at a high concentration throughout the white matterof the ipsilateral hemisphere with a rapid decline in concentration at gray-white interfaces Note that the distribution of Isovue is limited to theipsilateral hemisphere No neuropathologic changes were observed as a result of the infusion

shape and distribution of the edema are highly irregularand very dependent upon the brain structures encoun-tered Note that the edema uid moves into the internalcapsule but not into the thalamus or cortex Notice alsothat the edema uid does not cross the posterior limb ofthe corpus callosum although it comes into contact withit at several levels

A dominant feature of CED is that distribution of thedrug is determined by the infusion parameters and thetype of tissue being infused whereas the concentration ofdrug in the infusate is unrelated to these parameters Themaximum concentration of the drug in the infusate willbe determined by neurotoxicity within the central con-vective component of the infusion with variable concen-trations in the anisotropic brain surrounding the centralcomponent and concentrations 50 to 100 times lower inthe diffusional component furthermost from the infusioncatheter The ability to manipulate drug concentration inbrain and tumor tissue with such ease and to maintainthese manipulations for long periods of time (Groothuiset al 1999) brings a need for the development ofentirely new concepts For example the limits withregard to the composition of the infusate need to beexplored such as pH osmolarity and ionic compositionFurthermore how does one reliably study neurotoxicityAre infusions into animal models going to be reliable pre-dictors of toxicity in humans Will differential neurotox-icity become an issue It is possible that neuronsastrocytes and oligodendrogliocytes will display differ-ent levels of susceptibility to many drugs Will we seedelayed neurotoxicity such as that seen with radiation Itwill probably be best for many of the variables associatedwith neurotoxicity to be explored in animals before sub-jecting humans to CED infusions

Summary of Drug Delivery that Circumvents the BBB

There has been an explosion of methodology involvinglocal tissue drug delivery during the past 10 years Someof these techniques such as CED fulll the promise tocircumvent the BBB entirely In fullling this promisethese methods introduce new problems such as the rela-tionship between the spatial distribution of drugs andneurotoxicity that need to be understood before thesetechniques are widely used However for viral vectorsmonoclonal antibodies antisense oligonucleotides andother therapeutic agents that will be unable to cross theBBB in therapeutically effective amounts CED may pro-vide the necessary delivery tool

The State of Drug Delivery to Brain Tumorsndash1999

If we return to the statement of Vick et al at the begin-ning of this review we can now state with certainty thatthe BBB and BTB are major factors limiting the access ofmany therapeutic agents to brain tumors and these bar-riers will become even more signicant with the develop-ment of new molecular biological therapies that willinvolve large molecules viruses and even cells It is alsoremarkable how prescient those authors were in identify-ing the other parts of the drug delivery process that mustnow be understood and how little progress we havemade in elucidating ldquotumor cell uptake metabolic fatewithin tumor cells and the washout or sink effect of theextracellular space and CSFrdquo (Vick et al 1977) Formost of the past 20 years we have focused so much on

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 55

Fig 5 Infusion of 14C-sucrose into an RG-2 rat glioma An infusion cannula was placed into the caudate nucleus of a Fisher-344 rat (targetshown in Fig 2A) and RG-2 cells were injected Ten days later 14C-sucrose was infused at a rate of 10 microlh for 7 days Autoradiographic imageswere made (right images) and the sections were then stained with hematoxylin-eosin (left images) The top two images show that the 14C-sucrose distributes in high concentration throughout the RG-2 tumor extending into the edematous external capsule (E) However thelower two sections taken from 24 mm behind the infusion site still contain tumor and show that the concentration of 14C-sucrose is 100000times lower These sections illustrate the problems associated with controlling drug delivery by convection-enhanceddelivery

ways to increase delivery across the BBB that we havefailed to address these other issues which now becomethe forefront in drug delivery methods such as CED Thearsenal from which to choose a drug delivery method istruly impressive We must now study the other factorsthat limit effective brain tumor therapy Perhaps the mostpressing of these includes the need to develop effective

laboratory tools for identifying the relationship betweenthe concentration-time product of therapeutic agents andtumor cell kill rather than relying on empirical testing inhuman subjects It is most likely that the new century willsee the development and application of tools for individ-ualizing chemotherapy for brain tumors and other solidhuman tumors

Ali-Osman F (1999) Human glioma cultures and in vitro analysis of therapeu-

tic response In Berger MS and Wilson CB (Eds)The Gliomas

Philadelphia WB Saunders Co pp 134ndash142

Allen TM and Moase EH (1996) Therapeutic opportunities for targeted

liposomal drug delivery Adv Drug Deliv Rev 21 117ndash133

Arafat T Hentschel P Madajewicz S Manzione J Chowhan N Davis

R Roche P Af I Roque C Meek A and Magdy S (1999) Toxicities

related to intraarterial infusion of cisplatin and etoposide in patients with

brain tumors J Neurooncol 42 73ndash77

Bartus RT (1999) The blood-brain barrier as a target for phamacological

modulation Curr Opin Drug Discov Dev 2 152ndash167

Bickel U Yoshikawa T Landaw EM Faull KF and Pardridge WM

(1993) Pharmacologic effects in vivo in brain by vector-mediated peptide

delivery Proc Natl Acad Sci USA 90 2618ndash2622

Bigner DD Brown MT Friedman AH Coleman RE Akabani G Fried-

man HS Thorstad WL McLendon RE Bigner SH Zhao XG

Pegram CN Wikstrand CJ Herndon JE II Vick NA Paleologos N

Cokgor I Provenzale JM and Zalutsky MR (1998) Iodine-131-labeled

antitenascin monoclonal antibody 81C6 treatment of patients with recur-

rent malignant gliomas Phase I trial results J Clin Oncol 16 2202ndash2212

Black KL (1995) Biochemical opening of the blood-brain barrier Adv Drug

Deliv Rev 15 37ndash52

Neuro-Oncology n JANUARY 200056

DR Groothuis Review of BBB and BTB

Fig 6 Distribution of vasogenic edema in a malignant glioma There are four pairs of scans extending through a tumor in the left temporal lobeextending from the lowest (A) to the highest level (D) Each pair of scans contains a gadolinium-enhanced scan (left) and a T2-weighted image(right) The gadolinium-enhanced scans show the area of abnormally permeable BBB The T2-weighted images show the distribution of edema(water) emanating from the tumor The tumor may be considered a natural equivalent of a convection-enhanced delivery infusion The resultingedema spreads extensively through white matter in the ipsilateral hemisphere reaching forward in the internal capsule (levels C and D) but notpenetrating gray matter structures such as the thalamus (levels C and D) or cortex (levels AndashD) Note that the edema contacts the posterior limbof the corpus callosum in levels C and D but does not extend into the corpus callosum suggesting that there is a physiological barrier to the bulkmovement of uid in that direction

References

CSF The major source of CSF is from the choroidplexuses of the lateral and 4th ventricles as much asanother 30 may originate from the brain extracellularuid The direction of the ow is from the lateral ventri-cles into the 3rd and then 4th ventricles out through theforamina of Luschke and Magendie into the basal cis-terns around the convexities and out of the subarach-noid space through the arachnoid granulations In ratswhen 14C-sucrose was injected into the lateral ventriclethe sucrose distributed rapidly within 5 min it was foundin the basal cisterns and by 1 h the sucrose had largely leftthe subarachnoid space (Ghersi-Egea et al 1996) Whenthe injected agent is metabolically active it may enterselected cells (Rubertone et al 1993) In humans the nor-mal volume of CSF produced is between 400 and 500 mlper day Since the CSF spaces within the brain are about150 ml in volume there is a considerable bulk ow withinthis system although the velocity varies tremendouslyfrom site to site When isotopes are injected into the lum-bar subarachnoid space they travel up the spinal sub-arachnoid space and are found in the basal cisterns by 3 hBy 24 h they are found over the cerebral convexities CSFow from the lateral ventricles is directional isotopesinjected into the lumbar CSF do not normally enter thelateral ventricles As a result of these bulk ow pathwaysmaterials injected into the CSF will attain initial concen-trations that are directly proportional to the concentra-tion in the infusate This route of administration is idealfor situations in which the target is within the subarach-noid space (for example carcinomatous meningitis) or inwhich the target is close to the CSF brain interface Carci-nomatous meningitis has long been treated with intrathe-cal infusions including the more recent use of monoclonalantibodies (Brown et al 1996) However the capacity ofdrugs to enter the brain extracellular space from the CSFis limited Numerous studies have shown that entry intothe brain is diffusional and that concentrations declineexponentially from the brain surface (Blasberg 1977Blasberg et al 1977 Groothuis and Levy 1997 Patlak

and Fenstermacher 1975) This route of administration isboth impractical and inefcient for intraparenchymalbrain tumors

Intratumoral and Intracavitary Injection

These are discussed together and briey because they areboth limiting cases of convection-enhanced deliverywhich is discussed below These both represent localforms of drug administration in which the distributionwill be determined initially by the rate and duration ofthe injection and secondly by tissue bulk ow pathwaysand diffusion The history of intratumoral injection wasdiscussed by Tomita (1991) who recognized the poten-tial usefulness of this approach as well as the importanceof neurotoxicity as the limiting factor Intratumoral andintracavitary injections are being widely explored asmeans of delivering large agents such as viral vectorsgrowth factors and cells (Bigner et al 1998 Farkkila etal 1994 Hsiao et al 1997 Puri et al 1996 Ram et al1997 Yang et al 1997) All of the attributes discussed inthe section on CED will also apply to these forms ofdrug delivery

Microdialysis

Microdialysis is a method that employs the passive diffu-sion of a drug across a semipermeable membrane (deLange et al 1997 de Lange et al 1999 Parsons andJustice 1994) It can be used to both sample and deliverdrugs to the surrounding tissue and has been used tosample extracellular drug concentrations in experimentalbrain tumors (Devineni et al 1996 Nakashima et al1997) The principle features of microdialysis when usedas a drug delivery methodology are that the tissue con-centrations are a function of the drug concentration inthe dialysate (which places control of the concentrationin hands of the user) and that distribution of drug awayfrom the dialysis catheter occurs by diffusion (Dykstra et

Neuro-Oncology n JANUARY 200052

DR Groothuis Review of BBB and BTB

Table 3 Comparison of different methods for local drug delivery to brain tumors

Method of circumventing the BBB Principle advantages Principle disadvantages Limiting factor(s)

Intrathecal Easy access ideal for therapy of Not useful for parenchymal Bulk ow rate of CSFmeningeal disease disease

Intraventricular administration Easy access ideal for therapy of Not useful for parenchymal Bulk ow rate of CSFmeningeal disease disease

Intratumoral injection User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Intracavitary injection User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Microdialysis User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Biodegradable polymers User control over administered Invasive distribution is diffusional Unpredictable distribution dose 100 reaches target neurotoxicity

Convection-enhanced delivery User control over administered Invasive Unpredictable distribution dose 100 reaches target neurotoxicity

Abbreviations BBB blood-brain barrier CSF cerebrospinal uid

This table lists the different approaches that are available to deliver therapeutic drugs and agents by local administration The advantages and disadvantages are an expression of theauthorrsquos opinion about the most prominent feature of each method Details about each method may be found in the text

al 1992) However the normal BBB is disrupted imme-diately around the catheter and stays disrupted for sometime after catheter insertion making drug efux in thisvicinity an additional problem when trying to understandlocal delivery kinetics (Groothuis et al 1998 Morgan etal 1996 Westergren et al 1995) Because the principlefactors in local tissue concentrations are diffusion andorlocal brain bulk flow patterns the volume of brainreached by microdialysis will be relatively small anddrug concentrations will in general decline exponen-tially away from the dialysis cannula This method willbe applicable when the volume of tissue that needs to bereached is small The effect of normal brain bulk owpathways on this delivery method remains to be studied

Biodegradable Polymers

This represents a local delivery system in whichbiodegradable polymers are loaded with drug andimplanted into brain or tumor tissue resulting in timed-release of drug (Cao and Shoichet 1999 Menei et al1997) The principle features of this system are that mul-tiple polymer pellets or wafers can be implanted whichgives control over the spatial distribution and that drugdistribution away from the source occurs by diffusionwith a variable contribution from the inherent bulk owpathways of the brain (Fung et al 1998) This approachhas been used in patients with malignant gliomas whereits safety and efcacy have been demonstrated (Brem etal 1995 Olivi and Brem 1994) As with microdialysisthe principle limiting feature of this method of drug deliv-ery is that drug distribution occurs mainly by diffusionin which drug concentrations fall exponentially awayfrom the source and are generally limited to a few mil-limeters of tissue penetration (Fung et al 1998)

Convection-Enhanced Delivery

This is a relatively new method of drug delivery to braintumors The principle feature of this delivery method isthat a drug solution is infused directly into the brain Theconcentration of the drug is independent of the infusionrate and duration and the distribution of the drug is afunction of the hydrostatic pressure of the infusion aswell as diffusion into surrounding tissue From a deliveryperspective the only limiting factor to the drug concen-tration that can be delivered is the solubility of the drugCED has been discussed from a theoretical perspective(Morrison et al 1994) it has been explored in severalanimal models (Bobo et al 1994 Groothuis et al 1999Laske et al 1997a Lieberman et al 1995 Viola et al1995) and it has been used for delivery of a targetedtoxin in human gliomas (Laske et al 1997b) The distri-bution of the infused drug is highly dependent uponwhether the drug is infused into a homogeneous graymatter structure or a structure containing ber pathwaysIn the case of the infusion of a drug-containing solutioninto a homogeneous brain structure the infusion initiallyproduces a spherical volume in which the concentrationof the drug is directly proportional to that of the infusate(Fig 2b) After the infusion reaches equilibrium at whichpoint the amount of drug leaving across brain capillaries

is equal to the amount being infused the drug begins todiffuse away from the edge of the sphere at concentra-tions that decline exponentially in accordance with themathematics of diffusion (Fig 2b) If the infusionencounters an organized white matter pathway the rateof movement is either increased or decreased (dependingupon the direction of the inherent bulk ow within thatpathway) (Fig 2c) Morrison stated that when the infu-sion is being made into a homogeneous isotropicmedium the radius of the central spherical componentmdashthe convective componentmdashcan be related to the infusionrate and the efux constant of the drug being infused

rp 5 3 2qv (4p K) (Eq 7)

where qv is the infusion rate and K represents thesteady-state efux rate constant Fig 3 shows a plot of theradius of the convective component as a function of infu-sion rate and efux constant From reviewing Fig 3

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 53

Fig 2 The distribution of 14C-sucrose in rat brain by convection-enhanced delivery The diagram in the upper left (A) shows the targetlocation (center of the caudate nucleus) as well as the organization ofthe brain at the target level After a 1-h 05-microlmin infusion the iso-tope is found in a spherical shape with even distribution across (B) By8 h the isotope is moving into the external capsule (C) A similarappearance is found after a constant infusion over 7 days the centralconvective component is clearly visible surrounded by decreasingconcentration from the edge (D) Note the isotopersquos rapid lateralmovement down the external capsule (probably representing a nor-mal brain bulk uid pathway) but not into the corpus callosum Infu-sion into rat brain for 7 days produced no neuropathologic changesother than those associated with catheter insertion

which extends from infusion rates provided by osmoticmini-pumps (1 microlh) to high infusion rates (10 microlmin) it is

apparent that the volume of affected brain remains quitesmall for most infusion rates Similarly as the efux con-stant decreases indicating slower rates of crossing theBBB the convection volume increases Fig 3 illustratesthat CED is best suited for delivery of large compounds(with low efux rates) and delivery at moderately highinfusion rates if the intent is to reach large brain volumes

The shape of the volume of distribution at steady statewill be highly dependent on the location in the brain thatis being infused Fig 4 shows a CT scan of a dog brainreceiving an infusion of iodinated contrast into whitematter The iodinated contrast rapidly distributes in thewhite matter of the ipsilateral hemisphere with rapidlydecreasing concentrations in the cortex on the affectedside Note that in both Figs 2 and 4 the infused materialdoes not cross the midline through the corpus callosumThis suggests that the direction of the normal bulk owpathway within the corpus callosum is away from themidline thus opposing the CED infusion component Incontrast to intravascularly administered drugs whereincreased capillary permeability results in increased tissueconcentrations (Table 1) the opposite effect will occur inCED infusions Increased efux across permeable tumorcapillaries will reduce both the extracellular drug concen-tration and the volume of distribution In Fig 5 highconcentrations of 14C-sucrose were achieved by a CEDinfusion into an RG-2 glioma but at a distance of 24mm from the infusion site tissue concentrationsdecreased by several logarithmic units Although studieshave not been done in humans it is easy to anticipatethat the volume of distribution from a CED infusion willfollow similar principles Fig 6 shows several levels of anMRI scan through a malignant glioma Each set of scansshows a gadolinium-enhanced tumor margin (which canbe considered naturersquos equivalent to a CED infusion) anda T2-weighted image (which corresponds to the extracel-lular water originating from the tumor source) The

Neuro-Oncology n JANUARY 200054

DR Groothuis Review of BBB and BTB

Fig 3 Relationship between radius of convective CED componentinfusion rate and efux rate The radius of the inection pointbetween the convective and diffusional components of CED infusionsis shown for infusion into a homogeneous brain structure (Equation7) The values on the vertical axis represent the radius inside whichthe local concentrations remain uniform and outside of which drugconcentrations decline exponentially Changes for each 5-cm incre-ment are represented by different shades of gray The infusion ratewas chosen from 1 acute 10ndash6 to 1 acute 10ndash2 ml per minndash1 The lower valuesare those seen with infusions from osmotic mini-pumps while thehigher values represent the upper limit of CED infusion rates in dogsThe efux rate was chosen to vary from 1 acute 10ndash1 to 1 acute 10ndash7 minndash1The volume of the convective component increases as a function ofboth increasing infusion rate and decreasing efux rate

Fig 4 Illustration of CED infusion in a dog The gure on the left shows the CT scan at the start of a 3 microl minndash1 infusion of Isovue (iopamidol)into the hemispheric white matter of a dog while that on the right shows the same coronal level on the same dog after 10 days of infusion Thescan on the left contains an air bubble in the lateral ventricle the cannula tip contains a metal marker (the high density circle) and a small amountof Isovue surrounding the catheter tip In the scan on the right the Isovue has distributed at a high concentration throughout the white matterof the ipsilateral hemisphere with a rapid decline in concentration at gray-white interfaces Note that the distribution of Isovue is limited to theipsilateral hemisphere No neuropathologic changes were observed as a result of the infusion

shape and distribution of the edema are highly irregularand very dependent upon the brain structures encoun-tered Note that the edema uid moves into the internalcapsule but not into the thalamus or cortex Notice alsothat the edema uid does not cross the posterior limb ofthe corpus callosum although it comes into contact withit at several levels

A dominant feature of CED is that distribution of thedrug is determined by the infusion parameters and thetype of tissue being infused whereas the concentration ofdrug in the infusate is unrelated to these parameters Themaximum concentration of the drug in the infusate willbe determined by neurotoxicity within the central con-vective component of the infusion with variable concen-trations in the anisotropic brain surrounding the centralcomponent and concentrations 50 to 100 times lower inthe diffusional component furthermost from the infusioncatheter The ability to manipulate drug concentration inbrain and tumor tissue with such ease and to maintainthese manipulations for long periods of time (Groothuiset al 1999) brings a need for the development ofentirely new concepts For example the limits withregard to the composition of the infusate need to beexplored such as pH osmolarity and ionic compositionFurthermore how does one reliably study neurotoxicityAre infusions into animal models going to be reliable pre-dictors of toxicity in humans Will differential neurotox-icity become an issue It is possible that neuronsastrocytes and oligodendrogliocytes will display differ-ent levels of susceptibility to many drugs Will we seedelayed neurotoxicity such as that seen with radiation Itwill probably be best for many of the variables associatedwith neurotoxicity to be explored in animals before sub-jecting humans to CED infusions

Summary of Drug Delivery that Circumvents the BBB

There has been an explosion of methodology involvinglocal tissue drug delivery during the past 10 years Someof these techniques such as CED fulll the promise tocircumvent the BBB entirely In fullling this promisethese methods introduce new problems such as the rela-tionship between the spatial distribution of drugs andneurotoxicity that need to be understood before thesetechniques are widely used However for viral vectorsmonoclonal antibodies antisense oligonucleotides andother therapeutic agents that will be unable to cross theBBB in therapeutically effective amounts CED may pro-vide the necessary delivery tool

The State of Drug Delivery to Brain Tumorsndash1999

If we return to the statement of Vick et al at the begin-ning of this review we can now state with certainty thatthe BBB and BTB are major factors limiting the access ofmany therapeutic agents to brain tumors and these bar-riers will become even more signicant with the develop-ment of new molecular biological therapies that willinvolve large molecules viruses and even cells It is alsoremarkable how prescient those authors were in identify-ing the other parts of the drug delivery process that mustnow be understood and how little progress we havemade in elucidating ldquotumor cell uptake metabolic fatewithin tumor cells and the washout or sink effect of theextracellular space and CSFrdquo (Vick et al 1977) Formost of the past 20 years we have focused so much on

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 55

Fig 5 Infusion of 14C-sucrose into an RG-2 rat glioma An infusion cannula was placed into the caudate nucleus of a Fisher-344 rat (targetshown in Fig 2A) and RG-2 cells were injected Ten days later 14C-sucrose was infused at a rate of 10 microlh for 7 days Autoradiographic imageswere made (right images) and the sections were then stained with hematoxylin-eosin (left images) The top two images show that the 14C-sucrose distributes in high concentration throughout the RG-2 tumor extending into the edematous external capsule (E) However thelower two sections taken from 24 mm behind the infusion site still contain tumor and show that the concentration of 14C-sucrose is 100000times lower These sections illustrate the problems associated with controlling drug delivery by convection-enhanceddelivery

ways to increase delivery across the BBB that we havefailed to address these other issues which now becomethe forefront in drug delivery methods such as CED Thearsenal from which to choose a drug delivery method istruly impressive We must now study the other factorsthat limit effective brain tumor therapy Perhaps the mostpressing of these includes the need to develop effective

laboratory tools for identifying the relationship betweenthe concentration-time product of therapeutic agents andtumor cell kill rather than relying on empirical testing inhuman subjects It is most likely that the new century willsee the development and application of tools for individ-ualizing chemotherapy for brain tumors and other solidhuman tumors

Ali-Osman F (1999) Human glioma cultures and in vitro analysis of therapeu-

tic response In Berger MS and Wilson CB (Eds)The Gliomas

Philadelphia WB Saunders Co pp 134ndash142

Allen TM and Moase EH (1996) Therapeutic opportunities for targeted

liposomal drug delivery Adv Drug Deliv Rev 21 117ndash133

Arafat T Hentschel P Madajewicz S Manzione J Chowhan N Davis

R Roche P Af I Roque C Meek A and Magdy S (1999) Toxicities

related to intraarterial infusion of cisplatin and etoposide in patients with

brain tumors J Neurooncol 42 73ndash77

Bartus RT (1999) The blood-brain barrier as a target for phamacological

modulation Curr Opin Drug Discov Dev 2 152ndash167

Bickel U Yoshikawa T Landaw EM Faull KF and Pardridge WM

(1993) Pharmacologic effects in vivo in brain by vector-mediated peptide

delivery Proc Natl Acad Sci USA 90 2618ndash2622

Bigner DD Brown MT Friedman AH Coleman RE Akabani G Fried-

man HS Thorstad WL McLendon RE Bigner SH Zhao XG

Pegram CN Wikstrand CJ Herndon JE II Vick NA Paleologos N

Cokgor I Provenzale JM and Zalutsky MR (1998) Iodine-131-labeled

antitenascin monoclonal antibody 81C6 treatment of patients with recur-

rent malignant gliomas Phase I trial results J Clin Oncol 16 2202ndash2212

Black KL (1995) Biochemical opening of the blood-brain barrier Adv Drug

Deliv Rev 15 37ndash52

Neuro-Oncology n JANUARY 200056

DR Groothuis Review of BBB and BTB

Fig 6 Distribution of vasogenic edema in a malignant glioma There are four pairs of scans extending through a tumor in the left temporal lobeextending from the lowest (A) to the highest level (D) Each pair of scans contains a gadolinium-enhanced scan (left) and a T2-weighted image(right) The gadolinium-enhanced scans show the area of abnormally permeable BBB The T2-weighted images show the distribution of edema(water) emanating from the tumor The tumor may be considered a natural equivalent of a convection-enhanced delivery infusion The resultingedema spreads extensively through white matter in the ipsilateral hemisphere reaching forward in the internal capsule (levels C and D) but notpenetrating gray matter structures such as the thalamus (levels C and D) or cortex (levels AndashD) Note that the edema contacts the posterior limbof the corpus callosum in levels C and D but does not extend into the corpus callosum suggesting that there is a physiological barrier to the bulkmovement of uid in that direction

References

al 1992) However the normal BBB is disrupted imme-diately around the catheter and stays disrupted for sometime after catheter insertion making drug efux in thisvicinity an additional problem when trying to understandlocal delivery kinetics (Groothuis et al 1998 Morgan etal 1996 Westergren et al 1995) Because the principlefactors in local tissue concentrations are diffusion andorlocal brain bulk flow patterns the volume of brainreached by microdialysis will be relatively small anddrug concentrations will in general decline exponen-tially away from the dialysis cannula This method willbe applicable when the volume of tissue that needs to bereached is small The effect of normal brain bulk owpathways on this delivery method remains to be studied

Biodegradable Polymers

This represents a local delivery system in whichbiodegradable polymers are loaded with drug andimplanted into brain or tumor tissue resulting in timed-release of drug (Cao and Shoichet 1999 Menei et al1997) The principle features of this system are that mul-tiple polymer pellets or wafers can be implanted whichgives control over the spatial distribution and that drugdistribution away from the source occurs by diffusionwith a variable contribution from the inherent bulk owpathways of the brain (Fung et al 1998) This approachhas been used in patients with malignant gliomas whereits safety and efcacy have been demonstrated (Brem etal 1995 Olivi and Brem 1994) As with microdialysisthe principle limiting feature of this method of drug deliv-ery is that drug distribution occurs mainly by diffusionin which drug concentrations fall exponentially awayfrom the source and are generally limited to a few mil-limeters of tissue penetration (Fung et al 1998)

Convection-Enhanced Delivery

This is a relatively new method of drug delivery to braintumors The principle feature of this delivery method isthat a drug solution is infused directly into the brain Theconcentration of the drug is independent of the infusionrate and duration and the distribution of the drug is afunction of the hydrostatic pressure of the infusion aswell as diffusion into surrounding tissue From a deliveryperspective the only limiting factor to the drug concen-tration that can be delivered is the solubility of the drugCED has been discussed from a theoretical perspective(Morrison et al 1994) it has been explored in severalanimal models (Bobo et al 1994 Groothuis et al 1999Laske et al 1997a Lieberman et al 1995 Viola et al1995) and it has been used for delivery of a targetedtoxin in human gliomas (Laske et al 1997b) The distri-bution of the infused drug is highly dependent uponwhether the drug is infused into a homogeneous graymatter structure or a structure containing ber pathwaysIn the case of the infusion of a drug-containing solutioninto a homogeneous brain structure the infusion initiallyproduces a spherical volume in which the concentrationof the drug is directly proportional to that of the infusate(Fig 2b) After the infusion reaches equilibrium at whichpoint the amount of drug leaving across brain capillaries

is equal to the amount being infused the drug begins todiffuse away from the edge of the sphere at concentra-tions that decline exponentially in accordance with themathematics of diffusion (Fig 2b) If the infusionencounters an organized white matter pathway the rateof movement is either increased or decreased (dependingupon the direction of the inherent bulk ow within thatpathway) (Fig 2c) Morrison stated that when the infu-sion is being made into a homogeneous isotropicmedium the radius of the central spherical componentmdashthe convective componentmdashcan be related to the infusionrate and the efux constant of the drug being infused

rp 5 3 2qv (4p K) (Eq 7)

where qv is the infusion rate and K represents thesteady-state efux rate constant Fig 3 shows a plot of theradius of the convective component as a function of infu-sion rate and efux constant From reviewing Fig 3

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 53

Fig 2 The distribution of 14C-sucrose in rat brain by convection-enhanced delivery The diagram in the upper left (A) shows the targetlocation (center of the caudate nucleus) as well as the organization ofthe brain at the target level After a 1-h 05-microlmin infusion the iso-tope is found in a spherical shape with even distribution across (B) By8 h the isotope is moving into the external capsule (C) A similarappearance is found after a constant infusion over 7 days the centralconvective component is clearly visible surrounded by decreasingconcentration from the edge (D) Note the isotopersquos rapid lateralmovement down the external capsule (probably representing a nor-mal brain bulk uid pathway) but not into the corpus callosum Infu-sion into rat brain for 7 days produced no neuropathologic changesother than those associated with catheter insertion

which extends from infusion rates provided by osmoticmini-pumps (1 microlh) to high infusion rates (10 microlmin) it is

apparent that the volume of affected brain remains quitesmall for most infusion rates Similarly as the efux con-stant decreases indicating slower rates of crossing theBBB the convection volume increases Fig 3 illustratesthat CED is best suited for delivery of large compounds(with low efux rates) and delivery at moderately highinfusion rates if the intent is to reach large brain volumes

The shape of the volume of distribution at steady statewill be highly dependent on the location in the brain thatis being infused Fig 4 shows a CT scan of a dog brainreceiving an infusion of iodinated contrast into whitematter The iodinated contrast rapidly distributes in thewhite matter of the ipsilateral hemisphere with rapidlydecreasing concentrations in the cortex on the affectedside Note that in both Figs 2 and 4 the infused materialdoes not cross the midline through the corpus callosumThis suggests that the direction of the normal bulk owpathway within the corpus callosum is away from themidline thus opposing the CED infusion component Incontrast to intravascularly administered drugs whereincreased capillary permeability results in increased tissueconcentrations (Table 1) the opposite effect will occur inCED infusions Increased efux across permeable tumorcapillaries will reduce both the extracellular drug concen-tration and the volume of distribution In Fig 5 highconcentrations of 14C-sucrose were achieved by a CEDinfusion into an RG-2 glioma but at a distance of 24mm from the infusion site tissue concentrationsdecreased by several logarithmic units Although studieshave not been done in humans it is easy to anticipatethat the volume of distribution from a CED infusion willfollow similar principles Fig 6 shows several levels of anMRI scan through a malignant glioma Each set of scansshows a gadolinium-enhanced tumor margin (which canbe considered naturersquos equivalent to a CED infusion) anda T2-weighted image (which corresponds to the extracel-lular water originating from the tumor source) The

Neuro-Oncology n JANUARY 200054

DR Groothuis Review of BBB and BTB

Fig 3 Relationship between radius of convective CED componentinfusion rate and efux rate The radius of the inection pointbetween the convective and diffusional components of CED infusionsis shown for infusion into a homogeneous brain structure (Equation7) The values on the vertical axis represent the radius inside whichthe local concentrations remain uniform and outside of which drugconcentrations decline exponentially Changes for each 5-cm incre-ment are represented by different shades of gray The infusion ratewas chosen from 1 acute 10ndash6 to 1 acute 10ndash2 ml per minndash1 The lower valuesare those seen with infusions from osmotic mini-pumps while thehigher values represent the upper limit of CED infusion rates in dogsThe efux rate was chosen to vary from 1 acute 10ndash1 to 1 acute 10ndash7 minndash1The volume of the convective component increases as a function ofboth increasing infusion rate and decreasing efux rate

Fig 4 Illustration of CED infusion in a dog The gure on the left shows the CT scan at the start of a 3 microl minndash1 infusion of Isovue (iopamidol)into the hemispheric white matter of a dog while that on the right shows the same coronal level on the same dog after 10 days of infusion Thescan on the left contains an air bubble in the lateral ventricle the cannula tip contains a metal marker (the high density circle) and a small amountof Isovue surrounding the catheter tip In the scan on the right the Isovue has distributed at a high concentration throughout the white matterof the ipsilateral hemisphere with a rapid decline in concentration at gray-white interfaces Note that the distribution of Isovue is limited to theipsilateral hemisphere No neuropathologic changes were observed as a result of the infusion

shape and distribution of the edema are highly irregularand very dependent upon the brain structures encoun-tered Note that the edema uid moves into the internalcapsule but not into the thalamus or cortex Notice alsothat the edema uid does not cross the posterior limb ofthe corpus callosum although it comes into contact withit at several levels

A dominant feature of CED is that distribution of thedrug is determined by the infusion parameters and thetype of tissue being infused whereas the concentration ofdrug in the infusate is unrelated to these parameters Themaximum concentration of the drug in the infusate willbe determined by neurotoxicity within the central con-vective component of the infusion with variable concen-trations in the anisotropic brain surrounding the centralcomponent and concentrations 50 to 100 times lower inthe diffusional component furthermost from the infusioncatheter The ability to manipulate drug concentration inbrain and tumor tissue with such ease and to maintainthese manipulations for long periods of time (Groothuiset al 1999) brings a need for the development ofentirely new concepts For example the limits withregard to the composition of the infusate need to beexplored such as pH osmolarity and ionic compositionFurthermore how does one reliably study neurotoxicityAre infusions into animal models going to be reliable pre-dictors of toxicity in humans Will differential neurotox-icity become an issue It is possible that neuronsastrocytes and oligodendrogliocytes will display differ-ent levels of susceptibility to many drugs Will we seedelayed neurotoxicity such as that seen with radiation Itwill probably be best for many of the variables associatedwith neurotoxicity to be explored in animals before sub-jecting humans to CED infusions

Summary of Drug Delivery that Circumvents the BBB

There has been an explosion of methodology involvinglocal tissue drug delivery during the past 10 years Someof these techniques such as CED fulll the promise tocircumvent the BBB entirely In fullling this promisethese methods introduce new problems such as the rela-tionship between the spatial distribution of drugs andneurotoxicity that need to be understood before thesetechniques are widely used However for viral vectorsmonoclonal antibodies antisense oligonucleotides andother therapeutic agents that will be unable to cross theBBB in therapeutically effective amounts CED may pro-vide the necessary delivery tool

The State of Drug Delivery to Brain Tumorsndash1999

If we return to the statement of Vick et al at the begin-ning of this review we can now state with certainty thatthe BBB and BTB are major factors limiting the access ofmany therapeutic agents to brain tumors and these bar-riers will become even more signicant with the develop-ment of new molecular biological therapies that willinvolve large molecules viruses and even cells It is alsoremarkable how prescient those authors were in identify-ing the other parts of the drug delivery process that mustnow be understood and how little progress we havemade in elucidating ldquotumor cell uptake metabolic fatewithin tumor cells and the washout or sink effect of theextracellular space and CSFrdquo (Vick et al 1977) Formost of the past 20 years we have focused so much on

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 55

Fig 5 Infusion of 14C-sucrose into an RG-2 rat glioma An infusion cannula was placed into the caudate nucleus of a Fisher-344 rat (targetshown in Fig 2A) and RG-2 cells were injected Ten days later 14C-sucrose was infused at a rate of 10 microlh for 7 days Autoradiographic imageswere made (right images) and the sections were then stained with hematoxylin-eosin (left images) The top two images show that the 14C-sucrose distributes in high concentration throughout the RG-2 tumor extending into the edematous external capsule (E) However thelower two sections taken from 24 mm behind the infusion site still contain tumor and show that the concentration of 14C-sucrose is 100000times lower These sections illustrate the problems associated with controlling drug delivery by convection-enhanceddelivery

ways to increase delivery across the BBB that we havefailed to address these other issues which now becomethe forefront in drug delivery methods such as CED Thearsenal from which to choose a drug delivery method istruly impressive We must now study the other factorsthat limit effective brain tumor therapy Perhaps the mostpressing of these includes the need to develop effective

laboratory tools for identifying the relationship betweenthe concentration-time product of therapeutic agents andtumor cell kill rather than relying on empirical testing inhuman subjects It is most likely that the new century willsee the development and application of tools for individ-ualizing chemotherapy for brain tumors and other solidhuman tumors

Ali-Osman F (1999) Human glioma cultures and in vitro analysis of therapeu-

tic response In Berger MS and Wilson CB (Eds)The Gliomas

Philadelphia WB Saunders Co pp 134ndash142

Allen TM and Moase EH (1996) Therapeutic opportunities for targeted

liposomal drug delivery Adv Drug Deliv Rev 21 117ndash133

Arafat T Hentschel P Madajewicz S Manzione J Chowhan N Davis

R Roche P Af I Roque C Meek A and Magdy S (1999) Toxicities

related to intraarterial infusion of cisplatin and etoposide in patients with

brain tumors J Neurooncol 42 73ndash77

Bartus RT (1999) The blood-brain barrier as a target for phamacological

modulation Curr Opin Drug Discov Dev 2 152ndash167

Bickel U Yoshikawa T Landaw EM Faull KF and Pardridge WM

(1993) Pharmacologic effects in vivo in brain by vector-mediated peptide

delivery Proc Natl Acad Sci USA 90 2618ndash2622

Bigner DD Brown MT Friedman AH Coleman RE Akabani G Fried-

man HS Thorstad WL McLendon RE Bigner SH Zhao XG

Pegram CN Wikstrand CJ Herndon JE II Vick NA Paleologos N

Cokgor I Provenzale JM and Zalutsky MR (1998) Iodine-131-labeled

antitenascin monoclonal antibody 81C6 treatment of patients with recur-

rent malignant gliomas Phase I trial results J Clin Oncol 16 2202ndash2212

Black KL (1995) Biochemical opening of the blood-brain barrier Adv Drug

Deliv Rev 15 37ndash52

Neuro-Oncology n JANUARY 200056

DR Groothuis Review of BBB and BTB

Fig 6 Distribution of vasogenic edema in a malignant glioma There are four pairs of scans extending through a tumor in the left temporal lobeextending from the lowest (A) to the highest level (D) Each pair of scans contains a gadolinium-enhanced scan (left) and a T2-weighted image(right) The gadolinium-enhanced scans show the area of abnormally permeable BBB The T2-weighted images show the distribution of edema(water) emanating from the tumor The tumor may be considered a natural equivalent of a convection-enhanced delivery infusion The resultingedema spreads extensively through white matter in the ipsilateral hemisphere reaching forward in the internal capsule (levels C and D) but notpenetrating gray matter structures such as the thalamus (levels C and D) or cortex (levels AndashD) Note that the edema contacts the posterior limbof the corpus callosum in levels C and D but does not extend into the corpus callosum suggesting that there is a physiological barrier to the bulkmovement of uid in that direction

References

which extends from infusion rates provided by osmoticmini-pumps (1 microlh) to high infusion rates (10 microlmin) it is

apparent that the volume of affected brain remains quitesmall for most infusion rates Similarly as the efux con-stant decreases indicating slower rates of crossing theBBB the convection volume increases Fig 3 illustratesthat CED is best suited for delivery of large compounds(with low efux rates) and delivery at moderately highinfusion rates if the intent is to reach large brain volumes

The shape of the volume of distribution at steady statewill be highly dependent on the location in the brain thatis being infused Fig 4 shows a CT scan of a dog brainreceiving an infusion of iodinated contrast into whitematter The iodinated contrast rapidly distributes in thewhite matter of the ipsilateral hemisphere with rapidlydecreasing concentrations in the cortex on the affectedside Note that in both Figs 2 and 4 the infused materialdoes not cross the midline through the corpus callosumThis suggests that the direction of the normal bulk owpathway within the corpus callosum is away from themidline thus opposing the CED infusion component Incontrast to intravascularly administered drugs whereincreased capillary permeability results in increased tissueconcentrations (Table 1) the opposite effect will occur inCED infusions Increased efux across permeable tumorcapillaries will reduce both the extracellular drug concen-tration and the volume of distribution In Fig 5 highconcentrations of 14C-sucrose were achieved by a CEDinfusion into an RG-2 glioma but at a distance of 24mm from the infusion site tissue concentrationsdecreased by several logarithmic units Although studieshave not been done in humans it is easy to anticipatethat the volume of distribution from a CED infusion willfollow similar principles Fig 6 shows several levels of anMRI scan through a malignant glioma Each set of scansshows a gadolinium-enhanced tumor margin (which canbe considered naturersquos equivalent to a CED infusion) anda T2-weighted image (which corresponds to the extracel-lular water originating from the tumor source) The

Neuro-Oncology n JANUARY 200054

DR Groothuis Review of BBB and BTB

Fig 3 Relationship between radius of convective CED componentinfusion rate and efux rate The radius of the inection pointbetween the convective and diffusional components of CED infusionsis shown for infusion into a homogeneous brain structure (Equation7) The values on the vertical axis represent the radius inside whichthe local concentrations remain uniform and outside of which drugconcentrations decline exponentially Changes for each 5-cm incre-ment are represented by different shades of gray The infusion ratewas chosen from 1 acute 10ndash6 to 1 acute 10ndash2 ml per minndash1 The lower valuesare those seen with infusions from osmotic mini-pumps while thehigher values represent the upper limit of CED infusion rates in dogsThe efux rate was chosen to vary from 1 acute 10ndash1 to 1 acute 10ndash7 minndash1The volume of the convective component increases as a function ofboth increasing infusion rate and decreasing efux rate

Fig 4 Illustration of CED infusion in a dog The gure on the left shows the CT scan at the start of a 3 microl minndash1 infusion of Isovue (iopamidol)into the hemispheric white matter of a dog while that on the right shows the same coronal level on the same dog after 10 days of infusion Thescan on the left contains an air bubble in the lateral ventricle the cannula tip contains a metal marker (the high density circle) and a small amountof Isovue surrounding the catheter tip In the scan on the right the Isovue has distributed at a high concentration throughout the white matterof the ipsilateral hemisphere with a rapid decline in concentration at gray-white interfaces Note that the distribution of Isovue is limited to theipsilateral hemisphere No neuropathologic changes were observed as a result of the infusion

shape and distribution of the edema are highly irregularand very dependent upon the brain structures encoun-tered Note that the edema uid moves into the internalcapsule but not into the thalamus or cortex Notice alsothat the edema uid does not cross the posterior limb ofthe corpus callosum although it comes into contact withit at several levels

A dominant feature of CED is that distribution of thedrug is determined by the infusion parameters and thetype of tissue being infused whereas the concentration ofdrug in the infusate is unrelated to these parameters Themaximum concentration of the drug in the infusate willbe determined by neurotoxicity within the central con-vective component of the infusion with variable concen-trations in the anisotropic brain surrounding the centralcomponent and concentrations 50 to 100 times lower inthe diffusional component furthermost from the infusioncatheter The ability to manipulate drug concentration inbrain and tumor tissue with such ease and to maintainthese manipulations for long periods of time (Groothuiset al 1999) brings a need for the development ofentirely new concepts For example the limits withregard to the composition of the infusate need to beexplored such as pH osmolarity and ionic compositionFurthermore how does one reliably study neurotoxicityAre infusions into animal models going to be reliable pre-dictors of toxicity in humans Will differential neurotox-icity become an issue It is possible that neuronsastrocytes and oligodendrogliocytes will display differ-ent levels of susceptibility to many drugs Will we seedelayed neurotoxicity such as that seen with radiation Itwill probably be best for many of the variables associatedwith neurotoxicity to be explored in animals before sub-jecting humans to CED infusions

Summary of Drug Delivery that Circumvents the BBB

There has been an explosion of methodology involvinglocal tissue drug delivery during the past 10 years Someof these techniques such as CED fulll the promise tocircumvent the BBB entirely In fullling this promisethese methods introduce new problems such as the rela-tionship between the spatial distribution of drugs andneurotoxicity that need to be understood before thesetechniques are widely used However for viral vectorsmonoclonal antibodies antisense oligonucleotides andother therapeutic agents that will be unable to cross theBBB in therapeutically effective amounts CED may pro-vide the necessary delivery tool

The State of Drug Delivery to Brain Tumorsndash1999

If we return to the statement of Vick et al at the begin-ning of this review we can now state with certainty thatthe BBB and BTB are major factors limiting the access ofmany therapeutic agents to brain tumors and these bar-riers will become even more signicant with the develop-ment of new molecular biological therapies that willinvolve large molecules viruses and even cells It is alsoremarkable how prescient those authors were in identify-ing the other parts of the drug delivery process that mustnow be understood and how little progress we havemade in elucidating ldquotumor cell uptake metabolic fatewithin tumor cells and the washout or sink effect of theextracellular space and CSFrdquo (Vick et al 1977) Formost of the past 20 years we have focused so much on

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 55

Fig 5 Infusion of 14C-sucrose into an RG-2 rat glioma An infusion cannula was placed into the caudate nucleus of a Fisher-344 rat (targetshown in Fig 2A) and RG-2 cells were injected Ten days later 14C-sucrose was infused at a rate of 10 microlh for 7 days Autoradiographic imageswere made (right images) and the sections were then stained with hematoxylin-eosin (left images) The top two images show that the 14C-sucrose distributes in high concentration throughout the RG-2 tumor extending into the edematous external capsule (E) However thelower two sections taken from 24 mm behind the infusion site still contain tumor and show that the concentration of 14C-sucrose is 100000times lower These sections illustrate the problems associated with controlling drug delivery by convection-enhanceddelivery

ways to increase delivery across the BBB that we havefailed to address these other issues which now becomethe forefront in drug delivery methods such as CED Thearsenal from which to choose a drug delivery method istruly impressive We must now study the other factorsthat limit effective brain tumor therapy Perhaps the mostpressing of these includes the need to develop effective

laboratory tools for identifying the relationship betweenthe concentration-time product of therapeutic agents andtumor cell kill rather than relying on empirical testing inhuman subjects It is most likely that the new century willsee the development and application of tools for individ-ualizing chemotherapy for brain tumors and other solidhuman tumors

Ali-Osman F (1999) Human glioma cultures and in vitro analysis of therapeu-

tic response In Berger MS and Wilson CB (Eds)The Gliomas

Philadelphia WB Saunders Co pp 134ndash142

Allen TM and Moase EH (1996) Therapeutic opportunities for targeted

liposomal drug delivery Adv Drug Deliv Rev 21 117ndash133

Arafat T Hentschel P Madajewicz S Manzione J Chowhan N Davis

R Roche P Af I Roque C Meek A and Magdy S (1999) Toxicities

related to intraarterial infusion of cisplatin and etoposide in patients with

brain tumors J Neurooncol 42 73ndash77

Bartus RT (1999) The blood-brain barrier as a target for phamacological

modulation Curr Opin Drug Discov Dev 2 152ndash167

Bickel U Yoshikawa T Landaw EM Faull KF and Pardridge WM

(1993) Pharmacologic effects in vivo in brain by vector-mediated peptide

delivery Proc Natl Acad Sci USA 90 2618ndash2622

Bigner DD Brown MT Friedman AH Coleman RE Akabani G Fried-

man HS Thorstad WL McLendon RE Bigner SH Zhao XG

Pegram CN Wikstrand CJ Herndon JE II Vick NA Paleologos N

Cokgor I Provenzale JM and Zalutsky MR (1998) Iodine-131-labeled

antitenascin monoclonal antibody 81C6 treatment of patients with recur-

rent malignant gliomas Phase I trial results J Clin Oncol 16 2202ndash2212

Black KL (1995) Biochemical opening of the blood-brain barrier Adv Drug

Deliv Rev 15 37ndash52

Neuro-Oncology n JANUARY 200056

DR Groothuis Review of BBB and BTB

Fig 6 Distribution of vasogenic edema in a malignant glioma There are four pairs of scans extending through a tumor in the left temporal lobeextending from the lowest (A) to the highest level (D) Each pair of scans contains a gadolinium-enhanced scan (left) and a T2-weighted image(right) The gadolinium-enhanced scans show the area of abnormally permeable BBB The T2-weighted images show the distribution of edema(water) emanating from the tumor The tumor may be considered a natural equivalent of a convection-enhanced delivery infusion The resultingedema spreads extensively through white matter in the ipsilateral hemisphere reaching forward in the internal capsule (levels C and D) but notpenetrating gray matter structures such as the thalamus (levels C and D) or cortex (levels AndashD) Note that the edema contacts the posterior limbof the corpus callosum in levels C and D but does not extend into the corpus callosum suggesting that there is a physiological barrier to the bulkmovement of uid in that direction

References

shape and distribution of the edema are highly irregularand very dependent upon the brain structures encoun-tered Note that the edema uid moves into the internalcapsule but not into the thalamus or cortex Notice alsothat the edema uid does not cross the posterior limb ofthe corpus callosum although it comes into contact withit at several levels

A dominant feature of CED is that distribution of thedrug is determined by the infusion parameters and thetype of tissue being infused whereas the concentration ofdrug in the infusate is unrelated to these parameters Themaximum concentration of the drug in the infusate willbe determined by neurotoxicity within the central con-vective component of the infusion with variable concen-trations in the anisotropic brain surrounding the centralcomponent and concentrations 50 to 100 times lower inthe diffusional component furthermost from the infusioncatheter The ability to manipulate drug concentration inbrain and tumor tissue with such ease and to maintainthese manipulations for long periods of time (Groothuiset al 1999) brings a need for the development ofentirely new concepts For example the limits withregard to the composition of the infusate need to beexplored such as pH osmolarity and ionic compositionFurthermore how does one reliably study neurotoxicityAre infusions into animal models going to be reliable pre-dictors of toxicity in humans Will differential neurotox-icity become an issue It is possible that neuronsastrocytes and oligodendrogliocytes will display differ-ent levels of susceptibility to many drugs Will we seedelayed neurotoxicity such as that seen with radiation Itwill probably be best for many of the variables associatedwith neurotoxicity to be explored in animals before sub-jecting humans to CED infusions

Summary of Drug Delivery that Circumvents the BBB

There has been an explosion of methodology involvinglocal tissue drug delivery during the past 10 years Someof these techniques such as CED fulll the promise tocircumvent the BBB entirely In fullling this promisethese methods introduce new problems such as the rela-tionship between the spatial distribution of drugs andneurotoxicity that need to be understood before thesetechniques are widely used However for viral vectorsmonoclonal antibodies antisense oligonucleotides andother therapeutic agents that will be unable to cross theBBB in therapeutically effective amounts CED may pro-vide the necessary delivery tool

The State of Drug Delivery to Brain Tumorsndash1999

If we return to the statement of Vick et al at the begin-ning of this review we can now state with certainty thatthe BBB and BTB are major factors limiting the access ofmany therapeutic agents to brain tumors and these bar-riers will become even more signicant with the develop-ment of new molecular biological therapies that willinvolve large molecules viruses and even cells It is alsoremarkable how prescient those authors were in identify-ing the other parts of the drug delivery process that mustnow be understood and how little progress we havemade in elucidating ldquotumor cell uptake metabolic fatewithin tumor cells and the washout or sink effect of theextracellular space and CSFrdquo (Vick et al 1977) Formost of the past 20 years we have focused so much on

DR Groothuis Review of BBB and BTB

Neuro-Oncology n JANUARY 2000 55

Fig 5 Infusion of 14C-sucrose into an RG-2 rat glioma An infusion cannula was placed into the caudate nucleus of a Fisher-344 rat (targetshown in Fig 2A) and RG-2 cells were injected Ten days later 14C-sucrose was infused at a rate of 10 microlh for 7 days Autoradiographic imageswere made (right images) and the sections were then stained with hematoxylin-eosin (left images) The top two images show that the 14C-sucrose distributes in high concentration throughout the RG-2 tumor extending into the edematous external capsule (E) However thelower two sections taken from 24 mm behind the infusion site still contain tumor and show that the concentration of 14C-sucrose is 100000times lower These sections illustrate the problems associated with controlling drug delivery by convection-enhanceddelivery

ways to increase delivery across the BBB that we havefailed to address these other issues which now becomethe forefront in drug delivery methods such as CED Thearsenal from which to choose a drug delivery method istruly impressive We must now study the other factorsthat limit effective brain tumor therapy Perhaps the mostpressing of these includes the need to develop effective

laboratory tools for identifying the relationship betweenthe concentration-time product of therapeutic agents andtumor cell kill rather than relying on empirical testing inhuman subjects It is most likely that the new century willsee the development and application of tools for individ-ualizing chemotherapy for brain tumors and other solidhuman tumors

Ali-Osman F (1999) Human glioma cultures and in vitro analysis of therapeu-

tic response In Berger MS and Wilson CB (Eds)The Gliomas

Philadelphia WB Saunders Co pp 134ndash142

Allen TM and Moase EH (1996) Therapeutic opportunities for targeted

liposomal drug delivery Adv Drug Deliv Rev 21 117ndash133

Arafat T Hentschel P Madajewicz S Manzione J Chowhan N Davis

R Roche P Af I Roque C Meek A and Magdy S (1999) Toxicities

related to intraarterial infusion of cisplatin and etoposide in patients with

brain tumors J Neurooncol 42 73ndash77

Bartus RT (1999) The blood-brain barrier as a target for phamacological

modulation Curr Opin Drug Discov Dev 2 152ndash167

Bickel U Yoshikawa T Landaw EM Faull KF and Pardridge WM

(1993) Pharmacologic effects in vivo in brain by vector-mediated peptide

delivery Proc Natl Acad Sci USA 90 2618ndash2622

Bigner DD Brown MT Friedman AH Coleman RE Akabani G Fried-

man HS Thorstad WL McLendon RE Bigner SH Zhao XG

Pegram CN Wikstrand CJ Herndon JE II Vick NA Paleologos N

Cokgor I Provenzale JM and Zalutsky MR (1998) Iodine-131-labeled

antitenascin monoclonal antibody 81C6 treatment of patients with recur-

rent malignant gliomas Phase I trial results J Clin Oncol 16 2202ndash2212

Black KL (1995) Biochemical opening of the blood-brain barrier Adv Drug

Deliv Rev 15 37ndash52

Neuro-Oncology n JANUARY 200056

DR Groothuis Review of BBB and BTB

Fig 6 Distribution of vasogenic edema in a malignant glioma There are four pairs of scans extending through a tumor in the left temporal lobeextending from the lowest (A) to the highest level (D) Each pair of scans contains a gadolinium-enhanced scan (left) and a T2-weighted image(right) The gadolinium-enhanced scans show the area of abnormally permeable BBB The T2-weighted images show the distribution of edema(water) emanating from the tumor The tumor may be considered a natural equivalent of a convection-enhanced delivery infusion The resultingedema spreads extensively through white matter in the ipsilateral hemisphere reaching forward in the internal capsule (levels C and D) but notpenetrating gray matter structures such as the thalamus (levels C and D) or cortex (levels AndashD) Note that the edema contacts the posterior limbof the corpus callosum in levels C and D but does not extend into the corpus callosum suggesting that there is a physiological barrier to the bulkmovement of uid in that direction

References

ways to increase delivery across the BBB that we havefailed to address these other issues which now becomethe forefront in drug delivery methods such as CED Thearsenal from which to choose a drug delivery method istruly impressive We must now study the other factorsthat limit effective brain tumor therapy Perhaps the mostpressing of these includes the need to develop effective

laboratory tools for identifying the relationship betweenthe concentration-time product of therapeutic agents andtumor cell kill rather than relying on empirical testing inhuman subjects It is most likely that the new century willsee the development and application of tools for individ-ualizing chemotherapy for brain tumors and other solidhuman tumors

Ali-Osman F (1999) Human glioma cultures and in vitro analysis of therapeu-

tic response In Berger MS and Wilson CB (Eds)The Gliomas

Philadelphia WB Saunders Co pp 134ndash142

Allen TM and Moase EH (1996) Therapeutic opportunities for targeted

liposomal drug delivery Adv Drug Deliv Rev 21 117ndash133

Arafat T Hentschel P Madajewicz S Manzione J Chowhan N Davis

R Roche P Af I Roque C Meek A and Magdy S (1999) Toxicities

related to intraarterial infusion of cisplatin and etoposide in patients with

brain tumors J Neurooncol 42 73ndash77

Bartus RT (1999) The blood-brain barrier as a target for phamacological

modulation Curr Opin Drug Discov Dev 2 152ndash167

Bickel U Yoshikawa T Landaw EM Faull KF and Pardridge WM

(1993) Pharmacologic effects in vivo in brain by vector-mediated peptide

delivery Proc Natl Acad Sci USA 90 2618ndash2622

Bigner DD Brown MT Friedman AH Coleman RE Akabani G Fried-

man HS Thorstad WL McLendon RE Bigner SH Zhao XG

Pegram CN Wikstrand CJ Herndon JE II Vick NA Paleologos N

Cokgor I Provenzale JM and Zalutsky MR (1998) Iodine-131-labeled

antitenascin monoclonal antibody 81C6 treatment of patients with recur-

rent malignant gliomas Phase I trial results J Clin Oncol 16 2202ndash2212

Black KL (1995) Biochemical opening of the blood-brain barrier Adv Drug

Deliv Rev 15 37ndash52

Neuro-Oncology n JANUARY 200056

DR Groothuis Review of BBB and BTB

Fig 6 Distribution of vasogenic edema in a malignant glioma There are four pairs of scans extending through a tumor in the left temporal lobeextending from the lowest (A) to the highest level (D) Each pair of scans contains a gadolinium-enhanced scan (left) and a T2-weighted image(right) The gadolinium-enhanced scans show the area of abnormally permeable BBB The T2-weighted images show the distribution of edema(water) emanating from the tumor The tumor may be considered a natural equivalent of a convection-enhanced delivery infusion The resultingedema spreads extensively through white matter in the ipsilateral hemisphere reaching forward in the internal capsule (levels C and D) but notpenetrating gray matter structures such as the thalamus (levels C and D) or cortex (levels AndashD) Note that the edema contacts the posterior limbof the corpus callosum in levels C and D but does not extend into the corpus callosum suggesting that there is a physiological barrier to the bulkmovement of uid in that direction

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