monograph series

4NarcoticAntagonists:The Search forLong-ActingPreparations

The NIDA Research Monograph series is prepared by theResearch Division of the National Institute on Drug Abuse. Itsprimary objective is to provide critical reviews of research problemareas and techniques, the content of state-of-the-art conferences,integrative research reviews and significant original research. Itsdual publication emphasis is rapid and targeted dissemination tothe scientific and professional community.

EDITORIAL ADVISORY BOARD

Avram Goldstein, M.D. Addiction Research FoundationPalo Alto, California

Jerome Jaffee, M.D. College of Physicians and SurgeonsColumbia University, New York

Reese T. Jones, M.D. Langley Porter NeuropsychiatricInstituteUniversity of CaliforniaSan Francisco, California

William McGlothlin, Ph.D. Departrnent of Psychology, UCLALos Angeles, California

Jack Mendelson, M.D. Alcohol and Drug Abuse ResearchCenterHarvard Medical SchoolMcLean HospitalBelmont, Massachusetts

Helen Nowlis, Ph.D. Office of Drug Education, DHEWWashington, D.C.

Lee Robins, Ph.D. Washington University School ofMedicineSt. Louis, Missouri

NIDA RESEARCH MONOGRAPH series

Robert DuPont, M.D. DIRECTOR, NIDA

William Pollin, M.D. DIRECTOR, NIDA RESEARCH DIVISION

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monograph series

NarcoticAntagonists:The Search forLong-ActingPreparations

EditorRobert Willette, Ph.D.Division of ResearchNational Institute on Drug AbuseJanuary 1976

THE NATIONAL INSTITUTE ON DRUG ABUSE11400 Rockville PikeRockville, Maryland 20852

For sale by the Superintendent of Documents, U.S. Government Printing OfficeWashington, D.C. 20402 - Price $1.10

Stock Number 017-024-00488-0

DHEW Publication No. (ADM) 76-296Printed 1975

Library of Congress catalog number : 75-29949

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Molded beads of 25/75 lactic/glycolic copolymer50wt% naltrexone pamoate mixture. (mm)Dynatech R/D. (See first article).

Subcutaneous scapular implant site of Dl-6five weeks after implantation in rabbit.Note the nodular mass (arrow).

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Subcutaneous scapular implant site of D2-7five weeks after implantation.Note rod-shaped residues (arrow).

Subcutaneous scapular implant site of D2-8five weeks after implantation.Note rod-shaped residues (arrows).

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CONTENTS

INTRODUCTION

ABSTRACTS 7

DEVELOPMENT OF POLYLACTIC/GLYCOLIC ACIDDELIVERY SYSTEMS FOR USE IN TREATMENT OFNARCOTIC ADDICTION

A.D. Schwope, D.L. Wise, J.F. Howes 13

DEVELOPMENT OF INJECTABLE MICROCAPSULESFOR USE IN THE TREATMENT OF NARCOTICADDICTION

C. Thies 19

LONG-ACTING NARCOTIC ANTAGONIST COMPLEXESA.P. Gray, W.J. Guardina 21

SUSTAINED RELEASE OF NALTREXONEFROM GLYCERIDE IMPLANTS

M.F. Sullivan, D.R. Kalkwarf 27

USE OF SYNTHETIC POLYPEPTIDES IN THEPREPARATION OF BIODEGRADABLE DELIVERYVEHICLES FOR NARCOTIC ANTAGONISTS

K.R. Sidman, D.L. Arnold, W.D. Steber,L. Nelsen, F.E. Granchelli, P. Strong,S.G. Sheth 33

DEVELOPMENT OF CHRONOMERS™FOR NARCOTIC ANTAGONISTS

R.C. Capozza, E.E. Schmitt, L.R. Sendelbeck 39

TESTING OF DRUG DELIVERY SYSTEMSFOR USE IN TREATMENTOF NARCOTIC ADDICTION

R.H. Reuning, L. Malspeis, S. Frank,R.E. Notari 43

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INTRODUCTIONThe use of narcotic antagonists in the treatmentof opiate addiction is based on the concept of apharmaceutical agent capable of blocking the rein-forcing properties of a dose of opiate takenduring an addicts rehabilitation. The rationalefor use is that the antagonist blocks the opiate"high" and makes it pleasureless, thus removingthe addict's incentive for continued use. Earliersuccessful therapy with cyclazocine and naloxoneprompted the full-scale development of new andsuperior antagonists. Presently naltrexone isthe drug under the most intensive clinicalevaluation and appears to be a promising antag-onist candidate.

It was felt from the outset that a most desirablecomponent of antagonist therapy would be a long-acting drug, so that the need for an addict todecide to take his medication would be minimized.Naltrexone in oral doses of 70 mg. will provideadequate blocking protection for at least 48hours, or perhaps 72 hours in certain individuals.This is not felt to be a long enough interval be-tween dosages to aid the addict in becoming dis-sociated from his drug-taking behavior.

It was recognized very early that in order toachieve the desired one week, one month or longerduration between dosages, it would be necessaryto develop a long-acting drug delivery system ora sustained-release preparation of an acceptablebut short-acting antagonist. A "drug-deliverysystem" is the unwieldy but currently favoredexpression describing any pharmaceutical prepa-ration capable of providing a sustained or long-acting antagonistic effect. This effect may beachieved mechanically (e.g., by implanted discswith timed release capacity) or chemically (e.g.,

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microcapsules, tubes, solid balls, gelatinousmasses injected intramuscularly). Distinctfrom the problem not considered here, of find-ing an optimum antagonist, is the problem ofinventing suitable carriers for the antagonist,carriers that will deliver the antagonist, re-leasing it uniformly bit by bit over a periodof time.system."

Hence the barbarism, a "drug delivery

Efforts to achieve satisfactory drug deliverysystems were launched in the early 1970's bythe City of New York Public Health Departmentand by the Division of Narcotic Addiction andDrug Abuse, now the National Institute on DrugAbuse (NIDA).

During this early period, the pioneering efforts ofDr. Seymour Yolles, University of Delaware,demonstrated for the first time that a sustained-release of an antagonist could be obtained froma biodegradable polymer, i.e., polylactic acid.This success generated expanded and intensifiedefforts, a summary of which is the topic ofthis monograph.

At the present time, the program supported byNIDA includes six contracts that are concernedwith the development of new delivery systemsand three contracts that have the responsibilityof evaluating them for potential clinical trials.

Delivery Systems Development

Borrowing on its experience in the antifertilityarea supported by the Center for Population Re-search, at the National Institute for ChildHealth and Human Development, Dynatech Corpora-tion has had the major goal of developing bio-degradable devices from polyactic and poly-glycolic acids. This effort is now centered onrefining copolymer preparations with naltrexonethat are implantable and have either a one or sixmonth duration.

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Another project receiving early support has beenthat of Dr. curt Thies, Washington University,an effort to develop microencapsulated dosageforms. Working primarily with the polylactic andglycolic acids, Dr. Thies achieved early successof multiple-week release with the antagonistcyclazocine. As naltrexone became the drug ofchoice, it was substituted for cyclazocine. Thehighly experimental nature of these systems wasrevealed when it was quickly found that the phy-sicochemical properties of naltrexone posed newproblems. Among these was increased watersolubility. Notable progress has now been madein improving capsule quality to achieve longerrelease times.

One of the approaches supported originally byNew York City that has yielded very promisingsustained-release candidates has been directedby Dr. Alan Gray, Illinois Institute of Tech-nology Research. He has found that zinc andaluminum tannate complexes of naltrexone, in-jected in 2% aluminum monostearate in sesame oilgives sustained release from three to fourweeks. Advanced studies are now underway toevaluate its safety in order to initiateclinical trials.

Another early project that is now approachingfinal evaluation for clinical trials is basedon implantable pellets of naltrexone in poly-glycerides. This work has been conducted byDrs. Maurice Sullivan and Donald Kalkwarf, ofBattelle Pacific Northwest Laboratories. Thenatural polyglycerides used have shown excel-lent biocompatibility, that is, negligibleinflammation on insertion and little tissuereaction overall.

A newer contract with Arthur D. Little Inc.is concentrating on the develooment of new bio-degradable polymers based on polypeptides.Tubes of polyleucine-polyglutamate filled withnaltrexone appear to offer a promising newlead.

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One of the more unique approaches is that be-ing pursued by Alza Research. In contrast tothe other polymer based release systems, whichdiffuse drugs, the Alza chronomer erodes at arate equal to drug release. Interesting in-duction periods and solubility effects are cur-rently being studied to perfect a desirablerelease rate.

Evaluat ion

The evaluation of leading candidate devicesdeveloped by these contractors is initiatedby a group headed by Dr. Richard Reuning atOhio State University. The first series oftests consists of following the blocking ofmorphine-analgesia in mice by the test deviceto confirm any original test results. Next,the release of radiolabeled drug from thedevice is followed in rats and finally inmonkeys. In order to carry out this type ofevaluation, this group has conducted extensivepharmacokinetic studies on naltrexone. Thisinformation permits a determination of theactual rate of release of drug from the deviceunder testing.

Candidate devices that pass the first two testsat Ohio State are then submitted to IndustrialBio-Test, Inc., for evaluation of their tissuecompatability, and to Parke, Davis and Companyfor a rigorous final pharmacological evaluation.For the toxicity studies, mice, rats, rabbitsand monkeys receive various loadings of the testdevice and tissue reactions are checked, usuallyat 7 and 28 days. The pharmacological studiesare carried out in a group of monkeys trained tolever-press for injections of morphine. Asuccessful device will block this behavior aslong as drug is being released. Simultaneously,blood specimens are collected for the Ohio Stategroup to analyze for drug levels.

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As a result of these efforts to date, two orpossibly three candidates are being readied forclinical trials. The final preclinical worknecessary includes sterility checks, more de-tailed toxicity studies, and quality controlstandards. It is our hope to be able toevaluate at least one of these devices in manwithin a year.

Robert E. Willette, Ph.D.Division of ResearchNational Institute on Drug Abuse

September, 1975

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ABSTRACTRESEARCH MONOGRAPH #4, NATIONAL INSTITUTE ONDRUG ABUSE. Narcotic Antagonists: The Searchfor Long-Acting Preparations.DEVELOPMENT OF POLYLACTIC/GLYCOLIC ACIDDELIVERY SYSTEMS FOR USE IN TREATMENT OFNARCOTIC ADDICTION

in vivo efficacy has been-measured by directchallenge with morphine (Dewey-Harris mouse

Schwope, A.D., D.L. Wise, J.F. HowesImplantable polylactic/glycolic acid matrixsystems have successfully provided the sus-tained release of naltrexone to mice forperiods of up to 200 days. In vitro and invivo release rates have been determined bymeasuring chemical concentrations in pH 7buffer solution and urine, respectively, and

tail-flick test). Dosage forms of small im-plantable cylinders, 1/16" diameter, (25 mg/rod,one rod/mouse) containing 33% by weight nal-trexone pamoate in 90 L(+)/10 polylactic/glycolic acid have sustained the delivery ofchemical for 200 days. Delivery of chemicalfrom dosage forms of 1/16" diameter sphericalbeads (3 mg/bead, 3 beads/mouse) containing33% by weight naltrexone base in 90 L(+)/10polylactic/glycolic acid was sustained for 60days. Earlier a similar bead type dosage formof 75 L(+)/25 polylactic/glycolic acid contain-ing 50% by weight naltrexone base and coatedwith the pure polymer provided controlledrelease for 25 days.which incorporate the use of pharmacologicallysuitable catalysts and yield products repro-ducibly have been delineated. Techniques forsterilization of the final implant have beenscreened.

ABSTRACTRESEARCH MONOGRAPH #4, NATIONAL INSTITUTE ONDRUG ABUSE. Narcotic Antagonists: The Searchfor Long-Acting Preparations.DEVELOPMENT OF INJECTABLE MICROCAPSULESFOR USE IN THE TREATMENT OF NARCOTICADDICTION

Thies, C.

Injectible microcapsules containing narcoticantagonists have been prepared with dl-poly(lactic acid) as the coating material. Theencapsulation technology has developed tothe point that high yields of less than180 µ capsules can be prepared routinely.Such capsules with an initial payload of50 wt. % naltrexone pwnoate provide 60-90%atagonism to the action of morphine 28 daysafter injection into mice as a peanut oil/aluminum monostearate suspension at a doselevel of 40 miligrams naltrexone pamoate/ kg.mouse.

ABSTRACTRESEARCH MONOGRAPH #4, NATIONAL INSTITUTE ONDRUG ABUSE. Narcotic Antagonists: The Searchfor Long-Acting Preparations.LONG-ACTING NARCOTIC ANTAWNIST COMPLEXES

Gray, A.P., W. J. Guardina

We evaluated the ability of close to 100organic acids to form water-soluble saltswith methadone, cyclazocine, naloxone,naltrexone and, more recently, diprenorphine.About half the acids yielded insoluble salts.Polybasic acids affording insoluble saltswere evaluated for their ability to formdrug:acid:metal complexes with the polyvalentmetal ions, Zn++, Al+++, Mg++ and Ca++.Optimum conditions for forming complexes havebeen developed and the consistency of theircomposition has been established.

Salts were analyzed spectrophotometricallyfor drug content, and complexes were analyzedfor drug and metal content. The in vitrodegree of dissociation at equilibriummeasured for the preparations suspended ina simulated physiological buffer, pH 7.3.

Preparations of the narcotic antagonistdrugs showing relatively low degrees ofdissociation in vitro, since it-earlyappeared that a high degree of dissociationcontraindicated a prolonged duration ofpharmacologica1 action, were evaluated inmice after intramuscular administration atseveral dose 1evels by the mouse tail-flicktest for the potency and duration of theirmorphine antagonist activity.

Our most promising preparations to date,showing the most prolonged durations ofaction without evidence of gross toxicity,are naltrexone zinc tannate and naltrexonealuminum tannate. These are undergoingdetailed evaluation as potential clinicalcandidates. Thus far, the most useful ofseveral dosage forms studied is a sus-pension in an aluminum monostearate gel.

The abstracts are provided assummary and may be extractedfor reference and filing con-venience.

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ABSTRACTRESEARCH MONOGRAPH #4, NATIONAL INSTITUTE ONDRUG ABUSE. Narcotic Antagonist's: The Searchfor Long-Acting Preparations.SUSTAINED RELEASE OF NALTREXONEFROM GLYCERIDE IMPLANTSSullivan, M. F., D. R. KalkwarfSolid dispersions of naltrexone in naturalglycerides were used to form injectable im-plants which continuously release narcoticantagonists in vivo. The dispersions wereformed and tested either as small cylindri-Cal pellets, e.g. lx3.0 mm in size, oras particles with diameters in size rangesbetween 125-250 µ, that are suspended inan aqueous methyl cellulose solution.Both types of implants delivered naltrex-one to mice at rates that were effectivein blocking the antiociceptive action ofmorphine for at least one month. The rateof naltrexone release was controlled byaltering its concentration in the disper-sion and by varying the glyceride composi-tion. Degradation and absorption of theimplants were found to depend on theircomposition, dimensions and location inthe body. No appreciable tissue incompati-bility was seen in mice, rats, rabbits,monkeys and swine, even when long-lastingpreparations were removed a year aftertreatment.

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ABSTRACTRESEARCH MONOGRAPH #4, NATIONAL INSTITUTE ONDRUG ABUSE. Narcotic Antagonists: The Searchfor Long-Acting Preparations.USE OF SYNTHETIC POLYPEPTIDES IN THEPREPARATION OF BIODEGRADABLE DELIVERYVEHICLES FOR NARCOTIC ANTAGONISTSSidman, K.R., D.L. Arnold, W.D. Steber,L. Nelsen, F.E. Granchelli, P. Strong,S.G. ShethSynthetic polypeptides consisting of copoly-mers of glutamic acid and leucine have beenshown to be useful materials for the fabri-cation of practical, biodegradable deliveryvehicles for narcotic antagonists.

Model delivery vehicles in film form were pre-pared from copolymers containing 10 mole per-cent to 40 mole percent glutamic acid, andloaded with 10% to 40% naltrexone by weight.The naltrexone was found to be released bydiffusion, exhibiting diffusion coefficientsthat varied as a function of the glutamic acidcontent and the initial naltrexone loading.A wide range in diffusion coefficients wereachieved (0.31 x 10-7 cm2/hr to 120 x 10-7 cm2/hr), leading to release rates within practicalranges of interest for meeting the programgoals.

We have demonstrated that the polypeptidescan be fabricated into dosage forms that areamenable to administration by trochar. Forexample, rods 0.4 mm to 0.8 mm in diametercontaining as much as 40% naltrexone by weightwere extruded using a simple compression moldand die arrangement. An in vitro evaluationof the rods showed that antagonist is releasedby diffusion at a continuously decreasingrate, a behavior similar to that observed withthe film devices that were, nonetheless,capable of blocking an AD80 challenge ofmorphine sulfate in mice for more than 30days.

One of the most promising delivery vehiclesthat we have developed to date consists of apolypeptide tube filled with a naltrexone/polypeptide core. Preliminary experimentshave shown that these devices may be capableof administering high, constant rates of re-lease for prolonged periods of time. Addi-tional work, however, is required to developtechniques for the preparation of reproducibledelivery vehicles.

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A B S T R A C TRESEARCH MONOGRAPH #4, NATIONAL INSTITUTE ONDRUG ABUSE. Narcotic Antagonists: The Searchfor Long-Acting Preparations.

DEVELOPMENT OF CHRONOMERS™ FORNARCOTIC ANTAGONISTS

Capozza, R.C., E.E. Schmitt, L.R. Sendelbeck

The object of this program is to prepare abioerodable naltrexone deLivery system whichcan be implanted subcutaneously in humans andwhich can relieve the narcotic antagonistover 1-6 months at relatively constant andsufficient rates to block the euphoric effectof morphine based drugs. The system is com-posed of naltrexone uniformly dispersed in asolid hydropholic CHRONOMER™ matrix whichundergoes predictable surface erosion whenexposed to an aqueous medium. Kinetic studiesin vitro have been carried out during thecourse of the program to determine the bestcomposition for the system.

Toxilogical studies conducted at ALZA duringthe past 2 years have not revealed limitingadverse effects of either the CHRONOMER™materials or their hydrolysis products. Thetail-flick test procedure was used to measurethe effectiveness of naltrexone to antagonizethe analgesis of morphine in rats. Naltrexoneinfused intravenously at doses of 4 and 16ug/kg/hr resulted in, after 6 hours, 54 and89% antagonism, respectively, against a63.5% effective dose of morphine.

Preliminary Sterilization studies; showed thatno adverse effects to CHRONOMER™/naltrexonesystems occurred after exposure to 2.5 or 5.0mrads of 60Co irradiation.

ABSTRACTRESEARCH MONOGRAPH #4, NATIONAL INSTITUTE ONDRUG ABUSE. Narcotic Antagonists: The Searchfor Long-Acting Preparations.TESTING OF DRUG DELIVERY SYSTEMS FOR USE INTREATMENT OF NARCOTIC ADDICTION

Reuning, R.H., L. Malspeis, S. Frank,R.E. NotariThe evaluation of the drug release character-istic of four naltrexone delivery systems hasbeen carried out together with the developmentof analytical techniques and an investigationof the metabolic profile of naltrexone. Pharma-cologic evaluation of the four delivery systemsin the mouse indicated significant analgesicantagonism for a period of from 16-22 days.Further evaluation of one of these systems bymeasurement of the rate of excretion of radio--activity after administration of radiolabellednaltrexone in the delivery system confirmed thatsignificant release occurs for a time period ofabout 15 days. Electron capture gas-liquidchromatographic assays for naltrexone and nal-oxone in plasma or urine have been developedthat yield linear calibration curves and aresensitive to one ng/ml. Studies on naltrexonedisposition indicate that (a) binding to plasmaproteins in several species varies from 20-26%,(b) distribution of drug from blood is extremelyrapid and extensive, (c) -naltrexol is a majormetabolite of naltrexone in man, monkey andguinea pig among six species studied, whereasa-naltrexol is a minor metabolite in the monkeyand guinea pig only, and (d) metabolic reductionof naltrexone occurs in the 100,000 x g super-natant of guinea pig liver. Pharmacokineticstudies of naltrexone in the dog and monkeyindicate that the drug is rapidly distributedand eliminated, has a very large apparent volumeof distribution and a total body clearancegreater than the rate of liver blood flow.

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DEVELOPMENT OFPOLYLACTIC/GlYCOLIC ACIDDELIVERY SYSTEMSFOR USE IN TREATMENT OFNARCOTIC ADDICTION

Arthur D. SchwopeDonald L. Wise, Ph. D.

Dynatech R/D CompanyJohn F. Howes, Ph. D.

Sharps Associates

INTRODUCTION

A problem universal to chemotherapeutic This is achieved by injecting the agentprograms is the administration and in a carrier solution (e.g., oil andmaintenance of drug(s) at safe and ef- saline) and by encapsulating or sur-fective levels for extended periods. rounding the chemical with an absorb-Much technical activity has been able, release-regulating material.devoted to devising improved and differ-ent means for regulating drug delivery. In this study, polymers of lactide and

glycolide were evaluated as the release-One important category of dosage forms regulating materials. Polylacticlunder investigation is implantable, glycolic acids are tissue compatibletissue absorbable preparations which and hydrolyze to the metabolites, lacticrelease chemical (e.g., medicaments, and glycolic acids. The rate offertilizers, and pesticides) at a con- hydrolysis is controllable and dependentstant rate to biological systems. There on the, lactic/glycolic ratio. Physicalare two fundamentally different ap- mixtures, matrices, of narcotic antago-proaches to achieve that end. One nist - naltrexone - and polymer wererelies on chemical alteration of the prepared, formed and evaluated in vitroagent so that adsorption by the system and in vivo as long term, constantis delayed. In the second method, the delivery, dosage systems.active agent remains chemically un-changed, but its availability to thebody is restricted by physical means.

This paper describes the procedureswhich have been used to prepare and

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evaluate drug/polymer matrix systemswhich will provide the sustainedrelease of naltrexone for one month andand for six months. Results of theevaluations of selected systems arepresented.

METHODS AND MATERIALS

Sample preparation

L(+)-lactide and glycolide, synthesizedfrom the respective acids, were poly-merized into polylactic acid and poly-lactic/glycolic acid copolymers usingtriethylaluminum and suitable cata-lysts. Polymers ranging from 75 wt %L(+)lactide/25% G to 100% L(+) havebeen prepared. Molecular weights ofthe polymers determined by gel permea-tion chromatography, membrane osmometryand light scattering ranged from 40,000- 200,000. Intrinsic viscositiesmeasured at 37°C in tetrahydrofuranranged from 0.4 to 1.0.

Naltrexones base and pamoate (suppliedby NIDA) were used. The base wastritium labeled at the 15 and 16 posi-tions.

Chemical and polymer were blended intoan intimate, uniform mixture using thesolvents tetrahydrofuran and methylenechloride. The solution was cast andthe solvent removed by evacuation(<1 mm Hg) at 50°C for at least 24hours. After drying, the chemical/polymer matrix was formed into implant-able shapes. Beads 1/16" diameter wereprepared by transfer molding and 1/16"diameter rods by extrusion. A thin,polymer coating was added to selectedsamples by dipping the beads or rodsinto a 10 wt % polymer solution. Uponremoval from the dip-coat bath, thesample was dried under vacuum at 50°Cfor 48 hours. The resultant coatingwas estimated to be 0.001 to 0.003 ofan inch thick.

In vitro analysis

The in vitro release rate of chemicalinto 50 ml of 37°C, pH 7 phosphatebuffer was determined by measuring theconcentration of naltrexone in the buf-fer daily. The sample was suspended inthe solution inside a Whatman extrac-tion thimble. The 50 ml buffer solutionwas changed weekly in order to maintainthe agent concentration below 20% ofsaturation. The naltrexone base concen-tration was determined by means of a

Beckman LS100 liquid scintillator.R i a f l u o r ® (New England Nuclear) wasused as the cocktail. Naltrexone pam-oate concentration was measured with aHilger-Watts spectrophotometer.

In vivo analysis

Male albino Charles River mice (18-22 g)were used throughout the study. Themice were anesthetized with Penthrane(Abbott) and a small slit was made inthe scapular region. A single rod orthree beads were inserted below the slitand the wound was closed by a silksuture. The animals were allowed torecover. Groups of ten animals werethen placed in cages and allowed to de-velop normally until the test date.Ten individual mice were placed in sep-arate metabolism cages for collectionof urine.

The urine was collected daily for theduration of the experiment. The cageswere rinsed with distilled water andthe washings were added to the urine.The combine and washings were urinemade up to 10 ml with water. A 1.0 mlsample was taken and added to 10.0 mlof Aquasol liquid scintillation cock-tail (New England Nuclear) in a vial.The radioactivity was measured using aBeckman LS-230 liquid scintillationcounter. A quench curve was constructedusing a series of 3H quench standardsand this curve was used to correct allcounts.

The mouse tail flick procedure was usedthroughout this study. A ten secondcut-off time was employed, and a controlreaction time of two to four secondswas used. A rheostat, incorporated intothe instrument was used to adjust theintensity of the light falling on thetail of the mouse such that each reac-tion time fell within the stated range.Animals with a control reaction timeoutside the stated range were rejected.The rheostat adjustment was only madeif a significant proportion (more thantwo out of every ten mice) of thereaction times were outside the rangeof two to four sec. Groups of ten micewere medicated with an ED80 dose(14.0 mg/kg ip) of morphine sulphateintraperitoneally and re-exposed to thenoxious stimulus twenty minutes later.The analgesic response was calculatedas the percentage of the maximum pos-sible response time (Harris and Pierson,1964). The following formula was usedfor these calculations: (test-control/

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10-control) x 100 = % maximum possibleef fect .

Each group of animals was tested by thenarcotic antagonist procedure only once.Groups were planned so that testing wasperformed every 7 or 10 days.

At the termination of each experiment,the animals were sacrificed and theimplant site was examined. The condi-tion of the implanted material was notedand signs of encapsulation, injectionor irritation.

RESULTS AND DISCUSSION

The objectives of this ongoing study areto develop implantable, biodegradablenaltrexone - polylacticlqlycolic acidmatrix systems which will sustain thedelivery of chemical to a biologicalsystem for both one and six months.Primary to achieving these objectivesare the development of reproduciblepolymerization procedures using pharma-cologicallv suitable catalysts. thefabrication of matrices from-polymerswhich hydrolyze within specified times,and the selection of a practical steri-lization process. Also, as the studyproceeded, it was expected that a rela-tionship between the in vitro and invivo performances of the implants wouldbe developed.

Experiments were performed to ascertainthe effects on chemical release rateand duration of the

degree of chemical loading,solubility of the chemical inboth the surrounding environ-ment and the polymer,surface area/weight ratio andporosity of the matrix, andhydrolysis rate of the polymer(i.e., composition of the polymer).

Quantitative evaluations of most ofthese parameters have been completed.

The degree of chemical loading and thesolubilities have been identified asmost influential on the performance ofthe implant. The duration of chemicaldelivery is inversely proportional tothe drug loading level and the solu-bility of the chemical in the surround-ing environment. Thus, matricesdesigned to meet the six-month goal areprepared with naltrexone pamoate whichis less soluble in aqueous solutionsthan naltrexone base. The duration isdirectly proportional to the solubilityof the drug in the polymer. Chemical/

polymer solubility effects are especial-ly important in this study - the baseform of naltrexone forms a solid solu-tion with polylactic/glycolic acids.This behavior permits the use of thevery water soluble naltrexone base forlong duration implants.

In other experiments it has been deter-mined that dip-coating the implantswith a thin layer of polylacticlglycolicacid reduced the initial surge of chem-ical from the surface of the implantand extended the duration of delivery.The releases of chemical from rods,spheres, and micron-size particle ma-trices have been measured. As expectedthe rate is directly proportional tothe surface area/unit weight of implant.

Based on the above findings, matriceshave been fabricated specifically tomeet the forementioned goals. Thesesystems have been tested in vitro andin vivo. One matrix system, dip-coatedrods of 33% by weight naltrexone pam-oate in 90 L(+)/10 polylactic/glycolicacid, has proved antagonistic to mor-phine challenge for 180 days in mice( 8 mg naltrexone pamoate/mouse).Using the one-to-one in vitro-to-invivo relationships which has been-observed throughout the study, it isanticipated that this system will con-tinue to release chemical at an antag-onistic level for 20 more days.Inspection of the implant site after180 days found no signs of foreign bodyreact ion ( i .e . , no inflamation orwalling-off of the implant) and a rodwhich crumbled when probed'with forceps.It is concluded that the formulation ofthese rods is ideal for a 6-month nal-trexone delivery system.

With another system, uncoated beads of33% by weight 3H-naltrexone base in90 L(+)/10 polylactic/glycolic acid,chemical releases were measured bothin vivo (urine of mouse) and in vitroand significant levels of antagonism tomorphine challenge was realize; for55-60 days. Plots of the cumulativeresults are presented in Figure 1. Thedaily levels of 3H in the urine reportedas percentage of total 3H implanted arepresented graphically in Figure 2. Inthis test, the release rate approxi-mated zero-order throughout a majorportion of the delivery period; theone-to-one in vitro-to-in vivo relation-ship was clearly demonstrated: andsimilar to other tests in this studyroughly 50 percent of the implanted 3H

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was found in the urine. Since therelease duration is inversely propor-tional to loading, it is hypothesizedthat an uncoated bead matrix loaded to50% by weight with naltrexone base willmeet the 30-day goal.

The above two systems exemplify typicalsample evaluation procedures andresults. The samples were preparedwith polymers synthesized from lacticand glycolic acids. Extensive study ofthe polymerization has identified re-actant purity and ambient atmosphere asthe most influential factors affectingthe molecular weight of the polymer.Reactants with sharp melting points andan oxygen and moisture-free atmosphere(e .g . , <1 mm Hg vacuum) are required.Using infrared analysis, the composi-tions of the copolymers were determinedto be similar to the reactant ratiocharged to the reaction vessel. Thispoint is important to note since somecatalysts react preferentially with onereactant over another. It has also beenconcluded that the product is a random,straight-chained polymer.

In anticipation of more intensive ani-mal tests, and ultimately, clinicaltr ials , sterilization procedures havebeen screened to select a method appli-cable to the polylactic/glycolic acidmatrix drug delivery system. Normalmethods such as autoclaving and hightemperature, dry heat cannot be used

since the temperatures are above themelting point of the polymer. And be-cause the implant is porous, ethyleneoxide should not be used because com-plete degassing cannot be ensured.Therefore, radiation sterilization wasselected for evaluation. Several typesof irradiation (e.g., high energy elec-trons and gamma ray) are under evalua-tion.

SUMMARY

The utility of the implantable poly-lactic/glycolic acid matrix system as ameans of providing the sustained deliv-ery of naltrexone for periods up to 200days has been demonstrated. Concurrentwith drug delivery, the polymer hydro-lyzes to non-toxic metabolites whichare naturally eliminated. No foreignbody reaction to the implants has beenobserved in mice. A one-to-one corre-lation between in vitro and in vivoperformance (i.e., release of chemical)has been established.

Pharmacologically suitable catalystshave been used to reproducibly preparepolymers under controlled reactionconditions. Polymers have been char-acterized using standard techniques anda relationship between intrinsic vis-cosity and weight average molecularweight established. Irradiation hasbeen selected as the most practicalmethod of sterilization for polylacticlglycolic acid implants.

18

DEVELOPMENT OFINJECTABLE MICROCAPSULESFOR USE IN THE TREATMENTOF NARCOTIC ADDICTION

Curt Thios, Ph.D.

Department of Chemical Engineering,Washington University,St. Louis, Missouri 63130

Long-acting injectable narcotic antagonistformulations represent a potentially usefulmeans of treating heroin addicts. Conceptu-ally, the necessary slow release drug formu-lations can be prepared by microencapsulatingthe antagonist and then simply injecting thecapsules. The capsule wall should cause thecapsules to release their payload slowly,thereby providing prolonged antagonisticaction. This project is concerned with re-ducing the above concept to practice. Theoverall goal is to synthesize injectablemicrocapsules that will last one to six monthsin vivo.

The concept of an injectable microcapsule isrelatively simple, but the capsules usedmust meet a number of stringent requirements.Naturally, a prime requirement is that theyrelease their drug payload slowly and performfor four weeks or more in vivo. This requiresdeposition of relatively defect-free capsulewalls around individual drug particles. Thecoating used to form the microcapsule wallmust be biocompatible and bioabsorbable. Therate of bioabsorption preferably must besuch that the capsule wall will be completelyabsorbed within six to twelve months afterinjection.

The capsules must be small enough to freelypass through a 21 gauge needle. This re-quires capsules smaller than 150 to 200µ.

The capsules preferably will be injected asa saline dispersion and the maximum volumeof capsule dispersion administered as a sin-gle injection should be one to two cc.

In an effort to prepare capsules that meetthe above requirements, several new encapsu-lation procedures have been developed.D1-poly(lactic acid) (d1-PLA) and lactide/glycolide copolymers are used to form thecapsules because they are established bio-compatible and bioabsorbable polymers. Narcoticantagonists encapsulated successfully includecyclazocine free base, naltrexone free baseand naltrexone pamoate. Active contents ofmost capsules isolated range from 50 to 75wt. percent. At least 40 to 50 percent ofthe capsules isolated from a given capsulebatch are less than 300µ diameter; in sev-eral cases, up to 85% of the capsules iso-lated from a given batch are below 180µdiameter. Although capsules with lactide/glycolide coating have been made, thus farcapsules which release their antagonist pay-load the slowest have a d1-PLA wall.

Capsule quality in vitro is established byextracting the capsules in a rotating bottleextractor (37°C; pH 7.4 phosphate buffer)and spectroscopically assaying the extractsfor the amount of antagonist released. Re-lease curves obtained in this manner are notzero order. The capsules consistently show

19

a relatively high initial rate of antagonistrelease which declines steadily as extractioncontinues. This behavior is attributed toa distribution in quality of the capsulesbeing evaluated. Scanning electron micro-graphs of capsule surfaces reveal that thewalls of some capsules contain numerousmacroscopic defects like craters and pitholes while other capsules are relativelyfree of such defects. It is hypothesizedthat the capsules with numerous defects re-lease their payload rapidly upon immersionin the extraction medium thereby yieldingthe observed burst of antagonist release.Once the defective capsules are empty, anta-gonist release continues as a steadily de-clining rate from capsules with progressivelyfewer coating defects.

Support for the above hypothesis is providedby a recent advance in encapsulation tech-nology. This advance enables one to isolatehigh yields of <180µ spherical capsules.When viewed under a scanning electron micro-scope, the walls of such capsules appear tohave substantially fewer defects than pre-vious capsules. The latest capsules releasenaltrexone free base over more than a two-week period in vitro and appear to be thebest capsules made to date. In vivo evalua-tions are now in progress, so their in vivolifetimes are unknown. Nevertheless, it isrelevant to note that the best previousnaltrexone pamoate capsules had an in vitrolifetime of about two weeks. They provided60 to 90% antagonism in vivo for 28 days- . -after injection when injected into micethrough an 18 gauge needle as a peanut oil/aluminum monostearate suspension at a 40 mgantagonist/kg mouse dose level. Percent

antagonism was established by the mousetail-flick test procedure with 10 mg mor-phine/kg mouse as the challenge dose.

Significantly, the above in vivo experimentwas not terminated because the capsules ceasedto function. Termination occurred becauseonly four groups of test animals were innocu-lated for the experiment and data pointswere obtained 7, 14, 21 and 28 days afterinjection. The capsules provided 80% anta-gonism on day 28 after injection, so it isclear that they still were releasing nal-trexone free base at that time. How muchlonger the capsules would have been effectiveis unknown. Current in vivo experiments arebeing carried out for longer periods and aredesigned to answer this question. The <180µnaltrexone pamoate capsules isolated recentlyare expected to last at least 30 - 35 daysand perhaps longer when injected as a peanutoil suspension. The latest capsules alsoare being injected as a suspension in salinecontaining 0.1 wt.% methylcellulose. Theultimate goal is to eliminate the methyl-cellulose entirely and simply use saline.

Naltrexone free base is far more water-soluble than naltrexone pamoate. Thus, ithas been more difficult to synthesize nal-trexone free base capsules capable of re-leasing their payload over a prolongedperiod. The best naltrexone free base cap-sules made to date provide 75 to 95% antago-nism for 14 days after injection. The in-jection medium was peanut oil/alminummonostearate; the dose administered was 40 mgantagonist/kg mouse. Recent encapsulationwork has focused exclusively on naltrexonepamoate, so it is not known yet if the latestencapsulation technology will yield longer-lasting naltrexone free base capsules.

20

LONG-ACTlNG NARCOTICANTAGONIST COMPLEXES

Allan P. Gray, Ph.D.

William J. Guordina, Ph.D.

IIT Research Institute

INTRODUCTION

The overall goal of this research is thedevelopmnt of improved drug therapy for thetreatment of narcotic dependence.

Narcotic antagonist drugs are of potentiaivalue for this purpose but suffer fran thedrawback of being too short-acting. Thecurrent consensus is that durations of theorder of atleast l-3months are desirable.Although such durations would not be accept-able for narcotic substitutes such asmethadone, renewed interest has recentlybeen expressed in methadone preparationseffective for periods of several days.1

Our approach to achieving the goal has in-volved the preparation, in vitro and in vivostudy of sparingly soluble salts and ionic-complexes of potentially useful drugs sincethere are mple precedents in othertherapeutic areas showing that intramuscularinjection suspensions of such preparationscan provide slow release of drug and a usefulprolongation of action (Gray, Robinson, 1972;

1974; Gray, 1974). Our earlier work, whichhas been reviewed (Gray et al., loc. cit.),cocentrated on complexes of narcoticantaganist drugs (cycclazocine, naloxone,naltrexone and, more recently, diprenor-phine). We are, however, currently turningsome of our attention back to the study ofrelated complexes of the narcotic replacementdrug, methadone.

Two of our preparations, naltrexone zinctannate (Gray, Robinson, 1974) and nal-trexone aluminum tannate (Gray, 1974) appearmost promising. Administered in an aluniummonostearate gel suspension by intramuscularinjection to mice, antagonist activityremains at a constant high level and then,drops off rapidly at the end of the period(Gray et al. loc. cit.; Reuing2). Durationso f activity in miceof 3-4 weeks have beenobserved. Toxicological studies have thusfar revealed no contra-indicative effects.3

The pharmacokinetics of these preparationsare being evaluated,2 as well as theireffects in addicted monkeys4.

21

REVIEW OF EARLIER WORK

We have evaluated the ability of close to 100mono- and polybasic organic acids to formwater-insoluble salts with methadone and oneor the other of the narcotic antagonist drugs,cyclazocine,nalcmone,naltrexoneand,morerecently, diprenorphine. About half theacids yielded insoluble salts. Polybasicacids affording insoluble salts wereevaluated for their ability to form drug:metal; acid complexes with the polyvalentmetal ions, Zn++, Al++,Mg++and Ca++.Optimum conditions for forming complexes havebeen developed and the consistencyoftheircomposition has been established.

Salts were analyzed spectrophotometrically fordrug content, and complexes were analyzed fordrug and metal content. The in vitro degreeof dissociation at equilibrium was determinedfor the preparations-suspended in a simulatedphysiological buffer, pH 7.3, at 37°.

RECENT IN VITRO STUDIES

The work reported here has been carried outin the period January l-April 15, 1975.

Naltrexone Complexes

Additional 15,16-3H-naltrexone aluminumtannate (6) was prepared. 15,16-3H-naltrexone base, specific activity 47.7mCi/mg,5 was diluted in methanol solutionwith cold naltrexone hydrochloride to give15,16-3H-naltrexone hydrochloride, specificactivity 0.63 µCi/mg.

To a stirred aqueous solution of 360 mg(10.6 meq) of tannic acid neutralized with10.6 meq of 0.5 N NaOH was added 10.6 mqof a freshly prepared 3 N aluminum nitratesolution followed by an aqueous solution of200 mg (5.3 mmole) of the 15,16-3H-naltrexonehydrochloride. The mixture was stirred forone hour, allowed to stand for 16 hours andfiltered. The water-washed precipitate wasdried to constant weight in vacuum, first atroom temperature and then at 60°. A yieldof 3.14 g of complex was obtained havingthe following analysis:

% Naltrexone 19.0% (spectrophoto-metric assay)

% Aluminum 2.4% (Eriochrome cyanine Rdye method)

% Dissociation 10.1% (at equilibrium inphysiological buffer, pH 7.3, at37°)

Preparations showing relatively low degreesof dissociation in vitro, since it earlyappeared that a high of dissociationcontraindicated a prolonged duration ofpharmacological action, were evaluated inmice after intramucular administration atseveral dose levels by the mouse tail-flicktest for the potency and duration of theirmorphine antagonist activity.

Cur most promising preparations to date,showing the most prolonged durations ofaction without evidence of gross toxicity, havebeen zinc and aluminum tannate comlexes. Inparticular, the zinc and aluminum tannatecomplexes of naltrexone are undergoing de-tailed evaluation as potential clinical candi-dates. Thus far, the most useful of severalinjectable dosage forms studied is asuspension in an aluminum monostearate gel inpeanut oil. The particle size of these com-plexes in suspension has been shown to bedistributed predominantly in a range below16µ.

Because of the interest in the naltrexonecomplexes, zinc and aluminum tannates ofradiolabeled (15,16-3H) naltrexone5 havebeen prepared and are being evaluated forMaintenance of blood and urine levels and

at Ohio State University.2for their pharmocokinetic behavior here and

Specific activity 0.7 µCi/mg drug base(by scintillation counting).

Naltrexone preparations supplied for outsideevaluation during this period are listed inTable 1.

Additional supplies of naltrexone zinc andaluminum tannate complexes have been pre-pared and analyzed.

Methadone Complexes

Methadone zinc tannate and methadone alumi-num tannate were prepared by the samemethod (method 6) described for naltrexonecomplexes by use of 2 equivalents ofneutralized tannic acid and 2 equivalents ofmetal salt (zinc sulfate or aluninumnitrate) per mole of methadone hydrochloride.

22

Table 1

LotDate Naltrexone Complex No Wt, g

3/4-3/18/75 Zinc tannate (6) C21885 750 mgE-8-20

3/4-3/24/75 Aluminum tannate (6) C21950 750 mgE-10-21

3/4/75 Zinc tannate (6) C22089 2.5 gE-14-13

3/4/75 Aluminum tannate (6) C22040 6.6 gE-12-10

4/16/75 15,16-3H-labeled C22142 1.25 gAluminum tannate (6) E-16-17

Recipient Study

D. A. McCarthy AddictedParke-Davis monkeys

D. A. McCarthyParke-Davis

C. W. MastriIndustrial Biotest

C. W. MastriIndustrial Biotest

R. H. ReuningOSU

Addictedmonkeys

Toxicology

Toxicology

Pbarmaco-kinetics

Analytical data are given in Table 2.

Table 2

Zinc Tannate Aluminum Tannate

% Methadone 31.9 27.6

% Metal 4.7a 2.0

% Dissociation 4.1 4.3

aBy atomic absorption.

To assist in the in vivo evaluation ofmethadone complexes,their radiolabeledcounterparts have been prepared. o,o'-3H-Methadone base, specific activity 53.9 mCi/mg,5was diluted in ethanol solution with coldmethadone hydrochloride to give o,o'-3H-methadone hydrochloride, specific activity1.2 µCi/mg. o,o'-3H-Methadone zinc tannateand aluminum tannate complexes have beenprepared fran the tritium-labeled hydrochlorideby method 6; analysis is not yet complete.

Thin-Layer Chranatography of RadiolabeledHydrochloride Salts

The purity and stability of 15,16-3H-nal-trexone hydrochloride and o,o'-3H-methadonehydrochloride are being studied by thin-layerchromatography. After experimentation withthe cold salts, identifying spots by UV-fluorescence and by various spot reagents,

we have elected to work primarily with thefollowing two solvent systems:

a) chloroform:methanol:ammonia (90:10:1)

b) n-butanol:acetic acid:water (60:15:30)

With the tritim-labeled bases and hydro-chloride salts, chromatograms were developedon Polygram Sil G sheets. The sheets were cutinto l-cm-wide strips and the radioactivityof each strip was determined by scintillationcounting. Significant amounts of impuritieswere noted in the radiolabeled hydrochloridesamples. Further investigation is in progress.

Dissociation Under Simulated Sink Conditions

It had previously been noted that when anamount of naltrexone zinc or alluminum tannatecomplex calculated to contain 15 mg of drugbase was magnetically stirred in 10 ml ofisotonic phosphate buffer, pH 7.3, at 37°,equilibrium dissociation of the complex wasattained in less than 15 minutes. Therefore,in order to simulate sink conditions, thefollowing procedure was adopted.

An amount of complex calculated to contain15 mg of naltrexone base was stirred in 10 mlof buffer, pH 7.3, for 15 minutes at 37+0.l°.The mixture was centrifuged, an 8-ml aliquotof the supernatant removed and replaced by 8ml of fresh butter. The process was repeated.The concent ration of naltrexone in eachremoved aliquot was determined spectrophoto-metrically in 0.1 N hydrochloric acid in theusual way.

23

Results for naltrexone zinc tannate are shown The amount of drug remaining in the residualin Figure 1. A total of 10.7 mg, approximate- complex was determined spectrophotometri-ly 70%, of the naltrexone had been removed cally to be 2.3 mg.by 26 portions of buffer. Although, as can beseen from Figure 1, the amount of drug re- For comparison, a parallel experiment isleased with each portion of buffer was not being carried out with naltrexone aluminumconstant, it does follow a smooth curve which tannate.appears to be approaching zero releaseasymptotically.

RECENT IN VIVO STUDIES

Diprenorphine Complexes

ED80 Determinations

The ED80 dosages for diprenorphine HC1,diprenorphine zinc tannate, and di-prenorphine aluminum tannate in peanut oilare 0.03 mg/kg, 0.048 mg/kg, and 0.092 mg/kg,respectively. The antagonistic effect ofdiprenorphine to the analgesic effect ofmorphine (4Omg/kg/ip) on the tail flicktest was measured 40 minutes after theintramuscular injection of diprenorphine and30 minutes after the intraperitonealinjection of morphine. The ED8O dosages forthe diprenorphine preparations are somewhat

24

greater than those for the long actingpreparations of naltrexone possibly becausediprenorphine is released at a slower ratefromthe site of injection. The duration ofantagonism of analgesia by the ED80 doseof diprenorphine hydrochloride is shown inTable 3.

Antagonism of Analgesia

Table 4 shows the duration of the antagonismto morphine of the long acting preparationsof diprenorphine (4 mg/kg in peanut oil)compared to that of the hydrohloride.

The diprenorphine complexes exhibiteddurations of action comparable to those ofthe corresponding naltrexone complexes at

Table 3

ANTAGONISM OF ANALGESIA BY THE ED80 DOSEOF DIPRENORPHINE

%Antagonism Produced by a DoseContaining 0.03 Mg/kg. Drug at:

Drug Hours(in peanut oil) 1 3 5

Diprenorphine HCl 82 64 24

Table 4

ANTAGONISM OF ANALGESIAa

Drug(in peanut oil)

Diprenorphine HCl

%Antagdsm Produced by a DoseContaining 4.0 mg/kg. Drug at:

Hours

94 95 83 16 - - -

2 4 5 24 48 72 96

Diprenorphine ZincTannate - - 98 63 14 19 -

Diprenorphine AluminumTannate - - 97 78 64 42 16

aMorphine 40 mg/kg/ip injected 30 min before the tail flicktest.

this dose level in peanut oil. Diprenor-phine appears to offer no advantage induration of action and, moreover, exhibitsdefinite agonist properties.

Methadone Complexes

Measurement of the duration of action of longacting preparations of methadone is beingattempted by radiolabeling as well as pharm-cologicaltechniques. In one series ofexperiments which will begin shortly, radio-labeled methadone HC1, methadone zinc tannate,or methadone aluminum tannate will be in-jected intramuscularly in mice. The plasmaand brain tissue levels of radioactivity fromthese injections will then be measured overseveral days.

In another series of experiments, methadoneHCl, methadone zinc tannate, ormethadonealuminum tannate (4O mg/kg methadone base)is substituted for morphine in morphinedependent mice. Mice are addicted tomorphine by a series of intraperitonealinjections of morphine given at one hourintervals as described (Iorio et al., 1975).

The following five doses of morphine areadministered in ascending order: 12.5, 25,50, 50, and 50 mg/kg/ip. An intramuscularinjection methadone preparation is sub-stituted for morphine at the sixth injection.The duration of the addiction in these miceis tested by the presence of jumpingprecipitated by naloxone (25 mg/kg/ip).

Groups of mice were tested at 1, 24, 48, and72 hours after the intramuscular injection.Mice receiving the vehicle (peanut oil) ormethadone (40 mg/kg) jumped at nearly thesame rate following an injection of naloxone(25 mg/kg) at each of the test periods.These studies are being continued.

In another group of experiments, mice wereaddicted to morphine with injections giventwice daily for two weeks; they receivedmorphine (150 mg/kg/ip) on the last twodays. These studies showed that methadone(4O mg/kg) zinc tannate in peanut oilvehicle prolonged naloxone (10 mg/kg/ip)induced jumping by two days as compared tovehicle and methadcne HC1 treated mice.

25

Radiolabeled Naltrexone3H Naltrexone HC1 (4mg/kg) in the aluminummonostearate vehicle was injected intra-umscularly in mice. Mice were sacrificed bydecapitation 1, 2, 4, and 24 hr after theinjection. The brains of the mice wereremoved and immediately digested in NCS tissuesolubilizer. The plasma fraction of theblood was separated via centrifugation. Thelevelofradioactivity in each sample asmeasured 24 hrs later (see Table 5).

The plasma levels of radioactivity followingan intramuscular injection of naltrexone HClin an aluminum monostearate gel vehicledecreased steadily over the 24 hr samplingperiod. In contrast, the brain levels ofradioactivity did not decrease during thesame period of time. These results suggestthat the plasma levels of naltrexone do notdirectly relate to the brain levels ofnaltrexone. The work is being continued.

Table 5

PLASMA AND BRAIN LEVELS AFTER INTRAMUSCULAR INJECTIONIN MICE OF RADIOLABELED NALTREX0NE HC1IN ALUMINUM MONOSTEARATE GEL VEHICLE

(Hours)

Level of Radioactivity in Plasma and Brainas ng/ml and ng/g, Respectively,

Time 0f NaltrexonePlasma Brain

1 408 86

2 380 77

4 200 79

24 128 a

48 a a

aExperiments in progress.

FOOTNOTES REFERENCES

1. R. E. Willette, personal communication.

2. R. H. Reuning, NIDA Progress Reports,persona1 communications.

3. C. W.Mastri, personal communication.

4. D. A. McCarthy, Parke-Davis.

5. Obtained from J.A. Kepler, ResearchTriangle Institute.

Gray, A. P., Conference on BiomedicalResearch in Narcotic Abuse Problems,Vancouver, B. C., November 1974; Proceedingsin press.

Gray, A. P. and Robinson, D. S., FirstInternational Conference on NarcoticAntagonists, Airlie House, Warrenton, Va.,Nov. 1972; Proceedings: NarcoticAntagonists, M. C. Braude, L. S. Harris,E. L. May, J. P. Smith and J. E.Villarreal, Ed., Advances in BiochemicalPsychopharmacology, Vol. 8, Raven Press,New York, 1974, pp: 555-568.

Gray, A. P. and Robinson, D. S., J. Pharm.Sci., 63, 159 (1974).

Iorio, L. C., Deacon, M. A. and Ryan, E. A.,J. Pharmacol. Exptl. Therap., 192, 58 (1975).

26

SUSTAINED RELEASE OFNALTREXONE FROMGLYCERIDE IMPLANTS

M.F. Sullivan Ph.D.D. R. Kalkwarf, Ph. D.

Battelle, Pacific NorthwestLaboratories

INTRODUCTION

This research is concerned with de-veloping injectable and biodegradablesystems that release the narcoticantagonist, naltrexone, in vivo forat least 30 days at pharmacologicallyeffective rates. Such preparationsshould improve the efficiency oftreatment of heroin addicts withnaltrexone by compensating for therapid metabolism of this antagonist.Natural glycerides that are solids atbody temperature yet can be meltedand molded at relatively low tempera-tures appear to be appropriatevehicles for delivering the drug ateffective and constant rates. Thesesubstances provide a reservoir fromwhich the drug can diffuse graduallyduring treatment and that is itselfeventually absorbed by the body withnegligible toxic effects, This re-port describes their utilization innaltrexone-delivery systems.

DEVELOPMENT AND TESTING PROCEDURES

Implant preparation:

Small pellets and particles were pre-pared containing naltrexone dispersedhomogeneously in various glycerides.

Crystals of naltrexone (diameter <125µ)were added with vigorous stirring tomolten combination of a-monopalmitin,1,2- and 1,3- dipalmitin and tripal-mitin at 80 C. To form pellets,the liquid suspensions were drawn -in-to cylindrical molds, 1.7-mm indiameter, and cooled rapidly untilthey solidified. The cylinders wereextruded, cut into 3-mm lengths foruse in mice and into 10-mm lengths foruse in rats, rabbits, monkeys andswine. Particles were prepared in twosize ranges, 125-250 µ and < 125 µ bygrinding and sieving the solid dis-persions, They were then suspendedin an aqueous solution containing1.5% methyl cellulose at a concentra-tion that would allow injection ofthe desired amount of naltrexone ina 0.25 ml volume.

In Vitro Evaluation

The release rates of naltrexone fromthe preparations were measured invitro to determine if the releasecharacteristics justified furthertesting in vivo. The in vitroevaluation was done by placing asample in a continuous flow of iso-

27

tonic saline solution at 37 ºC using from the interior, The rate of drugthe apparatus shown in Figure 1. release into this "perfect sink" wasThe flow rate was increased until determined from spectrophotometricnaltrexone was removed from the measurements on the effluent solu-surface as fast as it diffused there tion at 281 nm,

FIGURE 1. CONTINUOUS FLOW SYSTEM FOR DETERMINING DRUG RELEASERATES IN VITRO

In Vivo Evaluation according to the procedure of D'Amourand Smith, 1941 as modified for mice

The mice used in these studies were by Harris et al., 1969, At least sixof the Swiss-Webster strain weigh- antagonist-implanted mice, twoing between 20-30 gm, Experimentaland control animals were maintainedin groups of 15 and fed Purina mousepellets ad libitum. Their weightswere determined weekly, The in vivo

vehicle controls and two normal con-trols were used for each test, unlessotherwise specified,

At the conclusion of the pharmacologicmeasurement, the mice were anesthe-tized, the pellets were removed, andthe implantation sites were inspectedfor any gross signs of tissue incom-patibi l i ty . Residual naltrexone ineach pellet was determined spectro-photometrically after dissolving itin chloroform. Residual glyceridewas determined by esterifying asample of the chloroform solutionwith methyl alcohol in the presenceof boron trifluride and determiningthe amount of ester with a gaschromatograph.

measurement of the pharmacologiceffectiveness of the various prepara-tions was done by either implantingpellets subcutaneously into the napeof the neck with a 12-gauge trocaror by injecting a suspension ofnaltrexone loaded particles in 1.5%methylcellulose as the suspendingagent. Both vehicle-implanted andnormal mice were used as morphine-injected controls, A dose of 15 mg/kgof morphine sulphate was injectedsubcutaneously and the tail-flick re-sponse was measured 30 minutes later

28

FIGURE 2. RELEASE OF NALTREXONE FROM 1.7 x 10.0 mm PELLETSCONTAINING 30% NALTREXONE IN 25:75 DIPALMITIN:TRIPALMITIN.

FIGURE 4. RELEASE OF NALTREXONE IN MICE FROM A 1.7 x 3.0 mmPELLET CONTAlNlNG 30% FREE BASE IN 25:75 DIPALMITIN:TRIPALMITIN AND COMPARISON WITH THE REACTION TIMEOF THE MICE IN THE TAIL-FLICK TEST.

FIGURE 3. RELEASE OF NALTREXONE IN MICE FROM A 1.7 x 3.0 mmPELLET CONTAINING 0.3% FREE BASE IN 25:75 DIPALMITIN:TRIPALMITIN AND COMPARISON WITH THE REACTION TIMEOF THE MICE IN THE TAIL-FLICK TEST.

FlGURE 5. RELEASE OF NALTREXONEAND RESULTINGANTAGONISTIC EFFECT FROM A 1.7 x 3.0 mmPELLET CONTAINING 30% NALTREXONE (FREEBASE IN DIPALMITIN AND IMPLANTED S.C.IN MICE

29

RESULTS

An early preparation that we evalu-ated was a solid implant, 1.7 x 10mm, that was loaded with 30 percentnaltrexone. The pellet matrix wasdipalmitin:tripalmitin in theratio of 25:75. The release rateinto isotonic saline at 37 C under"perfect sink" conditions isshown in Figure 2 along with thein vivo release that occurred fromsubcutaneous implants that wereremoved in groups of 3 at intervalsafter treatment. The release rateof naltrexone in vitro graduallydecreased with time as its concen-tration near the surface diminished;however, the release rate in vivoremained constant, indicating thatbiodegradation had continually ex-posed fresh surface, Rats that wereimplanted with a single pellet wereunaffected by morphine for 33 daysafter treatment as judged by theirtail-flick response.

Small pellets (1.7 x 3.0 mm) wereimplanted in mice to determine theeffect of the naltrexone dose onthe pharmacologic response tomorphine analgesia. The naltrexone-release rate and the duration ofaction of the implants is partiallydependent on the concentration ofthe antagonist in the glyceridematrix. This can be seen inFigure 3 and 4 where a 0.3% naltrex-one loading resulted in the releaseof 2.0 µg/day. However, bothpreparations were ineffective forantagonizing morphine beyond 2 weeksafter they were implanted.

The duration of naltrexone effec-tiveness by a single injection isextended when it is administered asnaltrexone pamoate. Pellets contain-ing various combinations of dipal-mitin and tripalmitin along withnaltrexone pamoate were tested fortheir release characteristics invitro. Naltrexone pamoate wasreleased from a dipalmitin-tripal-mitin matrix more rapidly than whenit was present as the free base,This formulation appeared to be un-suitable for providing 30-days ofeffective antagonism.

Pellets loaded with 30% naltrexonein an equal mixture of 1,2- and1,3- dipalmitin were tested bothin vitro and in vivo and were found

to release the antagonist at pharma-cologically effective rates for 30days, This is shown in Figure 5.Three of the ten mice tested at eachtime interval were implanted withpellets containing 3H-naltrexone.Urine collected daily from theseanimals during the 2 days prior tothe tail-flick test showed thaturinary excretion of tritium paral-leled the in vitro release ofnaltrexone as shown in Figure 6.Both types of data show that therelease rate of naltrexone was highinitially but decreased rapidly toa constant value, Our earlierstudies with 3H-naltrexone showedthat approximately 50% of the dosecould be recovered from the urineof mice after administration bysingle injection or by continuousinfusion. It was on this basisthat the quantity of naltrexonemetabolized daily was calculated.The dis appearance of drug anddipalmitin from 1.7 x 3.0 mm pelletsis shown in Figure 7. Earlier testsshowed that 23 of 29 1.7 x 10 mmdipalmitin implants disappeared com-pletely from intramuscular sites inthe buttocks of rabbits within 3months. The data points in Figure7 are not sufficient to predict thedisappearance rate of the 1.7 x3.0 mm pellets in mice accurately,but a line is drawn through them topresent a "best estimate" of thebiodegradation profile.

In an effort to obtain pellets thatbiodegraded more rapidly, glyceridemixtures containing either 10% -monopalmitin and 90% dipalmitin or20% a-monopalmitin and 80%-dipalmi-tin were loaded with 30% naltrexoneand tested, The biodegradationrates of the preparations did appearto be more rapid, but the releaserates of naltrexone were also great-er, as shown in Figure 8. Conse-quently, the drug contents of thepellets were depleted within a fewdays.

In a further effort to obtain effec-tive preparations that biodegradedmore rapidly, particles containing30% naltrexone dispersed in eitherdipalmitin or tripalmitin were pre-pared and tested. Suspensions ofthese particles in aqueous methylcellulose solution were injectedsubcutaneously in mice and testedat intervals thereafter for theirantagonist potency by the tail-

30

FIGURE 7.

FIGURE 6. COMPARISON OF NALTREXONE RELEASE RATESFROM A PELLET IN VITRO AND AS DETERMINEDBY URINARY EXCRETION OF 3H-NALTREXONEFROM MICE. PELLETS WERE 1.7 x 3.0 mm INSIZE AND CONTAINED 30% NALTREXONE INDIPALMITIN

LOSS OF NALTREXONE AND GLYCERIDE FROMA 17 x 3.0 mm, 7.8 mg PELLET IMPLANTEDSUBCUTANEOUSLY AND CONTAINING 30%NALTREXONE (FREE BASE) IN DIPALMITIN

FIGURE 8.

FIGURE 9. EFFECTIVENESS OF 8.0 mg OF TRIPALMITIN( )OR DIPALMITIN ( ) PARTICLESOF DESIGNATED SIZE CONTAINING 30%NALTREXONE FOR ANTAGONIZING MORPHINE(15 mg/kg) USING THE MOUSE TAIL-FLICKTEST

COMPARISON OF IN VIVO AND IN VITRORELEASE OF NALTREXONE FROM 1.7 x 3.0 mmPELLETS CONTAINING 30% NALTREXONE(FREE BASE) IN DIPALMITIN: MONOPAL-MITIN MIXTURES, 100.0, 90:10, OR 80:20

31

flick response. The results areshown in Figure 9. This prelimin-ary evaluation in only a few miceshowed that particles less than125 µ and dipalmitin particles wererelatively ineffective for anyprolonged duration of action. Onthe other hand evaluation in atleast 6 mice per interval showedthat tripalmitin particles in the125-250 µ range provided at leastone month of antagonist activity.Further refinement of these par-ticles is now in progress to providea more uniform glyceride coatingthat may extend the life of thispreparation.

No appreciable tissue response wasfound after implantation of eitherdipalmitin or tripalmitin in rats,rabbits, or swine at intramuscular,subcutaneous, or intraperitonealsites. No significant amount offibrosis was seen as long as 1 yearafter treatment with the slowerbiodegrading glycerides. There hasbeen no gross indication that anyof our more recent preparations aretoxic.

REFERENCES

1. D’Amour, F. E. and Smith, D. L.A Method for Determining Loss ofPain Sensation, J. PharmacoL. andExptl. Therap. 72, 74-79, 1941.

2. Harris, L. S., Dewey, D. L., Howes,J., Kennedy, J. S., and Pars, H.Narcotic Antagonist Analgetics,Possible Cholinergetic Mechanisms.J. Pharmacol and Expt'l Therap.169, 17-22, 1969.

CONCLUSIONS

These studies have shown thatglycerides and naltrexone can becombined to regulate the release ofthat antagonist in-laboratoryanimals for periods of at least onemonth. The dose released is depen-dent on the drug concentration with-in the matrix of the release ve-hicle and on the glyceride composi-tion of the matrix itself. The rateof biodegradability of a solid im-plant can be increased by increas-ing the proportion of the lowermolecular weight glyceride in thematrix. However, this change alsospeeds up the drug release ratethereby limiting the usefulness ofthis manipulation. Hopefully, thebiodegradation rate may also beincreased by administering slow-release vehicles in particle formdue to the increased surface area.

This form of the vehicle would beless retrievable by an addictthat wished to remove the implantand may be preferred by thetherapist. Our current efforts aredirected toward improving both theparticles and the pellet implants inorder to offer a range of slow re-lease devices that can be chosen tofit the needs of the addict beingtreated with narcotic antagonists.

32

USE OF SYNTHETIC POLYPEPTIDESIN THE PREPARATION OFBIODEGRADABLE DELIVERY VEHICLESFOR NARCOTIC ANTAGONISTS

Kenneth R. SidmanDouglas L. Arnold, Ph.D.William D. SteberLita NelsenF.E. Granchelli, Ph.D.

Suresh G. ShethARTHUR D. LITTLE, INC.

INTRODUCTION

Under contract No. ADM-45-74-218 we have beenworking on the development of practical,clinically acceptable narcotic antagonistdelivery vehicles prepared from syntheticpolypeptides such as glutamic acid/leucinecopolymers. The synthetic polypeptides werechosen because they potentially exhibitdiffusional release rates and biodegradationrates in the regions of interest, and havebeen shown to be biocompatible. Two types ofsystems are being sought: One capable ofadministering naltrexone for one month andanother for approximately six months. It isdesired that these systems biodegrade at asufficiently rapid rate that they are essen-tially "absorbed" as soon after they areexhausted of antagonist as possible.

In addition, in view of the constantly evolv-ing concepts advanced in the areas of antagon-ist development, psychopharmacology, andprotocols for patient treatment, it isimportant that the delivery system developed

33

in this program be amenable to other antagon-ists, and be capable of being fabricated intoa variety of dosage forms, including integralshapes such as rods and fibers, as well asdiscontinuous forms such as microcapsules andmicrobeads.

Although the antagonist may be released fromthe matrix by diffusion, by the degradationof the matrix, or by a combination of bothmechanisms, we are pursuing the developmentof a system which releases naltrexone essentiallyby diffusion alone because, at this stage, sucha system can most easily be fabricated. Biode-gradation subsequently removes the implant matrix.

METHODS

Preparation of Polymers

The glutamic acid/leucine (Glu/Leu) copolymerswere prepared following the procedure of Fasmanet.al. (Fasman, 1964). The polymers were charac-

terized by infrared spectroscopy, ultravioletspectroscopy, intrinsic viscosity, and solubilityin N, N dimethy l formamide(DMF).

Preparation of Delivery Vehicles

Film-shaped implants were prepared by casting aDMF solution of the copolymer and naltrexone(labelled with tritium and exhibiting a specificactivity of 0.48 mCi/gram) into a teflon mold.By alternately casting and drying (at 40°C-60°C)thin layers of the solution, films 0.007cm to0.07Ocm thick were built up. The films were thenplaced in a vacuum oven at 50°C-70°C, 30 in. Hgvacuum for twenty-four hours to remove the lasttraces of solvent. Individual devices exhibitingtotal surface areas of one to two cm were cutfrom the films.

Rod-shaped implants 0.04cm to 0.08cm in diameterwere prepared by extrusion through a die. Typical-ly, finely ground polymer was dry-blended in thedesired proportion with the tritium-labellednaltrexone. Sufficient solvent was added to makea 50% solids paste. This was then placed in asteel compression mold that had a O.lcm diameterhole bored in its side (later the mold was modi-fied by drilling and tapping so that it wouldaccept a screw-in die). Rod was extruded at roomtemperature by exerting moderate pressure on theplunger. The rod was then dried to remove thesolvent.

Tube devices with an inside diameter of 0.18cmand wall thicknesses ranging from 0.0026cm to0.012cm were prepared by casting a solution ofthe polymer onto a rotating glass mandrel. Extru-ded naltrexone/polymer cores consisting of 80%naltrexone by weight were inserted into the tubes.Pure polymer plugs were then inserted and cementedinto the ends of the tubes.

In Vitro Evaluation of Naltrexone Release Rates

The delivery vehicles were rinsed briefly withFarle's Balanced Salt Solution (IX - withoutphenol red, Grand Island Biological Company)andwere placed in stoppered test tubes containing10 ml of the Farle's solution. The tubes wereplaced in a shaker water bath at 37°C and weregently agitated. Drug concentration of thesolution was periodically assayed by scintillationcounting techniques to permit an evaluation ofthe in vitro drug-release behavior of the copoly-mers. (The solutions were replaced sufficientlyfrequently that the naltrexone concentrationremained far below saturation, i.e. below

460 mg/100 ml).

Charles River CD #l mice were prepared for deviceevaluation by implanting the delivery vehiclesusing the following protocol:

IN VITRO EVALUATION

Surgery: Under pentobarbital Na(60 mg/kg) anes-thesia the drug release devices were implantedsubcutaneously on the back of a mouse.

Apparatus: Immediately after surgery, the micewere fitted with coprophagy cups and placed inmetabolic cages. The coprophagy cups assuredcomplete separation of urine and feces and thusallowed for an accurate evaluation of the relativemagnitudes of excretion by both routes. Urineand feces were collected daily for the firstweek. and two days per week thereafter.

The wire floors and draining portions of thecages were flushed to dissolve dried urine andassure a thorough removal of radioactivity fromthe cages. Feces samples were removed from thetail cups and held in air-tight vials untilanalyzed.

Analysis of Excreta: Small aliquots of dilutedurine (urine and rinse) were analyzed directlyby scintillation methodology. Radioactivity inthe feces was determined by homogenizing thesamples and then analyzing them using scintil-lation counting procedures.

Tail Flick Bioassay: A Dewey-Harris tail-flickapparatus constructed by Dr. William L. Deweyand his associates was used in this program.

Experimental animals were weighed and controltail-flick times were determined prior to anytreatment or manipulation. Animals whose tail-flick times were outside the range of 2-4 secondswere discarded. At various times after implan-tation, test tail-flick times were taken thirtyminutes following an AD80 dose of morphine -SO4 (17.0 mg/kg).

RESULTS

Films

Initially, model delivery systems in film formwere prepared using various naltrexone/polypep-tide compositions to assess the compatibility ofthe antagonist in the polymer. Naltrexone wasfound to be highly compatible with Glu/Leucopolymers, showing no tendency to bloom to thesurface of films containing at least 40% drugby weight. This result indicates that the

34

naltrexone/polypeptide compositions arecapable of being stored for prolonged periodsof time without changing their composition orstructure.

Films prepared from 40/60, (i.e., 40 mole per-cent leucine/60 mole percent glutamic acid),35/65, 20/80, and 20/80 Glu/Leu copolymerscontaining 10% to 40% naltrexone by weightwere evaluated in vitro in mice. The naltre-xone was found to be released by diffusion,exhibiting a diffusion coefficient that variedas a function of the copolymer composition andthe initial naltrexone loading. For example,the diffusion coefficient increases as theglutamic acid content is raised; and, at afixed glutamic acid content, it increases asthe initial naltrexone concentration is in-creased. A wide range in diffusion coeffi-cients can be achieved (see Table 1). These

TABLE 1

35/65 102040

20/80

10/90

values are within practical ranges of interestfor the development of delivery vehiclescapable of meeting the program goals.

The diffusion coefficients may be used topredict release rate versus time behaviorusing suitable equations. One useful expres-sion for release rate is the following.(Baker, 1974)

where D=diffusion coefficient, =thickness,t=time

EFFECT OF COPOLYMER COMPOSITION AND ANTAGONIST LOADINGON NALTREXONE DIFFUSION COEFFICIENTS

CopolymerComposition

(Glu/Leu)

40/60

NaltrexoneContent

Wt. %)

102040

2040

40

DiffusionCoefficient

(Cm2/Hr)

20 x 10-7

22 x 10-7

120 x 10-7

0.31 x 10-7

11.5 x 10-7

26 x 10-7

1.0 x 10-7

12 x 10-7

19 x 10-7

35

Time To ReleaseOne-Half of Initial Drug LoadingFrom Films With Thicknesses

Of

0.007 Cm20.070 Cm2

1.2 Hrs1.10.2

78 78002.1 210

0.9 90

24 24002.0 200

1.3

120 Hrs110

20

130

ImplantThickness

cm

0.061

0.056

0.048

0.046

0.043

0.043

InitialLoading

Implant ofDrugµg

5800

4900

6600

4050

4650

5500

TABLE 2

COMPARISON OF IN VIVO RELEASE BEHAVIOR OF 35/65GLU/LEU FILMS CONTAINING 20% NALTREXONE

Percent Antagonism*

Mouse 1 4 7 9 11 14 16 18 21 24 28-Days

GT - - 100 - - 100 - - 100 100 -

GL - 73 64 - 65 - - 100 100 - 100

G H - - 100 - - 100 - - 100 100 -

RL 100 - - 100 - - 100 - 100 100 -

G R - 100 - - 100 - - 100 100 - 100

RT 100 - - 100 - - 72 - 100 - 100

*Mice were challenged with 17 mg/kg morphine sulfateadministered i.p., and were tested after 30 min.

36

TABLE 3

CHARACSTICS OF 20/80 GLU/LEU RODSCONTAINING 40% NALTREXONE EVALUATED IN VITRO

Average DiffusionDiameter Weight Sample No. Coefficient

cm mg cm2/hr

Uncoated Rods

0.061 7.5 18546-75-40-78-3 1.3 x 10-8

0.065 7.3 18546-75-40-78-2

Rods with Three Coats

0.060 6.1 18546-76-3-40-79-l 1.0 x 10-8

0.068 6.5 18546-76-3-40-79-3

Rods with Five Coats

0.056 7.1

0.064 6.9

28546-76-5-40-80-3

18546-76-5-40-80-l0.38 x 10-8

37

With this expression, a comparison of thetheoretical release rate with the actualin vivo release rates may be made. Figure 1shows such a comparison for typical 35/65Glu/Leu films containing 20% naltrexone byweight. Here the actual urinary recovery ofradioactive drug and metabolites is plottedversus time. (In these experiments the urinaryradioactivity accounted for 80-90% of the totalexcreted, and, therefore, the urine dataprovided a relatively accurate indication ofthe rates of release from the implants). Notethat the actual release rates follow the theo-retical predictions quite well, confirming thatnaltrexone is released from films by diffusion.

Since a naltrexone dosage of l-2 µg/hr permouse is all that is necessary to block anAD8o dose of morphine sulfate, animals receivingthese film implants were blocked for more than28 days (see Table 2).

Rods

Solid rod shaped devices released naltrexonein a manner similar to that observed with thefilms, indicating that a prolonged releasecan be achieved, albeit at a rate that contin-uously decreases with tine (as predicted bystandard diffusion equations).

Rods prepared from 20/80 Glu/Leu, containing40% naltrexone by weight, were found to ex-hibit an unexpectedly high initial rate ofrelease, which could be substantially reducedby overcoating the rods with pure 20/80 Glu/Leu copolymer (see Table 3 and Figure 2). Rodsreceiving five coats of polymer exhibiteddiffusion coefficients that were constant withtime, and corresponding release rates thatwere constant when plotted versus the squareroot of tine. After approximately 70 days,10% of the initial drug was released fromthese devices.

Despite the prolonged release obtainable frommonolithic devices such as films and rods,it is clear that constant rates of releaseby diffusion cannot be achieved with thesedosage forms because the drug flux continu-ously decreases as the drug concentrationgradient within the device decreases.Tube forms, however, behave quite differently.

REFERENCESTubes

Tube devices filled with a saturated solutionof the antagonist (with excess solid present)will maintain the concentration gradient acrossthe tube wall at a fixed value. As a con-sequence, these devices are capable of deli-vering naltrexone at a constant rate over aprolonged period (until the solid phasedisappears).

38

Preliminary experiments with tubes have begun.In these early experiments, variations inrelease rate were encountered because of poortransport of naltrexone from the solid drugcore to the interior wall of the tube. Aspreviously stated, a constant release ratedepends on maintaining a saturated solutionof naltrexone in the interior of the tubes.The tubes in these experiments were onlypartially filled with liquid, resulting inonly partial utilization of the surface areaavailable for diffusive release.

We are currently studying techniques for pre-filling tubes with saline so that a saturatedsolution of naltrexone will be in contact withthe entire inner surface of the tubes. This willensure full utilization of the surface area fordiffusive release, and consequently higher andmore predictable release rates. Tubes with wallthicknesses ranging from 0.026mm to 0.12mm arebeing employed in this study so that an evalua-tion of the influence of wall thickness may bemade.

CONCLUSIONS

Synthetic polypeptides may be fabricated into avariety of naltrexone delivery vehicles capableof administering the antagonist for prolongedperiods of time. Tube dosage forms, in partic-ular, offer advantages for this application since

They my contain drug loadings as high as80% to 90% by weight, allowing a minimum overallsize of depot.

They would be expected to exhibit a con-stant rate of antagonist release (by diffusion),and hence a high utilization of the drug payload.

They would contain a minimum amount ofpolymer to be removed by biodegradation.

They may be administered by trocar injec-tion.

However, additional work is required to developtechniques for the preparation of vehicles thatrelease antagonist at a reproducible and constantrate.

Raker, R.W. and Lonsdale, H.K., "ControlledRelease: Mechanisms and Rates," ControlledRelease of Biologically Active Agents,Tbnquary, A.C. and Lacey, R.E. (Advances inExperimental Medicine and Biology,. 47)Plenum Press, New York, 41, 1974.

Fasman, G.D., Lindblow, C. and Bodenheimer, E.,Biochemistry, 3, 155, 1964.

DEVELOPMENT OF CHRONOMERS™FOR NARCOTIC ANTAGONISTS

Richard C. Capozza, Ph. D.Edward E. Schmitt, Ph. D.Lee Richardson Sendelbeck, M.S.

ALZA Corporation

BACKGROUND

The National Institute of Drug Abuse (NIDA)is sponsoring work to develop an erodible,long-acting delivery system for the con-trolled administration of a narcotic anta-gonist to assist in the management ofnarcotic addiction. Based on a favorablerelationship between potency and lack ofundesirable side effects, naltrexone hasbeen recommended as the drug of choice. Therequirements of the system are:

(1) Ability to release naltrexoneat prescribed rates for oneto six months.

(2) A shape and size which willpermit convenient parenteraladministration.

(3) Absence of significant ad-verse tissue response.

(4) Bioerodibility of the drug-delivering element.

ALZA has undertaken the task of developing asystem which is aimed at meeting theserequirements and which is based on a dis-

persion of naltrexone in one of ALZA'sbioerodible polymeric matrices.

OBJECTIVE

The objective of this program is to preparea bioerodible naltrexone delivery systemwhich can be implanted subcutaneously inhumans and which can release the narcoticantagonist over one to six months at rela-tively constant rates sufficient to block theeuphoric action of morphine-based drugs.

CHRONOMER™ MATRICES

These novel and proprietary polymeric systems,identified by the trademark CHRONOMER™, arebeing developed by ALZA Research, a divisionof ALZA Corporation, for use in the fabri-cation of drug delivery systems and aredesigned to undergo predictable erosion whenplaced in a biological environment. Ingeneral, the CHRONOMER™ matrix materials arehydrophobic and absorb little water. Withina given family of CHRONOMER™ matrix mate-rials, variations of molecular structure makepossible the preparation of polymers whichpossess a range of physical properties in-

39

cluding tough, glassy solids (C 101) and soft,compliant materials (C 101 ct).

The erosion process proceeds via hydrolysisat the surface of the CHRONOMER™ matrix.The drug contained within that surface layeris released into the surrounding tissues ata predetermined rate.the CHRONOMER™

The erosion process ofmatrices can be influenced by

incorporation of additives, particularlyalkaline materials, which modify the rate ofhydrolysis of the polymers. If a drug isuniformly dispersed within a CHRONOMER™matrix, the drug is released at a rate de-termined by (1) its concentration in thepolymer, (2) the surface area exposed towater, and (3) the pH of the microenvironmentat the interface between the matrix and itssurroundings.

The drug release rate at any time is propor-tional to the surface area of the matrix.With some system configurations, such as thindiscs, the surface area remains essentiallyconstant throughout the system lifetime andthus results in a constant drug deliveryrate. With other shapes, such as rods orspheres, the surface area decreases withtime, causing a decrease in drug deliveryrate.

The erosion rate of CHRONOMER" materials isinversely related to the pH of the aqueousphase in contact with the surface of thepolymer. If the drug is acidic, its releasecan increase the acidity of the environmentand hence can accelerate CHRONOMER™erosionand concomitant drug release. When the drugis basic, the local hydrogen ion concentra-tion is reduced and the CHRONOMER™ matrixerodes at a slower rate than predicted forthe pH of the bulk of the surrounding medium.

Since relatively slow release rates are re-quired for a naltrexone delivery system,Naltrexone free base, rather than naltrexonehydrochloride, is being used in CHRONOMER™/naltrexone composites, If necessary, releaserates can be modified by incorporating basesor buffers into the matrix.

Prior to the commencement of the present NIDAsponsored contract, CHRONOMER™ drug deliverysystems had been demonstrated to provide con-tinuous, and substantially constant, releasein vitro and in vivo. However, in some cases,there was a lag time of up to one week beforethe onset of the predicted rate of erosion.

CHRONOMER™ matrix materials are presentlymade in 150 g batches in a modified AtlanticResearch 2CV reactor. Our ability to controland prescribe reaction conditions leading toreproducible batches of polymer has improved

considerably during the last few years due toour independent effort to perfect these uniqueerodible matrix materials. For example,neither the molecular weight, which approaches100,000, nor the molecular weight distribution(Mw/Mn = 3.5) of these polymers has varied bygreater than 10% in the last five consecutiveruns. A larger Atlantic Research 1OCV hasbeen purchased by the Corporation and couldincrease our batch capacity by 200 fold. Theunit has just been received, is not installed,and is currently undergoing a thorough in-spection to evaluate compliance withspecifications.

Toxicological studies conducted at ALZA duringthe past two years have not revealed limitingadverse effects of either the CHRONOMERTM

materials or their hydrolysis products.

ANIMAL MODELS

The development of a long-acting, parenteraldelivery system for the controlled adminis-tration of naltrexone requires a demonstra-tion of efficacy in an experimental animal.The tail-flick test was recommended by NIDAas the means of evaluating the pharmacolog-ical activity of naltrexone. In this test,morphine delays the tail-flick response toexternally applied heat and the narcoticantagonist reduces or eliminates this delay.Either the rat or the mouse can be used inthis test; we selected the rat because it isthe more suitable for drug release kineticstudies in vivo.

The ability to perform satisfactorily thetail-flick test procedure was demonstrated.The test was used to measure the effectivenessof naltrexone to antagonize the analgesicaction of an 80% effective dose (ED80) ofmorphine (found to be 6.5 mg/kg body weight)in the rat. Single doses of naltrexoneranging between 2 and 20 µg/kg, injectedsubcutaneously in rats, were used to establisha single dose-response curve, A linearrelationship resulted when dose (log scale)was plotted against response (probit scale).An 88% antagonist effect was achieved with analtrexone dose of 20 µg/kg. An order-of-magnitude estimate was made to translate thesedata into a continuous drug delivery rate-response relationship. About 5 µg of nal-trexone/kg/hr was calculated to be necessaryto maintain > 80% antagonism.

Experiments to determine the parenteraldosage of continuously administered naltrexonerequired to antagonize the analgesic actionof an 80% effective dose (ED80) of morphine inthe rat were recently completed. Naltrexonewas infused intravenously at doses of 0, 4and 16 µg/kg/hr. The analgesic effect of an

40

expected ED80 dose of morphine was determinedat the end of six hours, by means of the tail-flick procedure.

Four and 16 µg/kg/hr naltrexone resulted in54 and 89% antagonism, respectively, againsta 63.5% effective dose of morphine. A moreeffective morphine challenge (EDBO) is ex-pected to lower the level of antagonism usingthese rates of naltrexone delivery.

DRUG RELEASE KINETICS

In order to (1) determine the naltrexone re-lease kinetics and shape stability ofCHRONOMER™/naltrexone composites in vitroand in vivo and (2) evaluate them with respectto the degree of tissue response, naltrexonewas incorporated into C 101 and C 101 ctmatrix materials at a level of 20 mg/g. Purematrix materials and those filled with 10%Na2C03 were used to afford a range of usefullifetimes and physical characteristics. Themixtures were fabricated into 1.3 mm thickfilms and cut into 1 cm diameter circles.The systems were chemically analyzed for nal-trexone after fabrication, and the drug wasshown to be unaffected by either the pro-cessing conditions or the CHRONOMERTM

matrices.

The systems were implanted subcutaneously inrats. Naltrexone release from the systemswas measured by analyzing the recovered sys-tems for drug. Naltrexone release fromsamples of common origin was also measuredin vitro (phosphate buffer), The results ofthose experiments are shown later in the text.

All drug release studies showed a high levelof scatter. Average values of naltrexonereleased in vitro from each of these systemsresulted in a release rate of 1.6 to 2.6 µg/hr for the first week, Systems containingNa2CO3 continued to release naltrexone at thisrate for about the next 2½ weeks. Systemswithout Na2CO3 released naltrexone at aconstant rate of 6.0 to 7.8 µg/hr after thefirst week of the study.

stability able able

C 101, C 1Ol ct,System C 101 Na 2CO 3 C 101 ct

Shape accept- good poor accept-

Na2CO3

Rate ofdrug

release(µg/hr)in vitro

1st week

3rd week

Tissueresponse*

2nd week

4th week

2.0 1.6 1.5 1.7

7.8 1.6 6.0 2.6

6 1 0 1

0 0 1 1

*Number of moderate or severe reactionsobserved.

Data obtained by measuring the naltrexone insubcutaneously implanted devices, recoveredtwo and four weeks post-implantation, showedgreater scatter than that obtained in vitro.The C 101 ct systems without alkaline excip-ients exhibited poor shape stability. BothNa2CO3-free CHRONOMER

TM matrices eroded with-in a few weeks time. Both CHRONOMERTM

matrices containing 10% Na2C03 had acceptableerosion characteristics, although they in-creased in size and weight after implantation,probably because of water absorption by theexcipient. The four CHRONOMER systemsproduced some moderate tissue reaction at twoweeks post-implantation. Less tissue reactionwas observed at four weeks post-implantation,when the systems were significantly smallerand smoother compared with the original im-plant. With regard to drug release kinetics,shape stability, tissue response and longev-ity, the candidate based on a C 101 matrixcontaining 10% Na2CO3 was favored.

Because of the scatter observed in the drugrelease data, a number of experiments wereinitiated to better define the homogenity ofthe CHRONOMER™/drug mix and the drug releasekinetics in vitro. A variety of matrix/drugformulations were prepared. Part of thesource of scatter was traced to inadequatemixing. The mixing problem has since beenobviated by using an inexpensive but moreefficient blender, designed and developed byALZA. The other source of scatter was foundto result from instability of naltrexone inthe erosion medium. Relatively small amounts

41

degradation products (< 3%) result in a largeincrease in the extinction coefficient of theultraviolet absorption spectrum of naltrexone(as much as 40%). An improved assay proce-dure (liquid chromatography) is now routinelyused and this technique provides measurementswith less than a few percent error.

Using the improved techniques described above,two matrix materials (C 101 and C 101 ct)containing naltrexone and excipient were re-evaluated. Of the formulations investigated,CHRONOMER™ 101 ct/naltrexone systems nowappear to be the most promising. After aninitial lag interval of about two weeks,during which little drug is released, theC 101 ct matrix begins eroding and releasesdrug in a zero order fashion at a rate of30 - 45 µg/hr/cm2. That these C 101 ct/naltrexone systems erode and release drugconcomitantly is evidenced by complete matrixerosion at the time drug has disappeared fromthe system. The concentration of suspendeddrug in the polymer does not appear to affectthe rate of drug release at least to a con-centration of 30%.

In contrast to the results with C 101 ct,C 101 failed to provide satisfactory func-tionality. The lag period preceeding drugrelease for CHRONOMER™ 1O1/naltrexone sys-tems increased with decreasing drug contentup to as long as 18 days at a loading of 2%(w/w). All C lOl/naltrexone systems hadsubstantial (50 - 80%) polymer residues re-

maining after complete drug release. Weconclude that C lOl/naltrexone systems donot erode and release drug concomitantly,and hence no further efforts will be expendedon C 101 systems.

Preliminary sterilization studies showed thatno adverse effects to CHRONOMER™/naltrexonesystems occurred after exposure to 2.5 or5.0 mrads of 60Co irradiation.

FUTURE PLANS

The proposed program utilizes the backgroundand experience gained in the earlier studies.The plans are directed to the development ofthe C 101 ct matrix system containing nal-trexone and excipient. Our interpretationof (1) the long lag period between exposureto moisture and establishment of steady statedrug release from these systems and (2) thelarge quantity of crystalline residue ofC 101 matrix which remains after all the drughas been released, is based on phenomenabrought about by dissolved naltrexone in thematrix, Basic materials are known to inhibitthe ability of these matrices to erode andplasticizers are known to facilitate crys-tallization, Naltrexone is believed to playboth roles. If a less soluble (in the matrix)form of the drug is to be used, the problem ofdelayed erosion may be significantly reduced.By shifting to C 101 ct, crystallization phe-nomena may be avoided completely.

42

In all three phases of this evaluation re-search, a substantial effort has been devotedto the development of the techniques andexpertise necessary to carry out the evalua-tion in each phase. Progress in this develop-

TESTING OF DRUG mental work has been such that all threephases of evaluation can now be carried out

DELIVERY SYSTEMS FOR for naltrexone delivery systems.

USE IN THE TREATMENT PHARMACOLOGIC CHARACTERIZATION OF PROLONGEDANTAGONIST RELEASE

OF NARCOTIC ADDlCTlON

Richard H. Reuning, Ph.D.College of Pharmacy, Ohio State Univ.

Louis Malspeis Ph.D.College of Pharmacy. Ohio State Univ.

Sylan Frank, Ph.D.College of Pharmacv. Ohio State Univ.

Robert E. Notari, Ph.D.College of Pharmacy, Ohio State Univ.

The research on pharmacologic characterizationof prolonged antagonist release has been con-cerned with both the development of method-ology and the actual evaluation of severalnaltrexone delivery systems. In this work, amodified version of the mouse tail-flick testfor narcotic antagonist activity that wasdeveloped permits a reproducible temperatureto be applied to a small portion of the mousetail and yields data with comparatively smallstandard deviations. The technique has beencritically tested and found to be an accuratemeasure of analgesia and analgesic antagonism.

INTRODUCTION

Several naltrexone delivery systems have beenevaluated for duration of analgesic, antagonismat a dose level of 40 mg naltrexone/kg usingthis modified mouse tail-flick procedure. Adelivery system consisting of 35% naltrexonein 95% polylactic acid. 5% Citroflex 4 yielded

The objective of this research is to evaluate a duration of analgesic antagonism of approxi-the drug release characteristics of drug mately 17 days, compared to 1/2 day for the samedelivery systems developed for narcotic antag- dose of naltrexone in aqueous solution.onists, particularly naltrexone and naloxone. Results from additional controls indicate thatThe procedure which has been developed to the prolongation of activity was due to slowaccomplish this objective is a series of three release of naltrexone from the polymer deliv-successive testing procedures, each more quan- ery system. A second delivery system consist-titative than the previous in describing the ing of naltrexone zinc tannate complex sus-release rate. The phase I evaluation consists pended in a vehicle of 2% aluminum monostear-of a test for the duration of analgesic antag- ate in peanut oil yielded significant anal-onism in mice using the mouse tail-flick test. gesic antagonism for 16 days whereas naltrex-Phase II of our evaluation scheme is concerned one hydrochloride suspended in the samewith quantitating the excretion of tritium in vehicle lasted only 4 days. Results fromurine and feces after administration of a drug additional controls indicate that the mechan-delivery system containing radiolabeled narco- ism responsible for the prolonged activity istic antagonist to rats or guinea pigs. The slow dissolution of naltrexone from the deliv-third phase of this evaluation work is a phar- ery system. A somewhat similar delivery sys-macokinetic description of the release rate of tem, i.e., naltrexone aluminum tannate suspend-narcotic antagonist from the delivery systems ed in a vehicle of 2% aluminum monostearate inadministered to dogs and monkeys. In this peanut oil, yielded significant analgesicphase III work the narcotic antagonist is antagonism for 22 days. The fourth deliveryadministered in unlabeled form and samples of system consisted of beads containing 50% nal-blood and/or urine are subsequently collected trexone in a co-polymer of L(+)-lactic acidand analyzed for concentration of the narcotic (75%)/glycolic acid (25%). This systemantagonist by electron capture gas-liquid yielded significant antagonism for 18 days atchromatography. Such concentration data a 50 mg/kg dose level of naltrexone and for atobtained as a function of time, together with least 21 days at a 100 mg/kg dosage. Again,appropriate control experiments, permit a comparison with appropriate controls indicatesdetermination of the release rate of the nar- that the prolonged analgesic antagonism is duecotic antagonist from the drug delivery sys- to slow release of naltrexone from the deliv-tem. ery system.

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SEMI-QUANTITATIVE CHARACTERIZATION OF PRO-LONGED ANTAGONIST RELEASE

In order to carry out the proposed research onestimating narcotic antagonist release ratesfrom delivery systems containing tritiatedantagonist, it has been necessary to spendconsiderable effort in developing methodologyprior to evaluating release of naltrexone froma sustained-release delivery system. Initial-ly, both the radiochemical purity of tritiatednaltrexone and the lack of exchange of labelwith water in biological systems was estab-lished for both 8-3H-naltrexone and 15,16-3Hnaltrexone. Stability of the tritiated nal-trexone was monitored periodically by thin-layer chromatography. Essentially quantita-tive recovery of the administered tritiatednaltrexone was obtained in urine and fecesafter parenteral administration of a solutionof the drug to rats and guinea pigs. Thequantitation achieved by combustion of urineand fecal samples followed by liquid scintil-lation counting indicates that, in the rat,about 60% of the administered radioactivity isexcreted in feces and 40% in urine. On theother hand, in the guinea pig only 14% of theadministered radioactivity was found in fecesand about 84% in urine.

The naltrexone zinc tannate delivery systemmentioned previously was evaluated by thisradioactivity excretion technique. 3H-15,16-naltrexone was administered via this dosageform intramuscularly (20 mg/kg) in guineapigs. The overall elimination of radioactiv-ity was found to be approximately exponentialin four guinea pigs with 50% of the dose ex-creted in 2.7-4.9 days and 90% in 11.6-15.3days. This compares favorably with the 16day duration of analgesic antagonism obtainedfor this delivery system by the pharmacologictechnique discussed in the previous section.

PHARMACOKINETIC EVALUATION OF PROLONGEDANTAGONIST RELEASE IN DOG,MONKEY AND MAN

Analytical Development

A major part of the work aimed at an eventualpharmacokinetic quantitation of narcoticantagonist release rates has been devoted tothe development of sensitive gas chromato-graphic-electron capture (GLC/EC) assays fornaltrexone and naloxone in human, dog andmonkey plasma. These assays for theunchanged narcotic antagonist, which canreadily be adapted to assays for drug inurine as well, permit a pharmacokinetic quan-titation of the disposition of naltrexone andnaloxone in the dog, monkey or man, as wellas a quantitation of the release rate ofthese compounds from antagonist delivery

systems. The assays for naltrexone and nal-oxone generally involve addition of an inter-nal standard to the plasma sample, extractionof the basified plasma sample with benzene,back extraction into 0.1 N sulfuric acid,basification, re-extraction into benzene,evaporation, derivatization in benzene withpentafluoropropionic anhydride or heptafluoro-butyric anhydride in benzene, washing thereaction mixture with sodium tetraborate, andinjecting a portion of the derivatized mater-ial onto an OV-17 column. The assays fornaloxone and naltrexone appear to be specificfor unchanged drug and to clearly separate theunchanged drug from known metabolites. Thesensitivity limit is 5-10 ng of antagonist ina 0.5 ml sample of plasma. We have extendedthe sensitivity of the assay of naltrexone indog and monkey plasma to 1 ng/ml of plasma bythe use of 2-3 ml of plasma. This appears tobe the lower sensitivity limit of the GLC/ECtechnique. Linear calibration curves havebeen obtained for naloxone and naltrexone indog and human plasma and for naltrexone inmonkey plasma and for naltrexone in both un-hydrolyzed and glusulase-hydrolyzed dog urine.Preliminary work on a GLC/EC assay for cycla-zocine indicates that it may be possible toanalyze this narcotic antagonist at concentra-tions less than 1 ng/ml.

Biologic Disposition of Naltrexone

In order to determine the rate of release ofnarcotic antagonist from a delivery system bypharmacokinetic techniques, it is necessarythat the assay utilized to measure the concen-tration of antagonist in plasma and urine bespecific for the unchanged drug. Thus, it isessential that a knowledge of the basic meta-bolic pathways of the narcotic antagonist bedeveloped and that samples of authentic meta-bolites be synthesized or isolated and testedfor interference with the assay procedure.In addition, a knowledge of the physiologicdistribution of the narcotic antagonist canaid in explaining the relationship between theconcentration of antagonist measured in theplasma and the amount of drug remaining inother parts of the body. Thus far, our workon biologic disposition has been limited tonaltrexone.

Our research on the physiologic distributionof naltrexone has included a determination ofthe extent of binding of the drug in plasmafrom several species. It was found by equi-librium dialysis that naltrexone is bound inplasma to an extent of 2O-26% in man, monkey,dog, guinea pig, rat and mouse. The extentof binding was independent of naltrexone con-centration over a wide concentration rangein all species. A second approach to theinvestigation of naltrexone distributioninvolved a determination of the

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tissue levels of radioactivity in mice at 1,5, and 15 minutes after intravenous adminis-tration of tritiated naltrexone. The resultsof this study indicate that the radioactivityis very rapidly distributed from plasma totissues, concentrating in the liver, kidneyand gastrointestinal tract, with less than 4%of the dose being present in plasma 1 minuteafter administration.

Considerable progress has also been made con-cerning the metabolism of naltrexone in sev-eral species. Two metabolites of naltrexone,

a n d naltrexol, as well as two metabolitesof naloxone, and naloxol, have been syn-thesized by sodium borohydride reduction ofnaltrexone and naloxone, respectively. Eachpair of epimers was separated by preparativethin-layer chromatography and the structureswere determined by spectral techniques. Useof these authentic metabolites has permittedthe development of a qualitative GLC/EC methodfor distinguishing either or naltrexolin urine in the presence of large amounts ofnaltrexone . After parenteral administrationof naltrexone substantial quantities ofnaltrexol and/or its conjugates were found inenzymatically hydrolyzed urine samples fromman, monkey and guinea pig whereas smallerquantities were excreted by the rabbit, mouse,rat and dog. Trace amounts of the 6 -hydroxyepimer, a-naltrexol, were detected in theurine of monkeys and guinea pigs only. Invitro experiments with guinea pig liver indi-cate that the enzymes responsible for the re-duction of naltrexone to both andnaltrexol are present in the 100,000 x gsupernatant of the liver homogenate.

The isolation of a n d naltrexol has alsopermitted the development of a GLC/EC analyt-ical procedure for each of these metabolites.Linear calibration curves were obtained afterextraction of these metabolites from dogplasma containing 20 to 150 ng of eithero r naltrexol. As mentioned previously, wehave also confirmed that these metabolites donot interfere with the GLC/EC assay for nal-trexone.

Pharmacokinetics of Naltrexone

The characterization of the phannacokineticsof a narcotic antagonist is a necessary pre-requisite to the determination of the rate ofrelease of that antagonist from a deliverysystem. Since the species most likely to beuseful for testing narcotic antagonist deliv-ery systems are the dog and monkey, thepharmacokinetics of naltrexone has beenstudied in these species.

Plasma concentration-time data for intraven-ous naltrexone in the dog, obtained by the

GLC/EC analytical technique, indicate that atleast two “compartments” are necessary toaccount for the data. The half-life of theprincipal exponential phase of the plasmalevel-time curve was 32 and 36 min in two fox-hounds, and the apparent volume of distribu-tion was 3.2 and 3.6 liters/kg. In similari.v. bolus experiments using four monkeys,the plasma level-time half-life for naltrexonewas 40-82 minutes (mean, 59 min) and theapparent volume of distribution was 4.4-10.1liters/kg (mean, 6.4 liters/kg). These dataindicate that naltrexone is distributed fromblood to the rest of the body to an unusuallygreat extent. Also, the drug appears to berapidly eliminated in both the dog and themonkey. Since the clearance of naltrexone isgreater than liver blood flow, the data indi-cate that some extrahepatic elimination occurswith this drug. Most importantly, however,the phannacokinetic data from these experi-ments will permit a determination of therelease rate of naltrexone from a naltrexonedelivery system administered to the dog ormonkey in a subsequent experiment. Such anexperiment in the monkey is currently inprogress.

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