cell junctions and mdr

8/11/2019 Cell Junctions and MDR

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Epithelial Cell Junctions

&

Efflux Transporter Systems

I n relevance to

Oral Drug Absorption and Bioavailabi l i ty

Chandra Teja U

Department of Pharmaceutics

KVSR SCOPS

By

Y13MPPC140002


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What They Are;

How They are Important;

Their Types; and

Relevance in Oral Drug Absorption

We Will be iscussing


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Epithelial Cell Junctions


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Cell Junctions are the structures where long term association

between neighbouring cells and cells with ECM are established.

They

Hold the cells togetherwith enough stability to compose tissue.

Are composed of Membrane-Associated Structures, usually

Transmembrane Proteins.

Also help in communicationbetween cells.

What Are Cell Junctions!?


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How Many Types Are There!?

Cell Junctions are of the following types

1. Tight/Occluding junctions: Seal cells together into sheets.

(forming an impermeable barrier)

2. Adhering/Anchoring junctions: Attach cells to other cells or

ECM. (providing mechanical support)3. Gap/Communicating junctions: Allow exchange of chemical or

electrical information between cells


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Tight or Occluding JunctionsThe junction forms a

band around the cell

fusing cells togetherand closing

intercellular space.

The number of fusionsites often correlate

to the leakiness of the

tissue.

In renal tubule of the kidney (filtrating area) few fusion sites are

present enabling leakability.

In urinary bladder (contents should not leak out) numerous tight

junctionsare present to form a tight seal between cells.


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These junction greatly restricts the passage of water, electrolytes and

other small moleculesacross the epithelium.

They block the movement

of integral membraneproteins (red and green)

between the apical and baso-

lateral surfaces of the cell.

Thus the special functions ofeach surface, like

Receptor-mediated

endocytosisat the apical

surface

Exocytosisat the

basolateral surface

can be preserved.


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Anchoring JunctionsCells must be anchored to one another and to components of the ECM.

Junctional complexes called anchoring proteinsextend through theplasma membrane to link cytoskeletal proteins in one cell to that in

other cells as well as to the ECM.

Three types of anchoring junctions are observed: Adherens Junctions

Desmosomes

Hemidesmosomes

Anchoring junctions are most abundant in tissues that are subjected to

severe mechanical stress, such as heart, muscle, and epidermis.


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a)Adherens Junctions or Zonula Adherens

Situated atclose proximity to the

apexbut somewhat below the tight

junctions.

They are relatively permeable as

there are no fusion sites that

completely seal off the paracellularspace.

They often form a continuous

adhesion belt/band.

Their transmembrane anchors are composed of Cadherinsto anchor with

other cells andIntegrinsto anchor with ECM.

Adherens junctions share the characteristic of anchoring cells through

their cytoplasmic contractile bundle of actin filaments.


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b)Desmosomes. a.k.a. Macula AdherensCan be visualized as rivetsthrough

the plasma membrane of adjacent cells.

Can besituated anywhere on the lateral

surface of the cell.

It is a disk-shaped structurematched with

an identical structure at the surface of theadjacent cell.

Their transmembrane anchors areCadherins.

Its adhesion function is further empowered by groups of intermediate

Keratin filamentswhich provides firm adhesion.

The membranes in this region are usually further apart (30nm).


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c)Hemidesmosomes

Like desmosomes, they tie to intermediate filaments in the cytoplasm, but

their transmembrane anchors are Integrinsrather than Cadherins.

Hemidesmosomes are like Desmosomes, but form links betweenCells

and the basal laminathat underlie epithelia.


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Gap Junctions

Aresimply gaps(~2nm)betweenthe neighbouring cell membranes.

Containsprotein units called

Connexinwith a hydrophilic pore

of 1.5nm diameter that acts as anion channelbetween cells.

Present along the lateral surfaceof the epithelial cell.

A Gap-Junction is made up of six Transmembrane Connexin Subunits.

Inorganic ions, andsmall water soluble molecules(


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Efflux Transporter Systems

(Multi Drug Resistance)


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MDR Transporters belong to the family of the ATP Binding Cassette

(ABC) proteins& are present in all living organisms.

The key physiological taskof the MDR-ABC transporter network is to

provide general xenobiotic resistance, probably evolved as Complex

Cellular Defence Systems.

Hundreds of structurally diverse therapeutic agents are substrates totheseand they impedes the absorption, permeability, and retention of the

drugs, extruding them out of the cells.


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In humans, 3 major types of MDR proteins are present, that are the

members of the subfamilies

The ABCB (P-glycoprotein/ABCB1/MDR1),

The ABCC (ABCC1/MRP1, ABCC2/MRP2), and

The ABCG (ABCG2/MXR/BCRP).

The major practical causes for studying MDR-ABC transporter

expression and function are

Predicting the fate of pharmaceutical agents in our body.

The relevance of in vitro experiments to the in vivo role of

MDR-ABC transporters.

Related to cancer & infection drug resistance.


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MDR1 (P-gp) preferentially extrudes large hydrophobic molecules,

while ABCC1 and ABCG2 can transport both hydrophobic drugs and

intracel lularly formed metaboli tes, e.g., Drug Conjugates.


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P-glycoprotein is primarily found in epithelial cells lining the Small

Intestine, Colon, pancreatic & bile ducts, kidney proximal tubules, and

also in the BBB.

The highest MRP1 levels are reported at testis, cardiomyocytes,

placenta, prostate, lung, thymus and kidney. It is found in lower levels

along the GIT.

Significant expression of ABCG2 was observed in the Placental

Barrier, and at the blood-testis barrier, the BBB, and the stem cells.

Hence, P-gp is of greatest importance when talking about Oral BA of

Drugs.


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The P-GlycoProteinThe most extensively studied and experimented MDR Protein


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P-gp is encoded by theABCB1gene.

It contributes greatly in the extrusion of many drugs from the blood into

the intestinal lumen.

It is also responsible for enhancing the excretion of drugs out of

hepatocytes and renal tubules into bile and urine.

It can recognize and transport numerous structurally diverse drugs over

a molecular weight range of 250 g/mol (cimetidine) to 1200 g/mol

(cyclosporin).

Therefore, P-gp can potentially reduce the absorption and oral

bioavailabilityand decrease the retention time of a number of drugs.

The transporter is also overexpressed on the surface of many

neoplastic cellsand makes the chemotherapy almost ineffective in many

cases.


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Location of P-gp Function

GI Epithelial CellsEfflux of Cellular waste and Drugs

into lumen

Hepatocytes

Secretion of drugs into bile and re-

entry into Intestine

(Entero-Hepatic Circulation)

Renal Epithelial CellsEfflux of Drugs into Tubular

Lumen, easily Excreted

Placenta, Brain, Testis Barrier Action for Entry of Drugs

Cancer Cells Resistance to Agents


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Interactions of P-gp with Drugs

The substrate specificity of P-gp and CYP450 enzyme system has

overlap extensively.

Clinically Significant reduction in Oral Absorption and BA is seen with

Antacids

Cimetidine

Ranitidine

Antibiotics

Erythromycin

Levofloxacine

Cardiac Drugs

Digoxin

Antitumour Agents

Placitaxel

Doxorubicin

Vinblastin

Ca2+

Channel Blockers

Diltiazem

Immunosuppressants

Cyclosporine A

Anti-retrovirals

Ritonavir

Indinavir

Steroids

DexamethasoneOpioids

Morphine

Fentanyl


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P-gp also has Inducers and Inhibitors

An I nducer fur ther reduces the absorptionof other substrates.

An I nhibitor generally increases the absorption of other drugs byreducing P-gp activity.

I nducersAvasimibe

Carbamazepine

Phenytoin

Rifampin

Morphine

Dexamithazone

InhibitorsQuinidine

Reserpine

Cyclosporine

Verapamil

Azithromycin

Atorvastatin


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P-gp Follows Saturable Kinetics

In the case offast absorbing drugs having larger doses

, efflux byP-gp poses less impact on drug absorption and therefore is not

important in terms of bioavailability or pharmacokinetic properties.

This is because the transport activity of P-gp becomes saturated by

high concentrations of drugin the intestinal lumen.

This is best explained by Michaelis Menten Plot & Equation.


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Enhancement of BA by P-gp Inhibition

P-gp mediated drug efflux greatly interferes with the delivery of Potent

Drugs or the drugs with slow dissolution and dif fusion rates.

This decrease in drug absorptionof those small amounts of drugs can be

life threatening at times as sufficient plasma levels of drug cannot be

reached.

Moreover, it can make the sustained release dosage forms of the

substrates completely ineffectiveby limiting their absorptions.

This is even more important in case of MDR exhibiting Cancer.

Hence, P-gp inhibition has gained a lot of importance in Research field

and many approaches are being postulated.


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In general, P-gp can be inhibited by three mechanisms:

(i) blocking the pump competitively or non-competitively;

(ii) interfering with ATP hydrolysis; and

(iii) altering integrity of cell membrane lipids.


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P-gp inhibitors are classified into three generations based on their specificity,

affinity, and toxicity.

1stgen.inhibitors arepharmacologically active substances which are

clinically used but have the ability to inhibit P-gp.

2ndgen.inhibitorspharmacologically inactive. But, they inhibit CYP

enzymes and ABC transporters leading to complicated pharmacokineticalterations. (EgDexverapamil, valspodar, etc)

3rdgen. inhibitors are under clinical development, aiming for higher

specificity and low toxicity(EgLaniquidar, Tariquidar, etc)

Other approaches likeLiposomes (Active Targeting), Use ofP-gp bypassing

Polymers,Drug-Macromoecule Conjugates, etc are being studied.


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References

Google Images

http://www.histology.leeds.ac.uk/

http://the-aps.org

http://en.wikipedia.org

http://www.nature.com/

PubMed NCBI Articles

SlideShare and AuthorStream Presentations
http://www.histology.leeds.ac.uk/http://the-aps.org/http://en.wikipedia.org/http://www.nature.com/http://www.nature.com/http://www.nature.com/http://www.nature.com/http://www.nature.com/http://www.nature.com/http://www.nature.com/http://www.nature.com/http://en.wikipedia.org/http://en.wikipedia.org/http://en.wikipedia.org/http://en.wikipedia.org/http://en.wikipedia.org/http://en.wikipedia.org/http://en.wikipedia.org/http://en.wikipedia.org/http://the-aps.org/http://the-aps.org/http://the-aps.org/http://the-aps.org/http://the-aps.org/http://the-aps.org/http://the-aps.org/http://www.histology.leeds.ac.uk/http://www.histology.leeds.ac.uk/http://www.histology.leeds.ac.uk/http://www.histology.leeds.ac.uk/http://www.histology.leeds.ac.uk/http://www.histology.leeds.ac.uk/http://www.histology.leeds.ac.uk/http://www.histology.leeds.ac.uk/http://www.histology.leeds.ac.uk/http://www.histology.leeds.ac.uk/http://www.histology.leeds.ac.uk/


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