ABC transporters,SLC transporters

1. MEMBRANE transporters
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2. CONTENTS
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Introduction
Membrane Transporters in therapeutic and adverse drug responses
Basic transporter mechanisms
Transporter structure and mechanism
Vectorial transport
Transporter superfamilies in the human genome
Transporters involved in pharmacokinetics
Transporters and pharmacodynamics: drug action in the brain
The blood-brain barrier: a pharmacological view
Transporters in regulatory sciences

3. INTRODUCTION
• Membrane transport refers to the collection
of mechanisms that regulate the passage
of solutes such
as ions and molecules through biological
membranes.
• Regulation of passage is due to selective
membrane permeability
• Membrane permeability allows them to
separate substances of distinct chemical nature.

4. LIPID PERMEABILITY
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5. INTRODUCTION
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Membrane transport proteins are present in all organisms.
They control the influx of essential nutrients and ions and
the efflux of cellular waste, environmental toxins, drugs.
Membrane transporters function may be facilitated or
active.
Two major super families ,ABC (ATP binding casette)and
SLC (solute carrier) transporters.

6. MEMBRANE
TRANSPORTERS
IN
THERAPEUTIC
DRUG
RESPONSES
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Pharmacokinetics
Pharmacodynamics:
Transporters as Drug
Targets
Drug Resistance

7. PHARMACOKINETICS
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Transporters important in pharmacokinetics are located in
intestinal, renal, and hepatic epithelia.
They function in the selective absorption and elimination of
endogenous substances and xenobiotics ,including drugs.
They work with drug-metabolizing enzymes to eliminate
drugs and their metabolites.
Also serve as protective barriers to particular organs and cell
types.

8. Pharmacodynamics: Transporters as Drug Targets
◈ Membrane transporters are the targets of many
clinically used drugs.
◈ SERT/5HTT(sodium-dependent serotonin
transporter ) is a target for a major class of
antidepressant drugs, the SSRIs.
◈ These transporters also may be involved in the
pathogenesis of neuropsychiatric disorders,
including Alzheimer and Parkinson diseases.
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9. DRUG RESISTANCE
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Membrane transporters play critical roles in the
development of resistance to anticancer drugs,
antiviral agents, and anticonvulsants.
• Decreased uptake of drugs.
• Enhanced efflux of hydrophobic drugs
• Overexpression of MRP4 (multi drug resistant protein)
• Modulation of MDR1 expression and activity

10. /
MEMBRANE TRANSPORTERSANDADVERSE DRUG
RESPONSES 10
Transporters play crucial
roles in the cellular
activities and toxicities of
these agents.
Transporter-mediated
adverse drug responses
can be classified into
three categories
Decreased uptake at
clearance organs
Increased uptake at target
organs
Altered transport of
endogenous compounds
at target organs

11. DECREASED UPTAKE AT CLEARANCE ORGANS
Transporters in liver ,kidney and metabolic enzymes
key determinants of drug exposure in blood
affecting exposure and toxicity in all organs
due to a decrease in the uptake or secretion in clearance organs
(e.g., liver and kidney)
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12. DECREASED UPTAKE AT CLEARANCE ORGANS
Example:
After oral administration of an HMG-CoA reductase inhibitor
first-pass hepatic uptake of the drug by the SLC OATP1B1
maximizes the effects of such drugs on hepatic HMG-CoA
reductase
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13. MEMBRANE TRANSPORTERS AND ADVERSE DRUG
RESPONSES
Transporters expressed in tissues (e.g., brain) or in barriers to such
tissues (e.g., the BBB) that may be targets for drug toxicity
tightly control local drug concentrations
thus control the exposure of these tissues to the drug.
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14. CONTROL OF EXPOSURE OF TISSUES TO THE DRUG
Loperamide is a substrate of P-glycoprotein(Pgp)
Pgp prevents accumulation of Loperamide
Inhibition of Pgp–mediated efflux (Quinidine)in the BBB
Increase in the concentration of Loperamide in the CNS
Respiratory depression
◈ OAT1 (organic anion transporter), OCT1(organic cation
transporter), and OCT2 provide other examples of
transporter -related toxicity.14

15. DRUGS MAY MODULATE TRANSPORTERS
Drug inhibits influx of endogenous compound in eliminating or target organ.
Increase in the plasma concentration of an endogenous compound (e.g., a bile
acid)
Increase in concentration of the endogenous compound in the target organ
Drug-inhibited efflux of the endogenous compound
Adverse effects(cholestasis or hyperbilirubinemia)
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16. BASIC
MECHANISMS
OF
MEMBRANE
TRANSPORT
◈ Transporters Versus Channels
◈ Transporters
■ Passive Diffusion
■ Facilitated Diffusion
■ Active Transport
Ø Primary Active Transport
Ø Secondary Active Transport
 Symport
 Antiport
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17. CHANNELS VERSUS TRANSPORTERS
CHANNELS
Two primary states, open and closed
Open state act as pores for selected
ions flowing down an
electrochemical gradient.
Return to the closed state as a function
of time.
Drugs termed as potentiators (e.g.,
ivacaftor) may increase the probability
that a channel is in open state.
TRANSPORTERS
Transporter forms an intermediate
complex with the substrate (solute)
Conformational change in transporter
Induces translocation of substrate to other
side of the membrane.
Transporter-mediated membrane transport
is characterized by saturability and
inhibition by substrate analogues.17

18. PASSIVE
DIFFUSION
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Passive Diffusion consists of
three processes:
• partition from the aqueous to the
lipid phase,
• diffusion across the lipid bilayer,
• repartition into the aqueous phase
on the opposite side.
Passive diffusion of any solute
(including drugs) occurs down
an electrochemical potential
gradient of the solute

19. FACILITATED DIFFUSION
◈ Passive movement of molecules
along their concentration gradient,
◈ Transport is guided by the presence
of an integral membrane protein
forming a pore or channel..
◈ Does not require energy input.
◈ Examples:glucose, sodium ions, and
potassium ions.
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20. ACTIVE
TRANSPORT
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Transport requires the input of energy.
Transport of solutes against their
electrochemical gradients
Concentration of solutes on one side
create potential energy
An important role in the uptake and
efflux of drugs and other solutes.
Depending on the driving force,
• Primary active transport
• Secondary active transport

21. ACTIVE
TRANSPORT
◈ Black squares - ion that
supplies the driving force
for transport (size is
proportional to the
concentration of the ion).
◈ Blue ovals - transport
proteins.
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22. PRIMARYACTIVE TRANSPORT
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Transport that directly couples with ATP hydrolysis is called
Primary active transport.
ABC transporters are examples of primary active
transporters.
ABC transporters mediate unidirectional efflux of solutes
across biological membranes.
Another example of primary active transport is the Na + K +
ATPase.

23. SECONDARY ACTIVE TRANSPORT
◈ For transport of a solute S1 against its concentration
gradient energy is driven by the transport of another
solute S2 in accordance with its electrochemical gradient.
◈ Depending on the transport direction classified as
◈ -.
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Antiport Symport
Transporter moves S2 and S1 in
opposite directions.
Transport S2 and S1 in the same
direction
Example: Na+/Ca++ exchanger Example: Na+-glucose transporter
(SGLT1)

24. TRANSPORT STRUCTURE AND
MECHANISM
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Hydropathy analysis
indicate that transporters in
SLC and ABC super
families are
multimembrane-spanning
proteins.
Hydropathy plots allow the
visualization of
hydrophobicity over the
length of a peptide
sequence.
Emerging crystals
structures are adding to
our ideas of the
mechanisms of transport
via these proteins.

25. CRYSTAL STRUCTURE OF
TRANSPORTERS
ABC TRANSPORTER SLC TRANSPORTER
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26. ABC TRANSPORTERS
(adenosine triphosphate -binding cassette )
◈ ABC superfamily includes 49 genes, each containing one or two conserved
ABC regions.
◈ ABC transporters are primary transporters that couple the energy stored in
adenosine triphosphate (ATP) to the movement of molecules across the
membrane.
◈ They can be divided into .
○ Importers -mediate the uptake of essential nutrients into cells and are
found predominantly in prokaryotes.
○ Exporters -transport molecules out of cells or into organelles and are
found in all organisms.(eukaryotes)
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27. ABC TRANSPORTERS MECHANISM
ABC transporters have NBDs(nucleotide-binding domains) on the
cytoplasmic side that contain conserved motifs which participate in
binding and hydrolysis of ATP.
◈ Crystal structures of all ABC transporters show two NBDs, which are
in contact with each other.
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In case of drug extrusion
ATP binds to NBD
transporters open to outside
releasing their substrates
to the extracellular media.
hydrolysis products dissociation
transporters return to inward-
facing conformation,
permit binding of ATP and
substrate.

28. ABC TRANSPORTERS
Transporter accepts a solute molecule at the cytoplasmic membrane surface
When its nucleotide NBDs are fully charged with ATP.
Sequential hydrolysis of the ATP molecules
Produces steric change leading to translocation
Release of the solute at the exterior surface.
Exchange of ADP for ATP on both NBDs completes the cycle
Restores the system for readiness to transport another solute molecule
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29. ABC TRANSPORTERS
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30. SLC TRANSPORTERS
◈ SLC superfamily includes Channels, Facilitators and Secondary active
transporters .
SLC uses gated pore mechanism,
substrate binding
structural changes in carrier protein,
reorienting the opening of the binding site to the opposite side
substrate dissociates from the transport site
allowing another substrate to be bound
transported in the opposite direction.
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31. SLC TRANSPORTERS MECHANISM
GATED PORE
•The gated pore represents the
model for SGLT
•The rotation of two broken
helices facilitates access of
substrates to the intracellular and
extracellular sides of the plasma
membrane.
ROCKER SWITCH
•Mechanism thought to underlie
the transport of most
MFS transport is classically
described as the "rocker-switch"
mechanism
•This example models a facilitated
glucose transporter, glut2.
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32. VECTORIAL
TRANSPORT
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Asymmetrical transport across a
monolayer of polarized cells, such as
epithelial and endothelial cells of brain
capillaries, is called vectorial transport.
Important for the absorption of
nutrients,bile acids and drugs in the
intestine.
Plays a major role in hepatobiliary and
urinary excretion of drugs from blood
to lumen

33. VECTORIAL TRANSPORT
Efflux of drugs from the brain involves vectorial transport.
ABC transporters mediate only unidirectional efflux,
SLC transporters mediate either drug uptake or drug efflux.
 ABC transporters alone are able to achieve vectorial transport
without the help of influx transporters
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34. VECTORIAL
TRANSPORT
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In the liver , transporters located on the
sinusoidal membrane (facing blood)
transport bile acids, organic anions and
organic cations into the hepatocytes.
Similarly ABC transporters on the
canalicular membrane (facing bile)
export such compounds into the bile
Similar transport systems also are present
in the intestine, renal tubules, and
endothelial cells of the brain capillaries
Transporter expression can be regulated
transcriptionally in response to drug
treatment and pathophysiological
conditions, resulting in induction or
downregulation of transporter mRNAs

35. TRANSPORTER SUPERFAMILIES
SLC superfamily
◈ Includes 52 families and represents about 395 genes in the
human genome.
◈ They are multi-membrane spanning proteins.
◈ Associated with genetic diseases.
◈ Highly selective transporters that interact with structurally
similar molecules.
◈ Mostly facilitative transporters, although some are secondary
active transporters
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36. SLC LINKED DISEASES
 Hypophosphatemic nephrolithiasis
Primary bile acid malabsorption
Hereditary hemochromatosis
 Gitelman syndrome
Multiple epiphyseal dysplasia
Hypermanganesemia with dystonia
 Hypophosphatemic nephrolithiasis
 Primary bile acid malabsorption
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37. SLC SUPERFAMILY
Physiological role
Transport of amino acids
critical in protein synthesis
Absorption, elimination and
Cellular distribution
Transport water-soluble
vitamins.
Transport neurotransmitters
Pharmacological role
Drug absorption
 Elimination
Tissue distribution
Mediators of drug-drug
interactions
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38. ABC SUPER FAMILY
◈ There are 49 known ABC transporters present in humans,
which are classified into seven families by the Human
Genome Organization.
◈ ABC transporters are essential for many cellular processes
and mutations
◈ ABC transporters contribute to human genetic disorders.
◈ In particular,MDR1/ABCB1, MRP2/ABCC2, and
BCRP/ABCG2 are involved in overall drug disposition.
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39. TISSUE
DISTRIBUTION
OF ABC
TRANSPORTERS
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Expressed in the intestinal epithelia, where they
serve to pump out xenobiotics,including many
orally administered drugs.
They are also expressed in the polarized tissues
of kidney and liver.
Some ABC transporters are expressed
specifically on the blood side of the endothelial
or epithelial cells form barriers to the free
entrance of toxic compounds into tissues:
• Blood brain barrier,
• Blood-CSF barrier ,
• Blood-testis barrier
• Blood-placenta barrier.

40. ABC LINKED HUMAN DISEASES
◈ Tangier disease (defect in cholesterol transport)
◈ Stargardt syndrome (defect in retinal metabolism)
◈ Dubin-johnson syndrome (defect in biliary bilirubin
glucuronide excretion)
◈ Pseudoxanthoma (unknown mechanism)
◈ Cystic fibrosis (defect in cl− channel regulation)
◈ Adrenoleukodystrophy (result of fatty acid buildup)
◈ Sitosterolemia
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41. PHYSIOLOGICAL ROLES OF ABC TRANSPORTERS
llustrated by studies involving knockout animals or patients with genetic
defects in these transporters.
For instance, mice deficient in MDR1 function
are viable and fertile
no phenotypic abnormalities
but show hypersensitivity to the toxicity of drugs
◈ Complete absence of these drug-related ABC transporters is not lethal.
◈ Inhibition of physiologically important ABC transporters (especially those
related directly to the genetic diseases) by drugs should be avoided to
reduce the incidence of drug-induced side effects.
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42. ABC TRANSPORTERS IN DRUG ABSORPTION AND
ELIMINATION
◈ MDR1 is the most renowned ABC transporter yet identified.
◈ MDR1 is expressed on the brush border membrane of renal epithelia,
function can be monitored using digoxin (> 70% excreted in the urine).
MDR1 inhibitors (e.g., quinidine,verapamil,clarithromycin)
reduce renal excretion of digoxin
◈ Drugs with narrow therapeutic windows(e.g., digoxin, cyclosporine,
tacrolimus) should be used with great care.
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43. ABC TRANSPORTERS IN DRUG ABSORPTION AND
ELIMINATION
MRP3 mediate intestinal absorption in conjunction
MRP3 mediates sinusoidal efflux in the liver
Decreases excretion of metabolites, particularly glucuronide
conjugates.
Dysfunction of mrp3
Shortening of the elimination t1/2.
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44. TRANSPORTERS INVOLVED IN PHARMACOKINETICS
SLC TRANSPORTERS
•Located in sinusoidal
membrane of hepatocytes.
•Mediates hepatic uptake of
organic anions, cations, and
bile salts.
•Facilitated or Secondary Active
mechanisms.
•OATPs (SLCO), OCTs
(SLC22),
ABC TRANSPORTERS
•Bile canalicular membrane of
hepatocytes.
•Efflux (excretion) of drugs
and their metabolites, bile
salts, and phospholipids from
liver to bile.
•Primary active transport.
•MRP2, MDR1, BCRP, and
MDR2
45
Transporters in the liver and kidney have important roles in
removal of drugs from the blood and hence in metabolism and
excretion.

45. TRANSPORTERS INVOLVED IN
PHARMACOKINETICS
◈ Vectorial transport-circulates drugs from blood to the bile
using an uptake transporter (OATP family) and an efflux
transporter (MRP2,BCRP) .
◈ Examples illustrate importance of vectorial transport in
determining
○ Drug exposure in the circulating blood and liver
○ Role of transporters in drug-drug interactions
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46. VECTORIALTRANSPORT EXAMPLES
◈ HMG-CoA reductase inhibitors
◈ Gemfibrozil
◈ Irinotecan
◈ Bosentan
◈ Temocapril and other ACE inhibitors
◈ Angiotensin II Receptor Antagonists
◈ Repaglinide and Nateglinide
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47. TRANSPORTERS INVOLVED IN
PHARMACOKINETICS
Gemfibrozil and its glucuronide
Inhibit the uptake of active hydroxy forms of statins into
hepatocytes by OATP1B1
Increase in the plasma concentration of the statin
Increase in toxicity.
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48. Transporters in the hepatocyte that function in the uptake and efflux of drugs
across the sinusoidal membrane and efflux of drugs into the bile across the
canalicular membrane. 49

49. RENAL TRANSPORTERS
ORGANIC CATION TRANSPORT:
◈ Organic cations are secreted in the proximal tubule
◈ Many secreted organic cations are endogenous compounds
(e.g., Choline,n-methylnicotinamide, and DA)
◈ Renal secretion helps
 In Eliminating excess concentrations of these substances.
 Ridding the body of xenobiotics, drugs and their
metabolites.
 Cations that are secreted may be either hydrophobic or
hydrophilic.
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50. RENAL TRANSPORTERS
For transepithelial flux of a compound
Compound traverses two membranes
sequentially
Basolateral membrane and the apical
membrane
By two distinct transporters in the SLC
family OCT2 and OCT3.
Transported across this membrane down an
electrochemical gradient.
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51. RENAL TRANSPORTERS
Transport from cell to tubular lumen
Through electroneutral proton–organic cation
Exchange is mediated by transporters in SLC47 family.
Bifunctional transporters
Involved in carnitine reabsorption also.
In reuptake mode, function as Na+ cotransporters, relying on
the inwardly driven Na+ gradient created by Na+K+ATPase
Move carnitine from tubular lumen to cell.
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52. RENAL TRANSPORTERS
ORGANIC ANION TRANSPORT
◈ Primary function is removal of xenobiotics from the body.
◈ Two primary transporters are OAT1 AND OAT3.
◈ Hydrophilic organic anions are transported across the
basolateral membrane against an electrochemical gradient.
◈ Exchanging with intracellular α-ketoglutarate which moves
down its concentration gradient from cytosol to blood.
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53. RENAL TRANSPORTERS
◈ Outwardly directed gradient of α-
ketoglutarate is maintained by
Na+dicarboxylate transporter.(NaDC3)
◈ Transport of low-molecular-weight
organic anions by the cloned transporters
can be driven by α-ketoglutarate;
◈ Coupled transport of α-ketoglutarate and
LMW organic anions occurs in
basolateral membrane vesicles.
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54. TRANSPORTERS AND PHARMACODYNAMICS:
DRUG
ACTION IN THE BRAIN
Neurotransmitters are packaged in vesicles in presynaptic
neurons
Released in the synapse by fusion of the vesicles with plasma
membrane
Then taken back into the presynaptic neurons or postsynaptic
cells
◈ Transporters involved are SLC1 and SLC6.
◈ Transporters in both families play roles in reuptake of
GABA, glutamate and the monoamine neurotransmitters
NE, 5HT, and DA. 55

55. TRANSPORTERS AND PHARMACODYNAMICS: DRUG
ACTION IN THE BRAIN
◈ Serve as pharmacologic targets for neuropsychiatric
drugs.
◈ SLC6 family members localized in the brain and are
secondary active transporters.
◈ Many of these transporters are present in other tissues (e.g.,
intestine, kidney, and platelets)
◈ Examples:
○ GABA Uptake: GAT1 (SLC6A1), GAT3
(SLC6A11),GAT2 (SLC6A13)
○ Catecholamine Uptake: NET (SLC6A2)
○ Dopamine Uptake: DAT (SLC6A3)
○ Serotonin Uptake: SERT (SLC6A4)56

56. THE BLOOD-BRAIN BARRIER: A
PHARMACOLOGICAL VIEW
◈ CNS is well protected from neurotransmitters, exclude
many toxins,bacteria, and xenobiotics by a barrier called
the BBB.
◈ System relies on selective permeability.
◈ endothelial cells in the CNS have tight junctions that
limit paracellular flow and generally have very low rates
of vesicular transport.
◈ There are transport systems: for ions; for nutrients etc
○ SLC2A1/GLUT1 (glucose),
○ SLC7A1and SLC7A5/LAT1 (amino acids); for
nucleosides;
○ SLC16A1.(Metabolic by-products such as lactate and
pyruvate)
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57. TRANSPORTERS IN REGULATORY
SCIENCES
◈ Major determinants of variation in therapeutic and
adverse drug reactions.
◈ Transporters may mediate drug-drug interactions that
result in drug safety issues.
Gemfibrozil glucuronide formed in hepatocytes
Reduces the hepatic uptake and metabolism of cerivastatin
High Cp for cerivastatin.
Elevated statin levels
Statin-induced myopathies(rhabdomyolysis)
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58. TRANSPORTERS IN REGULATORY SCIENCES
◈ U.S. FDA has issued a draft clinical pharmacology
guidance on performing drug-drug interaction studies
during clinical drug development to make decisions
about whether to conduct a clinical drug-drug interaction
study.
◈ If a new molecular entity (nme) inhibits the in vitro
uptake of a canonical substrate of oct2 at clinically
relevant (unbound)concentrations
◈ The guidance recommends to perform a clinical drug-
drug interaction study to determine whether the
◈ NME inhibits the renal clearance of an OCT2 substrate
(e.g., Metformin) in vivo.
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59. REFERENCES
◈ Chabner BA, Longo DL: Cancer Chemotherapy and Biotherapy:
Principles and Practice, 5th ed. Lippincott Williams & Wilkins, 2011.
◈ Brunton LL, Chabner BA, Knollman BJ, eds. Goodman & Gilman’s
the pharmacological basis of therapeutics. 12thed. New York:
McGraw-Hill; 2011. p. 1505-20.
◈ Pedersen PL. Transport ATPases into the year 2008: a brief overview
related to types, structures, functions and roles in health and disease.
◈ Hung LW, Wang IX, Nikaido K, Liu PQ, Ames GF, Kim SH. Crystal
structure of the ATP-binding subunit of an ABC
transporter. Nature. 1998;396:703–7. doi: 10.1038/25393. [PubMed]
◈ Redox Regulation of Multidrug Resistance in Cancer Chemotherapy:
Molecular Mechanisms and Therapeutic Opportunities-Macus Tien
Kuo-Antioxid Redox
Signal.2009Jan;11(1):99133.doi: 10.1089/ars.2008.209
◈ Kaspar P. Locher, Allen T. Lee and Douglas C. Rees, Caltech-ABC
Transporter architecture and mechanism
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60. THANK YOU
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