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Hello everyone, my name is Ritu Singh and I'm the group leader of the ADME
group at the Center for the Development of Therapeutics,
Broad Institute, in Cambridge, Massachusetts.
The topic of my presentation today is laboratory methods for the in vitro study of drug metabolism.
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Here is an overview of the presentation.
I'll begin with the background on the major classes of drug-metabolizing enzymes,
covering their subcellular localization and co-factor requirement for in vitro incubation.
Next, I'll talk about subcellular fractions and
whole-cell systems used for in vitro incubations,
followed by some practical aspects of conducting in vitro assays.
The second part of the talk will cover common applications such as clearance estimation,
kinetic parameter determination, enzyme inhibition,
enzyme induction, and reaction phenotyping studies.
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Drug-metabolizing enzymes are categorized into two classes,
phase I and phase II enzymes.
The purpose of drug-metabolizing enzymes is to transform
the drug to a polar form, so that it can be easily excreted.
Phase I enzymes mediate changes such as oxidation, reduction, and hydrolysis,
while phase II enzymes are involved in conjugation of phase I metabolites, to further increase their hydrophilicity.
Phase I enzymes include cytochrome P450,
flavin-containing monooxygenase, monoamine oxidase,
aldehyde oxidase, and carboxylesterase.
Of all these enzymes, cytochrome P450 (CYP) enzymes play a significant role in
phase I metabolism of drugs, and these enzymes will be the primary focus of the talk.
Phase II enzymes include UDP glucuronosyltransferase,
sulfotransferase, N-acetyl transferase, glutathione transferase, and methyltransferase.
Of all these enzymes, UDP glucuronosyltransferase (UGT) enzymes
play a significant role in phase II metabolism of drugs.
I have a simple example, shown on this slide, of a compound with
a methyl group that can undergo hydroxylation mediated by phase I enzymes,
which can then be glucuronidated by UGT enzymes as the phase II enzyme.
The resulting hydrophilic metabolite can then be excreted easily.
