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Risk Assessment of Drug-Drug Interactions

Drug-drug interactions (DDIs) cause unexpected adverse drug reactions and contribute to drug withdrawal from the market. Through inducing or inhibiting drug-processing proteins, a new drug could induce toxicity or diminish efficacy of other concomitant drugs, thus causing DDIs.

The major drug-processing proteins involved in DDIs include phase I drug-metabolizing enzymes: CYP1A2, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, and CYP3A; phase II enzymes: UGT1A and UGT2B; and drug transporters: P-gp (MDR1), BCRP, OATP1B1, OATP1B3, OCT2, OAT1, and OAT3 .

At the early stage of drug development, it is critical to address two main issues regarding DDIs: 1) What are the main metabolic-processing proteins responsible for the absorption, distribution, metabolism, and excretion of the new drug? 2) Does the new drug inhibit or induce major drug-processing proteins?

Following the initial step of identifying drug-processing proteins that contribute to ultimate drug clearance – usually by using human liver microsomes, primary hepatocytes, and/or cDNA-based plasmids that express specific drug-processing proteins – researchers can adopt the following approaches to evaluate the inhibitory or inductive properties of new drugs:

1. Inhibition studies

A few well-established assays are commonly conducted which target CYP inhibition using human liver microsomes, primary hepatocytes, or recombinant CYP isoenzymes along with enzyme-specific substrates.

For example, IC50 values are typically determined in a high-throughput format using fluorescence probe substrates and recombinant CYP isoenzymes (i.e., CYP1A2, 2A6, 2B6, 2C8, 2C9, 2C19, 2D6, 2E1, and 3A4) during the early drug discovery phase. Based on the IC50 results, acquiring additional kinetic parameters, such as Ki, will be useful to further assess the CYP inhibition mode.

2. Induction studies

Unlike enzyme inhibition, enzyme induction occurs at the transcription level predominantly through activating xenobiotic-sensing nuclear receptors PXR and CAR. Responding to ligand binding (or activation), these nuclear receptors function as transcription factors and induce expression of their downstream target genes, including multiple CYP genes (3A4, 3A5, 3A7, 2B6, 2C8, 2C9, and 2A6), multiple UDP-glucurosyltransferase genes (UGT1A1, 1A3, and 1A6), and MDR1.

A number of cell-based high throughput reporter assays focusing on PXR activation have been developed to test the ability of new drugs to bind to PXR and induce its above-mentioned target genes. Another approach is to examine expression levels of these genes, quantitated by real-time PCR, in response to new drugs in human primary hepatocytes.

3. In vivo animal models

In vitro assays provide fast and cost-effective methods to evaluate induction and inhibition profiles of new drugs. However, in vivo animal models are often necessary to further assess drug interaction properties in the relevant physiological system and to overcome the limitations in extrapolating the results of in vitro studies.

By exposing animals to a probe substrate with or without a test drug, pharmacokinetic parameters (e.g., Cmax and AUC) can be compared. Advanced animal tools have also been created to carry out complex experiments regarding DDIs. For example, genetically modified mice have become useful tools to investigate the impact of a specific drug-processing gene or nuclear receptor on drug interactions. In these mice, these genes are deleted (knockouts), are inserted and overexpressed (transgenics), or replace their mouse counterparts (humanized mice).

These experimental tools offer a quantitative risk assessment strategy, but we still have a knowledge gap for certain issues. For example, appropriate probe substrates are lacking for detecting the inhibitory activity of some drug transporters. Nevertheless, evaluation of potential DDIs should be a priority in the pre-clinical drug discovery phase and is an essential part of the regulatory review. To ensure the safety of new drugs, the FDA has issued and updated guidance for assessing potential DDIs.

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Categories: Toxicology and Pharmacology

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