Induction of drug metabolizing enzymes: A survey of in vitro methodologies and interpretations used in the pharmaceutical industry—Do they comply with FDA recommendations?

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Abstract

The FDA has published guidelines by which to carry out and interpret in vitro induction studies using hepatocytes but do researchers in pharmaceutical companies actually follow these to the letter? In a survey of 30 participants in the pharmaceutical industry, 19 questions were posed regarding the species investigated, methodologies and interpretations of the data. Also addressed was the in-house decision making processes as a result of in vitro induction data. The survey showed that, although the basic methods were similar, no two researchers carried out and interpreted induction assays in exactly the same way. No single method was superior but all included enzyme activities as the major end point. Hepatocytes from animal species were used to confirm animal in vivo data but only human hepatocytes were used to predict human induction responses. If a compound was found to be positive in an in vitro induction assay, few would halt the development of the compound. The majority would consider other properties of the compound (bioavailability, clearance and therapeutic concentrations) and follow the FDA recommendation to conduct clinical drug–drug interaction studies. Overall, the results from this survey indicate that there is no standard pharmaceutical industry method or evaluation criterion by which in vitro assays are carried out. Rather than adhering to the FDA guidelines, some adapt methods and interpretation according to their own experience and need (whether screening or lead optimisation). There was general consensus that studies using human hepatocyte cultures currently provide the best indication of the in vivo induction potential of NCEs. In addition, the assessment of in vitro induction data from the literature suggest that the two-fold induction threshold and the percent of positive control criteria may not be the best methods to accurately assess the in vivo induction potential of a drug. Although the two-fold induction criterion is now obsolete, more predictive models for determining the clinical induction potential are needed. Alternative models are proposed and discussed herein.

Introduction

Drug metabolizing enzyme induction can have significant implications to the pharmacokinetics and toxicity of drugs; therefore, it is important to establish the induction potential of new chemical entities (NCEs) before they move up through the development pipeline. Recently, the FDA has published a draft guidance document outlining guidelines by which researchers should design drug–drug interaction (DDI) assays [1]. In addition, there are three other main papers which describe in some detail the considerations for enzyme induction: (1) Tucker et al. [2], which was a summary report from a conference held in Basel in November 2000; (2) Bjornsson et al. [3], a comprehensive outline of experimental conditions and interpretation recommendations; (3) Huang [4] which was a preliminary concept paper to the 2006 guidelines. The aim of these summary papers is to help industry towards an accepted standardized method by which to carry out and interpret DDI assays. If pharmaceutical companies wish to submit induction data, it makes sense to follow guidelines set out by the FDA. However, different companies develop compounds from different chemical classes and for different therapeutic areas. Moreover, different sites of the same company may well have their own preferred assay, depending on the stage of drug development (screening, lead optimisation, etc.), for determining the induction potential of NCEs. With this in mind, a survey (carried out in November 2005) was constructed to determine (1) if the majority of pharmaceutical companies were strictly adhering to the 2004 FDA draft guidance [4] (since this was the latest version at that time), (2) how they interpreted the data and (3) what decisions were made according to the results. Data from the literature was collated to examine the predictive value of the two 2004 FDA criteria for assessing the induction potential, namely, fold induction and percentage of the positive control. In this assessment, induction of CYP3A4 was used as an example because it is the most cited of the cytochrome P450 (CYP) enzymes and, in most cases rifampin was used as the positive control. In contrast, reports on CYP1A2 induction compare test compound induction with a variety of positive controls, such as β-naphthoflavone (BNF), omeprazole and 3-methycholanthrine (3-MC), which made the comparison between the two criteria more difficult. The draft FDA guidelines have changed since the survey was carried out, therefore, the changes and their significance will also be discussed.

Section snippets

The survey

Thirty researchers from 27 companies were asked to complete a questionnaire containing 19 questions. Of the companies involved, 23 were European and 7 were US-based from a total of 9 countries. For three companies, two researchers were from the same company; in one case, the two researchers were from European and US sites of the same company. Responses were collected between November 2005 and February 2006. The questions that comprised the survey focused on three main areas:

  • 1.

    In vitro methods and

Choice of species and enzyme isoforms

All 30 participants determined induction potential in human hepatocytes but some also used hepatocytes from animal species such as rat (seven researchers), dog (three researchers), monkey (two researchers) and pig (one researcher). It is generally considered that human hepatocytes are the best model for predicting interactions in humans because there are known differences in the induction response between animal and human hepatocytes to the same inducers. The well-known example of this is

Acknowledgements

The authors would like to sincerely thank the participants of this survey, the identity of whom must remain undisclosed but you know who you are!

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