From Known Knowns to Known Unknowns: Predicting in Vivo Drug Metabolites

Abstract

'It is better to be useful than perfect'. This review attempts to critically cover and assess the currently available approaches and tools to answer the crucial question: Is it possible (and if it is, to what extent is it possible) to predict in vivo metabolites and their abundances on the basis of in vitro and preclinical animal studies? In preclinical drug development, it is possible to produce metabolite patterns from a candidate drug by virtual means (i.e., in silico models), but these are not yet validated. However, they may be useful to cover the potential range of metabolites. In vitro metabolite patterns and apparent relative abundances are produced by various in vitro systems employing tissue preparations (mainly liver) and in most cases using liquid chromatography-mass spectrometry analytical techniques for tentative identification. The pattern of the metabolites produced depends on the enzyme source; the most comprehensive source of drug-metabolizing enzymes is cultured human hepatocytes, followed by liver homogenate fortified with appropriate cofactors. For specific purposes, such as the identification of metabolizing enzyme(s), recombinant enzymes can be used. Metabolite data from animal in vitro and in vivo experiments, despite known species differences, may help pinpoint metabolites that are not apparently produced in in vitro human systems, or suggest alternative experimental approaches. The range of metabolites detected provides clues regarding the enzymes attacking the molecule under study. We also discuss established approaches to identify the major enzymes. The last question, regarding reliability and robustness of metabolite extrapolations from in vitro to in vivo, both qualitatively and quantitatively, cannot be easily answered. There are a number of examples in the literature suggesting that extrapolations are generally useful, but there are only a few systematic and comprehensive studies to validate in vitro-in vivo extrapolations. In conclusion, extrapolation from preclinical metabolite data to the in vivo situation is certainly useful, but it is not known to what extent.

Modeling of interactions between xenobiotics and cytochrome P450 (CYP) enzymes

“…The knowledge obtained in in silico metabolism of drugs has not yet been transferred to predict the rate of metabolism in PBPK models of toxicological interest because of the differences in the key CYPs involved in metabolism and physicochemical properties of target compounds. Consequently, predictability of hepatic clearance of toxic compounds is limited, mostly focusing of a chemical class or closely related chemicals (Pelkonen et al, 2009, 2011; Coecke et al, 2013; Bessems et al, 2014). …”

A roadmap for the development of alternative (non-animal) methods for systemic toxicity testing

“…While these designs are not yet available, they are technically within reach. Pelkonen et al, 2009) the researcher and analysts, so they will be able to augment each other. As recently explained by Jaworska and Hoffmann (2010) via the concept of Bayesian networks, the structure of the testing strategy matters and will influence the risk assessment process.…”

“…Mass spectrometry, in particular, has proven to be an optimal tool to determine metabolites. As Pelkonen and co-workers state (Pelkonen et al, 2009) "…in silico or in vitro, in conjunction with animal data, provide useful and necessary information, on which to base the first PK studies in humans. The prerequisite is to use appropriate and up-to-date techniques and biological preparations."…”

“…Rather simple approaches give valuable information (Pelkonen et al, 2009), but it seems unlikely that this is sufficient for exclusion of a hazard. Still, assays like the peptide reactivity assay might be explored as to their predictive value for carcinogenicity.…”

A Pharmacokinetic Modeling Approach to Predict the Contribution of Active Metabolites to Human Efficacious Dose

“…Nonetheless, regulatory guidelines exist for the characterization of human metabolites according to whether they are deemed to have been adequately tested during preclinical toxicology [Food and Drug Administration, 2012 (http://www.fda.gov/downloads/Drugs/GuidanceComplianceRegulatoryInformation/Guidances/ UCM079266.pdf); International Conference on Harmonization, 2012 (http://www.ich.org/products/guidelines/multidisciplinary/article/ multidisciplinary-guidelines.html)]. Partially as a consequence of these guidelines, there has been much research into the prediction, detection, and quantification of human circulating metabolites (Anderson et al, 2009;Dalvie et al, 2009;Leclercq et al, 2009;Pelkonen et al, 2009;Lutz et al, 2010;Luffer-Atlas, 2012;Loi et al, 2013). The theoretical basis for predicting metabolite exposure has been discussed by Lutz et al (2010).…”