A novel in vitro approach for risk assessment of reactive metabolites in Drug Discovery

The Biotransformation of drug molecules may result in the formation of stable as well as electrophilic reactive metabolites (RM). These chemically reactive species have the propensity to react with cellular macromolecules and have been implicated in an array of adverse drug reactions (ADRs), with the liver being the most frequently affected organ. Drug induced liver injury (DILI) accounts for over 50% of the cases of acute liver failure in the United States. Historically, the approach within the Pharmaceutical Industry to the threat from RMs has been one of minimizing RM formation. While this approach has been useful in numerous cases, it has had major drawbacks for use as a decision making tool for the progression of compounds. We have previously proposed that addressing potential RM risks needs to be done alongside other potential mechanisms of DILI. (1) We present now an approach for risk assessing compounds based on combining scores from two independent in vitro tools:

1.       Estimated RM Body Burden based on the fraction of metabolism leading to covalent binding in human hepatocytes and the estimated therapeutic dose.

2.       An in vitro hepatic screening panel composed of assays designed specifically to assess processes that have the potential to initiate liver injury. The panel consists of assays to assess i) cell toxicity - immortalized human hepatocyte derived cell lines (THLE) with no CYP activity or with 3A4 activity, ii) mitochondrial impairment – HepG2 cytotoxicity in galactose vs. glucose medium, iii) inhibition of hepatic biliary transporter activity – Bile Salt Export Pump (BSEP) and Multi-drug resistance protein Type 2 (Mrp2)

Using a set of marketed drugs we have shown that this combination approach gives a unique new way of assessing risk related to RMs. We have found that the use of a quantitative measure of the propensity to form RMs in combination with the in vitro hepatic screening panel assessing cellular processes with the potential to initiate liver injury, gives a much more holistic appreciation of potential hazards behind DILI. The results from this assessment and potential for use in the Pharmaceutical industry will be discussed.

(1) R.A. Thompson, E.M. Isin, Y. Li, R. Weaver, L. Weidolf, I. Wilson, A. Claesson, K. Page, H. Dolgos, J.G. Kenna, Risk assessment and mitigation strategies for reactive metabolites in drug discovery and development, Chem. Biol. Interact. 192 (2011) 65-71. doi:10.1016/j.cbi.2010.11.002.