Abstract

The gold standard for human drug metabolism studies is primary hepatocytes. However, availability is limited by donor organ scarcity. Therefore, efforts have been made to provide alternatives, e.g., the hepatocyte-like (NeoHep) cell type, which was generated from peripheral blood monocytes. In this study, expression and activity of phase I and phase II drug-metabolizing enzymes were investigated during transdifferentiation of NeoHep cells and compared with primary human hepatocytes. Important drug-metabolizing enzymes are cytochrome P450 isoforms (CYP1A1, 1A2, 2A6, 2B6, 2C8, 2C9, 2D6, 2E1, and 3A4), microsomal epoxide hydrolase 1, glutathione S-transferase A1 and M1, N-acetyltransferase 1, NAD(P)H menadione oxidoreductase 1, sulfotransferase 1A1, and UDP-glucuronosyltransferase 1A6. Monocytes and programmable cells of monocytic origin expressed only a few of the enzymes investigated. Throughout differentiation, NeoHep cells showed a continuously increasing expression of all drug-metabolizing enzymes investigated, resulting in stable basal activity after approximately 15 days. Fluorescence-based activity assays indicated that NeoHep cells and primary hepatocytes have similar enzyme kinetics, although the basal activities were significantly lower in NeoHep cells. Stimulation with 3-methylcholanthrene and rifampicin markedly increased CYP1A1/2 or CYP3A4 activities, which could be selectively inhibited by nifedipine, verapamil, ketoconazole, and quercetin. Our data reveal similarities in expression, activity, induction, and inhibition of drug-metabolizing enzymes between NeoHep cells and primary human hepatocytes and hence suggest that NeoHep cells are useful as an alternative to human hepatocytes for measuring bioactivation of substances.