Going Beyond Common Drug Metabolizing Enzymes: Case Studies of Biotransformation Involving Aldehyde Oxidase,  -Glutamyl Transpeptidase, Cathepsin B, Flavin-Containing Monooxygenase, and ADP-Ribosyltransferase

Abstract

The significant roles that cytochrome P450 (P450) and UDPglucuronosyl transferase (UGT) enzymes play in drug discovery cannot be ignored, and these enzyme systems are commonly examined during drug optimization using liver microsomes or hepatocytes. At the same time, other drug-metabolizing enzymes have a role in the metabolism of drugs and can lead to challenges in drug optimization that could be mitigated if the contributions of these enzymes were better understood. We present examples (mostly from Genentech) of five different non-P450 and non-UGT enzymes that contribute to the metabolic clearance or bioactivation of drugs and drug candidates. Aldehyde oxidase mediates a unique amide hydrolysis of ,7-tetrahydro-1-benzothiophene-2-carboxamide), leading to high clearance of the drug. Likewise, the rodent-specific ribose conjugation by ADP-ribosyltransferase leads to high clearance of an interleukin-2-inducible T-cell kinase inhibitor. Metabolic reactions by flavin-containing monooxygenases (FMO) are easily mistaken for P450-mediated metabolism such as oxidative defluorination of 4-fluoro-N-methylaniline by FMO. Gamma-glutamyl transpeptidase is involved in the initial hydrolysis of glutathione metabolites, leading to formation of proximate toxins and nephrotoxicity, as is observed with cisplatin in the clinic, or renal toxicity, as is observed with efavirenz in rodents. Finally, cathepsin B is a lysosomal enzyme that is highly expressed in human tumors and has been targeted to release potent cytotoxins, as in the case of brentuximab vedotin. These examples of non-P450-and non-UGT-mediated metabolism show that a more complete understanding of drug metabolizing enzymes allows for better insight into the fate of drugs and improved design strategies of molecules in drug discovery.

Metabolism of F18, a Derivative of Calanolide A, in Human Liver Microsomes and Cytosol

“…In general, FMOs typically catalyze the addition of an oxygen atom to substrates that contain a nucleophilic heteroatom, such as nitrogen, sulfur, phosphorus or selenium (Fan et al, 2016). However, due to the lack of effective and specific inhibitors, the contributions of FMOs cannot be distinguished from those of CYPs.…”

“…In contrast to CYPs, FMOs are not readily irreversibly inhibited by different chemicals (Kedderis and Rickert, 1985). Instead, because FMOs are thermally labile in the absence of NADPH, thermal inactivation studies are often performed to evaluate the involvement of FMOs in drug metabolism (Fan et al, 2016). After thermal inactivation, the formation of M1 was effectively inhibited, indicating the contribution of FMOs to the production of M1.…”

“…(2) The metabolism of benzydamine, an FMO substrate, was effectively inhibited in the incubation system. FMOs have been reported to catalyze oxidative defluorination (Fan et al, 2016). Our results might provide a new scenario in which FMOs catalyze oxidative dehalogenation.…”

“…Furthermore, both M3-1 and M3-2 were detected in the incubations with recombinant FMO1, FMO3, and FMO5. As described above, FMO superfamily members mainly catalyze heteroatom oxidation, particularly that of nucleophilic atoms (Fan et al, 2016). To the best of our knowledge, the carbonyl reduction reactions catalyzed by FMOs have not been described previously.…”

Toward an Understanding of Structural Insights of Xanthine and Aldehyde Oxidases: An Overview of their Inhibitors and Role in Various Diseases

“…However, only limited examples are available where AOX1 bio‐reduces compounds containing N ‐oxides, and sulfoxides . AOX1‐medicated metabolism of N ‐heterocyclic drug candidates has gained a significant interest at clinical level in the drug discovery recently as they were previously developed to avoid cytochrome P450‐mediated oxidation (phase 1 metabolism) . This leads to inactivation of N ‐heterocyclic drug candidates.…”