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Histone deacetylase inhibitors (HDACi) are part of a growing class of epigenetic therapies used for the treatment of cancer. Although HDACis are effective in the treatment of T-cell lymphomas, treatment of solid tumors with this class of drugs has not been successful. Overexpression of the multidrug resistance protein P-glycoprotein (P-gp), encoded by ABCB1, is known to confer resistance to the HDACi romidepsin in vitro, yet increased ABCB1 expression has not been associated with resistance in patients, suggesting that other mechanisms of resistance arise in the clinic. To identify alternative mechanisms of resistance to romidepsin, we selected MCF-7 breast cancer cells with romidepsin in the presence of the P-gp inhibitor verapamil to reduce the likelihood of P-gp-mediated resistance. The resulting cell line, MCF-7 DpVp300, does not express P-gp and was found to be selectively resistant to romidepsin but not to other HDACis such as belinostat, panobinostat, or vorinostat. RNA-sequencing analysis revealed upregulation of the mRNA coding for the putative methyltransferase, METTL7A, whose paralog, METTL7B, was previously shown to methylate thiol groups on hydrogen sulfide and captopril. As romidepsin has a thiol as the zinc-binding moiety, we hypothesized that METTL7A could inactivate romidepsin and other thiol-based HDACis via methylation of the thiol group. We demonstrate that expression of METTL7A or METTL7B confers resistance to thiol-based HDACis and that both enzymes are capable of methylating thiol-containing HDACis. We thus propose that METTL7A and METTL7B confer resistance to thiol-based HDACis by methylating and inactivating the zinc-binding thiol., (©2023 American Association for Cancer Research.)

Time-dependent inactivation (TDI) of cytochrome P450 (CYP) enzymes may result in clinical drug-drug interactions (DDIs). Therefore, designing out of CYP TDI prior to advancing a compound to clinical development is highly desirable. As TDI of CYP3A is a common occurrence in small molecule drug discovery, high-throughput methods are sought to help identify the mechanism of inactivation and enable design strategies to mitigate CYP3A TDI. CYP inactivation via modification or destruction of the prosthetic heme group results in loss of the ability of the enzyme to bind carbon monoxide. Additionally, formation of a tight binding complex with the heme iron, referred to as a metabolic intermediate (MI) complex, also results in enzyme inactivation. The methods described herein provide a high-throughput means of identifying and comparing compounds for their ability to inactivate via destruction/modification of the heme via loss of the ability to bind carbon monooxide, as well as via formation of an MI complex., (Copyright © 2023. Published by Elsevier Inc.)