Abstract

Gefitinib and erlotinib are epidermal growth factor receptor–tyrosine kinase inhibitors (EGFR-TKIs) with activity against metastatic non–small cell lung cancer. Aldehyde oxidase-1 (AOX1) is a cytosolic drug-metabolizing enzyme. We conducted an experimental and molecular docking study on the effect of gefitinib, erlotinib, and select metabolites on the in vitro catalytic activity of AOX1, as assessed by carbazeran 4-oxidation, and determined the impact of AOX1 inhibition on hepatic metabolism of zaleplon and methotrexate. Gefitinib, desmorpholinopropylgefitinib, erlotinib, desmethylerlotinib, and didesmethylerlotinib inhibited human hepatic cytosolic carbazeran 4-oxidation by a competitive mode, with inhibition constants in submicromolar or low micromolar concentrations. Desmethylgefitinib did not affect AOX1 catalytic activity. A similar pattern was obtained when investigated with human kidney cytosol or recombinant AOX1. The differential effect of gefitinib on human, rat, and mouse hepatic AOX1 catalytic activity suggests species-dependent chemical inhibition of AOX1. Erlotinib was considerably more potent than gefitinib in decreasing hepatic cytosolic zaleplon 5-oxidation and methotrexate 7-oxidation. Molecular docking analyses provided structural insights into the interaction between EGFR-TKIs and AOX1, with key residues and bonds identified, which provided favorable comparison and ranking of potential inhibitors. Based on the US Food and Drug Administration guidance to assess the risk of drug-drug interactions, the calculated R₁ values indicate that further investigations are warranted to determine whether gefitinib and erlotinib impact AOX1-mediated drug metabolism in vivo. Overall, erlotinib desmethylerlotinib, didesmethylerlotinib, gefitinib, and desmorpholinopropylgefitinib are potent inhibitors of human AOX1 catalytic function and hepatic metabolism of zaleplon and methotrexate, potentially affecting drug efficacy or toxicity.