Sunitinib is a multitargeted tyrosine kinase inhibitor associated with idiosyncratic hepatotoxicity. The mechanisms of this toxicity are unknown. We hypothesized that sunitinib undergoes metabolic activation to form chemically reactive, potentially toxic metabolites which may contribute to development of sunitinib-induced hepatotoxicity. The purpose of this study was to define the role of cytochrome P450 (P450) enzymes in sunitinib bioactivation. Metabolic incubations were performed using individual recombinant P450s, human liver microsomal fractions, and P450-selective chemical inhibitors. Glutathione (GSH) and dansylated GSH were used as trapping agents to detect reactive metabolite formation. Sunitinib metabolites were analyzed by liquid chromatography–tandem mass spectrometry. A putative quinoneimine–GSH conjugate (M5) of sunitinib was detected from trapping studies with GSH and dansyl–GSH in human liver microsomal incubations, and M5 was formed in an NADPH-dependent manner. Recombinant P450 1A2 generated the highest levels of defluorinated sunitinib (M3) and M5, with less formation by P450 3A4 and 2D6. P450 3A4 was the major enzyme forming the primary active metabolite N-desethylsunitinib (M1). In human liver microsomal incubations, P450 3A inhibitor ketoconazole reduced formation of M1 by 88%, while P450 1A2 inhibitor furafylline decreased generation of M5 by 62% compared to control levels. P450 2D6 and P450 3A inhibition also decreased M5 by 54 and 52%, respectively, compared to control. In kinetic assays, recombinant P450 1A2 showed greater efficiency for generation of M3 and M5 compared to that of P450 3A4 and 2D6. Moreover, M5 formation was 2.7-fold more efficient in human liver microsomal preparations from an individual donor with high P450 1A2 activity compared to a donor with low P450 1A2 activity. Collectively, these data suggest that P450 1A2 and 3A4 contribute to oxidative defluorination of sunitinib to generate a reactive, potentially toxic quinoneimine. Factors that alter P450 1A2 and 3A activity may affect patient risk for sunitinib toxicity.

Tyrosine kinase inhibitors are a rapidly expanding class of molecular targeted therapies for the treatment of various types of cancer and other diseases. An increasing number of clinically important small molecule tyrosine kinase inhibitors have been shown to undergo cytochrome P450-mediated bioactivation to form chemically reactive, potentially toxic products. Metabolic activation of tyrosine kinase inhibitors is proposed to contribute to the development of serious adverse reactions, including idiosyncratic hepatotoxicity. This article will review recent findings and ongoing studies to elucidate the link between drug metabolism and tyrosine kinase inhibitor-associated hepatotoxicity.

Purpose: Sunitinib is a multi-targeted tyrosine kinase inhibitor approved for the treatment of renal cell carcinoma, gastrointestinal stromal tumors, and pancreatic neuroendocrine tumors. However, clinical use of sunitinib is associated with potentially fatal idiosyncratic liver injury. The mechanisms of this toxicity are unknown. We hypothesized that sunitinib undergoes metabolic activation to form chemically reactive, potentially toxic metabolites, which may contribute to the development of sunitinib-induced liver injury. The purpose of this study was to define the role of cytochrome P450 (CYP) enzymes in the metabolism and bioactivation of sunitinib to provide insight into the mechanisms of drug toxicity. Methods: Metabolic incubations were performed using individual recombinant P450 enzymes, human liver microsomal fractions, and P450-selective chemical inhibitors. To assess reactive metabolite formation, glutathione (GSH) and a chemical derivative of GSH, dansyl-GSH, were used as trapping agents to detect reactive metabolite-GSH conjugates. Sunitinib metabolites were analyzed by liquid chromatography-tandem mass spectrometry. Results: Recombinant CYP3A4 was the major enzyme to form the primary active metabolite N-desethyl-sunitinib. Recombinant CYP1A2 generated the highest levels of reactive metabolites trapped as GSH conjugates, with minor contributions from CYP2D6 and CYP3A4. Detection of reactive metabolite dansyl-GSH conjugates from incubations with dansyl-GSH confirmed the generation of reactive metabolites by CYP1A2. In human liver microsomal incubations, the CYP3A inhibitor ketoconazole reduced formation of N-desethyl-sunitinib by 88%, while the CYP1A2 inhibitor furafylline decreased generation of reactive metabolite-GSH conjugates by 62%, compared to control levels. Quinidine (CYP2D6 inhibitor) and ketoconazole (CYP3A inhibitor) also decreased GSH conjugate formation by 54 and 52%, respectively, compared to control. Kinetic assays with recombinant P450s revealed that the overall enzyme efficiency (kcat/Km) for converting sunitinib to reactive metabolite-GSH conjugates was in the following order: CYP1A2 > CYP3A4 > CYP2D6. Conclusions: Collectively, these data indicate that CYP1A2 and CYP3A4 are important contributors to metabolic activation of sunitinib to generate reactive, potentially toxic metabolites. Factors that alter CYP1A2 and CYP3A4 activity may affect patient risk for sunitinib toxicity. Future studies will focus on examining the impact of P450 genetic variations and environmental factors on sunitinib metabolism and toxicity. Citation Format: Klarissa D. Jackson, Gracia M. Amaya, Rebecca Durandis, Kahari J. Wines, Arsany A. Abouda, Samuel A. Starks, R. Nathan Daniels. Metabolic activation of sunitinib: Implications for sunitinib-induced toxicities [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4911.