Data from: Identification of the additional mitochondrial liabilities of 2-hydroxyflutamide when compared to its parent compound, flutamide in HepG2 cells

The androgen receptor antagonist, flutamide, is strongly associated with idiosyncratic drug-induced liver injury (DILI). Following administration, flutamide undergoes extensive first-pass metabolism to its primary metabolite, 2-hydroxyflutamide. Flutamide is a known mitochondrial toxicant; however there has been limited investigation into the potential mitochondrial toxicity of 2-hydroxyflutamide and its contribution to flutamide-induced liver injury. In this study we have used the acute glucose or galactose-conditioning of HepG2 cells to compare the mitochondrial toxicity of flutamide, 2-hydroxyflutamide and the structurally-related, non-hepatotoxic androgen receptor antagonist, bicalutamide. Compound–induced changes in mitochondrial oxygen consumption rate were assessed using Seahorse technology. Permeabilisation of cells and delivery of specific substrates and inhibitors of the various respiratory complexes provided more detailed information on the origin of mitochondrial perturbations. These analyses were supported by assessment of downstream impacts including changes in cellular NAD+/NADH ratio. Bicalutamide was not found to be a mitochondrial toxicant, yet flutamide and 2-hydroxyflutamide significantly reduced basal and maximal respiration. Both flutamide and 2-hydroxyflutamide significantly reduced respiratory complex I-linked respiration, though 2-hydroxyflutamide also significantly decreased complex II and V-linked respiration; liabilities not demonstrated by the parent compound. This study has identified for the first time, the additional mitochondrial liabilities of the major metabolite, 2-hydroxyflutamide compared with its parent drug, flutamide. Given the rapid production of this metabolite upon administration of flutamide, but not bicalutamide, we propose that the additional mitochondrial toxicity of 2-hydroxyflutamide may fundamentally contribute to the idiosyncratic DILI seen in flutamide-treated, but not bicalutamide-treated patients.

雄激素受体拮抗剂（androgen receptor antagonist）氟他胺（flutamide）与特发性药物性肝损伤（idiosyncratic drug-induced liver injury, DILI）密切相关。给药后，氟他胺会经历广泛的首过代谢（first-pass metabolism），转化为其主要代谢产物2-羟基氟他胺（2-hydroxyflutamide）。氟他胺是已知的线粒体毒物，但目前针对2-羟基氟他胺的潜在线粒体毒性及其在氟他胺诱导肝损伤中的作用的研究仍较为有限。本研究采用HepG2细胞（HepG2 cells）的急性葡萄糖或半乳糖条件培养模型，对比氟他胺、2-羟基氟他胺以及结构相关、无肝毒性的雄激素受体拮抗剂比卡鲁胺（bicalutamide）的线粒体毒性。采用Seahorse细胞能量代谢分析仪技术（Seahorse technology）检测化合物诱导的线粒体氧消耗速率变化。通过细胞透化处理并递送各类呼吸复合物（respiratory complexes）的特异性底物与抑制剂，可更深入解析线粒体功能紊乱的来源。上述分析辅以下游效应评估，包括细胞内NAD+/NADH比值（NAD+/NADH ratio）的变化。研究结果显示，比卡鲁胺未表现出线粒体毒性，而氟他胺与2-羟基氟他胺均显著降低了基础呼吸与最大呼吸速率。氟他胺与2-羟基氟他胺均可显著降低呼吸复合物I（complex I）依赖的呼吸活性，但2-羟基氟他胺还可进一步降低呼吸复合物II（complex II）与呼吸复合物V（complex V）依赖的呼吸活性——这两项毒性效应未在母体化合物（parent compound）氟他胺中观察到。本研究首次证实，相较于母体药物氟他胺，其主要代谢产物2-羟基氟他胺具有额外的线粒体毒性。由于氟他胺给药后可快速生成该代谢物，而比卡鲁胺则无此代谢特征，我们推测2-羟基氟他胺额外的线粒体毒性可能是氟他胺（而非比卡鲁胺）治疗患者出现特发性药物性肝损伤的核心致病因素。