Mitochondrial dysfunction and apoptosis underlie the hepatotoxicity of perhexiline.

Perhexiline is an anti-anginal drug developed in the late 1960s. Despite its therapeutic success, it caused severe hepatoxicity in selective patients, which resulted in its withdrawal from the market. In the current study we explored the molecular mechanisms underlying the cytotoxicity of perhexiline. In primary human hepatocytes, HepaRG cells, and HepG2 cells, perhexiline induced cell death in a concentration- and time-dependent manner. Perhexiline treatment also caused a significant increase in caspase 3/7 activity at 2 h and 4 h. Pretreatment with specific caspase inhibitors suggested that both intrinsic and extrinsic apoptotic pathways contributed to perhexiline-induced cytotoxicity, which was confirmed by increased expression of TNF-α, cleavage of caspase 3 and 9 upon perhexiline treatment. Moreover, perhexiline caused mitochondrial dysfunction, demonstrated by the classic glucose-galactose assay at 4 h and 24 h. Results from JC-1 staining suggested perhexiline caused loss of mitochondrial potential. Blocking mitochondrial permeability transition pore using inhibitor bongkrekic acid attenuated the cytotoxicity of perhexiline. Western blotting analysis also showed decreased expression level of pro-survival proteins Bcl-2 and Mcl-1, and increased expression of pro-apoptotic protein Bad. Direct measurement of the activity of individual components of the mitochondrial respiratory complex demonstrated that perhexiline strongly inhibited Complex IV and Complex V and moderately inhibited Complex II and Complex II + III. Overall, our data demonstrated that both mitochondrial dysfunction and apoptosis underlies perhexiline-induced hepatotoxicity. Published by Elsevier Ltd.