Untargeted metabolomics reveals anaerobic glycolysis as a novel target of the hepatotoxic antidepressant nefazodone.

Mitochondrial damage is considered a hallmark of drug-induced liver injury (DILI). However, despite the common molecular etiology, the evolution of the injury is usually unpredictable, with some cases that are mild and reversible upon discontinuation of the treatment and others characterized by irreversible acute liver failure. This suggests that additional mechanisms of damage play a role in determining the progression of the initial insult. To uncover novel pathways potentially involved in DILI, we investigated in vitro the metabolic perturbations associated with nefazodone, an antidepressant associated with acute liver failure. Several pathways associated with ATP production, including gluconeogenesis, anaerobic glycolysis and oxidative phosphorylation, were altered in human hepatocellular carcinoma-derived (Huh7) cells after two hour-exposure to a 50 μM extracellular concentration of nefazodone. In the presence or absence of glucose, ATP production of Huh7 cells was glycolysis- and oxidative phosphorylation-dependent, respectively. In glucose-containing medium, nefazodone-induced ATP depletion from Huh7 cells was biphasic. Huh7 cells in glucose-free medium were more sensitive to nefazodone than those in glucose-containing medium, losing the biphasic inhibition. Primary cultured mouse hepatocytes, mainly dependent on oxidative phosphorylation, were more sensitive than Huh7 cells to nefazodone exposure in glucose-containing medium. At lower extracellular concentrations, nefazodone inhibited the oxygen consumption of Huh7 cells, whereas, at higher extracellular concentrations, it also inhibited the extracellular acidification. ATP content was rescued by increasing the extracellular concentration of glucose. In conclusion, nefazodone has a dual inhibitory effect on mitochondrial-dependent and mitochondrial-independent ATP production. Significance Statement Mitochondrial damage is a hallmark of drug-induced liver injury, yet other collateral alterations might contribute to the severity and evolution of the injury. Our in vitro study supports previous results arguing that a deficit in hepatic glucose metabolism, concomitant to the mitochondrial injury, might be cardinal in the prognosis of the initial insult to the liver. From a drug development standpoint, coupling anaerobic glycolysis and mitochondrial function assessment might increase the drug-induced liver injury preclinical screening performance.