Idiosyncratic drug-induced liver injury (IDILI) is an adverse and typically rare response to drugs that are safe in most people. Although infrequent, the response to these reactions can be severe and result in liver failure and death. Despite extensive research, the underlying mechanisms responsible for IDILI are currently unknown. One theory suggests that a modest inflammatory stress can render individuals susceptible to IDILI. Animal models based on this hypothesis demonstrated that administration of a nontoxic dose of lipopolysaccharide (LPS) to induce modest inflammation interacts with drugs associated with causing IDILI to precipitate liver injury in rodents. Trovafloxacin (TVX) is a fluoroquinolone antibiotic associated with IDILI. In mice, TVX was not hepatotoxic but became injurious during concomitant inflammation induced by LPS. The pathogenesis of this liver injury was critically mediated by the proinflammatory cytokine tumor necrosis factor-alpha (TNF). The focus of this dissertation was to test the hypothesis that TVX and TNF interact directly to cause hepatocyte cell death, and to identify critical signaling mechanisms involved in cytotoxicity. In human derived HepG2 cells, neither TVX nor TNF alone induced any cytotoxicity in vitro. However, in combination TVX/TNF treatment resulted in cytotoxicity in a concentration-dependent manner. The hepatocellular death was dependent on activated caspases and involved a prolonged activation of c-Jun N-terminal kinase (JNK). TVX has the potential to inhibit mammalian topoisomerase enzymes, which can lead to DNA damage. In hepatocytes, TVX produced DNA damage and resulted in cell cycle arrest and decreased cell proliferation. TVX also activated extracellular signal-regulated kinases (ERK) and ataxia telangiectasia and Rad3 related (ATR), both of which can be activated in response to genotoxic stress. However, none of the above events evoked cytotoxicity unless cells were also exposed to TNF. Activated ERK and ATR promoted cytotoxicity in hepatocytes exposed to both TVX and TNF. Collectively, these results contribute to increased understanding of the molecular mechanisms underlying the cytotoxic interaction of TVX and TNF. If similar mechanisms are involved in response to other IDILI associated drugs interacting with TNF to kill hepatocytes, this information could be used to develop high-throughput screening assays capable of discerning the potential of new drug candidates to cause IDILI.
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