A mechanistic biomarker investigation of fialuridine hepatotoxicity using the chimeric TK-NOG Hu-liver mouse model and in vitro micropatterned hepatocyte cocultures

Abstract

Abstract Fialuridine (FIAU) is a nucleoside-based drug that caused liver failure and deaths in a human clinical trial that were not predicted by nonclinical safety studies. A recent report concluded that a TK-NOG humanized liver (hu-liver) mouse model detected human-specific FIAU liver toxicity, and broader use of that model could improve drug safety testing. We further evaluated this model at similar dose levels to assess FIAU sensitivity and potential mechanistic biomarkers. Although we were unable to reproduce the marked acute liver toxicity with two separate studies (including one with a “sensitized” donor), we identified molecular biomarkers reflecting the early stages of FIAU mitochondrial toxicity, which were not seen with its stereoisomer (FIRU). Dose dependent FIAU-induced changes in hu-liver mice included more pronounced reductions in mitochondrial to nuclear DNA (mtDNA/nucDNA) ratios in human hepatocytes compared to mouse hepatocytes and kidneys of the same animals. FIAU treatment also triggered a p53 transcriptional response and opposing changes in transcripts of nuclear- and mitochondrial-encoded mitochondrial proteins. The time dependent accumulation of FIAU into mtDNA is consistent with the ≥9-week latency of liver toxicity observed for FIAU in the clinic. Similar changes were observed in an in vitro micro-patterned hepatocyte coculture system. In addition, FIAU-dependent mtDNA/nucDNA ratio and transcriptional alterations, especially reductions in mitochondrially encoded transcripts, were seen in livers of non-engrafted TK-NOG and CD-1 mice dosed for a shorter period. Conclusion: These mechanistic biomarker findings can be leveraged in an in vitro model and in a more routine preclinical model (CD-1 mice) to identify nucleosides with such a FIAU-like mitochondrial toxicity mechanistic liability potential. Further optimization of the TK-NOG hu-liver mouse model is necessary before broader adoption for drug safety testing.