CYP3A Mediated Metabolism of Tyrosine Kinase Inhibitor Pexidartinib and Alleviated its Cytotoxicity to HepG2 Cells

Abstract

Pexidartinib (PLX, TURALIOTM) is a novel small molecule tyrosine‐kinase inhibitor with highly selective activity against colony‐stimulating factor‐1 receptor. PLX was developed for the treatment of symptomatic tenosynovial giant cell tumor in adult patients and approved by the US FDA in 2019. Despite its effectivity, frequently reported clinical cases with severe hepatotoxicity led to a black‐box warning issued to PLX by the FDA. However, the mechanisms of PLX‐induced hepatotoxicity remain largely unknown and the roles of PLX metabolism in its toxicity have not been investigated. Using liquid chromatography/mass spectrometry‐based metabolomic approaches, this study revealed 36 PLX metabolites in human liver microsome (HLM) and mouse liver microsome (MLM), including characterized 11 glutathione (GSH) ‐PLX adducts and 3 methoxyamine‐trapped aldehydes (hydrazones). Using recombinant cytochrome P450 enzymes, CYP3A4 and CYP3A5 were identified as the primary enzymes contributing to the formation of major metabolites, GSH‐PLX adducts, and methoxyamine‐trapped hydrazones. Their formation was significantly suppressed by the CYP3A inhibitor ketoconazole and in the liver S9 fraction of Cyp3a‐null mice, further confirming the role of CYP3A in PLX Phase I metabolism. More interestingly, we found that PLX is less toxic to CYP3A4‐ or CYP3A5‐overexpressed HepG2 cells compared to their counterpart HepG2 cells transduced with empty vector, indicating that CYP3A‐mediated metabolism attenuates PLX cytotoxicity to HepG2. In summary, 36 PLX metabolites including 11 GSH adducts and 3 hydrazones were identified in HLM and MLM. CYP3A4 and CYP3A5 are responsible for the Phase I biotransformation of PLX and alleviate its cytotoxicity in HepG2. Further studies are warranted to elucidate the role of metabolism in PLX hepatotoxicity and the mechanism associated with PLX‐induced liver injury.

Use of Structural Alerts for Reactive Metabolites in the Application SpotRM

“…Formation of a large range of GSH and methoxyamine adducts was recently established; metabolism in addition involved cleavage of the pyrrole ring. 37 For fedratinib (Entry 17), the analysis implies that there are risks of forming a quinone imine methide on the pyrimidine. There is precedence for pyrimidine methides from trimethoprim (TMP; Scheme 1) 23 and from a compound, BMS-932481 (Chart 1), reported in the literature.…”