CYP3A Mediates an Unusual C(sp²)-C(sp³) Bond Cleavage via Ipso-Addition of Oxygen in Drug Metabolism

Mammalian cytochrome P450 drug-metabolizing enzymes rarely cleave carbon-carbon (C-C) bonds and the mechanisms of such cleavages are largely unknown. We identified two unusual cleavages of non-polar, unstrained C(sp²)-C(sp³) bonds in the FDA-approved tyrosine kinase inhibitor pexidartinib that are mediated by CYP3A4/5, the major human phase I drug metabolizing enzymes. Using a synthetic ketone, we rule out the Baeyer-Villiger oxidation mechanism that is commonly invoked to address P450-mediated C-C bond cleavages. Our studies in ¹⁸O₂ and H₂ ¹⁸O enriched systems reveal two unusual distinct mechanisms of C-C bond cleavage: one bond is cleaved by CYP3A-mediated ipso-addition of oxygen to a C(sp²) site of N-protected pyridin-2-amines, and the other occurs by a pseudo-retro-aldol reaction after hydroxylation of a C(sp³) site. This is the first report of CYP3A-mediated C-C bond cleavage in drug metabolism via ipso-addition of oxygen mediated mechanism. CYP3A-mediated ipso-addition is also implicated in the regioselective C-C cleavages of several pexidartinib analogs. The regiospecificity of CYP3A-catalyzed oxygen ipso-addition under environmentally friendly conditions may be attractive and inspire biomimetic or P450-engineering methods to address the challenging task of C-C bond cleavages.