Metabolomics reveals a unique CYP3A-mediated C(sp3)-C(sp2) bond cleavage via ipso-addition reaction in drug metabolism

05 June 2023, Version 1
This content is a preprint and has not undergone peer review at the time of posting.


Cytochrome P450-mediated carbon–carbon (C-C) cleavages are unusual, especially for mammalian drug metabolizing enzymes. Revealing the unusual reactions in biological system is very arduous and selectively oxidative C-C cleavage is also a long-standing challenge in chemistry and biology. We herein present a rapid and efficient metabolomic-based approach to uncover human CYP3A-mediated non-polar, unstrained C(sp2)-C(sp3) bond cleavage in the CSF-1R inhibitor pexidartinib. Using synthetic metabolites, 18O2, and H218O, we demonstrate that one unique cleavage is via the ipso-addition reaction. This is the first report of CYP3A-mediated ipso-addition reaction to the 5-alkylated N-protected pyridin-2-amines. We have expanded the range of substrates undergoing CYP3A-mediated ipso-addition reactions beyond para-phenols to include N-protected alkylated pyridine-2-amines. Our metabolomic-based approach also successfully discovered the CYP3A-mediated C(sp2)-C(sp3) bond cleavage of PEX analogs as well as the antidepressant nefazodone. This work established an efficient strategy to identify the uncommon reactions in drug metabolism using a metabolomic strategy. More importantly, the environmentally friendly conditions of CYP3A-catalyzed unusual ispo-addition reactions hold the potential to inspire future exploration of biomimetic P450-reprogramming methods for addressing the challenging task of unactivated C-C bond cleavage in the field.


carbon-carbon bond cleavage
ipso addition reaction

Supplementary materials

Supplementary materials_Pexidartinib
Supplementary materials of Pexidartinib


Comments are not moderated before they are posted, but they can be removed by the site moderators if they are found to be in contravention of our Commenting Policy [opens in a new tab] - please read this policy before you post. Comments should be used for scholarly discussion of the content in question. You can find more information about how to use the commenting feature here [opens in a new tab] .
This site is protected by reCAPTCHA and the Google Privacy Policy [opens in a new tab] and Terms of Service [opens in a new tab] apply.