Human cytochrome P450 17A1 structures with metabolites of prostate cancer drug abiraterone reveal substrate-binding plasticity and a second binding site

Abiraterone acetate is a first-line therapy for castration -resistant prostate cancer. This prodrug is deacetylated in vivo to abiraterone, which is a potent and specific inhibitor of cy-tochrome P450 17A1 (CYP17A1). CYP17A1 performs two sequential steps that are required for the biosynthesis of an-drogens that drive prostate cancer proliferation, analogous to estrogens in breast cancer. Abiraterone can be further metab-olized in vivo on the steroid A ring to multiple metabolites that also inhibit CYP17A1. Despite its design as an active-site- directed substrate analog, abiraterone and its metabolites demonstrate mixed competitive/noncompetitive inhibition. To understand their binding, we solved the X-ray structures of CYP17A1 with three primary abiraterone metabolites. Despite different conformations of the steroid A ring and substituents, all three bound in the CYP17A1 active site with the steroid core packed against the I helix and the A ring C3 keto or hy-droxyl oxygen forming a hydrogen bond with N202 similar to abiraterone itself. The structure of CYP17A1 with 3-keto, 5 alpha- abiraterone was solved to 2.0 angstrom, the highest resolution to date for a CYP17A1 complex. This structure had additional electron density near the F/G loop, which is likely a second molecule of the inhibitor and which may explain the noncompetitive in-hibition. Mutation of the adjacent Asn52 to Tyr positions its side chain in this space, maintains enzyme activity, and pre-vents binding of the peripheral ligand. Collectively, our find-ings provide further insight into abiraterone metabolite binding and CYP17A1 function.