Metabolic Interactions of Side-chain Extended and Unsaturated Vitamin D Analogs with Cytochrome P450 Enzymes:...

The clinical use of 1,25-dihydroxycholecalciferol (1,25D3), the active form of vitamin D₃, is limited by its calcemic side effects and rapid metabolic degradation. To overcome these limitations, we designed novel vitamin D analogs with extended, rigidified, and branched side chains. Among them, PRI-1938, featuring a 5,6-trans triene system and 22,24-all-trans side-chain geometry, demonstrated markedly enhanced resistance to enzymatic catabolism. In vitro assays revealed that metabolic conversion of PRI-1938 by the nonselective cytochrome P450 3A4 (CYP3A4) enzyme was ca. 4-fold lower than that of the previously obtained PRI-1906 and over 9-fold lower than 1,25D3. All new analogs, including PRI-1927 and PRI-1937, exhibited significantly higher stability toward mitochondrial cytochrome P450 24A1 (CYP24A1), the vitamin D-selective catabolic enzyme, than that of 1,25D3. Molecular modeling and quantum mechanical calculations indicated that PRI-1938 adopts a highly stable conformation in the CYP24A1 active site, stabilized by four hydrogen bonds and multiple hydrophobic interactions. The spatially optimized interaction network reduces access to the catalytic heme, resulting in the lowest observed metabolic conversion. These findings highlight the critical role of the side-chain geometry in modulating metabolic stability and support the further development of PRI-1938 as a promising anticancer vitamin D analog.