Elucidation of metabolic pathways of 25-hydroxyvitamin D3 mediated by CYP24A1 and CYP3A using Cyp24a1 knockout rats generated by CRISPR/Cas9 system.

Abstract

CYP24A1-deficient (Cyp24a1 KO) rats were generated using the CRISPER/Cas9 system to investigate CYP24A1-dependent or -independent metabolism of 25(OH)D3, the prohormone of calcitriol. Plasma 25(OH)D3 concentrations in Cyp24a1 KO rats were approximately twofold higher than in wild-type rats. Wild-type rats showed five metabolites of 25(OH)D3 in plasma following oral administration of 25(OH)D3, and these metabolites were not detected in Cyp24a1 KO rats. Among these metabolites, 25(OH)D3-26,23-lactone was identified as the second major metabolite with a significantly higher Tmax value than others. When 23S,25(OH)2D3 was administered to Cyp24a1 KO rats, neither 23,25,26(OH)3D3 nor 25(OH)D3-26,23-lactone was observed. However, when 23S,25R,26(OH)3D3 was administered to Cyp24a1 KO rats, plasma 25(OH)D3-26,23-lactone was detected. These results suggested that CYP24A1 is responsible for the conversion of 25(OH)D3 to 23,25,26(OH)3D3 via 23,25(OH)2D3, but enzyme(s) other than CYP24A1 may be involved in the conversion of 23,25,26(OH)3D3 to 25(OH)D3-26,23-lactone. Enzymatic studies using recombinant human CYP species and the inhibitory effects of ketoconazole suggested that CYP3A plays an essential role in the conversion of 23,25,26(OH)3D3 into 25(OH)D3-26,23-lactone in both rats and humans. Taken together, our data indicate that Cyp24a1 KO rats are valuable for metabolic studies of vitamin D and its analogs. In addition, long-term administration of 25(OH)D3 to Cyp24a1 KO rats at 110 μg/kg body weight/day resulted in significant weight loss and ectopic calcification. Thus, Cyp24a1 KO rats could represent an important model for studying renal diseases originating from CYP24A1 dysfunction.

• Authors: Kaori Yasuda, Miyu Nishikawa, Kairi Okamoto, Kyohei Horibe, Hiroki Mano, Mana Yamaguchi, Risa Okon, Kimie Nakagawa, Naoko Tsugawa, Toshio Okano, Fumihiro Kawagoe, Atsushi Kittaka, Shinichi Ikushiro, Toshiyuki Sakaki
• Journal: The Journal of biological chemistry (J Biol Chem) 2021 Jan-Jun Vol. 296 Pg. 100668 ISSN: 1083-351X [Electronic] United States
• PMID: 33865853 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
• Copyright: Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.

Chemical References

• Vitamins
• Cytochrome P-450 CYP3A
• Cyp24a1 protein, rat
• Vitamin D3 24-Hydroxylase
• Calcifediol

Topics

• Animals
• Animals, Genetically Modified
• CRISPR-Cas Systems
• Calcifediol (administration & dosage, metabolism)
• Cytochrome P-450 CYP3A (metabolism)
• Metabolome (drug effects)
• Rats
• Vitamin D3 24-Hydroxylase (antagonists & inhibitors, genetics, metabolism)
• Vitamins (administration & dosage, metabolism)