Rat cytochrome P450C24 (CYP24) does not metabolize 1,25‐dihydroxyvitamin D₂ to calcitroic acid

Abstract

1alpha-Hydroxy-23 carboxy-24,25,26,27-tetranorvitamin D(3) (calcitroic acid) is known to be the major water-soluble metabolite produced during the deactivation of 1,25-(OH)(2)D(3). This deactivation process is carried out exclusively by the multicatalytic enzyme CYP24 and involves a series of oxidation reactions at C(24) and C(23) leading to side-chain cleavage and, ultimately, formation of the calcitroic acid. Like 1,25-(OH)(2)D(3), 1alpha,25-1,25-(OH)(2)D(2) is also known to undergo side-chain oxidation and side-chain cleavage to form calcitroic acid (Zimmerman et al. [2001]. 1,25-(OH)(2)D(2) differs from 1,25-(OH)(2)D(3) by the presence of a double bond at C(22) and a methyl group at C(24). To date, there have been no studies detailing the participation of CYP24 in the production of calcitroic acid from 1,25-(OH)(2)D(2). We, therefore, studied the metabolism of 1,25-(OH)(2)D(3) and 1,25-(OH)(2)D(2) using a purified rat CYP24 system. Lipid and aqueous-soluble metabolites were prepared for characterization. Aqueous-soluble metabolites were subjected to reverse-phase high-pressure liquid chromatography (HPLC) analysis. As expected, 1,23(OH)(2)-24,25,26,27-tetranor D and calcitroic acid were the major lipid and aqueous-soluble metabolites, respectively, when 1,25-(OH)(2)D(3) was used as substrate. However, when 1,25-(OH)(2)D(2) was used as substrate, 1,24(R),25-(OH)(3)D(2) was the major lipid-soluble metabolite with no evidence for the production of either 1,23(OH)(2)-24,25,26,27-tetranor D or calcitroic acid. Apparently, the CYP24 was able to 24-hydroxylate 1,25-(OH)(2)D(2), but was unable to effect further changes, which would result in side-chain cleavage. These data suggest that the presence of either the double bond at C(22) or the C(24) methyl group impedes the metabolism of 1,25-(OH)(2)D(2) to calcitroic acid by CYP24 and that enzymes other than CYP24 are required to effect this process.

Insights into Vitamin D metabolism using cyp24 over-expression and knockout systems in conjunction with liquid chromatography/mass spectrometry (LC/MS)

“…Another aspect of this story is that CYP24-containing tissues such as primary neonatal keratinocytes do generate 1␣,24S-(OH) 2 D 2 and its further catabolites from 1␣-OH-D 2 in vitro. The production is abolished in CYP24-XO keratinocytes leaving one to conclude that CYP24 must be involved in both the step of activation to 1␣,24S-(OH) 2 D 2 and steps of its catabolism to polyhydroxylated inactive products [4,10]. The new knowledge will be useful to rationalize the mechanism of 1␣-OH-D 2 prodrug activation and to the continued development of synthetic 1␣,24S-(OH) 2 D 2 as a drug.…”

Hypergravity modulates vitamin D receptor target gene mRNA expression in mice

“…Dietary vitamin D 2 (ergocalciferol) and vitamin D 3 undergo the same activation process, involving 25-hydroxylation in the liver and subsequent 1␣-hydroxylation in the kidney. 1,25(OH) 2 D 2 is catabolized differently from 1,25(OH) 2 D 3 (12), and vitamin D 2 is less effective than vitamin D 3 in maintaining serum 25(OH)D levels (1). Vitamin D deficiency, mainly caused by inadequate sun exposure, results in rickets and osteomalacia, and is also associated with increased risk of cancer, autoimmune disease, infection, and cardiovascular disease (10).…”

The Bile Acid Derivatives Lithocholic Acid Acetate and Lithocholic Acid Propionate are Functionally Selective Vitamin D Receptor Ligands

“…Dietary vitamin D 2 (ergocalciferol) and vitamin D 3 are metabolized through the same activation process, involving 25-hydroxylation in the liver and subsequent 1a-hydroxylation in the kidney [6]. 1,25(OH) 2 D 2 has a different catabolic pathway than 1,25(OH) 2 D 3 [7], and vitamin D 2 is less effective than vitamin D 3 in maintaining serum 25(OH)D levels [8]. Vitamin D deficiency, caused primarily by inadequate sun exposure, results in rickets and osteomalacia, and is also associated with increased risk of cancer, autoimmune disease, infection, and cardiovascular disease [2,9] (see Sections X, XI, and XII of this volume).…”