Metabolism of 26,26,26,27,27,27-F6-1alpha,25-dihydroxyvitamin D3 by CYP24: species-based difference between humans and rats.
The compound 26,26,26,27,27,27-F(6)-1alpha,25(OH)(2)D(3) is a hexafluorinated analog of the active form of Vitamin D(3). The enhanced biological activity of F(6)-1alpha,25(OH)(2)D(3) is considered to be related to a decreased metabolic inactivation of the compound in target tissues such as the kidneys, small intestine, and bones. Our previous study demonstrated that CYP24 is responsible for the metabolism of F(6)-1alpha,25(OH)(2)D(3) in the target tissues. In this study, we compared the human and rat CYP24-dependent metabolism of F(6)-1alpha,25(OH)(2)D(3) by using the Escherichia coli expression system. In the recombinant E. coli cells expressing human CYP24, bovine adrenodoxin and NADPH-adrenodoxin reductase, F(6)-1alpha,25(OH)(2)D(3) was successively converted to F(6)-1alpha,23S,25(OH)(3)D(3), F(6)-23-oxo-1alpha,25(OH)(2)D(3), and the putative ether compound with the same molecular mass as F(6)-1alpha,25(OH)(2)D(3). The putative ether was not observed in the recombinant E. coli cells expressing rat CYP24. These results indicate species-based difference between human and rat CYP24 in the metabolism of F(6)-1alpha,25(OH)(2)D(3). In addition, the metabolite with a cleavage at the C(24)z.sbnd;C(25) bond of F(6)-1alpha,25(OH)(2)D(3) was detected as a minor metabolite in both human and rat CYP24. Although F(6)-1alpha,23S,25(OH)(3)D(3) and F(6)-23-oxo-1alpha,25(OH)(2)D(3) had a high affinity for Vitamin D receptor, the side-chain cleaved metabolite and the putative ether showed extremely low affinity for Vitamin D receptor. These findings indicate that human CYP24 has a dual pathway for metabolic inactivation of F(6)-1alpha,25(OH)(2)D(3) while rat CYP24 has only one pathway. Judging from the fact that metabolism of F(6)-1alpha,25(OH)(2)D(3) in rat CYP24-harboring E. coli cells is quite similar to that in the target tissues of rat, the metabolism seen in human CYP24-harboring E. coli cells appear to exhibit the same metabolism as in human target tissues. Thus, this recombinant system harboring human CYP24 appears quite useful for predicting the metabolism and efficacy of Vitamin D analogs in human target tissues before clinical trials.
Sakaki
Toshiyuki
T
Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Kitashirakawa, Oiwake-cho, Sakyo-ku, Japan. tsakaki@kais.kyoto-u.ac.jp
Sawada
Natsumi
N
Abe
Daisuke
D
Komai
Koichiro
K
Shiozawa
Shunichi
S
Nonaka
Yasuki
Y
Nakagawa
Kimie
K
Okano
Toshio
T
Ohta
Miho
M
Inouye
Kuniyo
K
D000818:Animals; D002417:Cattle; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D004926:Escherichia coli; D005461:Fluorine / Q000378:metabolism*; D008401:Gas Chromatography-Mass Spectrometry; D006801:Humans; D006900:Hydroxylation; D051381:Rats; D018167:Receptors, Calcitriol / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D013045:Species Specificity; D013250:Steroid Hydroxylases / Q000378:metabolism*; D013379:Substrate Specificity; D014807:Vitamin D / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism*; D065668:Vitamin D3 24-Hydroxylase
D018167:Receptors, Calcitriol; D011994:Recombinant Proteins; C118756:dihydroxy-vitamin D3; D014807:Vitamin D; D005461:Fluorine; D003577:Cytochrome P-450 Enzyme System; D013250:Steroid Hydroxylases; D065668:Vitamin D3 24-Hydroxylase