Vitamin D metabolism in human colon adenocarcinoma-derived Caco-2 cells: expression of 25-hydroxyvitamin D3-1alpha-hydroxylase activity and regulation of side-chain metabolism

J Steroid Biochem Mol Biol. 1997 May;62(1):21-8. doi: 10.1016/s0960-0760(97)00020-4.


1Alpha,25-dihydroxyvitamin D3 (1alpha,25(OH)2D3) and its synthetic analogues exhibit structure-related variations in their growth inhibitory actions in human colon adenocarcinoma-derived Caco-2 cells. Because this might be caused by differences in resistance against metabolic degradation, we used high performance liquid chromatography (HPLC) analysis to investigate pathways of vitamin D metabolism in two different Caco-2 cell clones. Importantly, when Caco-2 cells were incubated with tritium-labelled 25-hydroxyvitamin D3 (25(OH)D3) for up to 2 h they produced almost exclusively a metabolite, which was identified as 1alpha,25(OH)2D3 by co-chromatography with the synthetic standard in two different HPLC systems, and by a radioligand assay showing an identical binding affinity to the intestinal nuclear vitamin D receptor. Expression of the 25(OH)D3-1alpha-hydroxylase appears to be constitutive because almost identical enzyme activities are observed in any growth phase. 1Alpha,25(OH)2D3 can also activate side chain metabolism in Caco-2 cells: thereby, 1alpha,25(OH)2D3 or 25(OH)D3 are metabolized through the C-24 oxidative pathway into 1alpha,24(R),25(OH)3D3 and 24(R),25(OH)2D3, respectively, which undergo sequential metabolism into 1alpha,25(OH)2-24oxo-D3 and 24-oxo-25(OH)D3. Through C-23 oxidation these intermediary metabolites are further converted into 1alpha,23,25(OH)3-24-oxo-D3 and 23,25(OH)2-24-oxo-D3. Also direct C-23 oxidation of the substrates 1alpha,25(OH)2D3 and 25(OH)D3 generates 1alpha,23(S),25(OH)3D3 and 23(S),25(OH)2D3, respectively. In summary, our results demonstrated the presence of distinct pathways of vitamin D metabolism in Caco-2 cells: apart from metabolizing 1alpha,25(OH)2D3 along the C-24 and C-23 oxidative pathways, Caco-2 cells are able to synthesize 1alpha,25(OH)2D3 from 25(OH)D3 through constitutive expression of 25(OH)D3-1alpha-hydroxylase activity. The relevance of this finding for the intrinsic growth control of neoplastic colonocytes is discussed.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Caco-2 Cells
  • Calcifediol / metabolism
  • Calcitriol / metabolism
  • Cholestanetriol 26-Monooxygenase
  • Chromatography, High Pressure Liquid
  • Clone Cells
  • Gene Expression Regulation, Enzymologic
  • Gene Expression Regulation, Neoplastic
  • Humans
  • Kinetics
  • Models, Chemical
  • Receptors, Calcitriol / metabolism
  • Steroid Hydroxylases / biosynthesis*
  • Vitamin D / analogs & derivatives
  • Vitamin D / metabolism*


  • Receptors, Calcitriol
  • Vitamin D
  • Steroid Hydroxylases
  • CYP27A1 protein, human
  • Cholestanetriol 26-Monooxygenase
  • Calcitriol
  • Calcifediol