Differentiation-Related Pathways of 1α,25-Dihydroxycholecalciferol Metabolism in Human Colon Adenocarcinoma-Derived Caco-2 Cells: Production of 1α,25-Dihydroxy-3epi-cholecalciferol

Abstract

We used the human colon adenocarcinoma-derived cell line Caco-2, which spontaneously differentiatesin vitro,as a model system to investigate the metabolism of 1α,25-dihydroxycholecalciferol in colon cancer cells. Subconfluent proliferating and confluent differentiating cells were incubated with 1 μM 1α,25-dihydroxycholecalciferol for a period of 24 to 48 h. HPLC analysis of the lipid extract of both cells and media was performed to isolate and identify the various metabolites of 1α,25-dihydroxycholecalciferol. Undifferentiated, highly proliferating Caco-2 cells metabolized 1α,25-dihydroxycholecalciferol into several side chain modified metabolites formed through the C-24 oxidation pathway. In contrast, no metabolites of the C-24 oxidation pathway were identified in differentiated Caco-2 cells. However, differentiated cells produced significant amounts of a metabolite which was less polar than 1α,25-dihydroxycholecalciferol on a straight phase HPLC system. This metabolite was identified as 1α,25-dihydroxy-3α-cholecalciferol by comigration with a synthetic standard on two different HPLC systems and gas chromatography–mass spectrometry. Thus, we were able to demonstrate that the state of differentiation has a profound influence on 1α,25-dihydroxycholecalciferol metabolism in colon cancer cells.