Endocrine Abstracts

The active form of vitamin D, 1,25-dihydroxyvitamin D₃ (1,25D₃) is a potent antiproliferative agent with putative applications in the treatment of common cancers. However, doses of 1,25D₃ required to achieve tangible anticancer responses also stimulate unwanted calciotropic effects. Data suggest that this is due, in part, to acquired resistance to 1,25D₃ in cancer cells, particularly in more aggressive tumours. To investigate this further we have assessed patterns of gene expression in MCF-7 breast cancer cells and a 1,25D₃ resistant variant of these cells (MCF-7res) cloned by extended culture in high concentrations (10^-6M) of 1,25D₃. In experiments MCF-7 and MCF-7res cells were cultured in the presence or absence of 1,25D₃ (100 nM) for 6 hours and RNA from n=3 separate experiments were isolated. These preparations of RNA were pooled and used in DNA array analyses to determine differences in gene expression between untreated cells and those exposed to 1,25D₃. After treatment with 1,25D₃ genetic profiles of the two cell lines were completely different with the exception of CYP24, the gene encoding 1,25D₃-inactivating enzyme 24-hydroxylase, which was upregulated in both wild type MCF-7 and MCF-7res cells. This finding emphasises the importance of this pathway in defining vitamin D responsiveness in breast tumours. Amongst the genes that were upregulated in MCF-7 but not MCF-7res was transforming growth factor beta 2 (TGFbeta2). TGFbeta2 is a negative growth regulator of epithelial cells and interestingly contains a Vitamin D response element in its promoter region. Further studies showed that following 12 hours of 1,25D₃ treatment, TGFbeta2 mRNA expression was upregulated in MCF-12A normal breast epithelial cells (8 fold, p<0001) and 1,25D₃-sensitive MCF-7 cells (4 fold, p<0.02) but not in 1,25D₃-insensitive cells such as MCF-7res and MDA-MB-231. These data suggest that activation of TGFbeta2 is an important step in mediating effects of 1,25D₃ on breast cancer cells.