Calcitriol supplementation effects on Ki67 expression and transcriptional profile of breast cancer specimens from post-menopausal patients

Background: Breast cancer patients present lower 1,25(OH)2D3 or 25(OH)D3 serum levels than unaffected women. Although 1,25(OH)2D3 pharmacological concentrations of 1,25(OH)2D3 may exert antiproliferative effects in breast cancer cell lines, much uncertainty remains about the effects of calcitriol supplementation in tumor specimens in vivo. We have evaluated tumor dimension (ultrassonography), proliferative index (Ki67 expression), 25(OH)D3 serum concentration and gene expression profile, before and after a short term calcitriol supplementation (dose to prevent osteoporosis) to post-menopausal patients. Results: Thirty three patients with operable disease had tumor samples evaluated. Most of them (87.5%) presented 25(OH)D3 insufficiency (<30 ng/mL). Median period of calcitriol supplementation was 30 days. Although tumor dimension did not vary, Ki67 immunoexpression decreased after supplementation. Transcriptional analysis of 15 matched pre/post-supplementation samples using U133 Plus 2.0 GeneChip (Affymetrix) revealed 18 genes over-expressed in post-supplementation tumors. As a technical validation procedure, expression of four genes was also determined by RT-qPCR and a direct correlation was observed between both methods (microarray vs PCR). To further explore the effects of near physiological concentrations of calcitriol on breast cancer samples, an ex vivo model of fresh tumor slices was utilized. Tumor samples from another 12 post-menopausal patients were sliced and treated in vitro with slightly high concentrations of calcitriol (0.5nM), that can be attained in vivo, for 24 hours In this model, expression of PBEF1, EGR1, ATF3, FOS and RGS1 was not induced after a short exposure to calcitriol. Conclusions: In our work, most post-menopausal breast cancer patients presented at least 25(OH)D3 insufficiency. In these patients, a short period of calcitriol supplementation may prevent tumor growth and reduce Ki67 expression, probably associated with discrete transcriptional changes. This observation deserves further investigation to better clarify calcitriol effects in tumor behavior under physiological conditions.

30

Urata YN, Lyra EC, Katayama ML, Basso RA, Assis PE, Cardoso AP, Roela RA, Nonogaki S, Sampaio Góes JC, Brentani MM, Folgueira MA