24R,25-Dihydroxyvitamin D₃ regulates breast cancer cells in vitro and in vivo

Background: Epidemiological studies indicate high serum 25(OH)D₃ is associated with increased survival in breast cancer patients. Pre-clinical studies attributed this to anti-tumorigenic properties of its metabolite 1α,25(OH)₂D₃. However, 1α,25(OH)₂D₃ is highly calcemic and thus has a narrow therapeutic window. Here we propose another metabolite, 24R,25(OH)₂D₃, as an alternative non-calcemic vitamin D₃ supplement. Methods: NOD-SCID-IL2γR null female mice with MCF7 breast cancer xenografts in the mammary fat pad were treated with 24R,25(OH)₂D₃ and changes in tumor burden and metastases were assessed. ERα66+ MCF7 and T47D cells, and ERα66- HCC38 cells were treated with 24R,25(OH)₂D₃ in vitro to assess effects on proliferation and apoptosis. Effects on migration and metastatic markers were assessed in MCF7. Results: 24R,25(OH)₂D₃ reduced MCF7 tumor growth and metastasis in vivo. In vitro results indicate that this was not due to an anti-proliferative effect; 24R,25(OH)₂D₃ stimulated DNA synthesis in MCF7 and T47D. In contrast, markers of invasion and metastasis were decreased. 24R,25(OH)₂D₃ caused dose-dependent increases in apoptosis in MCF7 and T47D, but not HCC38 cells. Inhibitors to palmitoylation, caveolae integrity, phospholipase-D, and estrogen receptors (ER) demonstrate that 24R,25(OH)₂D₃ acts on MCF7 cells through caveolae-associated, phospholipase D-dependent mechanisms via cross-talk with ERs. Conclusion: These results indicate that 24R,25(OH)₂D₃ shows promise in treatment of breast cancer by stimulating tumor apoptosis and reducing metastasis. General significance: 24R,25(OH)₂D₃ regulates breast cancer cell survival through ER-associated mechanisms similar to 24R,25(OH)₂D₃ effects on chondrocytes. Thus, 24R,25(OH)₂D₃ may modulate cell survival in other estrogen-responsive cell types, and its therapeutic potential should be investigated in ER-associated pathologies.