Antitumoral effects and calcemic activity of the novel analogue of calcitriol ML-344

Abstract

The active form of vitamin D3 , calcitriol, has been shown to display antitumoral effects on various types of cancer. However, hypercal- cemia is observed when effective antitumoral doses of calcitriol are employed, thus precluding its therapeutic application. In collabora- tion with the laboratory of Organic Chemistry of the University of Vigo we synthesized an analogue of calcitriol, called ML-344, in order to obtain a compound that retain or even increase the anti- tumoral activity but prevent the side effects. In this work we aimed to evaluate the biological effects of ML-344 by employing in vitro, in vivo and in silico assays. We demonstrated that the analogue exerts a significant decrease in the viability of different cancer cell lines (p< 0.001): head and neck squamous cell carcinoma (HN12), glioblasto- ma multiforme (GL26 and U251) and breast adenocarcinoma (4T1 and LM3) cells. Importantly, the cellular viability of human primary astrocytes and murine non-malignant mammary epithelial HC11 cell line is not affected after ML-344 treatment. Also, the analogue re- tards cell migration of 4T1 and LM3 cells (16 h: p< 0.001; 21 h: p< 0.01) while it does not affect HC11 cell motility. In concordance with the antimigratory effects, ML-344 decreases LM3 invasive capacity (p< 0.01) and induces a rearrangement of actin cytoskeleton by a reduction in the amount of cells with stress fibers (p< 0.001). In vivo studies showed that the analogue, in contrast to calcitriol, does not cause hypercalcemic effects in CF1 mice administrated daily at 5 μg/Kg of body weight during 96 h. In addition, hematocrit remained within the normal levels and no changes in body weight were found. Finally, computational studies showed that ML-344 is able to bind to VDR with greater affinity than calcitriol (∆G= -82.5 and -73.5 Kcal/ mol, respectively). Altogether, these results suggest the potential use of this analogue as an antitumor drug with a differential effect between tumor and non-malignant cells.