Probing the structural requirements for vitamin D3 inhibition of the hedgehog signaling pathway

A structure–activity relationship study to elucidate the structural basis for hedgehog (Hh) signaling inhibition by vitamin D3 (VD3) was performed. Functional and non-functional regions of VD3 and VD2 were obtained through straightforward synthetic means and their biological activity was determined in a variety of cell-based assays. Several of these compounds inhibited Hh signaling at levels comparable to the parent VD3 with no effects on canonical vitamin D signaling. Most notably, compounds 5 and 9, demonstrated potent inhibition of the Hh pathway, exhibited no binding affinity for the vitamin D receptor (VDR), and did not activate VDR in cell culture. In addition, several compounds exhibited anti-proliferative activity against two human cancer cell lines through a mechanism distinct from the Hh or VDR pathways, suggesting a new cellular mechanism of action for this class of compounds.