Rational design of vitamin D3 analogues which selectively restore activity to a vitamin D receptor mutant associated with rickets.

[formula: see text] Vitamin D3-resistant rickets (VDRR) is associated with mutations to the Vitamin D receptor (VDR) which effect ligand-dependent transactivation. Some VDRR associated mutants directly disrupt ligand binding. Using the reported VDR-1,25-dihydroxy vitamin D3 (1,25(OH)2D3) cocrystal structure, three 1,25(OH)2D3 analogues were designed to uniquely complement the rickets associated mutant VDR(Arg274-->Leu). The three analogues were 17 to 286 times more potent than 1,25(OH)2D3 with the mutant in cell-based assays and did not substantially activate cellular calcium influx.

Structure-based design of selective agonists for a rickets-associated mutant of the vitamin d receptor.

The nuclear and steroid hormone receptors function as ligand-dependent transcriptional regulators of diverse sets of genes associated with development and homeostasis. Mutations to the vitamin D receptor (VDR), a member of the nuclear and steroid hormone receptor family, have been linked to human vitamin D-resistant rickets (hVDRR) and result in high serum 1,25(OH)(2)D(3) concentrations and severe bone underdevelopment. Several hVDRR-associated mutants have been localized to the ligand binding domain of VDR and cause a reduction in or loss of ligand binding and ligand-dependent transactivation function. The missense mutation Arg274 --> Leu causes a >1000-fold reduction in 1,25(OH)(2)D(3) responsiveness and is, therefore, no longer regulated by physiological concentrations of the hormone. In this study, computer-aided molecular design was used to generate a focused library of nonsteroidal analogues of the VDR agonist LG190155 that were uniquely designed to complement the Arg274 --> Leu associated with hVDRR. Half of the designed analogues exhibit substantial activity in the hVDRR-associated mutant, whereas none of the structurally similar control compounds exhibited significant activity. The seven most active designed analogues were more than 16 to 526 times more potent than 1,25(OH)(2)D(3) in the mutant receptor (EC(50) = 3.3-121 nM). Significantly, the analogues are selective for the nuclear VDR and did not stimulate cellular calcium influx, which is associated with activation of the membrane-associated vitamin D receptor.