Discovery of Small-Molecule Inhibitors of Receptor Activator of Nuclear Factor-κB Ligand with a Superior Therapeutic Index.

Receptor activator of nuclear factor-κB ligand (RANKL) constitutes the master mediator of osteoclastogenesis, while its pharmaceutical inhibition by a monoclonal antibody has been approved for the treatment of postmenopausal osteoporosis. To date, the pursuit of pharmacologically more favorable approaches using low-molecular-weight inhibitors has been hampered by low specificity and high toxicity issues. This study aimed to discover small-molecule inhibitors targeting RANKL trimer formation. Through a systematic screening of 39 analogues of SPD-304, a dual inhibitor of tumor necrosis factor (TNF) and RANKL trimerization, we identified four compounds ( 1b , 3b , 4a , and 4c ) that selectively inhibited RANKL-induced osteoclastogenesis in a dose-dependent manner, without affecting TNF activity or osteoblast differentiation. Based on structure-activity observations extracted from the most potent and less toxic inhibitors of RANKL-induced osteoclastogenesis, we synthesized a focused set of compounds that revealed three potent inhibitors ( 19a , 19b, and 20a ) with remarkably low cell-toxicity and improved therapeutic indexes as shown by the LC ₅₀ to IC ₅₀ ratio. These RANKL-selective inhibitors are an excellent starting point for the development of small-molecule therapeutics against osteolytic diseases.