Novel selective proline-based peptidomimetics for human cathepsin K inhibition.

Human cathepsin K (CatK) stands out as a promising target for the treatment of osteoporosis, considering its role in degrading the bone matrix. Given the small and shallow S2 subsite of CatK and considering its preference for proline or hydroxyproline, we now propose the rigidification of the leucine fragment found at the P2 position in a dipeptidyl-based inhibitor, generating rigid proline-based analogs. Accordingly, with these new proline-based peptidomimetics inhibitors, we selectively inhibited CatK against other human cathepsins (B, L and S). Among these new ligands, the most active one exhibited a high affinity (pKi = 7.3 - 50.1 nM) for CatK and no inhibition over the other cathepsins. This specific inhibitor harbors two novel substituents never employed in other CatK inhibitors: the trifluoromethylpyrazole and the 4-methylproline at P3 and P2 positions. These results broaden and advance the path toward new potent and selective inhibitors for CatK.

Identification of mouse cathepsin K structural elements that regulate the potency of odanacatib.

Cathepsin K (CatK) is the predominant mammalian bone-degrading protease and thus an ideal target for antiosteoporotic drug development. Rodent models of osteoporosis are preferred due to their close reflection of the human disease and their ease of handling, genetic manipulation and economic affordability. However, large differences in the potency of CatK inhibitors for the mouse/rat vs. the human protease orthologs have made it impossible to use rodent models. This is even more of a problem considering that the most advanced CatK inhibitors, including odanacatib (ODN) and balicatib, failed in human clinical trials due to side effects and rodent models are not available to investigate the mechanism of these failures. Here, we elucidated the structural elements of the potency differences between mouse and human CatK (hCatK) using ODN. We determined and compared the structures of inhibitor-free mouse CatK (mCatK), hCatK and ODN bound to hCatK. Two structural differences were identified and investigated by mutational analysis. Humanizing subsite 2 in mCatK led to a 5-fold improvement of ODN binding, whereas the replacement of Tyr61 in mCatK with Asp resulted in an hCatK with comparable ODN potency. Combining both sites further improved the inhibition of the mCatK variant. Similar results were obtained for balicatib. These findings will allow the generation of transgenic CatK mice that will facilitate the evaluation of CatK inhibitor adverse effects and to explore routes to avoid them.