ABSTRACT   The potent cathepsin K (CatK) inhibitor, Tanshinone IIA sulfonic sodium (T06), was tested for its in vitro and in vivo antiresorptive activities. T06 binds in an ectosteric site of CatK remote from its active site and selectively inhibits collagen degradation with an IC50 value of 2.7 ± 0.2 μM (CatK:T06 molar ratio of 1:5). However, it does not suppress fluorogenic peptide cleavage and gelatinolysis at a 2500-fold molar excess. Contrary to active site-directed CatK inhibitors, such as odanacatib, T06 suppresses bone resorption in both human and mouse osteoclasts equally well (IC50 value for human and mouse osteoclasts: 237 ± 60 nM and 245 ± 55 nM, respectively) and its antiresorptive activity is fully reversible in both cell types. Moreover, T06 affects neither the metabolic activity of osteoclasts nor osteoclastogenesis. In in vivo studies, 40 mg T06/kg/d given to 12-week-old ovariectomized (OVX) mice for 3 months reduced plasma CTx-1 by 20% and increased osteoblast numbers and plasma P1NP by ∼28% when compared with the OVX control. μCT analysis of T06-treated OVX mice showed a 35% increase in bone mineral density and other femoral trabecular bone parameters when compared with OVX animals. T06 did not alter the number of osteoclasts, had no estrogenic effect on the uterus, did not change plasma estradiol levels, and did not inhibit fibroblast-mediated TGF-ß1 processing or degradation and cognitive functions in OVX mice. This study indicates that the ectosteric inhibitor, T06, is a selective antiresorptive CatK inhibitor that may overcome the shortcomings of side effect–prone active site-directed drugs, which all failed in clinical trials. © 2017 American Society for Bone and Mineral Research.