Cathepsin K antisense oligodeoxynucleotide inhibits osteoclastic bone resorption

Cathepsin K is a recently identified cysteine protease which is abundantly and selectively expressed in osteoclasts, To evaluate the contribution of cathepsin K to bone resorption processes, we investigated the effect of cathepsin K antisense phosphothiorate oligodeoxynucleotide (S-ODN) on the bone-resorbing activity of osteoclasts. Rabbit osteoclasts were cultured on dentine slices for 24 h in the presence or absence of antisense S-ODN in a medium containing 100 nM Tfx(TM)-50, polycationic liposome, as a carrier of the S-ODN, Uptake of the S-ODN by osteoclasts was confirmed microscopically using fluorescein-labeled S-ODN, The treatment with antisense significantly decreased the amount of cathepsin K protein in osteoclasts. The antisense inhibited the osteoclastic pit formation in a concentration-dependent fashion, At 10 mu M the antisense reduced the total pit number and area and average pit depth by 46, 52, and 30%, respectively, The sense and mismatch S-ODNs, which were used as negative controls, had no effect on either the cathepsin K protein level or the pit formation. A nonspecific cysteine protease inhibitor, E-64, also reduced pit formation in a concentration-dependent manner with maximum reductions at 1 mu M of 46, 48, and 35% in the above pit parameters, The inhibitory effect of the antisense almost equal to that of E-64 demonstrates that cathepsin K is a cysteine protease playing a crucial role in osteoclastic bone resorption.