Discovery of novel cathepsin inhibitors with potent anti-metastatic effects in breast cancer cells.

It is still challenging to determine the potential targets of natural products, which is essential for further drug research and development. Due to its novel mechanism of action of inducing autophagy effects in breast cancer cells, asperphenamate has received our considerable attention. However, its unknown target inevitably impedes further study. In our previous work, the target enzyme of asperphenamate was predicted as cathepsin by the natural product consensus pharmacophore strategy. Then, asperphenamate and its three derivatives were chosen to study in detail by molecular docking calculations with AutoDock 4 suite. The docking results showed the three derivatives interacted more tightly with either cathepsin L or cathepsin S than with asperphenamate. The ortho-benzyloxyl phenylacetyl derivative 1 andp-toluenesulfonyl derivative 3 showed similar interactions with cathepsin L and adopted a better geometric shape within the binding pocket than did the N-CBZ-piperidyl analog 2. On the other hand, compound 2 formed more hydrogen bonds than 1 and 3 to make it tightly bind within cathepsin S. The cathepsin inhibitory activity assay verified the molecular simulation results. Compound 2 was remarkably less active than 1 and 3 against cathepsin L. However, compound 2 showed the strongest potency against cathepsin S with IC50 of 13.12 ± 0.29 μM. Considering that cathepsin S plays a vital role in the process of metastasis in breast cancer cells, the inhibitory effect of 2 on migration and invasion was further studied in human breast cancer MDA-MB-231 cells by wound healing and transwell chamber assays. The results illustrated that 2 exhibited an apparent inhibitory ability to the metastasis of MDA-MB-231 cells. Next, 2 will be chosen as a lead compound to develop novel double functional chemotherapeutic agents with both novel mechanisms of action against apoptosis-resistant cancer cells, such as inducing autophagy and inhibiting cancer metastasis.