Computational multi-target approach for Anti-osteoclastic Assay of natural substances

Abstract

Osteoporosis is the most common age-related condition characterized by bone mineral density and change in the microarchitecture of bone, consequently increasing bone fragility. In Thailand, osteoporosis is also considered a serious public health concern resulting from longer life expectancy. Osteoporosis is a bone disease caused by an imbalance of bone formation and bone resorption. Especially, the rate of bone resorption resulting from the overactive osteoclasts is greater than that of bone formation. Additionally, the resorption activity of osteoclasts requires several weeks, while the formation activity of osteoblasts requires months to construct a new bone. Therefore, osteoclastic activity inhibition is interestingly targeted for osteoporosis treatment. Not only the medications are used to treat the osteoporotic condition, but medicinal plants are also used as osteoporotic prevention. The phytochemical compounds possess the osteoprotective property, including flavones, flavonols, flavanones, isoflavones, phenylpropanols, lignans, monoterpenoid, diterpenoids, sesquiterpenoids, triterpenoids, and miscellaneous compounds. Hence, the herbal substances can be used to develop anti osteoporotic agents toward various targets in the osteoclast, namely αVβ3 integrin receptor, cathepsin K, and V-ATPase. In this study, the target-based virtual screening and the clustering approaches were carried out to identify the multi-target herbal substances toward these targets on osteoclasts. A result of target-based virtual screening indicated that four substances with strong binding affinity against both enzymes would be used in the molecular dynamics simulation, including rutin, sagittatosideA, icariin, and kaempferitrin. Further, the MD simulations exhibited that rutin demonstrated the highest binding affinity toward cathepsin K, followed by sagittatoside A, icariin, and kaempferitrin. Moreover, MD simulations of V-ATPase exhibited that icariin obtained a strong binding affinity toward V-ATPase, followed by sagittatoside A, kaempferitrin, and rutin, respectively. Additionally, the anti-cathepsin K activities of candidate compounds were assessed using zebrafish scales by a decrease in the fluorescence intensity. The in vitro result suggested that hesperidin showed slight inhibitory activity on cathepsin K at the concentration of 1, 10, and 100 µM. Interestingly, rutin showed the most potent inhibitory activity among the compounds at the concentration of 100 µM, followed by hesperidin and quercetin, respectively. To be concluded, rutin, sagittatosideA, icariin, and kaempferitrin, exhibited the multi-target activity on osteoclast by using the virtual screening MD simulations, and the anti-cathepsin K activity assay.