Screening Of Nuclear Factor-κB(NF-κB) Inhibitors And Studies On The Anti-tumor And Anti-osteoclastogenic Effects Of The Active Compound Maslinic Acid

AbstractNuclear Factor-KB (NF-κB), an well-known evolutionarily conserved transcriptional factor, which plays critical roles in many biological processes including inflammation, immunity, cell proliferation, survival and apoptosis through regulating related target genes expression. The activation of NF-κB, which is rigorously controlled in vivo, is essential for cellular physiological function. Once this activity is disturbed, many diseases, including cancer, arthrositis, periodontitis and bone degradaton are followed. Therefore, NF-κB has become a promising drug target for these diseases.The activation of NF-κB is a multiple steps progress, including IκBαphosphorylation and degradation, nuclear translocation of NF-κB subunits, binding with specific DNA sequences and finally regulating its target genes expression. According to these steps, we firstly established a systemic assays model to evaluate the activity of NF-κB. Secondly, we found some inhibitors of NF-κB signaling pathway by screening small molecular compounds, which come from traditional Chinese medicine(TCM) using these assay models. We then studied one of these active compounds, maslinic acid (MA), and discovered the functions of MA in cancer and osteoclastogenesis and also elucidated the exact mechanisms involved in these physiological functions.In this study, we firstly studied the antitumor effects of MA in pancreatic cancer, and we found the following results:1 MA could enhance TNFα-induced pancreatic cancer cell growth;2 MA could potentiates TNFα-induced pancreatic cancer cell apoptosis;3 MA could activate caspase-dependent apoptotic pathway;4 MA could inhibit TNFα-induced pancreatic cancer cell migration;5 MA blocked TNFα-induced NF-κB activation and genes expression;6 MA suppressed pancreatic tumor growth and induced apoptosis in vivo.Together, in this part of our study, for the first time, our results showed that MA can enhance the anti-tumor activities of TNFa and inhibit pancreatic tumor growth and invasion by activating caspase-dependent apoptotic pathway and by suppressing NF-κB activation and- its downstream genes expression. Therefore, we concluded that MA together with TNFa could be new promising agents in the treatment of pancreatic cancer.We next determined the effects of MA on osteoclast differentiation and functions. And we observed the following phenomena:1 MA suppressed osteoclast differentiation and these suppressions were only observed at early stage during ostoclastogenesis;2 MA had little effects on precursors of osteoclast when treated with the lower concentration used in differentiation assay;3 MA blocked RANKL-induced actin-ring formation;4 MA inhibited RANKL-induced bone resorption on spicula in pits assay;5 MA ameliorated ovariectomy-induced bone loss in vivo;We further studied the molecular mechanisms of these observations. And we found that:1 MA blocked NF-κB and MAPK-AP1 activation;2 MA suppressed NFAT-c1 expression but not its activation;3 MA down regulated some osteoclast related genes expression;4 Maslinic acid cannot break RANKL induced calcium signaling in RAW264.7 cells.Taken together, in this part of our study, our results firstly demonstrated that MA could suppress RANKL-induced osteoclastogenesis and function through suppression NF-κB and MAPKs/AP-1 signaling pathway. These results suggested that MA could be a new promising agent in the treatment of osteoclast related diseases such as osteoporosis.Taken all together, we discovered some novel inhibitors of NF-κB signaling pathway using a systemic study platform for NF-κB signaling pathway we established in our lab. Furthermore, we also studied the effects of maslinic acid (MA), one of our discovered active compounds from TCM, on its antitumor and antiosteoclastogenic activities in vitro and in vivo. We found that MA could potentiate TNFα-induced pancreatic cancer cell apoptosis and suppress RANKL-induced osteoclastogenesis by regulating NF-κB signaling pathway. These results supported that MA could be a new promising agent both in the treatment of cancer and osteoclast related diseases such as osteoporosis.