The Anti-melanoma Function And Related Mechanism Of Chinese Propolis And Its Flavonoid Monomers

Melanoma is a malignant skin disease that develops very rapidly.Due to its high metastatic characteristics and high recurrence rate,melanoma has the characteristics of rapid development,high malignancy,poor prognosis,and high mortality.The five-year survival rate of patients with early melanoma is as high as 92%,but once melanoma begins to metastasize,the 5-year survival rate of patients quickly drops to 15%.Although there are certain related drugs and treatments for melanoma,their efficacy are not significant,and are associated with serious side effects as well as drug resistance.Therefore,research of new melanoma-targeted therapeutic drugs and methods is still a focus and hot spot in cancer research.Propolis is a resinous product collected by the honey bee from plants and possesses a broad spectrum of biological activities,including anti-oxidant,anti-inflammatory,anti-tumor effect and so on.Previous studies have demonstrated that Chinese propolis could induce cell apoptosis and cell cycle arrest in different tumor models,such as breast cancer and colon cancer,demonstrating a possibility that propolis to be developed as an alternative treatment for cancers.In this study,we evaluated the anti-melanoma effect of Chinese propolis(CP)and its flavonoids(Pinocembrin and chrysin)in vivo and in vitro.We also uncovered that underlying molecular mechanisms of CP and its flavonoids.We aimed to reveal the anti-melanoma capability of CP,provide new insights for the application of propolis and the treatment of melanoma.The main research findings are summarized as follows:1 Chinese propolis exerts anti-proliferation effects in human melanoma cellsIn this study,we confirmed the pro-apoptosis activity and anti-metastasis activity of CP on human melanoma cell line A375,and uncovered its molecular mechanism.After CP treatment,A375 cells underwent intrinsic apoptosis and cell cycle arrest.Furthermore,we found that CP suppressed inflammation in A375 cells.NLRP1(NLR family pyrin domain containing 1),confirmed as a proinflammatory protein in melanoma progression,was downregulated significantly by CP,as were the NLRP1-related CARD proteins,including caspase-1 and caspase-4.Additionally,decreasing mRNA levels of IL-1?,IL-1?,and IL-18 further proved the negative regulation of CP on the melanoma inflammatory environment.We also discovered that CP induced autophagy in A375 cells and inhibiting autophagy in CP-treated cells diminished its antitumor effect,suggesting that the autophagy was attributed to CP-induced apoptosis.In addition,our results demonstrated that CP can inhibit the metastasis of melanoma.2 Characterization and screening of the major anti-melanoma components of CPThe components of Propolis vary based on its botanical resources.Chinese propolis is mainly collected from populus spp.plants.In this study,we identified the major chemical components of CP that was used in our study by using LC-MS.3-O-Acetyl pinobanksin,chrysin,pinocembrin are the 3 most abundant components in CP.And among all the components we identified in CP,pinocembrin,CAPE,Chrysin,Pinobanksin,Apigenin and 3,4-dimethoxycinnamic acid showed cytotoxic effect on melanoma cell viability.3 Pinocembrin induces ER stress mediated apoptosis and suppresses autophagy in melanoma cellsPinocembrin is a natural flavanone with versatile biological and pharmacological activities.Here,we evaluated the anti-tumor effects of pinocembrin against melanoma in vitro and in vivo.Pinocembrin inhibited the proliferation of melanoma cells(B16F10 and A375)in a dose-dependent manner.It induced endoplasmic reticulum stress via IRE 1?/Xbp1 pathway and triggered caspase-12/-4 mediated apoptosis in both cell lines.Furthermore,we discovered that pinocembrin suppressed autophagy through the activation of PI3K/Akt/mTOR pathway,which serves as a dual mechanism to enhance the pro-death effect of pinocembrin.In vivo,pinocembrin inhibited the growth of B16F10 in mice by inducing apoptosis.4 Chrysin inhibits melanoma tumor metastasis via interfering the FOXM1/?-catenin signalingChrysin,which is rich in various plants,has shown great inhibitory effect on melanoma proliferation.Here,we evaluated the metastasis suppressive effect of chrysin on melanoma in vivo and in vitro.In vitro,chrysin effectively inhibited ankios resistance,cell migration,invasion and tube formation capacity of melanoma cells from 5?M.We discovered that chrysin interfering mesenchymal-epithelial transition via regulating FOXM1/?-catenin signaling,as over-expression of FOXM1 will attenuated the anti-metastasis effect of chrysin.We also tested the effect of chrysin on pulmonary metastasis model,and we found fewer tumors in the lungs of chrysin treated mice.In addition,the expression of FOXM1 was also downregulated by chrysin in vivo.Taken together,we evaluated the anti-melanoma effect of CP and its flavonoids in vivo and in vitro.We found that CP and its flavonoids could induce apoptosis,cell cycle arrest,metastasis and interfering tumor microenvironment via different mechanism.We believe that this compelling evidence expands our understanding about the mechanism of the anti-melanoma ability of CP and its flavonoids,providing a new approach for CP to be applied in melanoma therapy,and provide new insights for melanoma drug development.