The Anti-Gastric Cancer Pharmacodynamic And Mechanism Study Of Emetine And The Anti-Gastric Cancer Study Of Targeting ACK1

Part I The anti-gastric cancer pharmacodynamic and mechanism study of emetineBackground:Gastric cancer（GC）is a malignant tumor with high morbidity and mortality.Despite great progress for GC clinical therapy,many sufferers cannot benefit from the existing anti-GC treatments.Therefore,there is still a great clinical need to develop novel anti-GC medicines.Emetine,a natural small-molecule drug isolated from Psychotria ipecacuanha,is used for the clinical treatment of malaria and emetic and also reported to be active in tumor treatment.Here,we conducted a comprehensive study on the anti-GC effects of emetine and the related mechanisms of action.Methods:The anti-GC cell viability of emetine was evaluated by MTT and colony formation assay;the cellular proliferation and apoptosis were analyzed by Edu assay and Annexin V-PI staining,respectively;wound healing assay and transwell invasion assay were conducted to detect the inhibitory activity of emetine against GC cell migration and invasion;the molecular mechanism involved in the anti-GC effects of emetine was explored by the RNA sequencing and functional enrichment analysis as well as western blot assay;moreover,the in vivo anti-tumor efficacy of emetine was evaluated in the MGC803 xenograft model.Results:MTT and colony formation assay showed that emetine could effectively inhibit GC cell growth,with IC₅₀ values of 0.0497μM and 0.0244μM on MGC803 and HGC-27 cells,respectively.Further pharmacodynamic studies revealed that emetine restrained the GC cell growth mainly by inhibiting cell proliferation and inducing apoptosis,with no significant effect on the cell cycle.Moreover,emetine also had the ability to block GC cell migration and invasion.The transcriptome sequencing and western blot analysis showed that emetine exerted anti-GC effects mainly through regulating MAPKs and inhibiting Wnt/β-catenin,PI3K/AKT and Hippo/YAP signaling cascades.The in vivo pharmacodynamic studies revealed that emetine could also effectively inhibit the growth of MGC803 xenografts in vivo,with the inhibition rate of57.52%.Meanwhile,emetine was well tolerated and showed no obvious toxic and side effects at the administered dose.Conclusion:Our data demonstrated that emetine was a multi-target drug that could potently inhibit GC cell growth in vitro and in vivo.It is a promising lead compound and even a drug candidate for GC therapy,deserving further structural optimization and development.Part II The anti-gastric cancer study of targeting ACK1Background:ACK1,a non-receptor tyrosine kinase,is considered as an oncoprotein,which is closely related to tumorigenesis and progression.However,its role in GC progression was poorly reported.Here,we conducted a preliminary study on the anti-GC bioactivity of targeting ACK1 to evaluate its feasibility as a target for GC targeted therapy.Methods:We conducted bioinformatics analysis using public database to explore ACK1 expression in GC and adjacent normal tissues and its correlation with the prognosis of GC patients.Then sh RNA was constructed and transfected into HGC-27 cells to knock down ACK1 expression,and western blot assay was used for verification.MTT and colony formation assay were conducted to evaluate the influence of ACK1 knockdown on GC cell viability;Edu incorporation assay and flow cytometry were used to detect the effects of ACK1 knockdown on GC cell proliferation,apoptosis and cell cycle arrest;moreover,wound healing assay and transwell invasion assay were performed to evaluate the effects of ACK1 knockdown on the migration and invasion of GC cells.Results:The results of bioinformatics analysis showed that ACK1 expression in GC tissues was significantly higher than that in adjacent normal tissues,and the high-expression of ACK1 was closely associated with the poor prognosis of GC patients.Additionally,sh-ACK1-RNAs were successfully constructed and transfected,and sh-ACK1-RNA2 had the highest transfection and knockdown efficiency,which could significantly decrease the expression of ACK1 in HGC-27 cells.Knockdown of ACK1significantly inhibited cell viability,colony formation and cell proliferation and induced apoptosis,but had little influence on the cell cycle.Meanwhile,ACK1 knockdown could also effectively inhibit the migration and invasion of GC cells.Conclusion:Our data indicated that the high-expression of ACK1 was significantly associated with poor prognosis of GC patients.ACK1 had important biological functions in GC cell proliferation,apoptosis,migration and invasion,and was a potential therapeutic target against GC.