Molecular Mechanism Of Evodiamine Inducing Apoptosis In Human Renal Cancer Cells

Renal cell carcinoma(RCC)is one of the most serious cancers in adults,accounting for～90%of all renal malignancies.The annual incidence of RCC is continually increasing.Because of its highly invasive and metastatic characteristics,the initial times of diagnoses are often delayed.For example,nearly 30%of patients have distant metastases and 25%have locally advanced disease when they are initially diagnosed.The survival rate of patients with metastatic stage RCC can be as low as 9.5%.Because RCC is insensitive to radiotherapy and chemotherapy,therapy with targeting agents(e.g.,interleukin-2,Temsirolimus,or Pazopanib)has proven to be superior compared to other treatments.However,not all of these treatments can produce better therapeutic effects or achieve complete responses.Therefore,an urgent need exists to develop novel therapeutics that overcome resistance to the currently administered anticancer drugs for RCC.Evodiamine(EVO)is an indoloquinazoline alkaloid isolated from fruit of Evodia rutaecarpa.For millennia,it has been considered an effective Chinese medicine that can be used for the treatment of gastropathy,hypertension,and eczema.In the previous decade,increasing evidence has revealed that EVO can exhibit various biological effects,including antibacterial,antiobesity,and antinociceptive activities.Recently,it has been established that EVO exhibits antitumor activity against gastric adenocarcinoma,colon(lovo cell line),colorectal carcinoma,and breast cancer cells.A previous study demonstrated that EVO can inhibit the proliferation,invasion and metastasis of cancer cells;however,the mechanisms of action against cancer cells remains to be elucidated and no studies on the function of EVO in human RCC in vitro have yet been reported.In this study,cytological experiments and transcriptome profiling study were performed to reveal the anticancer mechanism of EVO.First,a series of experiments on cell viability and proliferation of human renal carcinoma 786-0 cells and Caki-1 cells and the human renal epithelial cell line HK-2 were carried out under EVO treatment.Then,a transcriptome profiling study was used to analyse EVO-regulated genes and signalling pathways responsible for growth and apoptosis using RNA sequencing.In addition,we verified the molecular mechanisms whereby EVO inhibits growth and induces apoptosis using ultrastructural observations,flow cytometry analysis,and qPCR.We found that EVO could signally change the morphology and inhibit the viability and proliferation of cells in a time-and dose-dependent manner in vitro.In addition,transcriptome analysis indicated that EVO can modulate the transcriptome of the Caki-1 cell line.Totally,7,243 differentially expressed genes were founded,among which 3,347 downregulated genes and 3,896 upregulated genes were mainly refered to cell migration,apoptosis,cell cycle and DNA replication.Furthermore,we demonstrated that EVO can cuase apoptosis,arrest cells at G2/M period and regulated the gene expression of apoptosis-and cell cycle-related genes in the Caki-1 cell line.Our study perceived the anticancer effects of EVO with cellular and molecular data,and indicated potential uses of this compound as a resource to portraye the antitumor mechanisms of E.rutaecarpa.