The Mechanism Of Prostate Carcinogenesis In P53&EAF2Knock Out Mouse

Prostate cancer is still one of the major causes of death in the United States andWestern Europe. With the improvement of people’s health awareness and medicalscreening, the incidence of prostate cancer increased year by year in our country. Andit has become one of the major threats to the men’s health in our country. There aremany clinical features of prostate cancers, such as insidious onset and slow course ofdisease. At present, the main treatment is the radical prostatectomy. Furthermore,hormonal therapy, chemotherapy and targeted therapy are alterative treatments forprostate cancer patients. Although patients with early stage prostate cancer have abetter efficacy of treatment, but patients with advanced disease or recurrent diseasehave a poor prognosis. When the disease developed to the hormone-resistant prostatecancer, either chemotherapy or targeted therapy can not significantly prolong theoverall survival of patients or improve the quality of life. The carcinogenicmechanisms of prostate cancer are still not fully understood, which is one of thereasons for this predicament.The onset and progression of prostate cancer are often accompanied by loss of aplurality of tumor suppressor genes. And these genes can regulate differentiation andcarcinogenesis of the prostate epithelial cells by modulating other important signalingpathway or regulating other gene expression. Previous studies have found thatandrogen responsive genes EAF2often missing in the high-grade prostate cancer cells,and one of its functions is to inhibit cell proliferation and induce apoptosis. Knockoutof EAF2can induce prostate PIN lesions, but not sufficient to induce the formation ofcancer. p53is also an important tumor suppressor gene, and it is frequent absence inthe high-level or metastatic prostate cancer cells. But p53knockout mice did not have any abnormal changes in the prostate. Our previous studies have found EAF2caninhibit angiogenesis in mouse prostate through regulation of p53downstream geneTSP-1. Therefore, we have a hypothesis that these two tumor suppressor genes bothplay an important role in the development of prostate and may have some synergisticfunction in this process.Objective：1. To investigate the relationship between p53and EAF2in prostate cancer;2. To investigate the function of p53and EAF2in the development of prostatecancer.Methods：1. In the prostate cancer cell line C4-2, concurrent silence the expression of p53and EAF2, then check the proliferation and migration ability of cancer cells.2.Detect the p53and EAF2binding domain at protein level using co-immunoprecipitation.3. Analyze gene changes in p53and EAF2double knockout cells using genesequencing technology, and further investigate those important genes or signalingpathways participating in this tumor model.4. Breed p53and EAF2double knockout mice using existing transgenic hybridsp53knockout mice and EAF2knockout mice, and observe the changes in theirprostate;Results：1.The effect of double knockout p53and EAF2on cell migration andproliferation in C4-2cellsDouble knockout of p53and EAF2significantly enhanced the proliferation andmigration ability of prostate cancer cells. And these results were consistent with theobservations in animal models. 2. The binding domain of p53and EAF2at protein levelIt was showed that p53and EAF2can bind to each other at protein level afterCo-IP assay. It was found that p53DNA binding domain can bind to EAF2, and EAF2protein sequence161-260(C ’segment fragment) can bind to p53.3. Gene sequencing analysis of double knockout p53and EAF2in prostatecancer cells. And further investigation into the involved pathway in this context.After analysis of RNA-seq results, we found interferon-inducible genes(Interferon Stimulated Genes, ISGs) were significantly upregulated. And it is reportedthat STAT3is an important member of interferon-mediated signal transductionpathway, which the main function is the regulation of cell proliferation and inhibitionof apoptosis. Moreover, it is also reported that p53can inhibit the activation of STAT3signaling pathway in prostate cancer. Therefore, we further examined the status ofSTAT3signaling pathway in p53and EAF2double knockout model, and found thatthe pathway was significantly activation in this model.4. Establishment and observation of mouse models:In this study, we obtained the p53and EAF2double knockout mice, and foundthat prostate cancer occurs in the young (4-6months) mice prostate. However, nothingwas found in the prostate of single knockout p53mice. But in the single knockoutEAF2mice, we found PIN lesions in their prostate. These results were consistent withour previous hypothesis that both genes may play an important role in thedevelopment of prostate cancer.Conclusions：1. p53and EAF2play an important role in the development of prostate cancer.Double knockout these two genes can induce the formation of prostate cancer.2. p53and EAF2can bind to each other at protein level. It is found that the p53DNA binding domain can bind to EAF2, and the EAF2protein sequence161-260(C ’segment fragment) can bind to p53.3. STAT3signaling pathway is activated in p53and EAF2double knockoutmodel. And it may be one of the mechanisms for the development of prostate cancerin this context.