The Tumor Suppressor Gene RB1 Prevents The Progression Of Prostate Cancer By Regulating The DNA Damage Response

Purpose: prostate cancer is a common malignant tumor in elderly male,and castration resistance is the most important clinical feature in the advanced stage.Whole-genome sequencing confirm that tumor suppressor genes RB1 and TP53 are altered frequently in metastatic castrationresistant prostate cancer(m CRPC)and promote resistance to multiple cancer therapeutics.Therefore,to reveal the respective role of RB1 and TP53 in the progression of prostate cancer may provide a new strategy for treatment in castration-resistant prostate cancer(CRPC).Methods: lentiviral interference vector(sh RNA)were used to target silence RB1 gene in prostate cancer cell lines(PC).CCK8,EDU,β-galactosidase staining and flow cytometry assay were performed to detect the effect of RB1 deletion on PC.The relationship between RB1 and TP53 were verified by lentiviral double knockdown experiments,protein stability experiments,phosphorylation specific inhibitors and immunofluorescence experiments.The sensitivity of androgen receptor antagonist Enzalutamide was measured in PC with RB1 and TP53 deletion,and q RT-PCR,Western Blot and immunofluorescence experiments were performed to analyze the possible mechanism of promoting castration-resistance.RNA sequencing was used to determine the difference in gene expression of prostate cancer cells after RB1 and TP53 deletion,where the molecular mechanism of promoting cell growth were identified through enrichment analysis and target gene intervention.Results: in vitro experiments demonstrated that acute loss of RB1 increased the accumulation of DNA damage in PC and promoted cell senescence by a TP53-dependent pathway.Furthermore,the stabilization of TP53 was regulated by ATM-mediated phosphorylation of MDM2 at Ser395 after the acute deletion of RB1.However,the double deletion of RB1 and TP53 reversed cell senescence induced by DNA damage,and promoted the proliferation of PC and reduced the sensitivity of cells to Enzalutamide.Simultaneously,the expression of androgen receptor was decreased,but neuroendocrine marker SYP and NSE genes were up-regulated.In addition,RB1/TP53 deletion leads to the up-regulation of numerous genes expression in PC,in which those are involved in hypoxia response,cell stemness and epigenetic regulation,DNA damage response.Importantly,PC with RB1/TP53 loss depend on the DNA repair gene PARP1 to promote cell proliferation,and the PARP1 inhibitor Olaparib significantly inhibited the growth of cells.Conclusion: Tumor suppressor genes RB1 and TP53 maintain the stability of prostate cancer genome by regulating the DNA damage response;RB1 combined with TP53 deletion increases the vulnerability of prostate cancer cells to inhibitor of the DNA repair gene PARP1.