The Study Of Monoamine Oxidase A(MAOA) Driving Neuroendocrine Differentiation In Prostate Cancer

Objective:We explored to establish cell models and animal models of neuroendo-crine prostate cancer（NEPC）by inducing with androgen receptor inhibitor enzalutamide（Enz）,assessed gene changes and analyzed the role of monoamine oxidase A（MAOA）dur-ing the neuroendocrine differentiation.After the expression of MAOA were regulated by lentivirus and the small molecule inhibitor clorgyline（CLG）respectively,we further ob-serve the expression of neuroendocrine markers and explore its possible mechanism,in order to provide a theoretical basis for targeting MAOA and its related molecules to drive the progression of NEPC.Methods:The sensitivity of multiple androgen receptor（AR）positive expression PCa cell lines（22RV1,C4-2,LNCa P）to AR inhibitor Enz was detected by cell viability assay,and C4-2 was confirmed is the most sensitive among all cell lines.We culture C4-2cell in RPMI-1640 medium containing 10-20μM Enz for 6 months with gradient increas-ing and obtain the C4-2^ENZR drug-resistant cell model.The neuroendocrine differentiation features of C4-2^ENZR cells were characterized by the analysis of morphology,real-time quantitative PCR and western blotting（WB）.We make sure that C4-2^ENZR represent NEPC model according down-regulation of adenocarcinoma markers and the up-regulation of neuroendocrine markers.MAOA was identified as a differentially expressed target gene after Enz resistance by bioinformatics analysis,and it was confirmed by the experiment of WB,MAO-Glo enzyme activity and real-time quantitative PCR.Moreover,we used lenti-viral transfection technology to overexpress MAOA in 22RV1 cell line and m TOR and HIF-1αwere detected to evaluate the effect and to explore mechanism.Finally,we observe the therapeutic effect of MAOA-specific inhibitor CLG alone or in combination with Enz on prostate cancer in vivo and in vitro,and expect to provide a reference for the develop-ment of new PCa treatment strategies in the clinic.Results:We successfully established Enz-resistant NEPC model C4-2^ENZR.Com-pared with the parental cell C4-2,C4-2^ENZR was able to proliferated at higher concentra-tions of enzalutamide,showed Neuron-like Changes and displayed higher expression of neuroendocrine marker including SYP,NCAM1,EN02 and MAOA（P<0.05）.The expres-sion of MAOA gene in 22RV1 cell was increased using lentivirus transfection,named22RV1-MAOA cells.This cell line displayed more strong proliferation ability and more positive neuroendocrine marker expression than 22RV1 cells.Mechanistic studies revealed that MAOA was involved in mediating the neuroendocrine process by regulating the down-stream genes m TOR and HIF-1α.Antioxidant NAC can block HIF-1α-mediated elevation of SYP.MAOA-specific inhibitor CLG can significantly delay the neuroendocrine differ-entiation process in vivo and in vitro,so as to achieve a better therapeutic effect.Conclusions:The NEPC model was successfully constructed and identified.We re-veal that MAOA drives the neuroendocrine differentiation of PCa,which provide a new strategy for clinical treatment of PCa by using MAOA inhibitor CLG.