| BackgroundProstate cancer is the second most common cancer and the fifth leading cause of cancer death in men of the world.According to the data in 2022,the incidence and mortality of prostate cancer in men will rise sharply in China.Androgen receptor(AR)plays a key role in the occurrence and development of prostate cancer.As an endocrine-related tumor,prostate cancer is clinically treated by combining castration drugs and androgen receptor(AR antagonist)to block the AR signaling pathway.However,after the initial treatment had achieved success,almost all patients inevitably developed prostate cancer progression due to the reactivation of the AR signaling pathway and enter the stage of castration-resistant prostate cancer(CRPC).The prognosis of patients at this stage is very bad.At present,for the treatment of CRPC,the second-generation AR antagonist enzalutamide had been used to the patients of CRPC.Enzalutamide mainly inhibits the activation of AR signaling pathway by competitively antagonizing AR.Enzalutamide has been approved for the treatment of CRPC,because it can prolong the survival of patients of CRPC.However,patients with the treatment of enzalutamide will still develop drug resistance due to the emergence of multiple drug resistance mechanisms,especially the reactivation of the AR signaling pathway.These enzalutamide-resistant patients were often insensitive to subsequent active treatments.Therefore,in-depth exploration of the specific mechanism of enzalutamide leading to drug resistance in CRPC is particularly critical for finding new treatment options for CRPC.More and more studies have identified post-translational modification(PTM)as an important mechanism for regulating AR activity.At present,the phosphorylation of the 81st serine site in AR is the most studied,which is considered to promote the downstream signaling pathway of AR,while the functional mechanism of other modification sites is still unclear.After the treatment of enzalutamide,the increase of AR activity often leads to drug resistance.Therefore,the relationship between the post-translational modification site of AR and enzalutamide resistance should be explored in depth.The drug resistance of enzalutamide would be clarified from a new perspective,which would guide significance for the development of new drugs.MethodsIn this study,firstly,the phenomenon of enzalutamide promoting AR ligand-independent nuclear translocation in prostate cancer cell lines was observed by fluorescence experiment,nuclear cytoplasmic separation experiment and protein real-time molecular labeling techniques.Secondly,enzalutamide-resistant prostate cancer cell lines were successfully constructed by gradually increasing the concentration of enzalutamide.We performed immunofluorescence experiment,protein half-life experiment,and DHT-induced cell proliferation experiment to observe the functional changes of AR between enzalutamide-resistant cell lines and parental cell lines.Finally,for the identified phosphorylation site,a mutant plasmid of AR was constructed.We further identified the functional role of the phosphorylation site in drug-resistant cell lines.Results and ConclusionIn this study,it was the first time to confirm that enzalutamide can promote the ligand-independent nuclear translocation of AR and mediate the enrichment of AR in the nucleus.Secondly,through the construction of drug-resistant cell lines,the changes of prostate cancer cells in the process of castration resistance were observed,which confirmed the close relationship between AR nuclear enrichment and AR high stability and high sensitivity during disease progression.Finally,we confirmed that the phosphorylation of serine 94 of AR was not only involved in the drug resistance process of enzalutamide,but also an important regulator of AR ligand-independent nuclear translocation. |
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