CNR1 Reverses Progesterone Resistance In Endometrial Cancer Through The ERK Pathway

Background:Endometrial carcinoma(EC)is the most usual epithelial-derived reproductive tract pernicious tumors in women.In recent years,the occurrence of EC is increasing year by year due to the epidemic of obesity and diabetes and the incidence of EC tends to be younger.Endocrine therapy is an effective treatment for young patients who want to preserve reproductive function,but drug resistance limits the practical use of endocrine therapy.This study analyses the effect of CNR1 on the mechanism in progesterone resistance,which is helpful to understand the function channel of progesterone-resistance in EC,so as to improve the efficacy of endocrine therapy and expand the treatment population,which has momentous medical sense.Methods:The correlation between CNR1 and EC was preliminarily analyzed by using the database.EdU,Transwell,MTT,western blot and RT-PCR verified the migration and invasion,proliferation ability,progesterone resistance and the expression of CNR1 of progesterone resistant cells.Resistant cells were transfected with lentivirus to construct sh-CNR1 cell lines and sh-NC cell lines,respectively,and given a certain concentration of MPA.Effects of CNR1 on resistant cells’ migration,invasion were detected by Transwell,scratch,proliferation and sensitivity to progesterone were detected by MTT,EdU,monoclonal,flow cytometry and immunofluorescence.Western Blotting and RT-PCR were used to verify the effect of CNR1-knockout on ERK pathway phosphorylation.Subcutaneous tumor model of nude mice was established to demonstrate the influence of regulating CNR1 and ERK phosphorylation on tumor-forming ability of resistant cells as well as progesterone sensitivity.Results:Survival analysis displayed that the level of CNR1 in EC patients’ tissues was inversely correlated with OS and RFS.The biological characteristics of the two types of cells were compared,resistant IshikawaPR cells’ migration,invasion and proliferation were enhanced,the protein level of related markers was changed accordingly,and the sensitivity to progesterone was decreased.On the other hand,CNR1 was highly expressed in resistant cells at both protein and mRNA levels,and PGR has a inverse relationship with CNR1 expression at mRNA level.CNR1 knockdown can improve the progesterone resistance and reduce the migration and invasion,proliferation ability of resistant cells.Western Blot and RT-PCR confirmed that down-regulation of CNR1 could inhibit ERK pathway phosphorylation,and the use of ERK activator could partially improve the proliferation ability of shCNR1 resistant cells.Tumor formation experiments in nude mice confirmed that CNR1 was involved in progesterone resistance in EC.The Tumorigenic capacity of sh-CNR1 cells was obviously weaker than the sh-NC cells,and progesterone sensitivity was higher.Conclusions:the ability of migration and invasion,proliferation of progesterone resistant cells in EC were enhanced.Our study demonstrated for the first time that CNR1 plays an important role in progesterone resistance in endometrial cancer.Loss of CNR1 expression can regulate the ERK phosphorylation and further make a dent in the migration and proliferation ability of resistant cells and thus reverse EC progesterone resistance.