TROP2 Expression In Serous Ovarian Carcinoma And Its Effects On The Tumor Biological Behavior

Ovarian cancer(OC)is one of the common gynecological malignancies,although its incidence ranks third among gynecological reproductive tumors,it has the highest death rate.Due to the complexity of the pathogenesis of OC,effective diagnostic molecular markers and therapeutic targets are still lacking.Therefore,finding a unique molecular marker for ovarian cancer is still a major concern in the basic research of OC for the purpose to effectively improve the survival rate of patients and bring new hope for OC patients.Human trophoblast cell surface antigen-2(TROP2)is a transmembrane glycoprotein expressed in a variety of tumors,it’s encoded by the TACSTD2 gene and can participate in the activation of cell cycle related proteins and influence the proliferation,invasion,and apoptosis of tumor cells and malignant biological behavior by transmitting intracellular calcium signals and regulating ERK/MAPK signaling pathways.TROP2 was first discovered in the trophoblasts of the placenta and 1s expressed in stem-like cells.Studies have shown that TROP2 is overexpressed in diseases such as endometrioid adenocarcinoma,cervical cancer,breast cancer,laryngeal cancer and colorectal cancer.Overexpression of this protein has been associated with decreased survival in many patients with malignant tumors and increased tumor invasiveness and metastasis,and the overexpression of TROP2 in metastatic tissues make it an attractive and potential therapeutic target for the treatment of advanced malignancies.Serous ovarian cancer.(SOC)is the most common form of OC.In this study,we detected the high expression of TROP2 in SOC,evaluated its relationship with clinicopathological features and prognosis,and observed that the expression of TROP2 was significantly down-regulated after the interference of siRNA on OC cells.Our results indicated that TROP2 overexpression is closely related to the progression of SOC invasion and poor prognosis,and is an independent factor for evaluating the poor prognosis of SOC,besides,it could promote the proliferation,invasion and metastasis of OC cells.TROP2 might become a potential prognostic marker for SOC and a new target for targeted therapy.PART I Expression of TROP2 in serous ovarian carcinoma and its correlation with Ki-67Objective:To detect and analyze the expression of TROP2 in ovarian lesions and normal ovarian and fallopian tube tissues,and to explore the role of TROP2 expression in SOC cell proliferation and its correlation with clinicopathological features of patients.Methods:Immunohistochemistry was used to determine the expression of TROP2 and Ki-67 proteins in 126 patients with SOC,36 patients with borderline ovarian tumors,26 patients with ovarian serous cystadenoma,20 patients with normal ovaries,and 20 patients with normal fallopian tube parasol parafifin-embedded tissues.TROP2 expression levels were compared among different clinicopathological characteristics,and the correlation between TROP2 and Ki-67 expression was analyzed.Results:1.TROP2 was mainly expressed in the cell membrane of OC tissue.Normal ovarian tissue was used as the control,and the experimental group is ovarian low-grade serous adenocarcinoma.There were 12 negative cases(60%),7 weakly positive cases(35%),1 moderately positive case(5%),and no strong positive cases.And among the ovarian low-grade SOC group,6 cases were negative(10%),1 3 cases were weakly positive(2 1.7%),20 cases were moderately positive(5%),and 21 cases were strongly positive(35%).The expression of TROP2 protein was significantly higher than that of normal ovarian,ovarian serous cystadenoma and ovarian borderline tumor tissues(P<0.001).Normal ovarian tissue was used as the control,and the experimental group is ovarian high-grade serous adenocarcinoma.There were 10 negative cases(50%),8 weakly positive cases(40%),2 moderately positive case(10%),and no strong positive cases.And among the ovarian low-grade SOC group,8 cases were negative(12.1%),16 cases were weakly positive(24.2%).22 cases were moderately positive(33.3%),and 20 cases were strongly positive(30.3%).The expression of TROP2 protein was significantly higher than that of normal ovarian,ovarian serous cystadenoma and ovarian borderline tumor tissues(P<0.001).The expression of TROP2 protein was significantly higher than that of normal oviduct(P<0.001).2.TROP2 expression was closely correlated with SOC clinical stage,pathological grade,preoperative serum CA125 level,ascites and lymph node metastasis(P<0.05),but was not significantly correlated with age(P=0.280)or platinum resistance(P=0.944).3.Ki-67 expression in low grade and high grade SOC was significantly higher than that in the control group and benign lesion tissues(P<0.001).Spearman correlation analysis showed that the expression of TROP2 is associated with Ki-67 expression in SOC(rs=0.453,P<0.001).Conclusions:1.The expression of TROP2 in SOC was significantly higher than that in the control group and benign lesions,and the high expression of TROP2 was significantly associated with the adverse clinicopathological features of OC,indicating that TROP21s involved in the development of ovarian cancer.2.TROP2 was positively correlated with Ki-67 expression in the same SOC tissues,indicating that TROP2 might be involved in the growth and proliferation of OC cells.PART Ⅱ Clinical data statistics and survival analysis of 126 patients with SOCObjective:To further explore the factors affecting survival in patients with SOC in part Ⅰ of this study.To analyze the effects of TROP2 expression,FIGO clinical stage,tumor pathological grade,preoperative serum CA125 level,ascites,lymph node metastasis and age on the survival time of SOC patients.Methods:A total of 126 patients with SOC were enrolled with the same criteria as in part Ⅰ.The patients underwent surgery in Qilu Hospital of Shandong University and completed the treatment in time,the complete case records and follow-up data were saved.The follow-up time was 60 months.The follow-up information included total survival time(OS)and progression-free survival time(PFS).The survival time was compared by Kaplan-Meier method,and the survival curve was drawn.Univariate and multivariate Cox proportional risk regression analysis was used to screen the main factors affecting prognosis.Results:1.Univariate analysis showed that TROP2 protein overexpression was associated with PFS(P<0.001)and OS(P=0.017).The median survival of patients with FIGO clinical stage,tumor pathological grade,preoperative serum CA125 level,ascites and lymph node metastasis was statistically different(P<0.05),while there was no correlation between median survival and age group.2.Multivariate COX regression model analysis showed that TROP2 expression,FIGO clinical stage,preoperative serum CA125 level,ascites and lymph node metastasis were independent factors affeecting the five-year survival rate of SOC patients.Conclusions:1.High expression of TROP2 is positively correlated with poor prognosis of SOC,and TROP2 is an independent prognostic indicator for SOC.2.FIGO clinical stage,preoperative serum CA125 level,ascites and lymph node metastasis are also independent factors affecting the prognosis of SOC.PART Ⅲ Effects of TROP2 expression on biological behavior of OC cellsObjective:To observe the effects of changes in TROP2 expression levels on biological behaviors such as proliferation,apoptosis,invasion and migration of OC cells,and explore the role and possible mechanism of TROP2 in the development of OC.Methods:We synthesized two independent siRNA sequences siRNA-550 and siRNA-1100 to transfect OC cells and interfere with the endogenous expression of TROP2.The expression of TROP2 protein after transfection was determined by Western blot.The effect of TROP2 expression on the proliferation of ovarian cancer cells was determined by CCK-8 method.The migration and invasion abilities of cells were measured by scratch test and invasion test respectively.The expressions of apoptosis-related proteins Bax and Bcl-2 were determined by Western blot.Results:1.Immunofluorescence staining showed a red fluorescent stain on the surface of A2780,HO8910 and SK-OV-3 cells.Western blot analysis showed that TROP2 protein was expressed in all three kinds of cells,and the expression of TROP2 in A2780 cell line was higher than that of the other two.2.TROP2 protein expression was significantly decreased after transfecting siRNA-550 and siRNA-1100 in A2780 cell.3.CCK-8 showed that TROP2 gene silencing significantly decreased the proliferation ability of A2780 cells(P<0.05).4.Cell invasion experiments found that the number of OC cells passing through Transwell compartment after TROP2 gene silencing was significantly decreased(P<0.05),indicating that the ability of cell invasion was decreased.5.The cell scratch test indicated that after 24 and 48 hours of the cell scratch,the OC cells with TROP2 gene silencing showed a significantly slow scratch healing speed(P<0.05),indicating that the migration ability of the cells was significantly decreased.6.Western blot analysis showed that the expression of Bcl-2 protein in OC cells was significantly decreased after TROP2 gene silencing,while the expression of Bax protein was significantly enhanced.Conclusions:TROP2 expression is related to the proliferation,apoptosis,migration and invasion of OC cells,and it might induce apoptosis by disrupting the balance regulation between Bax/Bcl-2 family proteins.TROP2 plays an important role in the development and progression of OC,and it might become a new target for OC therapy and provide a theoretical basis for targeted therapy.