Epithelial-Mesenchymal Transition Biomarkers In Peripheral Blood Predict Docetaxel Efficacy In Patients With Metastatic Castration-Resistant Prostate Cancer

Objective:Prostate cancer is the most common malignant tumor of the male genitourinary system with the highest incidence in the United States.The incidence of prostate cancer in China is lower than that in European and American countries,but with the change of lifestyle and the improvement of detection rate,the incidence of prostate cancer in China has been on the rise.Prostate cancer is prone to metastasis.Endocrine therapy is the main method for the treatment of metastatic prostate cancer by reducing the level of androgen in the body and controlling tumor growth.However,as the disease progresses,the vast majority of patients become less sensitive to endocrine therapy and eventually develop into metastatic castration-resistant prostate cancer(mCPRC).Currently,the main treatment of mCRPC is combined systemic chemotherapy or novel endocrine therapy on the basis of maintenance endocrine therapy.Although there are many prognostic predictors for mCRPC,the lack of predictors of treatment outcome remains to be a problem.Epithelial-mesenchymal transition(EMT)is a common phenomenon of tumor cell metastasis,which usually occurs in epithelial cells and manifests as loss of polarity,decreased adhesion,and enhanced migration.Vimentin is a major component of the intermediate filament protein family,which is commonly expressed in mesenchymal cells and is known for maintaining cell integrity and resistance to stress.E-cadherin belongs to the transmembrane glycoprotein family and is responsible for calcium-dependent intercellular adhesion.Both of them are recognized as important markers of EMT.The characteristic changes during EMT mainly include down-regulation of the epithelial differentiation marker E-cadherin and up-regulation of the stromal marker Vimentin,and then epithelial tumor cells enter the blood circulation and induce metastasis,which is related to the drug resistance and poor prognosis of the tumors.Recent studies have also found that EMT is associated with the occurrence,progression and drug resistance of mCPRC.In addition,circulating tumor cells(CTC)are a general term for all tumor cells circulating in the peripheral blood.Upon entering the circulation,they develop incomplete EMT and express both epithelial and mesenchymal cell characteristics and markers.The number of CTC has been proved to affect the prognosis of patients in a variety of tumors including prostate cancer,breast cancer and lung cancer,so the detection of cells expressing both Vimentin and E-cadherin in peripheral blood may indirectly reflect the status of CTC.Based on the above theory,we hypothesized that the expression of Vimentin and e-cadherin in the peripheral blood of patients with MPRC could predict the resistance and efficacy of docetaxel therapy.Flow cytometry was used to detect the ratio of cells expressing Vimentin and E-cadherin(expressed as Vim/E-cad)and the proportion of cells expressing both Vimentin and E-cadherin(expressed as Double)in peripheral blood before the first chemotherapy and after 4 cycles of chemotherapy,to test whether there is a correlation between these markers and efficacy of docetaxel as well as explore the predictors of efficacy.Methods:This study was a prospective cohort study involving male patients who were diagnosed with mCRPC,met the inclusion criteria and completed at least 4 cycles of docetaxel chemotherapy from November 2015 to June 2019.7.5 mL of peripheral blood was respectively collected before the first chemoth erapy and after 4 cycles of chemotherapy.After processing the sample,Vimentin antibody and E-cadherin antibody were added to the sample for double fluorescence staining,and then flow cytometry was used to detect the proportion of cells expressing Vimentin or E-cadherin in blood samples.At the same time,we collected the baseline information of patients,including age,ECOG score,Gleason scores of prostate cancer,prostate cancer TNM stage,transfer sites,whether there is a serious pain,previous treatment,PSA before the first chemotherapy,hemoglobin(Hb),neutrophils,lymphocyte,the whole body imaging evaluation.The treatment information after 4 cycles of chemotherapy were than collected,including PSA and systemic imaging evaluation.Each patients were followed up to all study endpoints,including PSA response,PSA progression free survival(PSA-PFS),and clinical progression-free survival(cPFS).Univariate analysis,multivariate analysis and survival curve analysis were used to examine the correlation between baseline Vim/E-cad and Double in peripheral blood before chemotherapy and PSA response,PSA-PFS and cPFS after 4 cycles of chemotherapy,and the relationship between the changes of Vim/E-cad and Double for chemotherapy and PSA response,to explore the predictive value of Vimentin and e-cadherin expression in peripheral blood of mCRPC patients for the efficacy of docetaxel.Results:A total of 79 patients were enrolled in this study,among which 3 patients failed to complete 4 cycles of chemotherapy due to adverse events or personal wishes,and 2 patients failed to be tested successfully due to quality of blood samples.The total of effective patients enrolled was 74 cases.The cases with baseline Vim/E-cad?1 and Vim/E-cad>1 were 55 and 22.The cases with baseline Double?2%and Double>2%were 49 and 25.(1)PSA response occurred in 34 of 52 patients(65.38%)with baseline Vim/E-cad<1,and in 6 of 22 patients(27.27%)with baseline Vim/E-cad>1,PSA response occurred in 32 of 49 patients(65.31%)with baseline Double?2%,and in 8 of 25 patients(32.00%)with baseline Double>2%.In multivariate analysis,baseline Double was an independent predictor of PSA response after docetaxel treatment(OR0.129,95%CI:0.029-0.573;P=0.007),patients with baseline Double>2%tended to have a poor PSA response.(2)Patients with baseline im/E-cad>1 tended to have shorter PSA-PFS(median 2.5 months vs.6.0 months,P<0.001)and shorter CPFS(median 4.5 months vs.8.0 months,P<0.001)after docetaxel treatment.Similarly,patients with baseline Double>2%tended to have shorter PSA-PFS(median 3.0 months vs.6.0 months,P=0.012)and shorter CPFS(median 6.0 months vs.7.5 months,P=0.007)after docetaxel treatment.In multivariate analysis,baseline Vim/E-cad>1 was associated with shorter PSA-PFS after docetaxel treatment(HR=2.217,95%CI:1.287-3.821;P=0.004)and shorter CPFs(HR=2.632,95%CI:1.507-4.599;P=0.001).(3)In the analysis of the relationship between baseline Vimentin,E-cadherin and clinical features,Gleason score was correlated with baseline Vim/E-cad and Double and an independent influencing factor of baseline Vim/E-cad(HR0.228,95%CI:0.063-0.826;P=0.024).(4)In the analysis of the factors influencing the efficacy of docetaxel,previous administration of abiraterone(OR0.037,95%CI:0.003-0.447;P=0.009)and Hb(OR7.964,95%1.772-35.803;P=0.007)were independent influencing factors for PSA response after 4 cycles of chemotherapy.Patients without previous treatment or with Hb?120g/L tended to have better PSA response.(5)The changes of Vimentin and e-cadherin expression before and after chemotherapy were not related to PSA response and could not predict the efficacy of chemotherapy,and the baseline expression was more valuable for predicting efficacy.Conclusions:We established a reliable method with flow cytometry for the detection of Vimentin and E-cadherin in peripheral blood,and demonstrated that the expression of Vimentin and e-cadherin in peripheral blood of patients with mCRPC could predict the efficacy of docetaxel.The predictive efficacy was worthy of further exploration in large-scale clinical trials and long-term clinical observation.