Intratumoral Heterogeneity Of High Grade Serous Ovarian Cancer And Clinical Significance Of Circulating Tumor DNA In Monitoring The Efficacy Of Patients

Objective:Epithelial ovarian cancer(EOC)is the most deadly gynecological malignancy,since about 60%-70%of EOC patients have already been in advanced stage when they received treatment.Although majority of the patients are able to obtain complete clinical remission after tumor cytoreductive surgery and platinum-based combined chemotherapy,70%-80%of which will relapse within 1-2 years.High grade serous ovarian cancer(HGSOC),as the most common histological subtype of EOC,is a highly heterogeneous malignant tumor.Intratumoral heterogeneity may be the cause of chemotherapy resistance.In recent years,circulating tumor DNA(ctDNA)has shown certain clinical application value,and is expected to overcome the influence of intratumoral heterogeneity and to monitor tumor progression and the effect of chemotherapy in real time.This study aimed to examine the mutations of tumor susceptibility gene in HGSOC patients,analyze the intratumoral heterogeneity of HGSOC,and explore the clinical significance of monitoring the treatment response of HGSOC patients through using TP53 mutations in plasma ctDNA.Method:18 patients with pathological diagnosis of HGSOC were selected from the ovarian cancer patients hospitalized in the gynecology department of the Third Affiliated Hospital of Guangzhou Medical University from January 1st,2016 to January 1st,2019.We collected the primary tumor tissue,metastatic tumor tissue and recurrent tumor tissue of each HGSOC patient as much as possible.Each tumor tissue was equally dissected into six to eight pieces for independent DNA extraction.DNA from preoperative blood leukocytes was used as a control to perform next generation sequencing(NGS)on 18 tumor susceptibility genes and the results were analyzed.Based on the results of NGS,we designed droplet digital PCR(ddPCR)detection reagent for TP53 mutation,analyzed TP53 mutation in the ctDNA extracted from the plasma of HGSOC patients from pre-operation,post-operation and each post-chemotherapy,and then followed up the level of cancer antigen 125(CA125)and the imaging manifestations such as CT/MRI according to their treatment process.Results:Tumor tissue DNA from 18 HGSOC patients were analyzed by next generation sequencing.The results showed that 88.9%(16/18)of HGSOC patients had tumor susceptibility gene mutations,among which TP53 mutation rate was 83.3%(15/18),BRCA1 mutation rate was 22.2%(4/18),BRCA2 mutation rate was 11.1%(2/18),PTEN mutation rate was 5.6%(1/18).Interestingly,gene mutation frequency is significantly different in different regions from the same tumor tissue,suggesting prevalent intratumoral heterogeneity in HGSOC.In addition,mutation frequency changes at different clinical stages,confirming tumor heterogeneity is a dynamic process shaped by disease progression and treatment.Based on the results of NGS,a series of plasma ctDNA of 11 HGSOC patients were detected by ddPCR.The results showed that TP53 specific somatic mutations consistent with tumor tissue DNA were detected in plasma ctDNA of 8(72.7%)patients.In the course of treatment,the mutation level of TP53 in plasma ctDNA decreased,and the mutation level of TP53 increased when the disease progressed,which was consistent with the change of traditional EOC tumor biomarker CA125.Conclusions:Intratumural heterogeneity is highly dynamic in HGSOC tumors during disease progression and treatments,which may explain frequent emergence of resistance following repeated chemotherapies among HGSOC patients.TP53 mutation in ctDNA may be a potential tumor biomarker to monitor the response of HGSOC patients.