The Value Of Liquid Biopsy In Neoadjuvant Chemotherapy For Locally Advanced Breast Cancer

Neoadjuvant chemotherapy is the standard treatment for locally advanced breast cancer patients.The imaging evaluation has some defects in comprehensiveness and timeliness.Liquid biopsy technology has been widely used in the whole management of breast cancer patients due to its non-invasive,real-time and comprehensive characteristics,but its role in the efficacy evaluation of neoadjuvant chemotherapy for breast cancer remains to be determined.This study was divided into three parts to investigate the role of circulating tumor cells,aneuploidy circulating endothelial cells and cell-free DNA in the efficacy evaluation of neoadjuvant chemotherapy for breast cancer.A total of 61 patients with locally advanced breast cancer were enrolled in this study.All patients underwent core needle biopsy to obtain pathological results,including histological type,hormone receptor,HER-2 status,and Ki-67 index.All patients were treated with EC×4-T×4 NCT.（Epirubicin 90mg/m² IV d1,cyclophosphamide600mg/m² IV d1,21 days as one cycle,a total of 4 cycles;Docetaxel 100 mg/m² IV d1,21 days as one cycle,a total of 4 cycles）.Peripheral blood samples were collected before treatment（at the time of biopsy）and after the first and eighth courses of chemotherapy for testing.Patients’response to chemotherapy was assessed based on core needle biopsy specimens and surgical specimens at the end of chemotherapy,and the assessment was conducted in two different ways:1.Miller-Payne system 2.The change of Ki-67 index before and after chemotherapy.Patients were divided into high response group and low response group.In the first part,a total of 45 patients underwent CTC testing.Using the SE-i·FISH^?system,the total number of CTC and the changes of each subtype during neoadjuvant chemotherapy were evaluated in this study.The results showed that overall CTC and small-size CTC in the high-response group were generally stable during chemotherapy,and continued to increase in the low-response group.At the end of the eighth course of chemotherapy,the CTC level in the low-response group was significantly higher than that in the high-response group.The proportion of triploid and tetraploid CTC increased continuously in the low response group.The number of total CTC and subtypes at the end of the eighth cycle of chemotherapy has a certain ability to predict the response to chemotherapy.PFS and OS of patients with lower CTC were significantly better than those with higher CTC.In the second part,a total of 41 patients were able to obtain aneuploid CEC test data.In this part,the level changes of positive/aneuploid CEC during NCT were investigated under the detection of SE-i·FISH^?system.In the process of NCT,the overall CEC showed a trend of increasing at first and then decreasing.The aneuploid CEC increased first and then decreased in the high-response group,and continued to increase in the low-response group.There was no statistical difference in the change of diploid CEC during NCT.In this study,epithelial-endothelial fusion cells with Ep CAM+CD31+phenotype were detected.In the process of NCT,there was a significant positive correlation between aneuploid CEC and CTC.In the third part,a total of 61 patients underwent cf DNA detection of peripheral blood.Concentrations of 111bp and 260bp DNA fragments were detected by PCR to evaluate the level of cf DNA.Genomic DNA extracted from breast cancer cell line SUM-1315was used as a control to evaluate the integrity of plasma DNA.PCR results showed that there were no statistically significant differences in cf DNA levels at 3 time points between the high response group and the low response group.Before and after NCT,Alu-111bp fragment,an indicator of apoptosis,remained relatively stable in the low response group,but the level of Alu-111bp increased significantly in the high response group.There was no significant difference between Alu-260bp and CFDI among different chemotherapy response groups.In vitro experiments,breast cancer cell line MCF-7 and drug-resistant cell line MCF-7/ADR were added with epirubicin to simulate chemotherapy environment.The correlation between Alu-111bp fragment concentration and apoptosis in cell supernatant was found.