Detection of Tumor DNA in Peritoneal Lavage Samples and Plasma in Patients with Locally Advanced Rectal Cancer

Abstract

Introduction. Locally advanced rectal cancer (LARC) is an advanced disease of rectal cancer where the tumor grows through the rectum wall and into other organs. Patients with LARC are referred to the Norwegian Radium Hospital for evaluation. The standard of care for LARC is neoadjuvant treatment followed by surgery where the tumor and affected organs are resected. Most patients have curative surgery; still, some patients develop local recurrences and/or metastasis in the abdominal cavity. Additional diagnostic tools are needed to identify patients at risk of recurrence and metastasis, that might benefit from additional follow-up and treatment. Detection of exfoliated tumor cells in the abdominal cavity has been presented as a possible biomarker to predict recurrence and overall survival in CRC. This thesis aimed to investigate the prognostic value of detecting tumor DNA in peritoneal lavage samples collected from LARC patients during surgery. Additionally, we wanted to investigate the prognostic value of detecting circulating cell-free tumor DNA (ctDNA) in blood samples collected from patients before neoadjuvant treatment.

Materials and Methods. We processed and prepared patient samples for targeted sequencing and droplet-digital PCR (ddPCR). We used targeted Ion Torrent next-generation sequencing (NGS) to identify tumor-specific mutations in biopsies from LARC patients. The mutations were utilized as biomarkers for detecting tumor DNA in peritoneal lavage samples and ctDNA in plasma with ddPCR. Statistical analysis was performed to investigate if the detection of tumor DNA and ctDNA was associated with disease-free and overall survival.

Results. We analyzed peritoneal lavage samples from 68 patients and plasma samples from 58 patients with ddPCR using the tumor-specific mutation, identified with NGS, as biomarkers. Further, we detected tumor DNA in the peritoneal lavage in 44% of patients and ctDNA in 71% of patients. Preliminary statistical analysis showed no differences in clinical characteristics and disease-free or overall survival in the patients with and without detected tumor DNA.

Conclusion. We have shown that ddPCR can be used to detect tumor DNA from exfoliated tumor cells in the abdominal cavity and ctDNA in plasma by using tumor-specific biomarkers. Our preliminary results indicated that the detection of tumor DNA in peritoneal lavage and ctDNA in plasma might not be a prognostic biomarker for LARC patients. However, the analysis is based on a low number of patients, and the findings need to be verified by further analysis of the remaining patients in the cohort or larger study cohorts.