Circulating cell-free DNA as predictor of pathological complete response in locally advanced rectal cancer patients undergoing preoperative chemoradiotherapy

Background: The watch and wait (W&W) strategy is proposed for patients with locally advanced rectal cancer (LARC) achieving clinical complete response (cCR) after neoadjuvant radiotherapy. cCR is only in partial concordance with pathological complete response (pCR) due to persisting viable tumour cells. The aim was to investigate circulating-free-deoxyribonucleic-acid (cfDNA) as a biomarker for prediction of pCR. Materials and methods: Patients treated with neoadjuvant radiotherapy for LARC, were included in a prospective biomarker study in Aarhus, Denmark from 2017 to 2020. Plasma cfDNA levels were analysed by a direct fluorescent assay (DFA). Surgical specimens were reviewed by pathologists to categorize response to cytotoxic therapy. Results: In total, 76 patients were included with plasma available at baseline (n = 70), mid therapy (n = 50), and end of therapy (n = 54). Higher cfDNA levels were observed in LARC patients compared with healthy subjects (p < 0.01). By ROC analysis (AUC: 0.87 (95% CI, 0.81-0.92)) the optimal cut-off was 0.71 ng/mu L for differentiation between healthy subjects and LARC patients. Thirteen patients obtained pCR with a median cfDNA level of 0.57 ng/mu L at end of therapy. Patients with cfDNA levels at end of therapy below the cut-off (p < 0.02) and `cfDNA responders' with descending levels greater than the 75th percentile during therapy had a significantly higher chance of pCR (p < 0.01). Conclusion: This hypothesis generating study indicates that low cfDNA levels at end of treatment or ' cfDNA responders ' might be associated with pCR. Quantification of cfDNA by the rapid and feasible DFA analysis could potentially facilitate personalized follow-up as a complementary tool to identify candidates for a W&W strategy.