Circulating extrachromosomal circular DNA as a prognostic biomarker for colorectal cancer

Abstract Background Delayed detection of recurrence significantly contributes to colorectal cancer (CRC) mortality, underscoring the need for robust prognostic biomarkers. Although extrachromosomal circular DNA (eccDNA) is a known oncogenic driver, its prognostic utility in CRC remains largely unexplored. Methods In this 6-year prospective cohort study, full-length eccDNA profiling of 153 plasma samples was performed using Nanopore sequencing. Differential eccDNA signatures between recurrence (R, n = 20) and non-recurrence (NR, n = 133) patients enabled construction of predictive models for recurrence and mortality. Functional validation of eccDNAs was conducted in HCT116 cells. Results Compared to NR patients, R patients exhibited enrichment of eccDNAs derived from chromosome 9, shorter median eccDNA lengths, and reduced variability in eccDNA length. All 4.9–5.0 kb eccDNAs derived from CKM, while other eccDNAs showed a strong genomic distribution correlation between groups (Spearman’s ρ = 0.73). Promoter-derived eccDNAs were enriched in R patients, particularly from the promoter of CARD9 (eccPromoter-CARD9, 10.4-fold increase). Overexpression of eccPromoter-CARD9 significantly promoted CRC cell proliferation and migration. R patients exhibited elevated eccDNAs harboring the hsa-mir-374c cluster in plasma and tissues, and their corresponding miRNAs demonstrated exceptional diagnostic accuracy in CRC-related TCGA cohorts. An eccDNA-based random forest classifier achieved superior recurrence prediction accuracy (AUC > 0.8), correlating with shorter time-to-recurrence (HR = 3.79) and elevated CA125 and CEA levels. Additional eccDNA-based models effectively predicted recurrence-associated mortality (AUC ≥ 0.93). Conclusions The plasma eccDNA landscape may serve as an early and powerful non-invasive biomarker for CRC prognostication, optimizing risk stratification and guiding personalized treatment.