A comprehensive multi-center study published from Japan investigates the prevalence and chronology of colibactin-associated mutational processes in colorectal cancer (CRC). Evaluating 600 individuals—including 454 CRC patients and 146 healthy controls recruited between 2014 and 2025—researchers utilized whole-genome sequencing and whole-genome metagenomic sequencing to map microbial signatures and genetic alterations.
Inside the Clinical Cohorts and Institutional Framework
The research initiative was backed by formal approvals from the Research Ethics Committees of the National Cancer Center, the University of Osaka, and the Institute of Science Tokyo. Operating under institutional guidelines for medical and health research, the study secured written informed consent from all enrolled participants. Every dataset underwent strict de-identification and a double-anonymization procedure to safeguard participant privacy.
Cohort 1 consisted of prospectively collected cases specifically gathered for this protocol, while cohort 2 incorporated cases from previously published research. To maintain clinical focus, investigators deliberately excluded individuals presenting with hereditary CRC syndromes—such as familial adenomatous polyposis and hereditary non-polyposis CRC—along with patients diagnosed with inflammatory bowel disease.
Methodology: Connecting Stool Metagenomics and Tumor Genomes
Whole-genome sequencing data was successfully derived from primary CRC tissues across 200 cases in cohort 1. Meanwhile, whole-genome metagenomic sequencing data extracted from stool samples was compiled for a total of 395 CRC patients and 146 healthy controls across both cohorts. To capture lifestyle variables, participants completed a detailed, 25-page, self-administered questionnaire comprising 475 items based on the Japan Public Health Center-based Next Generation Study framework.
Stool collection protocols were tightly regulated. Healthy controls were identified as individuals who presented with a positive fecal occult blood test during a hospital visit, underwent a secondary screening colonoscopy, and showed no clinically significant findings. Stool specimens were acquired prior to any clinical intervention, including surgical resection or chemotherapy, utilizing the first stool passed at the hospital on the day of the colonoscopy. These samples were immediately frozen on dry ice and preserved at −80 degrees Celsius.
| Participant Category | Total Enrolled | WGS Data (Tumor Tissues) | WGMS Data (Stool Samples) |
|---|---|---|---|
| Colorectal Cancer (CRC) Patients | 454 | 200 (Cohort 1) | 395 |
| Healthy Controls (HC) | 146 | N/A | 146 |
| Combined Total | 600 | 200 | 541 |
Defining Endpoints and Clinical Tracking
Tumor tissues were harvested directly from surgically resected specimens, while matched non-tumor DNA was primarily obtained from peripheral blood lymphocytes. In select cases, non-neoplastic colonic mucosa sampled from anatomical sites distant from the primary tumor served as a normal control. Clinical outcomes were tracked using standardized definitions: overall survival was measured from the initiation of treatment to death from any cause, with right-censoring applied at the date of last confirmed follow-up for surviving patients. Progression-free survival was calculated from treatment initiation to the first documented radiographic evidence of disease progression.
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