Introduction
Colorectal cancer (CRC) is the third commonest malignancy globally & the second leading cause of cancer mortality. Prognosis is highly stage-dependent, with survival exceeding 85% for early-stage diagnoses but dropping sharply at advanced stages. While national screening programs have reduced mortality in older populations, incidence in younger individuals is rising, often accompanied by delays in diagnosis. Existing non-invasive approaches, such as quantitative faecal immunochemical (qFiT) testing & blood-based liquid biopsies, provide limited sensitivity, particularly for pre-malignant or early-stage disease, leaving colonoscopy as the invasive gold standard. Because CRC is a mucosal disease, rectal mucus represents an under-explored biospecimen enriched for exfoliated cells, DNA, & microbial material, potentially enabling early & minimally invasive detection.
Aims & Methods
We assessed rectal mucus sampling as a novel diagnostic approach for CRC & adenomatous polyps. Using the OriCol™ device, rectal mucus was retrieved from 800 participants from 4 NHS colorectal surgical outpatients clinics without the need for bowel preparation (July 2020 - October 2023). A hologenomic strategy was applied, integrating three complementary laboratory pipelines: error-corrected next-generation sequencing for somatic mutations (n=161), enzymatic methyl-sequencing for DNA methylation profiling (n=208), & whole-genome shotgun metagenomics for microbiome characterisation (n=379). Data underwent rigorous quality control, with feature selection & integrative multi-omics analysis used to identify clinically informative biomarkers, stratify pathology by site & stage, and evaluate diagnostic potential.
Results
Distribution of CRC was 24% right-colon, 23% hepatic-splenic flexure, 18% left-colon, 35% rectal. Somatic mutation analysis revealed pathogenic variants in well-known CRC drivers including TP53, APC, KRAS, BRAF, ERBB2, FBXW7, PIK3CA & SMAD4, with mutation allele frequencies representing disease state. DNA methylation profiling demonstrated widespread hypermethylation at CpG islands near promoters and transcription start sites, particularly in SDC2, LRRC4, PPP2R5C & RNF217, with gradients of signal strength from rectal to right-sided cancers & from CRC to advanced adenomas (>10mm +/- HG dysplasia), to early polyps/normal controls. Microbiome profiling identified 94 species associated with CRC, including previously reported taxa such as Fusobacterium nucleatum & Hungatella hathewayi, alongside novel associations with Clostridium scindens & Anaerotruncus colihominis. Multi-omics integration markedly improved diagnostic resolution, clustering CRC & advanced adenomas polyps separately from controls, with early-stage polyps occupying an intermediate position.
Conclusion
We established rectal mucus as a rich biospecimen for minimally invasive CRC diagnostics. By combining host genetic & epigenetic markers with microbial signatures, the hologenomic approach provides sensitive detection of pre-cancerous lesions & stratification by site/stage, outperforming single-modality assays. Importantly, local mucosal sampling enabled discovery of novel microbial-pathology associations not detectable in stool-based studies. These findings demonstrate the translational potential of rectal mucus hologenomics as a scalable diagnostic platform, offering a non-invasive alternative for triaging symptomatic patients & potentially extending to screening programmes. Further clinical validation is underway to refine performance metrics & benchmark against existing diagnostic standards (ORICOL-EGI-04 Study NCT06649123).
References
Complete references for this work are listed in the accepted Nature Communications paper
Disclosure
1. This work has been accepted for publication in Nature Communications and is currently embargoed whilst final editing is completed. A letter of confirmation of acceptance for publication can be provided.
2. The study is funded and performed through Origin Sciences Ltd with many of the staff holding share options in the company. The clinical research teams were provided with educational grants from Origin Sciences Ltd to fund the research and support research fellows and nurses.
3. Rectal mucus samples and tumour tissues were collected following UK Health Regulatory Authority (HRA) and Ethics Committee approvals (IRAS 263745, 19/EM/0266). Written informed consent was obtained from all donors. All samples were collected from patients enrolled in the ORI-EGI-02 study; Exploratory Study of Rectal Mucus for Diagnosing Disease (NCT04659590) from four NHS Hospitals.
4. The presenting author is employed by the NHS as a Consultant Colorectal Surgeon and is Chief Medical Officer, Origin Sciences and holds share options with the company.
