Profiling small RNAs in CRC screening samples such as the widely used fecal immunochemical test, is it possible?

Faecal microRNAs represent promising molecules with potential clinical interest as non-invasive diagnostic and prognostic biomarkers for their stability and detectability. Colorectal cancer (CRC) screening based on the fecal immunochemical test (FIT) is an effective tool for prevention of cancer development. However, due to the poor sensitivity of FIT for premalignant lesions, there is a need for implementation of complementary tests. Improving the identification of individuals who would benefit from further investigation with colonoscopy using molecular analysis, such as miRNA profiling of the FIT leftover buffer, would be ideal due to its widespread use. In the present study, we applied small RNA sequencing to FIT leftover samples collected from two European screening populations. We showed robust detection of miRNA and microbial profiles, which were similar to those obtained from specimens sampled using RNA stabilising buffers and archived fecal samples. Detected miRNAs exhibited differential abundance between CRC and control samples that was consistent between sampling methods, suggesting a promising potential to identify small RNA CRC biomarkers using FIT leftovers. We demonstrated that it is possible to analyse gut miRNAs in FIT leftover samples and envision that these potential biomarkers can complement the FIT in large scale screening settings. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement This work was supported by the Italian Institute for Genomic Medicine (IIGM) and Compagnia di San Paolo Torino, Italy (to AN, BP, and ST). This project has received funding from the European Union's Horizon 2020 research and innovation program under grant agreement No 825410 (ONCOBIOME project to AN, BP, and ST). The research leading to these results has received funding from AIRC under IG 2020, ID. 24882, P.I. Naccarati Alessio Gordon (to AN) and under IG2019 , ID. 23473, P.I. Senore Carlo (to CS). This project also received funding from the Norwegian Cancer Society open call 2017 (190179) to TBR and 2018 (198048) to TBR and from the Cancer Registry of Norways funds to TBR. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: For the MITOS cohort the study design and the informed consent were approved by the local Ethics committee (AOU Città della salute e della Scienza di Torino, Italy). For the NORCCAP and BCSN cohorts anonymized samples were used. This was approved by the Norwegian Regional Ethics Committee - South-East (REC) projects, S-98052 and 2011/1272, respectively. I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes The datasets used and/or analysed during the current study are available from the corresponding author on reasonable request.