An investigation of plasma cell-free RNA for the detection of colorectal cancer: From transcriptome marker selection to targeted validation.

Regular screening for colorectal cancer (CRC) is critical for early detection and long-term survival. Despite the current screening options available and advancements in therapies there will be around 53,000 CRC related deaths this year. There is great interest in non-invasive alternatives such as plasma cell-free RNA (cfRNA) for diagnostic, prognostic, and predictive applications. In the current study, our aim was to identify and validate potential cfRNA candidates to improve early CRC diagnosis. In phase 1 (n = 49; 25 controls, 24 cancers), discovery total RNA sequencing was performed. Select exons underwent validation in phase 2 (n = 73; 35 controls, 29 cancers, 9 adenomas) using targeted capture sequencing (n = 10,371 probes). In phase 3 (n = 57; 30 controls, 27 cancers), RT-qPCR was performed on previously identified candidates (n = 99). There were 895 exons that were differentially expressed (325 upregulated, 570 downregulated) among cancers versus controls. In phases 2 and 3, fewer markers were validated than expected in independent sets of patients, most of which were from previously published literature (FGA, FGB, GPR107, CDH3, and RP23AP7). In summary, we optimized laboratory processes and data analysis strategies which can serve as methodological framework for future plasma RNA studies beyond just the scope of CRC detection. Additionally, further exploration is needed in order to determine if the few cfRNA candidates identified in this study have clinical utility for early CRC detection. Over time, advancements in technologies, data analysis, and RNA preservation methods at time of collection may improve the biological and technical reproducibility of cfRNA biomarkers and enhance the feasibility of RNA-based liquid biopsies.
Competing Interests: I have read the journal’s policy and the authors of this manuscript have the following competing interests: Ms Berger, Messrs Taylor & Mahoney, and Dr. Kisiel are inventors of Mayo Clinic intellectual property licensed to Exact Sciences (Madison, WI) and may receive royalties, paid to Mayo Clinic. John Kisiel receives funding from a sponsored research agreement between Mayo Clinic and Exact Sciences. Because of the terms of the Sponsored Research Contract and Intellectual Property Development Agreement between Mayo Clinic and Exact Sciences, we have restrictions on sharing raw sequencing data publicly.
(Copyright: © 2024 Northrop-Albrecht et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)