Fecal HBA mRNA as an alternative to FIT for colorectal cancer screening: a computational and clinical validation study

Current US FDA-cleared stool-based colorectal cancer (CRC) molecular screening tests include fecal immunochemical tests (FIT) for occult blood detection; however, integration with nucleic acid-based biomarkers requires separate workflows, thereby increasing complexity and cost. Computational analysis of GTEx RNA-seq data identified blood-specific genes. Clinical validation used RT-qPCR on stool samples from 364 participants (CRC, advanced precancerous lesions [APL], non-APL, negatives). FIT analysis was performed in parallel using standard methods. Diagnostic performance was compared with that of FIT using random forest models and cross-validation. HBA1/HBA2 mRNA was identified as the best mRNA biomarker for detecting occult blood in stool due to its high blood-specificity and abundance. In clinical samples, HBA mRNA Ct values were inversely correlated with FIT hemoglobin concentration and achieved comparable CRC and APL sensitivity at matched specificities. Combining HBA mRNA with CRC-associated mRNA markers achieved CRC and APL sensitivities comparable to those obtained with FIT plus CRC-associated mRNA markers. Fecal HBA mRNA offers diagnostic accuracy comparable to FIT while simplifying workflows for nucleic acid-based screening for CRC and APL.