Methylation Status of Corticotropin-Releasing Factor (CRF) Receptor Genes in Colorectal Cancer

Abstract

The corticotropin-releasing factor (CRF) system has been strongly associated with gastrointestinal pathophysiology, including colorectal cancer (CRC). We previously showed that altered expression of CRF receptors (CRFRs) in the colon critically affects CRC progression and aggressiveness through regulation of colonic inflammation. Here, we aimed to assess the potential of <i>CRFR</i> methylation levels as putative biomarkers in CRC. In silico methylation analysis of CRF receptor 1 (<i>CRFR1</i>) and CRF receptor 2 (<i>CRFR2</i>) was performed using methylome data derived by CRC and Crohn's disease (CD) tissues and CRC-derived circulating cell-free DNAs (ccfDNAs). In total, 32 and 33 differentially methylated sites of CpGs (DMCs) emerged in <i>CRFR1</i> and <i>CRFR2</i>, respectively, between healthy and diseased tissues. The methylation patterns were verified in patient-derived ccfDNA samples by qMSP and associated with clinicopathological characteristics. An automated machine learning (AutoML) technology was applied to ccfDNA samples for classification analysis. In silico analysis revealed increased methylation of both <i>CRFRs</i> in CRC tissue and ccfDNA-derived datasets. <i>CRFR1</i> hypermethylation was also noticed in gene body DMCs of CD patients. <i>CRFR1</i> hypermethylation was further validated in CRC adjuvant-derived ccfDNA samples, whereas <i>CRFR1</i> hypomethylation, observed in metastasis-derived ccfDNAs, was correlated to disease aggressiveness and adverse prognostic characteristics. AutoML analysis based on <i>CRFRs</i> methylation status revealed a three-feature high-performing biosignature for CRC diagnosis with an estimated AUC of 0.929. Monitoring of <i>CRFRs</i> methylation-based signature in CRC tissues and ccfDNAs may be of high diagnostic and prognostic significance in CRC.