Abstract
Endoscopy is the gold standard for diagnosing gastric cancer (GC), but its invasiveness limits widespread participation and has not substantially reduced GC-related mortality. This study developed and validated a blood-based digital PCR assay for early GC detection using DNA methylation biomarkers. Genome-wide methylation profiles from over 10,000 samples were screened, and two candidates were validated in GC cell lines, tumors, matched non-cancerous tissues, and plasma. Plasma from 60 GC patients, including 38 with stage I disease, and 40 healthy controls was analyzed with a digital PCR assay targeting the selected biomarkers, using ACTB as a reference. GHR and GLRB methylation were identified as novel GC biomarkers, showing consistent hypermethylation in GC cell lines and tumor tissues. In plasma, the two-marker assay achieved 83.3% (95% CI 71.5%-91.7%) sensitivity and 90% (95% CI 76.3%-97.2%) specificity, clearly outperforming carcinoembryonic antigen (CEA) testing (10.0%; 95% CI 3.8%-20.5%). Incorporation of GATM methylation as a third marker increased sensitivity to 86.7% (95% CI 75.4%-94.1%) overall and 81.6% (95% CI 65.7%-92.3%) for stage I disease, while maintaining 90.0% specificity. This methylation-based digital PCR assay enabled accurate, noninvasive detection of GC, particularly at early stages, and may facilitate timely diagnosis and curative treatment.