A multiplex DNA methylation assay for noninvasive early detection of gastric cancer.

300 Background: Gastric cancer (GC) is the fifth leading cancer that ranked fourth in cancer mortality worldwide. Early screening and detection can significantly improve patients’ prognosis and survival. Although in standard GC diagnosis, gastroscopy provides sufficient diagnostic power for identifying GC in early stages, the invasive and costly procedure is not suitable for screening and has low compliance in countries with high GC incidence and large populations. Furthermore, the serum-based GC related biomarkers and non-invasive tests (serum pepsinogen I and II, gastrin-17, and anti-Helicobacter pylori IgG antibody tests) showed insufficient evidence as cost-effective GC screening approaches. A non-invasive DNA methylation assay with high sensitivity was aimed to develop and validate as a potential alternative for GC early detection. Methods: A multiplex quantitative PCR (qPCR) assay for detecting GC-specific DNA methylation markers in plasma was developed, in which the markers were further optimized from a reported seven-marker panel based on a pilot case-control study with 63 GC and 130 non-GC samples. The GC detection model of the assay was developed and locked down in the first cohort consisting of 54 GC and 79 non-GC samples. The second validation cohort with 117 GC and 309 non-GC samples was used to evaluate the assay performance. All plasma samples used in the two cohorts were prospectively collected from patients diagnosed with GC or other gastrointestinal diseases or from healthy donors. Results: The multiplex DNA methylation assay showed an overall sensitivity and specificity of 80% (95% CI: 77–83%) and 65% (95% CI: 63–66%) in the model development cohort, respectively, with a sensitivity of 80% (95% CI: 65–95%, n=10) in the detection of stage I GC. The assay revealed similar performance characteristics in the validation cohort, with an overall sensitivity of 82% (95% CI: 80–83%), a specificity of 69% (95% CI: 68–69%), and a sensitivity of 78% (95% CI: 74–82%, n=41) for the stage I GC detection. The DNA methylation assay outperformed the reported serum-based GC related biomarkers and non-invasive tests (sensitivity of 69.6-70.7% and specificity of 66.8-67.8%) with higher sensitivity, particularly in the stage I GC with a comparable specificity. Conclusions: The multiplex DNA methylation assay provides a potential cost-effective tool that features high sensitivity for GC early detection.