A novel DNA methylation signature associated with lymph node metastasis status in early gastric cancer

Abstract

Background: Lymph node metastasis (LNM) is an important factor for both treatment and prognosis of early gastric cancer (EGC). Current methods are insufficient to evaluate LNM in EGC due to suboptimal accuracy. Herein, we aim to identify methylation signatures for LNM of EGC, facilitate precision diagnosis, and guide treatment modalities. Methods: For marker discovery, genome-wide methylation sequencing was performed in a cohort (marker discovery) using 47 fresh frozen (FF) tissue samples. The identified signatures were subsequently characterized for model development using formalin-fixed paraffin-embedded (FFPE) samples by qPCR assay in a second cohort (model development cohort, n = 302, training set: n = 151, test set: n = 151). The performance of the established model was further validated using FFPE samples in a third cohorts (validation cohort, n = 130) and compared with image-based diagnostics, conventional clinicopathology-based model (conventional model), and current standard workups. Results: Fifty LNM-specific methylation signatures were identified de novo and technically validated. A derived 3-marker methylation model for LNM diagnosis was established that achieved an AUC of 0.87 and 0.88, corresponding to the specificity of 80.9% and 85.7%, sensitivity of 80.6% and 78.1%, and accuracy of 80.8% and 83.8% in the test set of model development cohort and validation cohort, respectively. Notably, this methylation model outperformed computed tomography (CT)-based imaging with a superior AUC (0.88 vs. 0.57, p < 0.0001) and individual clinicopathological features in the validation cohort. The model integrated with clinicopathological features demonstrated further enhanced AUCs of 0.89 in the same cohort. The 3-marker methylation model and integrated model reduced 39.4% and 41.5% overtreatment as compared to standard workups, respectively. Conclusions: A novel 3-marker methylation model was established and validated that shows diagnostic potential to identify LNM in EGC patients and thus reduce unnecessary gastrectomy in EGC.