Abstract LB256: Circulating methylated <i>GCM2</i> and <i>TMEM240</i> as novel biomarkers for dynamic monitoring of breast cancer progression and treatment response

Abstract Background Breast cancer remains a leading cause of cancer-related deaths, with 20-30% of early-stage cases progressing to metastasis. Current biomarkers, such as CA15-3 and CEA, are limited in detecting metastases, necessitating improved diagnostic tools. This study develops and validates a blood test using methylated GCM2 and TMEM240 for monitoring breast cancer progression, treatment response, tumor burden, and clinical parameters. Methods A prospective study in Taiwan (396 patients) and a retrospective cohort from Western populations (134 samples) were analyzed. cfDNA was extracted, bisulfite-converted, and GCM2 and TMEM240 methylation levels were measured using QMSP. Monte Carlo cross-validation (1,000 iterations) was performed to assess sensitivity, specificity, and accuracy. Methylation changes were further evaluated for correlations with treatment response, tumor burden, and clinical parameters. Results In the training set (n = 166, 13.3% recurrence/metastasis), biomarker cut-offs were established. Validation in an independent cohort (n = 325, 20% recurrence/metastasis) demonstrated significantly improved diagnostic performance compared to CA15-3 and CEA, with an accuracy of 95.1%, sensitivity of 89.4%, specificity of 96.5%, PPV of 86.8%, and NPV of 97.3%. Monte Carlo analysis of the training data yielded an average sensitivity of 95.7%, specificity of 90.3%, and accuracy of 91.5%, while validation data achieved sensitivity of 92.8%, specificity of 89.5%, and accuracy of 90.3% across 1,000 replicates.Critically, methylation levels of GCM2 and TMEM240 demonstrated high utility for monitoring treatment response and tumor burden. Decreased methylation levels were observed in patients responding favorably to treatment, signifying therapeutic success, whereas increased methylation levels were strongly associated with cancer progression, including recurrence and pre-metastatic events. Positive methylation levels were also significantly associated with advanced-stage disease (P &amp;lt; 0.001), larger tumor size (P = 0.002), and distant metastasis (P &amp;lt; 0.001). Conclusions The detection of methylated GCM2 and TMEM240 in plasma offers a highly accurate and noninvasive approach for monitoring breast cancer progression. The ability to dynamically track treatment response and tumor burden further underscores the clinical value of this novel biomarker assay in enhancing patient management and outcomes. Citation Format: Chin-sheng Hung, Hsieh-Tsung Shen, Ruo-Kai Lin. Circulating methylated GCM2 and TMEM240 as novel biomarkers for dynamic monitoring of breast cancer progression and treatment response [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_2):Abstract nr LB256.