B-381 A Multiplex Blood-based Assay Targeting DNA methylation in Peripheral Blood Mononuclear Cells Enables Early Detection of Breast Cancer

Abstract

Abstract Background The immune system can monitor the development of tumors, and the DNA methylation is involved in the body immune response to tumors. Therefore, the DNA from peripheral blood mononuclear cell (PBMC) that carries cancer-specific epigenetic aberrations may serve as a non-invasive marker for early detection of breast cancer (BC). Methods Genome-wide DNA methylation profiling on PBMC DNA from BC patients and healthy controls (Cohort I, n = 80) was performed in the discovery phase by Infinium 850K BeadChips. The candidate CpG sites were selected and validated in a multistep validation phase (Cohort II, n = 200) via pyrosequencing and targeted bisulfite sequencing. After that, the multiplex quantitative methylation-specific PCR assay (mBC-MSP) was established to detect BC, and the performance of this diagnostic method was analyzed in a multicenter cohort (Cohort III, n = 501). Results A total of 289 differentially methylated CpG positions (DMPs) between BC patients and healthy controls were identified. Among them, eight significant DMPs markers were selected and further validated using pyrosequencing and targeted bisulfite sequencing, and four hyper-methylated DMPs (cg18637238, cg16652347, cg13828440, cg11754974) passed the final validation. Then, we developed a mBC-MSP assay based on the four methylated markers. This assay exhibited excellent performance in distinguishing BC patients, with an AUC of 0.925, sensitivity of 83.1%, and specificity of 90.4%. More importantly, compared with CA153, CA125 and CEA, the mBC-MSP assay had higher sensitivity for early-stage BC (stage 0, 88.2% vs 0%, 0%, 5.9%) and minimal tumor with diameter ≤1.5 cm (91.7% vs 0%, 2.1%, 0%). Conclusions The mBC-MSP assay based on the four identified DNA methylation markers from PBMC can serve as a noninvasive, accurate, rapid, and high-throughput method for early diagnosis of BC, and it is more sensitive than traditional tumor markers for detecting early-stage BC and minimal tumor.