A multiplex methylation-specific PCR assay for the detection of early-stage ovarian cancer using cell-free serum DNA

- DNA methylation changes may have a future role in clinical practice by facilitating early-stage EOC diagnosis.
- We established a novel multiplex methylation-specific PCR assay to assess the methylation status of candidates from minimal cell-free serum DNA.
- The analysis reached higher levels of sensitivity and specificity compared with CA125 in the detection of early-stage EOC.

ObjectiveEpithelial ovarian cancer (EOC) remains the most lethal disease among gynecological malignancies. Prompt diagnosis is challenging because of the non-specific symptoms exhibited during the early stage of the disease. As a result, there is an urgent need for improved detection methods. In this study, we established a multiplex methylation-specific PCR (MSP) assay to improve the early detection of ovarian cancer, via identification of the methylation status of cell-free serum DNA.MethodsAfter screening, we chose seven candidate genes (APC, RASSF1A, CDH1, RUNX3, TFPI2, SFRP5 and OPCML) with a high frequency of methylation to construct the multiplex-MSP assay. When methylation of at least one of the seven genes was observed, the multiplex-MSP assay was considered positive. We performed retrospective and screening studies to verify the specificity and sensitivity of the assay in the detection of EOC.ResultsThe methylation status of cell-free serum DNA was examined in the preoperative serum of 202 patients, including 87 EOC patients (stage I, n = 41; stage II-IV, n = 46), 53 patients with benign ovarian tumors and 62 healthy controls. As expected, the multiplex MSP assay achieved a sensitivity of 85.3% and a specificity of 90.5% in stageI EOC, strikingly higher rates compared with a single CA125, which produced a sensitivity of 56.1% at 64.15% specificity [P = 0.0036].ConclusionA multiplex MSP assay that analyzes the methylation status of cell-free serum DNA is a suitable and reliable approach to improve the early detection of ovarian cancer, potentially benefiting a broad range of applications in clinical oncology.