title: A multiplex methylation-specific PCR assay for the detection of early-stage ovarian cancer using cell-free serum DNA.
abstract: OBJECTIVE
  Epithelial 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.
  
  
  METHODS
  After 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.
  
  
  RESULTS
  The 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].
  
  
  CONCLUSIONS
  A 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.
authors.affiliation: Department of Obstetrics and Gynecology, Qilu Hospital, Shandong University, Ji'nan, Shandong 250012, PR China.
authors.forename: Qing
authors.initials: Q
authors.lastname: Zhang
authors.forename: Guohong
authors.initials: G
authors.lastname: Hu
authors.forename: Qifeng
authors.initials: Q
authors.lastname: Yang
authors.forename: Ruifen
authors.initials: R
authors.lastname: Dong
authors.forename: Xing
authors.initials: X
authors.lastname: Xie
authors.forename: Ding
authors.initials: D
authors.lastname: Ma
authors.forename: Keng
authors.initials: K
authors.lastname: Shen
authors.forename: Beihua
authors.initials: B
authors.lastname: Kong
mesh_terms: D018394:CA-125 Antigen / Q000097:blood; D045744:Cell Line, Tumor; D019175:DNA Methylation; D004273:DNA, Neoplasm / Q000097:blood*; D055088:Early Detection of Cancer / Q000379:methods; D005260:Female; D006801:Humans; D008565:Membrane Proteins / Q000097:blood; D060787:Neoplasm Grading; D009367:Neoplasm Staging; D010051:Ovarian Neoplasms / Q000097:blood / Q000175:diagnosis* / Q000235:genetics* / Q000473:pathology; D016133:Polymerase Chain Reaction / Q000379:methods*; D012189:Retrospective Studies; D012680:Sensitivity and Specificity
chemical_list: D018394:CA-125 Antigen; D004273:DNA, Neoplasm; C451759:MUC16 protein, human; D008565:Membrane Proteins