Promoter hypermethylation of the tumor-suppressor genes ITIH5, DKK3, and RASSF1A as novel biomarkers for blood-based breast cancer screening

We examined promoter methylation of seven putative tumor-suppressor genes (SFRP1, SFRP2, SFRP5, ITIH5, WIF1, DKK3, and RASSF1A) in cfDNA extracted from serum. Clinical performance was first determined in a test set (n = 261 sera). In an independent validation set (n = 343 sera), we validated the most promising genes for further use in early breast cancer detection. Sera from 59 benign breast disease and 58 colon cancer patients were included for additional specificity testing.Based on the test set, we determined ITIH5 and DKK3 promoter methylation as candidate biomarkers with the best sensitivity and specificity. In both the test and validation set combined, ITIH5 and DKK3 methylation achieved 41% sensitivity with a specificity of 93% and 100% in healthy and benign disease controls, respectively. Combination of these genes with RASSF1A methylation increased the sensitivity to 67% with a specificity of 69% and 82% in healthy controls and benign disease controls, respectively.Tumor-specific methylation of the three-gene panel (ITIH5, DKK3, and RASSF1A) might be a valuable biomarker for the early detection of breast cancer.Breast cancer remains the most frequently diagnosed cancer and the leading cause of cancer deaths in European women [1]. According to the World Health Organization (WHO), more than 449,000 women in Europe will be diagnosed yearly with breast cancer, comprising approximately 28% of all cancers in female patients. Localized, early-stage breast cancer has a favorable prognosis, with a 5-year survival rate of up to 98%. However, the 5-year survival rate declines to 20% when the tumor has metastasized [2]. Although clinical examination and ultrasound provide a preliminary screening method, the two most sensitive and specific detection methods to date are mammography and magnetic resonance imaging (MRI). Mammography has become the standard of care in breast cancer screening. Mammograms are useful in that they are reliable and sensitive enough to detect duc