title: Methylation analysis of plasma cell-free DNA for breast cancer early detection using bisulfite next-generation sequencing.
abstract: Circulating cell-free DNA (cfDNA) has been considered as a potential biomarker for non-invasive cancer detection. To evaluate the methylation levels of six candidate genes (EGFR, GREM1, PDGFRB, PPM1E, SOX17, and WRN) in plasma cfDNA as biomarkers for breast cancer early detection, quantitative analysis of the promoter methylation of these genes from 86 breast cancer patients and 67 healthy controls was performed by using microfluidic-PCR-based target enrichment and next-generation bisulfite sequencing technology. The predictive performance of different logistic models based on methylation status of candidate genes was investigated by means of the area under the ROC curve (AUC) and odds ratio (OR) analysis. Results revealed that EGFR, PPM1E, and 8 gene-specific CpG sites showed significantly hypermethylation in cancer patients' plasma and significantly associated with breast cancer (OR ranging from 2.51 to 9.88). The AUC values for these biomarkers were ranging from 0.66 to 0.75. Combinations of multiple hypermethylated genes or CpG sites substantially improved the predictive performance for breast cancer detection. Our study demonstrated the feasibility of quantitative measurement of candidate gene methylation in cfDNA by using microfluidic-PCR-based target enrichment and bisulfite next-generation sequencing, which is worthy of further validation and potentially benefits a broad range of applications in clinical oncology practice. Quantitative analysis of methylation pattern of plasma cfDNA by next-generation sequencing might be a valuable non-invasive tool for early detection of breast cancer.
authors.affiliation: The State Key Laboratory of Medical Genetics and School of Life Sciences, Central South University, 172 Tongzipo Road, Changsha, Hunan, 410013, China.
authors.forename: Zibo
authors.initials: Z
authors.lastname: Li
authors.affiliation: Sanway Gene Technology Inc., Changsha, Hunan, 410205, China.
authors.forename: Xinwu
authors.initials: X
authors.lastname: Guo
authors.affiliation: Department of Breast Surgery, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China.
authors.forename: Lili
authors.initials: L
authors.lastname: Tang
authors.affiliation: Sanway Gene Technology Inc., Changsha, Hunan, 410205, China.
authors.forename: Limin
authors.initials: L
authors.lastname: Peng
authors.affiliation: Sanway Gene Technology Inc., Changsha, Hunan, 410205, China.
authors.forename: Ming
authors.initials: M
authors.lastname: Chen
authors.affiliation: Sanway Gene Technology Inc., Changsha, Hunan, 410205, China.
authors.forename: Xipeng
authors.initials: X
authors.lastname: Luo
authors.affiliation: Department of Breast Surgery, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China.
authors.forename: Shouman
authors.initials: S
authors.lastname: Wang
authors.affiliation: Department of Breast Surgery, Xiangya Hospital, Central South University, Changsha, Hunan, 410008, China.
authors.forename: Zhi
authors.initials: Z
authors.lastname: Xiao
authors.affiliation: Sanway Gene Technology Inc., Changsha, Hunan, 410205, China.|Research Center for Technologies in Nucleic Acid-Based Diagnostics, Changsha, Hunan, 410205, China.|Research Center for Technologies in Nucleic Acid-Based Diagnostics and Therapeutics, Changsha, Hunan, 410205, China.
authors.forename: Zhongping
authors.initials: Z
authors.lastname: Deng
authors.affiliation: Sanway Gene Technology Inc., Changsha, Hunan, 410205, China.|Research Center for Technologies in Nucleic Acid-Based Diagnostics, Changsha, Hunan, 410205, China.|Research Center for Technologies in Nucleic Acid-Based Diagnostics and Therapeutics, Changsha, Hunan, 410205, China.
authors.forename: Lizhong
authors.initials: L
authors.lastname: Dai
authors.affiliation: The State Key Laboratory of Medical Genetics and School of Life Sciences, Central South University, 172 Tongzipo Road, Changsha, Hunan, 410013, China.
authors.forename: Kun
authors.initials: K
authors.lastname: Xia
authors.affiliation: The State Key Laboratory of Medical Genetics and School of Life Sciences, Central South University, 172 Tongzipo Road, Changsha, Hunan, 410013, China. junwang@csu.edu.cn.
authors.forename: Jun
authors.initials: J
authors.lastname: Wang
mesh_terms: D014408:Biomarkers, Tumor / Q000097:blood* / Q000235:genetics*; D001943:Breast Neoplasms / Q000175:diagnosis* / Q000235:genetics; D016022:Case-Control Studies; D019175:DNA Methylation; D004273:DNA, Neoplasm / Q000097:blood / Q000235:genetics; D055088:Early Detection of Cancer; D005260:Female; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D008875:Middle Aged; D009367:Neoplasm Staging; D011379:Prognosis; D013447:Sulfites / Q000737:chemistry*
chemical_list: D014408:Biomarkers, Tumor; D004273:DNA, Neoplasm; D013447:Sulfites; C042345:hydrogen sulfite
keywords: Breast cancer; Cell-free DNA; Methylation; Microfludic PCR; Next-generation sequencing; Plasma