Single CpG hypermethylation, allele methylation errors, and decreased expression of multiple tumor suppressor genes in normal body cells of mutation‐negative early‐onset and high‐risk breast cancer patients

Abstract

To evaluate the role of constitutive epigenetic changes in normal body cells of BRCA1/BRCA2‐mutation negative patients, we have developed a deep bisulfite sequencing assay targeting the promoter regions of 8 tumor suppressor (TS) genes (BRCA1, BRCA2, RAD51C, ATM, PTEN, TP53, MLH1, RB1) and the estrogene receptor gene (ESR1), which plays a role in tumor progression. We analyzed blood samples of two breast cancer (BC) cohorts with early onset (EO) and high risk (HR) for a heterozygous mutation, respectively, along with age‐matched controls. Methylation analysis of up to 50,000 individual DNA molecules per gene and sample allowed quantification of epimutations (alleles with >50% methylated CpGs), which are associated with epigenetic silencing. Compared to ESR1, which is representative for an average promoter, TS genes were characterized by a very low (< 1%) average methylation level and a very low mean epimutation rate (EMR; < 0.0001% to 0.1%). With exception of BRCA1, which showed an increased EMR in BC (0.31% vs. 0.06%), there was no significant difference between patients and controls. One of 36 HR BC patients exhibited a dramatically increased EMR (14.7%) in BRCA1, consistent with a disease‐causing epimutation. Approximately one third (15 of 44) EO BC patients exhibited increased rates of single CpG methylation errors in multiple TS genes. Both EO and HR BC patients exhibited global underexpression of blood TS genes. We propose that epigenetic abnormalities in normal body cells are indicative of disturbed mechanisms for maintaining low methylation and appropriate expression levels and may be associated with an increased BC risk.

Hypermethylation of RAD9A intron 2 in childhood cancer patients, leukemia and tumor cell lines suggest a role for oncogenic transformation

“…Alleles with > 50% aberrantly (de)methylated CpGs in DBS are considered as functionally relevant epimutations. Consistent with an epimutation screen in breast cancer susceptibility genes [3], we considered EMRs > 1% as elevated and EMRs > 2.5% as likely pathogenic constitutive epimutations. Using the above-described classi cation, we did not detect any APC epimutations in broblasts of patient 1N08.…”

“…Next-generation sequencing (NGS) libraries for DBS were generated as described previously [3]. PCR ampli cation of APC, CDKN2A, TP53, and RAD9A was performed using primers containing a target-speci c part and partial adapter overhangs (supplementary Table 2).…”

“…In contrast, most hereditary forms of cancer are caused by germline mutations in tumor suppressor genes, predisposing patients to tumor development, which itself is triggered by inactivation of the second TS allele. Accumulating evidence suggests that similar to germline mutations, constitutive epimutations involving soma-wide hypermethylation of tumor suppressor genes in normal body cells, can cause phenocopies of cancer syndromes such as hereditary non-polyposis colon cancer (HNPCC) as well as breast-and ovarian cancer [3,4].…”

Big Data in 5G Distributed Applications

“…One of these communities is that of physicians involved in the diagnosis of genetic diseases and using the Gen-searchNGS tool [28]. This Java software analyzes next-generation sequencing data (NGS) to detect changes in DNA sequences for the diagnosis of genetic diseases [29,30]. In order to be able to easily integrate into any Java software, including Gen-searchNGS, TFC capabilities the POP-Java tool [31,32] was used.…”

Association between the Methylation of CpG Islands in JAK-STAT Pathway-Related Genes and Colorectal Cancer

“…Methylation of peripheral blood-derived DNA is related to the occurrence of tumors, and individual differences in methylation can re ect an individual's susceptibility to tumors [17,18]. In addition, it is reported that the effect of environmental factors on methylation status also found in peripheral blood and environmental factors contribute to CRC risk.…”