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Biomarker Detects Fatal Breast Cancer One Year Earlier

By LabMedica International staff writers
Posted on 03 Jan 2018
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Image: The High Sensitivity Large Fragment Analysis Kit is used for the automated sizing and quantification of large DNA fragments and smears (Photo courtesy of Advanced Analytical Technologies).
Image: The High Sensitivity Large Fragment Analysis Kit is used for the automated sizing and quantification of large DNA fragments and smears (Photo courtesy of Advanced Analytical Technologies).
DNA methylation is the addition of a methyl group to DNA, which often affects gene expression. Aberrant DNA methylation is common in human tumors and methylation changes occur very early in breast cancer development.

Over the last decade, DNA methylation (DNAme) has been shown to be a hallmark of cancer and occurs very early in breast cancer (BC) development. DNAme is centered on specific regions (CpG islands) and is chemically and biologically stable. This enables the development of early detection tools and personalized treatment, based upon the analysis of cell-free DNA contained within serum or plasma.

An international team of scientists working with experts at the Elizabeth Garrett Anderson Institute for Women’s Health (London, UK) used a total of 31 tissues and 1,869 serum samples in their study. In Phase 1, they analyzed breast cancer tissue and white blood cells (WBCs) in order to identify breast cancer specific DNAme markers. In Phase 2, they established serum DNAme assays using serum sets collected from women attending hospitals in London, Munich, and Prague. Phase 3 was initiated to validate the top marker performance by using serum samples from two large clinical studies.

DNA was isolated from tissue and serum samples; Serum DNA was quantified using the Agilent Fragment Analyzer and the High Sensitivity Large Fragment Analysis Kit and tissue DNA was quantified using NanoDrop and Qubit, and the size was assessed by agarose gel electrophoresis. Genome-wide methylation analysis was performed by reduced representation bisulfite sequencing (RRBS). The digested DNA was adapter-ligated, bisulfite-modified, and polymerase chain reaction (PCR)-amplified. The libraries were sequenced on a HiSeq 2500.

The authors first analyzed the biomarker EFC#93 DNA methylation in blood serum samples from 419 breast cancer patients taken at two time points: after surgery (before the start of chemotherapy), and after completion of chemotherapy. They demonstrated that aberrant DNA methylation in samples taken before chemotherapy was a marker for poor prognosis independent of the presence of circulating tumor cells (cells that have shed from the primary tumor into the blood or lymphatic system and circulate throughout the body).

To assess whether EFC#93 can diagnose women with a poor prognosis earlier (that is, before the cancer becomes detectable) the authors further analyzed serum samples of 925 healthy women, 229 of whom went on to develop fatal and 231 of whom went on to develop non-fatal breast cancer, within the first three years of donating serum samples.

Martin Widschwendter, MD, FRCOG, a professor of Woman’s Cancer and lead author of the study, said, “We found that the presence of EFC#93 DNA methylation in blood serum correctly identified 43% of women who went on to be diagnosed with fatal breast cancer within three to six months of giving serum samples, as well as 25% of women who went on to be diagnosed within six to twelve months of giving samples.” The study was published on December 22, 2017, in the journal Genome Medicine.

Related Links:
Elizabeth Garrett Anderson Institute for Women’s Health

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