Breast cancer (BC) heterogeneity calls for molecular subtyping that would assist in personalized treatment. An advantage of DNA methylation markers is that their detection in tumors is not compromised by the presence of normal tissues. With a perspective to develop methylation-based BC diagnostic markers, we have performed a genome-wide DNA methylation profiling of a collection of breast tissues and cell lines.
XmaI-RRBS method was used to profile DNA methylation of 170 BC samples obtained before chemotherapy, six BC cell lines, and 10 normal breast autopsy specimens. Unsupervised hierarchical cluster analysis was used to discern intrinsic DNA methylation BC subtypes; clustering uncertainty was assessed with pvclust R package using bootstrap permutation approach.
We have identified 10 epigenetic BC subtypes different in the DNA methylation profiles. Of these, BC cell lines constitute a separate extremely high methylated subtype clustering far from any tissues assessed. In turn, BC tissues are classified into two major epigenetic subtypes, high- and low-methylated at the promoter regions of genes. We identified 114 genes that distinguish between high- and low-methylated BC subtypes. Noteworthy are the genes of adenylate cyclases ADCY4, ADCY8 and adenylate cyclase stimulants ADORA2B, ADCYAP1; proteins of cell adhesion and extracellular matrix (CDH4, NRXN2, MXRA5, COMP, integrins A8 & A11, ADAM19; potassium channels (KCNH8, KCNJ2, KCNG1, KCNK10, KCNK17, ATP1A3). More than a third among differentially methylated are homeobox genes (VAX2, TLX3, GSX1, IRX1, FOXC2, FOXE3, NKX6-2, VSX1, SOX21, POU4F1) and genes encoding proteins involved in early development and morphogenesis (ZIC1, SPOCK2, DPYSL3, ATOH1, ITGA8). Expectedly, there is no statistically significant difference in methylation of the classical tumor suppressor genes between epigenetic subtypes of BC, as their abnormal methylation is ubiquitous in cancers and thus non-discriminative between tumor types.
Intrinsically epigenetically heterogeneous BC may be classified into a reasonable number of DNA methylation subtypes, promising the discovery of new diagnostic and prognostic markers, as well as of new therapeutic targets.
Clinical trial identification
Legal entity responsible for the study
Research Centre for Medical Genetics.
Russian Science Foundation (project No.18-15-00430).
All authors have declared no conflicts of interest.