Identification of predictive epigenetic biomarkers in breast cancer

The aim of the project was to investigate the relationship between DNA methylation levels in genes regulating cell growth, invasiveness and metastasis and advanced breast cancers and to evaluate the association between the methylation profiles and inhibition of protein expression. In our study we analyzed the methylation profiles in samples from primary tumours, lymph node metastases, plasma and blood cells from 221 patients in 11 genes using pyrosequencing. Protein expression in 206 patients was evaluated using immunohistochemistry. Invasive breast tumours showed hypermethylation of RASSF1A, APC, CXCL12 and ADAM23 (means 38.98%, 24.84%, 12.04% and 10.01%, respectively). Positive correlations were identified between methylations in tumours and lymph node metastases, but not between plasma and tumours. The cumulative methylation from 11 genes in tumours and lymph node metastases manifested a similar spectrums of methylated genes that indicate the maintaining of aberrant methylation during breast tumourigenesis. No association was observed between the increasing levels of DNA methylation and the decreasing of related protein expression in both tumours and lymph node metastases samples indicating that DNA methylation is not the only mechanism of tumour suppressor gene silencing and loss of relevant protein product. Significantly elevated methylation levels in RASSF1A, APC, CXCL12 and ADAM23 were found in oestrogen receptor positive breast cancers in comparing with oestrogen receptor negative cases; therefore, the evaluation of DNA methylation is more appropriate for testing of oestrogen receptor positive breast cancer patients. The risk for lymph node metastases and higher proliferation of cancer cells measured through Ki-67 expression was increased by hypermethylation of CXCL12 and ADAM23, respectively. Therefore, the quantification of CXCL12 and ADAM23 methylation could be useful markers for monitoring of metastatic potential in breast cancers.