Folate, one of the most studied dietary compounds, has recently become the main topic of debates on food fortification. Although low folate levels may be associated with increased risk of cancer development, simultaneously several reports indicate a detrimental effects mediated by high folate concentrations. Using the methylation sensitive restriction analysis (MSRA) and real-time RT-PCR we tested the effect of folic acid on DNA promoter methylation and expression of PTEN, APC and RARbeta2 tumour suppressor genes in MCF-7 and MDA-MB-231 breast cancer cell lines with different invasive capacity. The tested genes encode proteins involved in regulation of oncogenic intracellular signaling pathways. The results show that the increasing concentrations of folic acid lead to a dose-dependent down-regulation of tumour suppressor genes which may be linked to the increased DNA methylation detected within their promoter regions. The effects were more remarkable in non-invasive MCF-7 cells where we also observed 30% up-regulation of DNMT1 expression at the highest folate concentration used. Our findings show that caution need to be used when introducing folic acid supplementation since it may lead to cancer progression. (C) 2012 Elsevier Inc. All rights reserved.