BackgroundDNA methylation changes, frequent early events in cancer, can modulate the binding of transcription factors. RE1-silencing transcription factor (REST) plays a fundamental role in regulating the expression of neuronal genes, and in particular their silencing in non-neuronal tissues, by inducing chromatin modifications, including DNA methylation changes, not only in the proximity of its binding sites but also in the flanking regions. REST has been found aberrantly expressed in brain cancer and other cancer types. In this work, we investigated DNA methylation alterations at REST binding sites and their flanking regions in a brain cancer (pilocytic astrocytoma), two gastrointestinal tumours (colorectal cancer and biliary tract cancer) and a blood cancer (chronic lymphocytic leukemia).ResultsDifferential methylation analyses focused on REST binding sites and their flanking regions were conducted between tumour and normal samples from our experimental datasets analysed by Illumina microarrays and the identified alterations were validated using publicly available datasets. We discovered distinct DNA methylation patterns between pilocytic astrocytoma and the other cancer types in agreement with the opposite oncogenic and tumour suppressive role of REST in glioma and non-brain tumours.ConclusionsOur results suggest that these DNA methylation alterations in cancer may be associated with REST dysfunction opening the enthusiastic possibility to develop novel therapeutic interventions based on the modulation of this master regulator in order to restore the aberrant methylation of its target regions into a normal status.Supplementary InformationThe online version contains supplementary material available at 10.1186/s13148-023-01514-9.