DNA methyltransferases (DNMTs) are important enzymes involved in epigenetic control of gene expression and represent valuable targets in cancer chemotherapy. A number of nucleoside DNMT inhibitors (DNMTi) have been studied in cancer including cancer stem cells, and two of them (azacytidine and decitabine) have been approved for treatment of myelodysplastic syndromes. However, only few non-nucleoside DNMTi have been identified so far, and much less validated in cancer. Through a process of hit-to-lead optimization, we report here the discovery of compound 5 as potent non-nucleoside DNMTi, and selective towards other AdoMet-dependent protein methyltransferases. Compound 5 was potent at single-digit µM against a panel of cancer cells, and less toxic in peripheral blood mononuclear cells. In mouse medulloblastoma stem cells 5 inhibited cell growth while the related compound 2 showed high cell differentiation. To the best of our knowledge, 2 and 5 are the first non-nucleoside DNMTi tested in a cancer stem cell line.