Epigenetic changes play a role in the pathogenesis of myeloid malignancies, and hypomethylating agents have shown efficacy in these diseases. We studied the apoptotic effect, genome-wide methylation, and gene expression profiles in HL60 cells following 5-aza-2'-deoxycytidine (decitabine; DAC) treatment, using microarray technologies. Decitabine treatment resulted in a decrease in global DNA methylation, corresponding to 4876 probeset IDs with significantly reduced methylation levels, while the expression of 2583 IDs was modified. The integrated analysis identified 160 genes demethylated and up-regulated by decitabine, mainly including development and differentiation pathway genes. Gene targets of Polycomb group protein regulation were overrepresented in this group. Apoptosis was induced by decitabine, and apoptosis-specific PCR arrays more precisely indicated decitabine-induced up-regulation of 13 apoptosis-related genes, in particular DAP-kinase 1 and BCL2L10. Correspondingly, in primary patient samples, BCL2L10 was hypermethylated in 45% of AML, 43% of therapy-related myeloid neoplasms, 12% of MDS, and in none of the controls. In conclusion, decitabine induces global demethylation and gene expression, in particular of Polycomb target genes involved in development and differentiation pathways. The apoptotic gene BCL2L10 is a frequent target for aberrant promoter methylation in patients with acute leukemia, de novo and therapy-related.