This study aimed to investigate the efficacy and mechanism of decitabine (DAC) and all-trans retinoic acid (ATRA) in elderly acute myeloid leukemia (AML) patients and cultured cells. Our clinical trial enrolled 36 elderly patients who were judged ineligible for conventional chemotherapy, receiving DAC and ATRA regimen (DAC 20 mg/m² days 1–5; ATRA 20 mg/m² days 4–28 in the first cycle and days 1–28 in the subsequent cycle). Treated with a median of 3 cycles (range 1–6), 44.4 % of patients achieved complete remission (CR), 11.1 % achieved CR with incomplete peripheral count recovery (CRi) and 13.9 % achieved partial remission (PR). The median overall survival (OS) was 12.1 months; the 1-year and 2-year OS rates were 49.6 % and 17.2 %. In addition, our in vitro studies indicated that the antineoplastic activities of DAC and ATRA mutually reinforced, which induced growth inhibition, cell cycle arrest and apoptosis of AML cells. Meanwhile, we found DAC and ATRA inhibited DNMT1, activated miR-34a via promoter hypomethylation, down-regulated its target MYCN and thus exerted a synergistic antineoplastic effect. In conclusion, DAC plus ATRA regimen might be effective and well-tolerated for elderly patients partially through modulating miR-34a/MYCN axis.