Interferon (IFN)-gamma is the major mediator of anti-tumor immune responses; nevertheless, cancer cells use intrigue strategies to alter IFN-gamma signaling and avoid elimination. Understanding the immune regulatory mechanisms employed by acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) cells upon exposure to IFN-gamma is critical for development of immunotherapy and checkpoint blockade therapy approaches. This study aims to explore the influence of myeloid maturation on IFN-gamma-induced PD-L1 and PD-L2 expression and on pro-leukemogenic transcription factor STAT3 signaling in AML and MDS. Stimulation of myeloid blasts' maturation by all-trans retinoic acid (ATRA) or 1 alpha,25-dihydroxyvitamin D3 (vitamin D) increased the CD11b(+) fraction that expressed PD-1 ligands in response to IFN-gamma. Intriguingly, STAT3 pathway was potently induced by IFN-gamma and strengthened upon prolonged exposure. Nonetheless, STAT3-mediated atypical IFN-gamma signaling appeared as a negligible factor for PD-L1 and PD-L2 expression. These negative influences of IFN-gamma could be alleviated by a small-molecule inhibitor of STAT3, stattic, which also inhibited the upregulation of PD-L1. In conclusion, induction of myeloid maturation enhances the responsiveness of AML and MDS cells to IFN-gamma. However, these malignant myeloid cells can exploit both STAT3 pathway and PD-1 ligands to survive IFN-gamma-mediated immunity and maintain secondary immune resistance.