| dc.description.abstract | Immunotherapy and epigenetic therapy provide an alternative approach in leukemia treatment. The objective of this study was to investigate the effect of HDACi Bel in combination with HMTi DZNep and RA in vitro and in vivo using human APL cells NB4 and HL-60 and immunodeficient NOG mice bearing APL xenografts. In parallel, the effect of immunization with WT1 peptide was also evaluated. For in vitro studies certain gene and protein expression analysis, also ChIP-seq with HyperAc H4 were performed. In vivo APL model was established by NB4 cells injection into NOG mice. Peripheral blood (PB) counts, certain gene and protein expression in mice PB, organs and tumors were analyzed. Our in vitro studies demonstrated that in NB4 and HL-60 cells treatment with Belinostat and DZNep together with RA induced gene expression of granulocytic differentiation associated genes (C/EBPα, C/EBPε, PPAR and miR-223). Changes in gene and protein expression of epigenetic modifiers (PRC2, HDAC1, HDAC2), as well as in associated modifications (H3 K27me3, H3 K9me2, HyperAc H4) were also detected. ChIP-seq analysis revealed that used treatments induced HyperAc histone H4 association with genes involved in various processes: for example, the induction of apoptosis (TNFRSF14, LMTK3, PCBP1), immune response (ETV6, GAB2, CHUK), macrophage activation (COL4A2, COL11A2, BPI), epigenetic modifications (HDAC4, EPC1, ASH2L, SCMH1), etc. Our in vivo study showed that treatment with the inhibitors of epigenetic modifiers as well as with WT1 peptide prolonged APL xenografted mice survival, though the effect was more pronounced upon immunotherapy approach. In addition, differences in gene expression as well as in PB cell surface markers and PB cell counts were detected in untreated mice and mice treated with either Bel+DZNep+RA or with WT1 peptide. Furthermore, the presence of oncomarker WT1 in leukemia NB4 cells and in tumors of APL xenografted mice were demonstrated and the decrease in WT1 protein level in differentiating NB4 cells and cured APL xenografted mice were shown. In addition, the activity of epigenetic markers (HyperAc H4, H3 K4me3, H3 K9me3, H3 K27me3) was found to be different in treated and untreated APL xenografted mice as well as being dependent on the tissue specificity and functional activity. Summarizing, our pre-clinical studies showed that epigenetic and immunotherapy agents used in this study have a potential in cancer treatment. | eng |
