EMT TARGETING IN CML BLAST CRISIS K562 CELLS BY VITAMIN E: BLOCKING EMT-SNAIL TRANSCRIPTION FACTOR AND UNBLOCKING MYELOID MASTER REGULATOR CEBP ? TRANSCRIPTION FACTOR

L. P. Shvachko*

ABSTRACT

Chronic myeloid leukemia (CML) is a clonal hematopoietic stem cell disorder associated with the activity of BCR-ABL fusion oncogene due to the reciprocal translocation t(9;22)(q34;q11). The persistence of leukemic stem cells (LSCs) remains a major obstacle to cure CML. Epithelial mesenchymal transition (EMT) mechanism is known to contribute to LSC tumor progression. Although EMT has been studied in relation to epithelium-derived tumors, there is increasing evidence implicating the involvement of EMT activators in hematopoietic malignancies. The expression of some EMT modulators has been demonstrated in Ph+ leukemia cells. EMT-inducer Snai1 has the most important role is in maintaining stemness properties in tumor progression. Nevertheless, the role of Snai1 in hematopoiesis and leukemia remains obscure. Earlier, we revealed that alpha-tocopherol might be an effective inducer of mRNA CEBP alpha in CML blast crisis K562 cells in vitro (Shvachko L.P., et al., 2018). The aim: of present study is to determine the relationship between Snai1-EMT suppression and restored CEBP alpha myeloid differentiation potential in CML cells induced by vitamin E (alpha-tocopherol). Materials and Methods: K562 cells were exposed to alpha-tocopherol (100 μM) or metformin (4 mM) for 48 h. RNA was extracted and converted to cDNA; RT-PCR reactions were carried out according to SYBR Green protocol detection using HotStarTaq DNA polymerase with primers for Snai1 and C/EBPalpha genes. The gene expression was quantified using 2-ΔCt method with normalization to mRNA expression of GAPDH gene. Results: We have found highly detectable Snai1 mRNA expression and down-regulated CEBP alpha in K562 cells. Vitamin E, alpha-tocopherol, targeted suppressed Snail mRNA expression. Such suppression of Snail inversely is correlated with restored CEBP alpha mRNA expression by vitamin E and enhanced by metformin pointing to the possible synergistic effect with vitamin E effect. Conclusion: We observed that several modulators of gene expression affect Snail1 and CEBP alpha mRNA expression in K562 cells in different directions. In particular, vitamin E (alpha-tocopherol) down-regulates Snail1 and up-regulates CEBP alpha. One could suggest the causal relationship between Snai1 suppression and restoration of CEBP alpha expression that seems to contribute to recover myeloid differentiation potential of CML blast cells. From one side, this study have suggested that Snai1 EMT-inducer contribute to pathogenesis of CML blast crisis. On the other hand we revealed that vitamin E can restore normal HSC myelopoietic function during SNAIL repression. Therefore, we have proposed that vitamin E - alpha-tocopherol can be used in anti-EMT stemnes therapy for clinical CML blast crisis progression with LSC phenotype which recently poor resolving.

Keywords: .