BCR-ABL oncoprotein stimulates cell proliferation and inhibits apoptosis in chronic myeloid leukemia (CML). For cure, imatinib is a widely used tyrosine kinase inhibitor, but developing chemotherapeutic resistance has to be overcome. In this study, we aimed to determine differing genome-wide microRNA (miRNA) and messenger RNA (mRNA) expression profiles in imatinib resistant (K562/IMA-3 μM) and parental cells by targeting STAT5A via small interfering RNA (siRNA) applications. After determining possible therapeutic miRNAs, we aimed to check their effects upon cell viability and proliferation, apoptosis, and find a possible miRNA::mRNA interaction to discover the molecular basis of imatinib resistance. We detected that miR-2278 and miR-1245b-3p were most significantly regulated miRNAs according to miRNome array. Upregulating miR-2278 expression resulted in the inhibition of resistant leukemic cell proliferation and induced apoptosis, whereas miR-1245b-3p did not exhibit therapeutic results. Functional analyses indicated that AKT2, STAM2, and STAT5A mRNAs were functional targets for miR-2278 as mimic transfection decreased their expressions both at transcriptional and translational level, thus highlighting miR-2278 as a tumor suppressor. This study provides new insights in discovering the mechanism of imatinib resistance due to upregulating the tumor-suppressor hsa-miR-2278 which stands for a functional therapeutic approach, inhibited leukemic cell proliferation, induced apoptosis, and regain of chemotherapeutic drug response in CML therapy.

To determine the gene expression profiles of the JAK/STAT pathway members STAT3, STAT5A, STAT5B at both mRNA and protein levels in HL-60 and K-562 leukemia cells that were undergoing apoptosis following high-dose methylprednisolone (MP) treatment. HL-60 cells were treated with 0.1 mM MP and K-562 cells were treated with 0.4 mM MP according to their IC(50) values. STAT3, STAT5A, and STAT5B mRNA relative expression levels were determined by qRT-PCR whereas the protein levels were detected via western-blot analysis and apoptosis was evaluated by Annexin V method. A significant decrease was seen in STAT5A mRNA relative expression level at 48 hours of MP treatment (P < 0.05) both in HL-60 and K-562 cells. Other STATs showed a lower downregulation in their relative expressions at 48 hours at mRNA level for both of the cell lines. STAT proteins showed no expression change in K-562 cells in time course experiments but while STAT5A expression was downregulated; STAT5B showed an increase at 96 hours in HL-60 cells. Apoptosis was triggered by high-dose MP treatment that was evaluated by fluorescent microscopy. The JAK/STAT pathway components may play an important role in the apoptosis mechanism of leukemic cells under MP treatment in HL-60 and K-562 cells. Other pathways may also be involved with a post-translational modification seen in the HL-60 cell line, with both upregulation and downregulation of protein expression levels of STAT5B and STAT5A, respectively.

This study is the first to investigate the therapeutic potential of a mono/combination treatment with the selective phase III BCR::ABL1 tyrosine kinase inhibitor (TKI) asciminib (ABL001) and the c-MET inhibitor tivantinib (ARQ197) in hematopoietic stem cells (HSCs) and leukaemia stem cells (LSCs) in terms of cytotoxicity, apoptosis, target gene expression profiles, and bioinformatics in vitro. In silico studies were also performed for various tivantinib and asciminib derivatives. Asciminib and tivantinib exhibited significant antileukaemic effects, with tivantinib showing the strongest apoptotic impact in LSCs and asciminib in HSCs. Although their combination exhibited an additive effect on LSCs, enabling dose reduction and potentially minimizing side effects; the treatment succeeded in promoting apoptosis as effectively as tivatinib monotherapy. Gene expression analysis showed increased pro-apoptotic and tumour suppressor markers, whereas decreased oncogenes' expressions following mono/combined treatment in LSCs. Molecular docking showed that a new analogue of tivantinib (compound-1) and CHEMBL5274046 are promising c-MET and ABL1 inhibitors, respectively. The findings highlight the antileukaemic potential of initially tivantinib and asciminib in LSCs. Despite an additive effect, combination therapy allowed dose reduction, giving rise to a potential decrease in side effects. Targeting c-MET, alone or in combination with BCR::ABL1 inhibition, presents a compelling strategy for eradicating LSCs, which underpin resistance and relapse in CML therapy - thus marking a crucial advancement in the pursuit of more effective treatment paradigms. In silico studies indicated that various tivantinib and asciminib analogues could also have potential therapeutic effects as c-MET and ABL1 inhibitors for future anti-leukaemia research.