ABSTRACT
Chronic myeloid leukemia (CML) is a myeloproliferative disorder of hematopoietic stem cells carrying Philadelphia (Ph) chromosome and the oncogenic BCR-ABL1 fusion gene. microRNAs (miRNAs) belong to hematopoiesis transcription regulators and their deregulated expression associates with pathogenesis of CML. The current study assesses and validates expression profiles of selected oncogenic and tumor suppressing miRNAs that are associated with different imatinib mesylate (IM) response in CML patients carrying rare BCR-ABL variants. Microarray analysis has identified different expression of 70 miRNAs (46 up- and 24 down-regulated) when compared IM-resistant with IM-responsive patients carrying Ph chromosome. Significantly up-regulated expression of oncogenic miRNAs (miR-17, miR-18a, miR-20a, miR-21, miR-27a and miR-155) and significantly down-regulated expression of tumor supressing mRNAs (let-7d, miR-205, miR-320, miR-451 and miR-574) in IM-resistant compared to IM-responsive patients was confirmed and validated by qRT-PCR. This study confirms the involvement of the selected oncogenic and tumor suppressing miRNAs in CML pathogenesis and IM response and suggests that these miRNAs could be suitable biomarkers for differential diagnosis of CML patients carrying rare BCR-ABL transcripts, as well as for prediction of their IM response and therapy outcome.
ABSTRACT
The presence of BCR-ABL oncogene mutations in patients with chronic myeloid leukemia (CML) may be responsible for the failure of tyrosine kinase inhibitor (TKI) treatment. The aim of the study was to evaluate the frequency of BCR-ABL gene mutations in patients with CML treated with tyrosine kinase inhibitors. Our lab received 64 samples (34 women, 30 men) from patients with CML who failed or had suboptimal response to TKI treatment. The mutation analysis was performed in 61 patients with CML, 3 patients could not be tested because of inadequate RNA quality. An 866 base pair fragment containing the ABL kinase domain was amplified in a seminested RT (reverse transcriptase)-PCR and then sequenced using Applied Biosystems BigDye Terminator chemistry with two pairs of primers. We analyzed 61 patients with CML, 11 mutations were detected in 13 (21%) patients and SNP (single nucleotide polymorphism) in 6 patients (10%). In addition to 9 point mutations (G250E / F317L, F359V, L387M, Y253H, M388L, M244V, T315I, D276G), 35 bp insertion between exons 8 and 9 and deletion exon 7 were detected. Our results demonstrate that direct sequencing is suitable for routine clinical monitoring patients with CML and may be useful for optimizing therapy.
