FLT3 Is Up-Regulated in Blast Crisis of Chronic Myeloid Leukemia and Combination of Small Interference RNA of FLT3 and Gleevec (STI571) Synergistically Induces the Apoptosis in K562 Cells.

To identify a gene signature for prognostic markers at transition from chronic phase to blast crisis of chronic myeloid leukemia (CML), we have applied Affymetrix Genechips of 22,000 transcripts to analyze total RNA of CML cells from 12 patients with chronic phase and 12 patients with blast crisis. Data analysis using GeneSpring 6.0 generated a list of 143 differentially expressed genes. A total of 89 genes were up-regulated and 54 genes were down-regulated in blast crisis of CML, and vice versa in chronic phase of CML. Array data for 32 genes was validated using quantitative realtime PCR analysis. The expression levels of HSA6591, FLT3, NTE5, RSG1, LAF4, CPA3, ATF, FCGR3A, MYD88, IFIT1, TP73L, DTNA, MDA, and IL18R1 showed statistically significant difference (p < 0.05) between chronic phase and blast crisis. Since CML cells of blast crisis were generally unresponsive to STI571, we further analyzed roles of FLT3 which is known to be a poor prognositic marker in acute myeloid leukemia. For this experiment, K562 cells (CML blast cells) were transfected with small hairpin RNAs (shRNAs), also referred to as small interfering RNAs, to target human FLT3, resulting in the significant inhibition of FLT3 expression at mRNA and protein levels. MTT assay demonstrated that FLT3 knockdown K562 cells by shRNAs were more sensitive to STI571 compared to wild type of K562, although there was no difference at high concentration of STI571 (320 nM) between FLT3 knockdown K562 cells and wild type of K562 cells. The higher expression levels of apoptosis related genes (PARP, caspase-3, Bax) were observed in FLT3 knockdown K562 cells compared to wild type of K562 cells. Thus, RNA interference-directed targeting of FLT3 might be a novel treatment modality in STI571 refractory CML patients.