Down Regulation of TOPK, PBX3, SRPK, DDX21 and CLC Genes in Newly Diagnosed, Early-Chronic Phase Chronic Myeloid Leukemia Patients Treated with Imatinib Mesylate.

Background: Due to the BCR/ABL chimeric protein expression in chronic myeloid leukemia (CML), several genes and signaling pathways have been reported to be activated. These alterations in gene expression contribute to the pathophysiology of p210^BCR/ABL-mediated transformation. Some of the overexpressed genes are implicated in cellular processes known to be disturbed in CML, including the mitogen-activated protein kinase or the ubiquitin phatway, whereas overexpression of other genes may implicate new cellular pathways involved in CML. The characterization of new genes which could be implicated in the pathophysiology of CML, may lead to the identification of potentially novel therapeutic targets for CML. In a previous study we adopted a microarray analysis to study the gene expression profiling of CD34+ cells taken from 5 de novo CML patients and treated in vitro with Imatinib (data not shown). Five genes had shown a greater down-regulation and a possible involvement in the in vitro effects of Imatinib.

Aims: Aim of the present study was to confirm these data in vivo through a quantitative real time PCR analysis in 10 CML patients enrolled in a multicenter clinical trial of the GIMEMA CML working party (newly diagnosed, early-CP CML patients treated with Imatinib 400mg/die and peghilated interferon). Five patients were known to have chromosome 9 deletion.

Methods: this study was performed on 10 bone marrow samples from 10 consecutive patients enrolled in the protocol. Bone marrow bone was collected prior to treatment (baseline) and after 3 and 6 months of Imatinib therapy. Five genes (TOPK, PBX3, SRPK, DDX21 and CLC) were analyzed at the RNA levels by means of a quantitative RT-PCR analysis (TaqMan). β2 microglobuline was used as control gene. Results were expressed as a median ΔCt value. Expression of these 5 genes before treatment with Imatinib and after 6 months of therapy was compared. Statistical significance was tested by the student’s T-Test.

Results: Quantitative RT-PCR analysis confirmed that the 5 tested genes were downregulated after 6 months of treatment with Imatinib. About 1 log reduction was observed. The differences between the baseline median values and median values after 6 months of treatment were statistically significant (p>0.005). We were not able identify any differences between patients who showed chromosome 9 deletion and patients who did not.

Conclusions: treatment of CML patients with the ABL-specific tyrosine kinase inhibitor Imatinib decreased expression at the mRNA level of all the genes tested. This suggests that increased gene expression could be in some case tyrosine-kinase dependent and it could be implicated in the pathophisiology of CML. Study supported by: COFIN 2003 (M. Baccarani), AIRC, AIL, Fondazione del Monte di Bologna e Ravenna, FIRB 2001.