Combination of Imatinib and Hypusination Inhibitors Represents a Novel Therapeutic Strategy in Bcr-Abl Positive Leukemias.

Abstract

In order to identify new cellular Bcr-Abl downstream targets, we have recently detected differential regulation of eukaryotic initiation factor 5A (eIF-5A) in Bcr-Abl positive cells. EIF-5A represents the only known eukaryotic protein activated by post-translational hypusination. Hypusination inhibitors (HI) GC7 and ciclopirox alone exerted an anti-proliferative effect on Bcr-Abl positive (K562, BA/F3p210) and negative human leukemia cell lines (HL-60, BA/F3) in vitro. No direct effect of HI treatment on phosphorylation of Bcr-Abl downstream targets (global p-tyrosination, p-CRKL, p-STAT5) was observed. However, the synergistic dose-effect relationship for apoptosis induction, cytotoxicity and inhibition of proliferation found for the combination of Imatinib and HI was restricted to Bcr-Abl-positive cells. Furthermore, this synergistic effect was confirmed by cytotoxicity assays, cell cycle analysis and CFSE-labeling of primary CD34+ CML cells in comparison to CD34+ cells from healthy donors. In parallel to these findings, eIF-5A mRNA is expressed at clearly (if significant then say significant and add p-value, otherwise delete clearly) elevated levels in PBMCs from CML patients at different disease stages when compared with PBMCs from healthy donors. Specificity of this effect could be demonstrated by co-treatment of K562 cells with Imatinib and a stable lentiviral siRNA against eIF-5A. Based on this observation, we observed a strong antiprolifrative effect of HI as single agents on BA/F3p210 cells, including the Imatinib-resistant mutants T315I, M351T or E255K. Furthermore, a combination of Imatinib and HI had a synergistic effect in inducing cytotoxicity, apoptosis and inhibition of proliferation in leukemic cell lines with moderate resistance to imatinib (M351T). In contrast, in BA/F3p210T315I (T315I) cells, which are resistant to Imatinib as a single agent, the co-treatment of Imatinib and HI did not lead to additive or synergistic effects. In conclusion, we have identified inhibition of eIF-5A hypusination as a promising new approach for single agent or combination therapy in Bcr-Abl