In BCR-ABL-positive cells, STAT-5 tyrosine-phosphorylation integrates signals induced by imatinib mesylate and Ara-C

Abstract

In BCR-ABL-positive cells, the transcription factor STAT-5 is constitutively activated by tyrosine phosphorylation. activation results in upregulation of bcl-X L and increased resistance to induction of apoptosis. Here, we investigated the effects of imatinib mesylate and cytosine arabinoside (Ara-C) on STAT-5 tyrosine-phosphorylation, cellular proliferation and induction of apoptosis in cell lines and primary hematopoietic cells. Imatinib mesylate treatment strongly suppressed STAT-5 tyrosine-phosphorylation in K562 and primary CML blasts. In contrast to JAK-2 and PI-3-kinase inhibition, exposure of K562 cells to imatinib mesylate resulted in obvious suppression of proliferation. Reduced cell growth was due to specific induction of caspase activation followed by apoptotic cell death. In addition, we investigated the effects of Ara-C on STAT-5 tyrosine-phosphorylation. Exposure to Ara-C resulted in significant downregulation of STAT-5 tyrosine-phosphorylation and inhibition of DNA binding. Treatment of K562 cells with Ara-C in combination with imatinib mesylate revealed synergistic effects at the level of STAT-5 tyrosine-phosphorylation and DNA binding, Hck tyrosine-phosphorylation, cell growth and induction of apoptosis. Overall, in this report we demonstrate that STAT-5 tyrosine-phosphorylation is a specific target of imatinib mesylate and Ara-C. Our results suggest that, in combination therapy, inhibition of STAT-5 tyrosine-phosphorylation may be responsible for synergistic or additive effects on BCR-ABL-positive cells.

Novel pathway in Bcr-Abl signal transduction involves Akt-independent, PLC-γ1-driven activation of mTOR/p70S6-kinase pathway

“…The percentage of apoptotic cells was determined by measurement of the fraction with sub-G1 DNA content upon propidium iodide incorporation as described earlier (Kindler et al, 2003;Heidel et al, 2006). Briefly, 4 Â 10 4 cells were treated with various inhibitors for 24 or 48 h, collected and washed with PBS.…”

“…MNCs were isolated by Ficoll density-gradient centrifugation as described earlier (Kindler et al, 2003). For purification of CD34 þ cells, fresh leukapheresis, peripheral blood or bone marrow samples from CML patients or healthy donor were collected.…”

Efficacy and safety of imatinib in adult patients with c-kit–positive acute myeloid leukemia

“…Preparation of cellular lysates was performed as described. 48 Supernatants were shock frozen and stored at Ϫ80°C. Protein concentration was measured by using the Bradford assay.…”

Hematopoietic Stem Cells in Chronic Myeloid Leukemia

“…It competitively binds to the ATP-binding site of BCR.ABL and inhibits protein tyrosine phosphorylation in vitro and in vivo [66]. In vitro studies had shown that Imatinib is capable to inhibit cell proliferation of cell lines expressing Bcr-Abl [67][68][69], effect accomplished through JAK5-STAT and PI3 kinase signaling inhibition [70,71]. It has also been shown that STI571 can inhibit CML MNC, obtained both in chronic phase and blast crisis [71] and reduces the colony forming cells from Mobilized Peripheral Blood (MPB) and Bone Marrow from patients with CML in chronic phase [60].…”