Autophagy and glycogen synthase kinase-3 (GSK3) as molecular targets capable of increasing the activity of drugs used in the treatment of myeloid leukemias (acute and chronic)

Abstract

Although autophagy is well known as a physiological process in which the cellular components are degraded by lysosomes, current studies indicate that autophagy can also suppress or increase survival of tumor cells. In leukemia's, autophagy is generally unregulated. In addition, via GSK3, which seems to have a direct relationship with autophagy has been implicated as an important modulator of cell growth MLL leukemia, a subtype of acute leukemia extremely aggressive and of poor prognosis. Therefore, this project aims to understand autophagy and participation of GSK3 signaling pathway in death and/or survival of leukemia cells. For these purposes, using four cell lines with specific molecular features such as HL60 (promyelocytic-like), NB4 (myelocytic leukemia pro-) chronic myeloid leukemia K562 and Lucena (K-562 resistant to vincristine), we intend to evaluate how the modulation of autophagy and GSK-3 pathway alters the response of the first line to cytosine arabinoside, the second line to retinoic acid (ATRA) and the last two lines to imatinib mesylate (IM) and nilotinib. These studies will be performed by testing the cytotoxicity, cell death (labeling with annexin V/PI), cell cycle (using PI), proteins involved in apoptosis and autophagy such as LC3II/I ratio, 1 beclin-1, p62, p53 and Bcl2/Bax by immunoblotting. The methodology will also be used to assess total and phosphorylated GSK3 protein. The cell differentiation will be studied using antibodies specific for mature and progenitor cells such as CD34+ and CD11b+, respectively by flow cytometry. Cell proliferation will be evaluated by clonal culture in presence of specific growth factors to myeloid lineage as GM-CSF. Furthermore, we will evaluate the involvement of caspase-3 in ours groups of studied. We hopefully, with this work, improve a little more knowledge of the impact of autophagy pathway and GSK-3 in the treatment of leukemia focusing the development of new strategies therapeutic. (AU)