Suppression of Programmed Cell Death 4 (PDCD4) Protein Expression by BCR-ABL-regulated Engagement of the mTOR/p70 S6 Kinase Pathway

Abstract

There is accumulating evidence that mammalian target of rapamycin (mTOR)-activated pathways play important roles in cell growth and survival of BCR-ABL-transformed cells. We have previously shown that the mTOR/p70 S6 kinase (p70 S6K) pathway is constitutively activated in BCR-ABL transformed cells and that inhibition of BCR-ABL kinase activity by imatinib mesylate abrogates such activation. We now provide evidence for the existence of a novel regulatory mechanism by which BCR-ABL promotes cell proliferation, involving p70 S6K-mediated suppression of expression of programmed cell death 4 (PDCD4), a tumor suppressor protein that acts as an inhibitor of cap-dependent translation by blocking the translation initiation factor eIF4A. Our data also establish that second generation BCR-ABL kinase inhibitors block activation of p70 S6K and downstream engagement of the S6 ribosomal protein in BCR-ABL transformed cells. Moreover, PDCD4 protein expression is up-regulated by inhibition of the BCR-ABL kinase in K562 cells and BaF3/BCR-ABL transfectants, suggesting a mechanism for the generation of the proapoptotic effects of such inhibitors. Knockdown of PDCD4 expression results in reversal of the suppressive effects of nilotinib and imatinib mesylate on leukemic progenitor colony formation, suggesting an important role for this protein in the generation of antileukemic responses. Altogether, our studies identify a novel mechanism by which BCR-ABL may promote leukemic cell growth, involving sequential engagement of the mTOR/p70 S6K pathway and downstream suppression of PDCD4 expression. Chronic myeloid leukemia (CML)2 is a clonal myeloproliferative disorder, whose hallmark is the presence of the BCR-ABL oncoprotein, which results from the abnormal bcr-abl fusion oncogene (1-3). bcr-abl is created by a reciprocal translocation involving chromosomes 9 and 22, and its abnormal protein product plays a key and essential role in the pathogenesis of CML (1-4). The constitutive tyrosine kinase activity of BCR-ABL mediates phosphorylation of multiple downstream substrates and engagement of various mitogenic pathways that promote cell growth and survival (1-7). The introduction of imatinib mesylate in the management of chronic myelogenous leukemia has had a dramatic impact on the natural history of the disease (reviewed in Refs. 8 -11). This agent induces long lasting hematologic and cytogenetic responses in patients with early and late phase CML (8, 12, 13, 14, 16 -18) by its ability to block activation the BCR-ABL kinase and generation of mitogenic responses (8 -11).Although the introduction of imatinib mesylate was a major breakthrough in the management of CML, there has been emerging evidence for resistance to its antileukemic properties in vitro and in vivo (19 -22). Such resistance involves a variety of cellular mechanisms, including mutations of the bcr-abl gene, amplification of the bcr-abl gene locus, and activation of downstream signaling elements, such as Src-kinase-dependent pathways (19,(23)(24)(25)(26)(27)(28). Research ...

Regulatory Effects of Programmed Cell Death 4 (PDCD4) Protein in Interferon (IFN)-Stimulated Gene Expression and Generation of Type I IFN Responses

“…Clonogenic assays in methylcellulose to detect leukemic CFU-blast (CFU-L) colony formation from KT1 cells were performed essentially as previously described (3). The effects of IFN-␣ on CFU-L colony formation from KT1 cells transfected with control siRNA or siRNAs specific for PDCD4 or HA-tagged PDCD4(S67/71A) were determined essentially as previously described (3). Myeloid progenitor (granulocyte-macrophage CFU [CFU-GM]) colony formation from normal CD34 ϩ cells was assessed in clonogenic assays in methylcellulose, as previously described (45).…”

Interferon-Dependent Engagement of Eukaryotic Initiation Factor 4B via S6 Kinase (S6K)- and Ribosomal Protein S6K-Mediated Signals

“…Clonogenic assays in methylcellulose to detect leukemic CFU-blast (CFU-L) colony formation from KT1 cells were performed essentially as previously described (5). Assays for the effects of IFN-␣ on CFU-L colony formation from KT1 cells transfected with control siRNA or siRNAs specific for eIF4B or RSK1 were performed essentially as previously described (5,16,23,33).…”

Antileukemic effects of AMPK activators on BCR-ABL–expressing cells

“…1,2 The constitutive tyrosine kinase activity of BCR-ABL leads to engagement of a plethora of antiapoptotic and proproliferative effector cascades, including PI3K/mammalian target of rapamycin (mTOR) [3][4][5][6][7] and MAPK pathways. 8,9 Inhibition of mTOR and its effectors plays an important role in the generation of the antileukemic effects of BCR-ABL inhibitors, [3][4][5][6][7]10 whereas there is evidence that combinations of rapamycin with imatinib mesylate or nilotinib result in enhanced antileukemic responses. [3][4][5][6][7]10 The better understanding of the structure of distinct complexes formed by mTOR (mTORC1 and mTORC2) and the development of catalytic inhibitors of mTOR have led to efforts to overcome the resistance that BCR-ABL-expressing cells develop in many cases.…”

“…8,9 Inhibition of mTOR and its effectors plays an important role in the generation of the antileukemic effects of BCR-ABL inhibitors, [3][4][5][6][7]10 whereas there is evidence that combinations of rapamycin with imatinib mesylate or nilotinib result in enhanced antileukemic responses. [3][4][5][6][7]10 The better understanding of the structure of distinct complexes formed by mTOR (mTORC1 and mTORC2) and the development of catalytic inhibitors of mTOR have led to efforts to overcome the resistance that BCR-ABL-expressing cells develop in many cases. Two different catalytic inhibitors (PP242 and OSI-027) have been recently shown to block growth of BCR-ABL-expressing cells, including cells with BCR-ABL mutations that are resistant to nilotinib, imatinib, or dasatinib.…”