Activation of mTOR Signaling Pathway Contributes to Tumoral Cell Survival in ALK-Positive Anaplastic Large Cell Lymphoma.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2419-2419
Author(s):  
Francisco Vega ◽  
L. Jeffrey Medeiros ◽  
Coralyn Atwell ◽  
Jeong Hee Cho ◽  
Ling Tian ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Cell Lines ◽  
Anaplastic Large Cell Lymphoma ◽  
Cell Lymphoma ◽  
Large Cell ◽  
Sirna Treatment ◽  
Cycle Arrest ◽  
Large Cell Lymphoma ◽  
Alk Positive

Abstract Anaplastic lymphoma kinase (ALK)-positive anaplastic large cell lymphoma (ALCL) frequently carries the t(2;5)(p23;q35) resulting in aberrant expression of nucleophosmin (NPM)-ALK. Previously, NPM-ALK has been shown to activate phosphatidylinositol 3-kinase (PI3K) and its downstream effector, the serine/threonine kinase AKT. Recently, we have shown that mTOR signaling proteins are activated in ALK-positive ALCL tumors and that mTOR activation depends, at least in part, on activation of AKT (Lab Invest2005; 85: 255A). In this study, we investigate the biological effects of inhibition of mTOR on two ALK-positive ALCL cell lines, Karpas 299 and SU-DHL1. For this purpose, we used rapamycin to inhibit mTOR-raptor complex and mTOR-specific small interfering RNA (siRNA) to silence the endogenous mtor gene. Treatment with rapamycin, resulted in a marked concentration-dependent decrease of phosphorylated (p)-mTOR, and its downstream targets, p-p70S6K, p-S6K, p-4E-BP1 and total eIF4E. Similarly, silencing the expression of mtor resulted in a decrease in the activation/phosphorylation level of these proteins as well as in the level of p-AKT. Both treatments induced apoptosis and cell cycle arrest in both ALK-positive ALCL cell lines as demonstrated by trypan blue exclusion, annexin V staining, BrdU incorporation, and cell cycle studies. There was a concentration-dependent decrease in the anti-apoptotic proteins BCL-2, BCL-XL, MCL-1 and c-FLIP (L and S) with increasing concentrations of rapamycin or after mTOR siRNA treatment. The cyclin dependent kinase inhibitors p21waf1 and p27kip1 and underphosphorylated (Un-p)-RB protein were upregulated, after treatment with rapamycin or after mTOR siRNA treatment. In conclusion, we provide evidence that inhibition of mTOR induces cell cycle arrest and apoptosis in ALK-positive ALCL cells. The decrease of p-AKT by silencing mtor suggests that mTOR is necessary to activate AKT in ALK-positive ALCL, and thus, mTOR can function as a feedback signal activity of its own pathway.

scholarly journals Selective inhibition of STAT3 induces apoptosis and G1 cell cycle arrest in ALK-positive anaplastic large cell lymphoma

Oncogene ◽  
2004 ◽  
Vol 23 (32) ◽  
pp. 5426-5434 ◽  
Author(s):  
Hesham M Amin ◽  
Timothy J McDonnell ◽  
Yupo Ma ◽  
Quan Lin ◽  
Yasushi Fujio ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Anaplastic Large Cell Lymphoma ◽  
Cell Lymphoma ◽  
Large Cell ◽  
Selective Inhibition ◽  
Cycle Arrest ◽  
G1 Cell Cycle Arrest ◽  
Large Cell Lymphoma ◽  
Alk Positive

scholarly journals Autocrine release of interleukin-9 promotes Jak3-dependent survival of ALK+ anaplastic large-cell lymphoma cells

Blood ◽  
2006 ◽  
Vol 108 (7) ◽  
pp. 2407-2415 ◽  
Author(s):  
Lin Qiu ◽  
Raymond Lai ◽  
Quan Lin ◽  
Esther Lau ◽  
David M. Thomazy ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Cell Lines ◽  
Anaplastic Large Cell Lymphoma ◽  
Kinase Activity ◽  
Cell Lymphoma ◽  
Large Cell ◽  
Primary Tumors ◽  
Cycle Arrest ◽  
Large Cell Lymphoma

Abstract The aberrant fusion protein NPM-ALK plays an important pathogenetic role in ALK+ anaplastic large-cell lymphoma (ALCL). We previously demonstrated that Jak3 potentiates the activity of NPM-ALK. Jak3 activation is restricted to interleukins that recruit the common γ chain (γc) receptor, including IL-9. NPM-ALK was previously shown to promote widespread lymphomas in IL-9 transgenic mice by unknown mechanisms. We hypothesized that IL-9 plays an important role in ALK+ ALCL via Jak3 activation. Our studies demonstrate the expression of IL-9Rα and IL-9 in 3 ALK+ ALCL-cell lines and 75% and 83% of primary tumors, respectively. IL-9 was detected in serum-free culture medium harvested from ALK+ ALCL-cell lines, supporting autocrine release of IL-9. Treatment of these cells with an anti–IL-9–neutralizing antibody decreased pJak3 and its kinase activity, along with pStat3 and ALK kinase activity. These effects were associated with decreased cell proliferation and colony formation in soft agar and cell-cycle arrest. Evidence suggests that cell-cycle arrest can be attributed to up-regulation of p21 and down-regulation of Pim-1. Our results illustrate that IL-9/Jak3 signaling plays a significant role in the pathogenesis of ALK+ ALCL and that it represents a potential therapeutic target for treating patients with ALK+ ALCL.

scholarly journals Inhibition of Akt increases p27Kip1 levels and induces cell cycle arrest in anaplastic large cell lymphoma

Blood ◽  
2005 ◽  
Vol 105 (2) ◽  
pp. 827-829 ◽  
Author(s):  
George Z. Rassidakis ◽  
Marianna Feretzaki ◽  
Coralyn Atwell ◽  
Ioannis Grammatikakis ◽  
Quan Lin ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Anaplastic Large Cell Lymphoma ◽  
Cell Cycle Progression ◽  
Cell Lymphoma ◽  
Large Cell ◽  
Cycle Progression ◽  
Anaplastic Lymphoma ◽  
Cycle Arrest ◽  
Large Cell Lymphoma

Abstract Anaplastic large cell lymphoma (ALCL) is a highly proliferative neoplasm that frequently carries the t(2;5)(p23;q35) and aberrantly expresses nucleophosmin–anaplastic lymphoma kinase (NPM-ALK). Previously, NPM-ALK had been shown to activate the phosphatidylinositol 3 kinase (PI3K)/Akt pathway. As the cyclin-dependent kinase (CDK) inhibitor p27Kip1 (p27) is usually not expressed in ALCL, we hypothesized that activated Akt (pAkt) phosphorylates p27 resulting in increased p27 proteolysis and cell cycle progression. Here we demonstrate that inhibition of pAkt activity in ALCL decreases p27 phosphorylation and degradation, resulting in increased p27 levels and cell cycle arrest. Using immunohistochemistry, pAkt was detected in 24 (57%) of 42 ALCL tumors, including 8 (44%) of 18 ALK-positive tumors and 16 (67%) of 24 ALK-negative tumors, and was inversely correlated with p27 levels. The mean percentage of p27-positive tumor cells was 5% in the pAkt-positive group compared with 26% in the pAkt-negative group (P = .0076). These findings implicate that Akt activation promotes cell cycle progression through inactivation of p27 in ALCL.

scholarly journals CD30-mediated cell cycle arrest associated with induced expression of p21CIP1/WAF1 in the anaplastic large cell lymphoma cell line Karpas 299

Oncogene ◽  
2001 ◽  
Vol 20 (5) ◽  
pp. 590-598 ◽  
Author(s):  
Gabriele Hübinger ◽  
Elke Müller ◽  
Inka Scheffrahn ◽  
Christof Schneider ◽  
Eberhard Hildt ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Cell Line ◽  
Anaplastic Large Cell Lymphoma ◽  
Cell Lymphoma ◽  
Large Cell ◽  
Lymphoma Cell Line ◽  
Induced Expression ◽  
Cycle Arrest ◽  
Large Cell Lymphoma

scholarly journals Everolimus in Combination with Crizotinib Synergistically Inhibits the Growth of ALK-Positive Anaplastic Large Cell Lymphoma Cells

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 4488-4488
Author(s):  
Wendan Xu ◽  
Ji-Won Kim ◽  
Junglim Lee ◽  
Hyo Jung Kim ◽  
Hwi-Joong Yoon ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Signaling Pathways ◽  
Cell Lines ◽  
Combination Treatment ◽  
Anaplastic Large Cell Lymphoma ◽  
Fusion Gene ◽  
Cell Lymphoma ◽  
Large Cell ◽  
Large Cell Lymphoma ◽  
Alk Positive

Abstract More than a half of anaplastic large cell lymphoma (ALCL) harbors an aberrant NPM-ALK fusion gene, which activates a number of down-stream signaling pathways such as Ras/ERK, PI3K/AKT, and JAK3/STAT3. Through this mechanism, mTOR pathway is also activated in ALK-positive ALCL (Vega F, et al. Cancer Res 2006). Everolimus, an mTOR inhibitor, has shown promising anti-tumor activity in a variety of lymphomas (Jundt F, et al. Blood 2005; Wanner K, et al. Br J Haematol 2006; Haritunians T, et al. Leukemia 2007), although the clinical efficacy of everolimus monotherapy was not satisfactory, possibly due to activation of several pro-surviving signaling pathways. The combined effect of everolimus and crizotinib, an ALK inhibitor, has not yet been investigated in ALK-positive tumors so far. The aim of this study was to evaluate the effect of everolimus in combination with crizotinib in ALK-positive ALCL cell lines, K-299 and SU-DHL-1. We treated K-299 and SU-DHL-1 cells with various concentrations of everolimus and crizotinib at a fixed ratio of 1:40 (Figure 1). After 72 hours, the combination index (CI) values calculated by the Chou-Talalay method were less than 1 (range, 0.583-0.763 in K-299 cells and 0.271-0.616 in SU-DHL-1 cells) in all tested combinations, suggesting synergistic cytotoxicity of everolimus and crizotinib. The Western blot analysis (Figure 2) demonstrated that everolimus treatment up-regulated the phosphorylation of ERK Thr202/Tyr204 and AKT Thr308 and Ser473 in K-299 cells. However, this aberrant activation of ERK and AKT was attenuated by the addition of crizotinib. In addition, while everolimus selectively inhibited phosphorylation of mTOR Ser2448, a marker for mTORC1 activity, the combination treatment more potently inhibited mTOR Ser2448 phosphorylation and decreased phosphorylated mTOR at Ser2481, a marker for mTORC2, as well. In the cell-cycle analysis, the combination treatment induced G1 arrest. Everolimus treatment alone did not increase the fraction of cells in the sub-G1 region compared to the control (2.16% vs. 4.03% in K-299 and 1.34% vs. 1.68% in SU-DHL-1), while crizotinib monotherapy increased the sub-G1 population (11.88% vs. 4.03% in K-299 and 28.68% vs. 1.68% in SU-DHL-1). The combination of crizotinib and everolimus markedly increased the sub-G1 population in both ALK-positive ALCL cell lines (22.25% in K-299 and 46.40% in SU-DHL-1). PARP cleavage was also increased after the combination treatment. To test the hypothesis that our findings could be applyed to other ALK-positive malignancies, we treated NCI-H2228, a lung adenocarcinoma cell line that harbors an EML4-ALK fusion gene, with everolimus and crizotinib for 72 hours. The CI values were less than 1 in all tested combinations: 0.228 in 1 nM everolimus plus 80 nM crizotinib, 0.216 in 2 nM everolimus plus 160 nM crizotinib, and 0.349 in 4 nM everolimus and 320 nM crizotinib. In summary, everolimus combined with crizotinib synergistically inhibited the growth of ALK-positive ALCL cells. Crizotinib abrogated aberrant ERK and AKT signaling activation induced by everolimus and more potently inhibited both mTORC1 and mTORC2 activity when combined with everolimus, resulting in increased G1 cell-cycle arrest and apoptosis (Figure 3). Our findings may provide an evidence for future research using everolimus and crizotinib combination in ALK-positive ALCL and could be used to improve the therapeutic outcome in patients with ALK-positive ALCL. Figure 1 Figure 1. Figure 2 Figure 2. Figure 3 Figure 3. Disclosures No relevant conflicts of interest to declare.

Bortezomib Induces Caspase-9-Mediated Apoptosis through Activation of the BH3-Only Proteins Noxa, Bik and Puma In Both ALK-Positive and ALK-Negative Anaplastic Large Cell Lymphoma Cells

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2847-2847
Author(s):  
Saskia AGM Cillessen ◽  
Nathalie J Hijmering ◽  
Laura M Moesbergen ◽  
Gert J. Ossenkoppele ◽  
Joost J Oudejans ◽  
...  
Keyword(s):  
Cell Death ◽  
Cell Lines ◽  
Anaplastic Large Cell Lymphoma ◽  
Cell Lymphoma ◽  
Large Cell ◽  
Caspase 9 ◽  
Lymphoma Cells ◽  
Induction Of Apoptosis ◽  
Large Cell Lymphoma ◽  
Alk Positive

Abstract Abstract 2847 Anaplastic large cell lymphoma (ALCL) is a CD30 positive T-cell lymphoma that can be divided into a systemic and a primary cutaneous type. Systemic ALCL can be further divided into an anaplastic lymphoma kinase (ALK) expressing type and an ALK-negative type. Despite intensive treatment regimens, the disease will be fatal in 20–30% of the systemic ALK-positive and 50–70% of the systemic ALK-negative ALCL patients. A recent study in primary ALCL samples has demonstrated an increased expression of a fraction of NF-κB target genes, suggesting upregulation of NF-κB activity in ALCL tumor cells. NF-κB activity can be inhibited by the proteasome inhibitor bortezomib resulting in induction of apoptosis. In this study, we therefore investigated if bortezomib can induce apoptosis of cultured lymphoma cells of three systemic ALK-positive and three ALK-negative ALCL patients and seven ALCL cell lines and we examined the mechanisms by which bortezomib induced cytotoxicity in these ALCL cells. Treatment with bortezomib resulted in induction of apoptosis in all ALK-positive and ALK-negative ALCL patient samples and ALCL cell lines tested, when we compared the percentage cell death with the non-neoplastic CD4- and CD8-positive PBMC and tonsil T-cells from healthy donors. The lethal dose (LD50) varied between 54nM and more than 100nM after 24 hours and varied between 21nM and 52nM after 48 hours of exposure. ALK-negative ALCL cases were more sensitive to bortezomib and showed significant lower LD50 values than ALK-positive ALCL cells. We show that bortezomib-induced cell death in ALK-positive and ALK-negative ALCL is dependent on caspase-9 and/or caspase-8 mediated apoptosis and that bortezomib induces depolarization of the mitochondrial membrane. mRNA-expression and protein analysis revealed clearly upregulation of the BH3-only proteins Noxa, Bik and Puma, resulting in Bak and Bax release from the anti-apoptotic proteins Mcl-1 and Bcl-2. We also demonstrated that ALCL cells relatively resistant to bortezomib were characterized by high expression of Bcl-2A1, suggesting the possibility of pre-defining patients most likely to benefit from bortezomib therapy. Our preclinical data support the therapeutic application of bortezomib as potential drug in the treatment of ALCL, especially ALK-negative ALCL patients to improve their prognosis. Disclosures: No relevant conflicts of interest to declare.

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