An Oncogenic Metabolic Switch Mediates Resistance to NOTCH1 Inhibition in T-ALL

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 285-285
Author(s):  
Daniel Herranz ◽  
Valeria Tosello ◽  
Alberto Ambesi-Impiombato ◽  
Mireia Castillo ◽  
Carlos Cordon-Cardo ◽  
...  
Keyword(s):  
T Cell ◽  
Tumor Cells ◽  
Carbon Metabolism ◽  
Pten Loss ◽  
Notch1 Signaling ◽  
Induced Tumors ◽  
Notch Inhibition

Abstract Abstract 285 Activating mutations in NOTCH1 are common in T-cell lymphoblastic leukemias (T-ALL), making this receptor a promising target for drugs such as γ-secretase inhibitors (GSIs), which block a proteolytic cleavage required for NOTCH1 activation. Aberrant activation of the PI3K-AKT pathway due to mutational loss of PTEN is found in 20% of human T-ALLs and has been linked with in vitro resistance to GSIs therapy in T-ALL cell lines, suggesting that in the absence of PTEN, constitutive activation of the PI3K pathway may render T-cell lymphoblasts insensitive to NOTCH1 inhibition with GSIs. Still, cell lines frequently fail to recapitulate the biology of primary tumor cells, and in vitro studies fall short of addressing the role of the NOTCH-PI3K interaction in clinical resistance to GSI therapy, which is best defined as disease progression under treatment in vivo. Moreover, the specific role and mechanisms of GSI resistance downstream of PTEN loss in T-ALL remain to be elucidated. To address this question, and to analyze the actual significance of the interaction between NOTCH1 signaling and PTEN loss in T-cell transformation and therapy response we analyzed the response of NOTCH1 induced PTEN-positive and PTEN-deleted isogenic tumors to GSI therapy in vivo. Towards this goal we first generated NOTCH1 induced T-ALLs via bone marrow transplantation of tamoxifen-inducible conditional PTEN knockout (Rosa26TMCre PTEN flox/flox) hematopoietic progenitors infected with retroviruses expressing a mutant constitutively active form of the NOTCH1 receptor (NOTCH1 L1601P Δ-PEST). NOTCH1 L1601P Δ-PEST Rosa26TMCre PTEN flox/flox tumor cells injected into secondary recipients were treated with vehicle only or tamoxifen in order to generate PTEN-non-deleted and PTEN-deleted isogenic tumors, respectively. Treatment of PTEN-positive tumor bearing mice with DBZ, a highly active GSI, demonstrated marked responses to therapy by in vivo bioimaging compared with vehicle only treated controls, which translated into a significant improvement in survival (P < 0.005). In contrast, all mice harboring PTEN-deleted tumors failed to respond to DBZ treatment, showed overt progression under treatment and died of their disease demonstrating a direct role of PTEN loss in the development of resistance to inhibition of NOTCH1 signaling with GSIs in vivo. Moreover, limiting dilution analyses demonstrated that secondary loss of PTEN increased the leukemia initiating cell potential of NOTCH1 L1601P Δ-PEST induced tumors. Analysis of NOTCH1 signaling showed complete clearance of activated NOTCH1 protein and marked downregulation of Hes1 in both in PTEN-positive and PTEN-deleted NOTCH1 L1601P Δ-PEST induced tumors treated with DBZ compared with controls. However, global analysis of gene expression profiling with oligonucleotide microarrays showed that while NOTCH1 direct target genes are downregulated in both PTEN-positive and PTEN deleted tumors, there is a global reversal of much of the transcriptional effects of NOTCH inhibition, consisting of downregulation of genes involved in anabolic pathways and upregulation of genes involved in catabolic pathways and autophagy, upon PTEN loss. Consistently, electron microscopy analysis demonstrated increased autophagy in NOTCH1 induced tumors upon NOTCH1 inhibition, which was reversed upon PTEN deletion. Moreover, global metabolomic analyses of PTEN-non-deleted and PTEN-deleted NOTCH1 L1601P Δ-PEST induced tumors treated with DBZ compared with controls demonstrated that NOTCH inactivation induces a global anabolic shutdown in T-ALL with a marked block of glycolysis and glutaminolysis which renders NOTCH induced tumors dependent on branched amino acid catabolism to sustain their carbon metabolism. Notably these effects are globally rescued in PTEN deleted tumor cells, which show high basal levels of glycolysis and sustained glycolysis and glutaminolysis despite effective NOTCH1 inhibition with DBZ. Overall, these results formally demonstrate that loss of PTEN induces in vivo drug resistance to NOTCH inhibition in T-ALL; highlight the fundamental importance of NOTCH1 in the control of tumor cell metabolism; strongly suggest that increased glycolysis and sustained carbon metabolism can induce resistance to GSI therapy and provide the basis for the design of new therapeutic strategies targeting these metabolic pathways in T-ALL. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Inhibition of tumor growth by elimination of granulocytes.

1995 ◽  
Vol 181 (1) ◽  
pp. 435-440 ◽  
Author(s):  
L A Pekarek ◽  
B A Starr ◽  
A Y Toledano ◽  
H Schreiber
Keyword(s):  
T Cell ◽  
Tumor Cells ◽  
Specific Antigen ◽  
Induced Tumors ◽  
Inhibition Of Tumor Growth ◽  
Parental Tumor ◽  
Antigranulocyte Antibody ◽  
Tumor Specific Antigen

As observed for many types of cancers, heritable variants of ultraviolet light-induced tumors often grow more aggressively than the parental tumors. The aggressive growth of some variants is due to the loss of a T cell-recognized tumor-specific antigen; however, other variants retain such antigens. We have analyzed an antigen retention variant and found that the variant tumor cells grow at the same rate as the parental tumor cells in vitro, but grew more rapidly than the parental cells in the T cell-deficient host. The growth of the variant cells was stimulated in vitro by factors released from tumor-induced leukocytes and by several defined growth factors. In addition, the variant cancer cells actually attracted more leukocytes in vitro than the parental cells. Furthermore, elimination of granulocytes in vivo in nude mice by a specific antigranulocyte antibody inhibited the growth of the variant cancer, indicating that this tumor requires granulocytes for rapid growth.

Investigating Synthetic Lethality Between mTOR Hyperactivation and Cap-Dependent mRNA Translation In a Receptor Tyrosine Kinase Driven Model Of Acute T-Cell Leukemia

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 3913-3913
Author(s):  
Adrian Schwarzer ◽  
Helmut Holtmann ◽  
Celina Schauerte ◽  
Nomme Benn ◽  
Jan Völkel ◽  
...  
Keyword(s):  
T Cell ◽  
Tyrosine Kinase ◽  
Receptor Tyrosine Kinase ◽  
Initiation Complex ◽  
Pten Loss ◽  
Rtk Signaling ◽  
Induction Of Apoptosis

Abstract T-cell acute lymphoblastic leukemia (T-ALL) remains a therapeutic challenge. In particular, relapsed disease is refractory to further therapy and has a dismal outcome. High activation of the PI3K-AKT-mTOR pathway is a hallmark of T-ALL that has been linked to the resistance of T-ALL to glucocorticoids and chemotherapy. The master regulator of PI3K-AKT signaling is PTEN. PTEN loss (1) is accompanied by an adverse outcome, (2) occurs as secondary event in T-ALL relapses and (3) is selected for after xenotransplantation of human T-ALLs into NSG mice. Interestingly, even in the absence of PTEN, the PI3-Kinases γ and δ, which link receptor tyrosine kinases (RTK) and PIP3 signaling, are critical for T-ALL formation. In lung cancer, both PTEN loss and activation of RTK signaling are required for full activation of AKT and aggressive disease. However, whereas the role of PTEN in T-ALL is well appreciated, little is known about the contribution of RTK signaling. We previously demonstrated the aberrant expression of members of the Neurotrophin receptor tyrosine kinase family (tropomyosin-related kinases - TRKA/B/C) in primary human leukemias of myeloid and lymphoid origin. We detected high expression of TRKB in several T-ALL samples. Some samples also co-expressed intracellular BDNF (brain derived neurotrophic factor), the ligand for TRKB, suggesting the existence of autocrine loops between the receptor and its ligands. Expression of the TRKB/BDNF autocrine loop or of a constitutively active form of the human TRKA receptor (ΔTrkA) in murine hematopoietic stem cells elicited T-ALL with a mean latency of 100 days in our murine transplantation model. Here we dissect the downstream signaling cascades in BDNF/TRKB or ΔTrkA induced T-ALL clones and show acquisition of activating Notch1 mutations and loss of PTEN during clonal evolution of T-ALLs induced by deregulated TRK-signaling. All three events contribute independently to activation of mTORC1 and mTORC2, demonstrating a strong selective pressure for enhanced mTOR signaling in T-ALL. To investigate the role of both mTOR complexes and downstream effectors, we used an improved Tet-regulated miR30-shRNA system in conjunction with a FACS-based reporter assay, allowing the rapid identification of shRNAs that give powerful knockdown at the single copy level. We identified several potent RNAi triggers against Rictor (mTORC2), Raptor (mTORC1) and members of the cap-initiation complex. Knockdown of Rictor or Raptor alone caused a transient decrease of proliferation and viability. In contrast, complete inhibition of mTOR with Torin1 or knockdown of eIF4E, the rate limiting subunit of the cap initiation complex, resulted in strong induction of apoptosis. This demonstrates that increased cap-dependent translation is a key effector of oncogenic mTOR in TRK+Notch+PTEN- T-ALL. Next, we investigated the efficacy of 4EGI-1E, a novel isoform of the previously described inhibitor of cap-dependent translation 4EGI-1, in this T-ALL model. 4EGI-1E induced apoptosis of T-ALL blasts in vitro with an EC50 of 3.5 mM, whereas the EC50 for global inhibition of cap-dependent translation was 50 mM, suggesting that 4EGI-1E targets a subset of mRNAs that are regulated by cap-dependent translation and are crucial for T-ALL survival. Microarray analysis of polysome fractions from DMSO and 4EGI-1E-treated cells in vitro revealed that 4EGI-1E treatment decreased the active translation of mRNAs for genes observed to be upregulated in T-ALL. These genes included members of the translational apparatus, mitochondrial matrix proteins, cyclins, c-Myc and Bcl-2. Polysome profiling in vivo recapitulated the in vitro results, showing that 4EGI-1E caused a global decrease in the ribosomal occupancy of cellular mRNAs. Network analysis suggested c-Myc as a central node within the depleted mRNAs. Abrogation of c-myc expression and induction of apoptosis were observed as soon as 4 hours after injection of 4EGI-1E (1.5 mg) into leukemia bearing animals. Depletion of T-ALL blasts from the bone marrow was achieved after 5 days of daily 4EGI-1E treatment, T-ALL (DMSO: 39% (SD 18%) vs. 4EGI-1E 0.54% (SD 0.62%) p=0.003). Finally, we investigated the effects of 4EGI-1E on healthy hematopoiesis, demonstrating the existence of a therapeutic window for inhibition of cap-dependent translation in vivo. Hence, inhibiting eIF4E is a promising approach to target c-myc in a genetically complex T-ALL model. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Involvement of leukocyte function-associated antigen-1 (LFA-1) in the invasion of hepatocyte cultures by lymphoma and T-cell hybridoma cells.

1987 ◽  
Vol 105 (1) ◽  
pp. 553-559 ◽  
Author(s):  
E Roos ◽  
F F Roossien
Keyword(s):  
T Cell ◽  
Tumor Cells ◽  
Cell Types ◽  
Hybridoma Cells ◽  
Lymphoma Cells ◽  
Hepatocyte Cultures ◽  
Leukocyte Function ◽  
Vitro Model

We studied the interaction of MB6A lymphoma and TAM2D2 T cell hybridoma cells with hepatocyte cultures as an in vitro model for in vivo liver invasion by these tumor cells. A monoclonal antibody against leukocyte function-associated antigen-1 (LFA-1) inhibited adhesion of the tumor cells to the surface of hepatocytes and consequently strongly reduced invasion. This effect was specific since control antibodies, directed against Thy.1 and against T200, of the same isotype, similar affinity, and comparable binding to these cells, did not inhibit adhesion. This suggests that LFA-1 is involved in the formation of liver metastases by lymphoma cells. TAM2D2 T cell hybridoma cells were agglutinated by anti-LFA-1, but not by control antibodies. Reduction of adhesion was not due to this agglutination since monovalent Fab fragments inhibited adhesion as well, inhibition was also seen under conditions where agglutination was minimal, and anti-LFA-1 similarly affected adhesion of MB6A lymphoma cells that were not agglutinated. The two cell types differed in LFA-1 surface density. TAM2D2 cells exhibited 400,000 surface LFA-1 molecules, 10 times more than MB6A cells. Nevertheless, the level of adhesion and the extent of inhibition by the anti-LFA-1 antibody were only slightly larger for the TAM2D2 cells.

On the role of APC-activation for in vitro versus in vivo T cell priming

2003 ◽  
Vol 225 (1) ◽  
pp. 1-11 ◽  
Author(s):  
Tazio Storni ◽  
Martin F. Bachmann
Keyword(s):  
T Cell ◽  
T Cell Priming

scholarly journals Studying the immunosuppressive role of indoleamine 2,3-dioxygenase: tryptophan metabolites suppress rat allogeneic T-cell responses in vitro and in vivo

2005 ◽  
Vol 18 (1) ◽  
pp. 95-100 ◽  
Author(s):  
Thomas M. Bauer ◽  
Lucian P. Jiga ◽  
Jing-Jing Chuang ◽  
Marco Randazzo ◽  
Gerhard Opelz ◽  
...  
Keyword(s):  
T Cell ◽  
T Cell Responses ◽  
Cell Responses ◽  
Indoleamine 2,3 Dioxygenase ◽  
Tryptophan Metabolites

scholarly journals IL-27 promotes T cell–dependent colitis through multiple mechanisms

2010 ◽  
Vol 208 (1) ◽  
pp. 115-123 ◽  
Author(s):  
Jennifer H. Cox ◽  
Noelyn M. Kljavin ◽  
Nandhini Ramamoorthi ◽  
Lauri Diehl ◽  
Marcel Batten ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Intestinal Inflammation ◽  
Interferon Γ ◽  
Transfer Model ◽  
Proinflammatory Role ◽  
Immune Pathology

Interleukin-27 (IL-27) is a cytokine known to have both proinflammatory and immunoregulatory functions. The latter appear to dominate in vivo, where IL-27 suppresses TH17 responses and promotes the differentiation of Tr1 cells expressing interferon-γ and IL-10 and lacking forkhead box P3 (Foxp3). Accordingly, IL-27 receptor α (Il27ra)–deficient mice suffer from exacerbated immune pathology when infected with various parasites or challenged with autoantigens. Because the role of IL-27 in human and experimental mouse colitis is controversial, we studied the consequences of Il27ra deletion in the mouse T cell transfer model of colitis and unexpectedly discovered a proinflammatory role of IL-27. Absence of Il27ra on transferred T cells resulted in diminished weight loss and reduced colonic inflammation. A greater fraction of transferred T cells assumed a Foxp3+ phenotype in the absence of Il27ra, suggesting that IL-27 functions to restrain regulatory T cell (Treg) development. Indeed, IL-27 suppressed Foxp3 induction in vitro and in an ovalbumin-dependent tolerization model in vivo. Furthermore, effector cell proliferation and IFN-γ production were reduced in the absence of Il27ra. Collectively, we describe a proinflammatory role of IL-27 in T cell–dependent intestinal inflammation and provide a rationale for targeting this cytokine in pathological situations that result from a breakdown in peripheral immune tolerance.

scholarly journals Bacterial Pathogens Induce Abscess Formation by CD4+ T-Cell Activation via the CD28–B7-2 Costimulatory Pathway

2000 ◽  
Vol 68 (12) ◽  
pp. 6650-6655 ◽  
Author(s):  
Arthur O. Tzianabos ◽  
Anil Chandraker ◽  
Wiltrud Kalka-Moll ◽  
Francesca Stingele ◽  
Victor M. Dong ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Pathogenic Bacteria ◽  
Cell Activation ◽  
Bacterial Pathogens ◽  
Abscess Formation

ABSTRACT Abscesses are a classic host response to infection by many pathogenic bacteria. The immunopathogenesis of this tissue response to infection has not been fully elucidated. Previous studies have suggested that T cells are involved in the pathologic process, but the role of these cells remains unclear. To delineate the mechanism by which T cells mediate abscess formation associated with intra-abdominal sepsis, the role of T-cell activation and the contribution of antigen-presenting cells via CD28-B7 costimulation were investigated. T cells activated in vitro by zwitterionic bacterial polysaccharides (Zps) known to induce abscess formation required CD28-B7 costimulation and, when adoptively transferred to the peritoneal cavity of naı̈ve rats, promoted abscess formation. Blockade of T-cell activation via the CD28-B7 pathway in animals with CTLA4Ig prevented abscess formation following challenge with different bacterial pathogens, including Staphylococcus aureus,Bacteroides fragilis, and a combination ofEnterococcus faecium and Bacteroides distasonis. In contrast, these animals had an increased abscess rate following in vivo T-cell activation via CD28 signaling. Abscess formation in vivo and T-cell activation in vitro required costimulation by B7-2 but not B7-1. These results demonstrate that abscess formation by pathogenic bacteria is under the control of a common effector mechanism that requires T-cell activation via the CD28–B7-2 pathway.

Dendritic cell-derived IL-2 production is regulated by IL-15 in humans and in mice

Blood ◽  
2005 ◽  
Vol 105 (2) ◽  
pp. 697-702 ◽  
Author(s):  
Sonia Feau ◽  
Valeria Facchinetti ◽  
Francesca Granucci ◽  
Stefania Citterio ◽  
David Jarrossay ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
T Cell ◽  
Molecular Mechanisms ◽  
Interleukin 2 ◽  
Adaptive Immune ◽  
Deficient Mice ◽  
Mouse System

Abstract Dendritic cells (DCs) are involved in the initiation and regulation of innate and adaptive immune responses. Several molecular mechanisms regulate these diverse DC functions, and we have previously reported that mouse dendritic cells (mDCs) can produce interleukin-2 (IL-2) in vitro and in vivo, in response to microbial activation and T-cell-mediated stimuli. This property is shared by different DC subtypes, including Langerhans cells. Here we show that, on appropriate stimulation, human DCs, both plasmacytoid and myeloid subtypes, also express IL-2. Interestingly, the production of IL-2 by myeloid DCs is induced by T-cell-mediated stimuli and depends on the presence of IL-15. The key role of this cytokine in regulating IL-2 production was also confirmed in the mouse system. In particular, we could show that DCs from IL-15-deficient mice were strongly impaired in the ability to produce IL-2 after interactions with different microbial stimuli. Our results indicate that DC-produced IL-2 is tightly coregulated with the expression of IL-15.

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