Role of Spleen Tyrosine Kinase Signaling in Early B Cell Acute Lymphoblastic Leukemia.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3092-3092
Author(s):  
Tatiana Perova ◽  
Lauryl Nutter ◽  
Irina Matei ◽  
Ildiko Grandal ◽  
Polly Pine ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Tyrosine Kinase ◽  
B Cell ◽  
Molecular Mechanisms ◽  
Lymphoblastic Leukemia ◽  
Employment Equity ◽  
Cell Acute Lymphoblastic Leukemia ◽  
Equity Ownership ◽  
All Treatment

Abstract Abstract 3092 Poster Board III-29 Introduction Early B cell acute lymphoblastic leukemia (B-ALL) is the most common type of childhood malignancy, characterized by abnormal accumulation and proliferation of progenitor-B or precursor-B (pre-B) cells. Current challenges associated with B-ALL treatment include fatal relapses, treatment-related toxicities and long-term morbidities underscoring a need to develop new targeted therapies aimed at eradicating leukemia cells and their stem cells. To achieve this, a better understanding of molecular mechanisms involved in leukemia initiation and progression is required. Our laboratory developed p53-/- PrkdcSCID/SCID double mutant (DM) strain as a mouse model of early B-ALL. We showed that DM leukemias progress through discrete developmental stages of leukemogenesis despite the absence of a pre-B cell receptor (pre-BCR), a crucial checkpoint in B cell development. Spleen tyrosine kinase (SYK), a key proximal component of pre-BCR signaling, was activated in the DM leukemias despite the absence of pre-BCR and was required for their survival. Approximately 70% of pediatric pre-B-ALLs also do not express pre-BCR, which lead us to investigate SYK signaling in human pre-B-ALL and to test potential therapeutic application of SYK inhibition in these leukemias. Patients and Methods We examined 22 viably frozen primary pediatric pre-B-ALL bone marrow samples to test their responses to SYK inhibition in vitro and in vivo and have investigated the molecular basis for aberrant SYK-mediated signaling in B-ALL. Results Western blot analyses revealed that SYK and BLNK, a dominant target of SYK, were expressed in pre-B-ALL patient samples. The majority of human pre-B ALL samples tested (14/22) displayed significantly attenuated proliferation in the presence of SYK inhibitors suggesting that SYK is necessary for their survival and/or proliferation. Treatment with SYK inhibitor R406 prevented phosphorylation of downstream SYK targets including BLNK and PLC-γ2. We are continuing to study the effects of SYK inhibition using phospho-flow cytometry and genome wide expression arrays. Preliminary data will also be presented on therapeutic efficacy of an orally bioavailable form of R406-mediated SYK inhibition in vivo by xenotransplantation of human leukemias into immuno-deficient mice. Conclusions Understanding the molecular mechanisms of pre-BCR-independent SYK activation involved in proliferation and survival of leukemic blasts may provide a rational basis for development of effective treatment for ALL. Specifically, targeted therapeutic inhibition of SYK signaling may be effective B-ALL treatment that may improve outcomes of current treatment regiments with minimal additional treatment-related toxicity. Disclosures Pine: Rigel Pharmaceuticals: Employment, Equity Ownership. Hitoshi:Rigel Pharmaceuticals: Employment, Equity Ownership.

Rac2 GTPase Activation Is Necessary for Development of p190-BCR-ABL-Induced B-Cell Acute Lymphoblastic Leukemia

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3790-3790
Author(s):  
Abel Sanchez-Aguilera ◽  
Ami tava Sengupta ◽  
Joseph P Mastin ◽  
Kyung H Chang ◽  
David A Williams ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
B Cells ◽  
Tyrosine Kinase ◽  
B Cell ◽  
Rho Gtpases ◽  
Lymphoblastic Leukemia ◽  
Hematopoietic Stem ◽  
Cell Acute Lymphoblastic Leukemia

Abstract The fusion gene BCR-ABL, resulting from t(9;22) reciprocal chromosomal translocations, encodes a constitutively active tyrosine kinase. Two different isoforms of BCR-ABL, p190 and p210, are associated to two completely different diseases. In the tyrosine kinase inhibitor (TKI) era, while p210-BCR-ABL-induced CML is highly responsive to TKI, p190-BCR-ABL still induces a poor prognosis B-cell acute lymphoblastic leukemia (B-ALL). The only difference between these two forms of BCR-ABL is the existence of a DH/Cdc24/PH domain in p210-BCR-ABL, which acts as a guanine nucleotide exchange factor (GEF) able to activate Rho GTPases. Rac is a subfamily of Rho GTPases with regulatory activity on hematopoietic stem cell and progenitor (HSC/P) functions. We have previously shown that Rac2 and further the combination of Rac1 and Rac2 mediate downstream signals in p210 BCR-ABL-induced myeloproliferation (Thomas EK, et al., Cancer Cell, 2007). Interestingly, despite the absence of a GEF domain in p190-BCR-ABL, Rac is activated, suggesting the activation of other GEF(s). Here we have analyzed whether Vav and Rac family members are involved in p190-BCR-ABL-induced B-ALL. We have used a combination of in vitro (Ba/F3 pro-B cells transduced with p190 or p210 BCR-ABL) and in vivo (murine transduction-transplantation model of p190 BCR-ABL-induced B-ALL) approaches. In Ba/F3 cells, both p190 BCR-ABL and p210 BCR-ABL activated Rac and the Rac effector p21 activated kinase (PAK), and their proliferation and survival appeared severely decreased in response to the Rac activation inhibitor NSC23766. Stat3, Stat5 and Jnk, but not ERK, p38 or NF-kB, were constitutively hyperactivated in p190 BCRABL-expressing Ba/F3 cells and primary murine B-ALL cells. Intracellular flow cytometry analysis demonstrated that Stat5 was specifically activated in the pro/pre-B leukemic cell population, compared to normal B cells. In the murine model of B-ALL, loss of Rac2, but not Rac3, prolonged survival and impaired leukemia development. Like in Ba/F3 cells, primary B-CFU and outgrowth in Witte-Whitlock assays of leukemic primary cells from mice was severely decreased by the addition of NSC23766 to the culture. Although Vav was activated by both p190- and p210-BCR-ABL, since NSC23766 does not block the activation by Vav1, we hypothesized that other GEFs were involved. Indeed, the loss of Vav1 or even combined loss of Vav1 and Vav2 did not impair BCR-ABL-mediated lymphoid leukemogenesis in vivo. Vav3, another member in the Vav family which uses a different mechanism of activation of Rac GTPases was a likely candidate. In fact, loss of Vav3 alone was able to significantly prolong the survival and attenuate development of p190 BCR-ABL-driven B-ALL. In conclusion, the results of this study indicate that Rac activation is necessary for the development of B-ALL induced by p190-BCR-ABL in vitro and in vivo, and validate a new signaling pathway as a therapeutic target for BCR-ABL-induced B-ALL.

scholarly journals AT1412, a Patient-Derived Antibody in Development for the Treatment of CD9 Positive Precursor B-Acute Lymphoblastic Leukemia

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 4461-4461
Author(s):  
Greta De Jong ◽  
Sophie E Levie ◽  
Remko Schotte ◽  
Wouter Pos ◽  
Daniel Go ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Cell Death ◽  
Acute Lymphoblastic Leukemia ◽  
B Cell ◽  
Cell Lines ◽  
Lymphoblastic Leukemia ◽  
Melanoma Cells ◽  
Employment Equity ◽  
Equity Ownership

Despite rapid advances in immunotherapeutic options for precursor B-acute lymphoblastic leukemia (ALL), outcomes remain poor especially for adult ALL and relapsed pediatric ALL. With conventional chemotherapy, remission percentages in adult ALL range from 75 to 90%, but relapse rates are high and long-term leukemia-free survival ranges between 35-70% depending on age and risk group. The introduction of CD19 targeting immunotherapy has significantly improved patient outcomes in (relapsed) B-ALL. However, tumor escape via downregulation of CD19 occurs in a significant number of patients. Therefore an ongoing urgency remains for the identification of additional or alternative immunotherapeutic targets for the treatment of ALL. AT1412 is an antibody that was identified from the peripheral blood memory B cell pool of a patient cured of metastatic melanoma after adoptive T-cell therapy, using a B cell immortalization technology (AIMSelect) with ectopic Bcl-6 and Bcl-xL expression as described previously [Kwakkenbos et al. Nat. Med. 2010]. The antibody was selected based on differential binding to melanoma cells as compared to healthy melanocytes and was shown to be successful in killing melanoma cells in vitro and in vivo [manuscript submitted]. In addition to melanoma, AT1412 binds other tumor types including B-ALL, gastric, colon- and pancreatic cancer. The target of AT1412 is the tetraspanin CD9, which is expressed by more than half of all B-ALL. Expression of CD9 has been correlated with adverse prognosis [Liang et al. Cancer Biomark. 2018]. We assessed binding of this human CD9 antibody to a panel of ALL cell lines using flow cytometry. Binding of AT1412 to the B-ALL cell lines SUP-B15, MHH-CALL-2 and CCRF-SB varied as expected based on the CD9 levels that we detected using a commercial CD9 antibody. AT1412 induced antibody dependent cellular cytotoxicity (ADCC) on these cells, in line with the level of AT1412 binding. No binding was seen to the T-ALL cell line Jurkat. Importantly, these findings were confirmed in primary ALL samples, obtained prospectively at diagnosis from a cohort of patients with T- or B-ALL (n=30). AT1412 showed binding to 61% of B-ALL samples but not to T-ALL samples. The potential of AT1412 to induce ADCC was tested on patient samples from the same panel. Remarkably, AT1412 induced ADCC of all B-ALL samples it bound to (8 out of 14) and of none of the T-ALL samples. Cytotoxicity significantly correlated with the level of AT1412 binding. These findings were supported by the observation that AT1412 induced B-ALL cell death when a freshly drawn whole bone marrow sample from a patient with newly diagnosed B-ALL was cocultured with AT1412. AT1412-induced cell death of B-ALL blasts occurred without affecting the monocytic, granulocytic and lymphocytic populations. This cell death was not observed when this patient's ALL blasts were incubated with AML-targeting antibodies. Remarkably, AT1412 induced cell death in the absence of added effector cells or other (chemo)therapeutic agents, while the bone marrow sample contained over 80% blasts and as little as 3% lymphocytes. We are currently investigating the in vivo efficacy of the antibody in a humanized immune system mouse model with human B-ALL. Taken together, the majority of precursor B-ALL blasts express CD9 and expression of CD9 is associated with a dismal outcome. Our data demonstrate that CD9 can be successfully targeted by the human CD9 antibody AT1412, suggesting that AT1412 has the potential to be developed as a therapeutic antibody for B-ALL. AT1412 is currently being advanced through preclinical development. Disclosures De Jong: AIMM Therapeutics: Employment. Levie:AIMM Therapeutics: Employment. Schotte:AIMM Therapeutics: Employment, Equity Ownership, Patents & Royalties: Patent WO2017119811A1. Pos:AIMM Therapeutics: Patents & Royalties: Patent WO2017119811A1. Go:AIMM Therapeutics: Employment, Patents & Royalties: Patent WO2017119811A1. Yasuda:AIMM Therapeutics: Employment, Equity Ownership. Cercel:AIMM Therapeutics: Employment. van Hal-van Veen:AIMM Therapeutics: Employment. Frankin:AIMM Therapeutics: Employment. Villaudy:AIMM Therapeutics: Employment, Equity Ownership, Patents & Royalties: Patent WO2017119811A1. van Helden:AIMM Therapeutics: Employment, Equity Ownership, Patents & Royalties: Patent WO2017119811A1. van Eenennaam:AIMM Therapeutics: Employment. Spits:AIMM Therapeutics: Employment, Equity Ownership, Patents & Royalties: Patent WO2017119811A1. Hazenberg:AIMM Therapeutics: Other: Employment/equity of partner/spouse.

scholarly journals Non-Coding RNA Signatures of B-Cell Acute Lymphoblastic Leukemia

2021 ◽  
Vol 22 (5) ◽  
pp. 2683
Author(s):  
Princess D. Rodriguez ◽  
Hana Paculova ◽  
Sophie Kogut ◽  
Jessica Heath ◽  
Hilde Schjerven ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
B Cell ◽  
Molecular Mechanisms ◽  
Lymphoblastic Leukemia ◽  
Transcriptome Profiling ◽  
Rna Seq ◽  
Protein Coding ◽  
Non Coding Rna ◽  
Technological Developments ◽  
Cell Acute Lymphoblastic Leukemia

Non-coding RNAs (ncRNAs) comprise a diverse class of non-protein coding transcripts that regulate critical cellular processes associated with cancer. Advances in RNA-sequencing (RNA-Seq) have led to the characterization of non-coding RNA expression across different types of human cancers. Through comprehensive RNA-Seq profiling, a growing number of studies demonstrate that ncRNAs, including long non-coding RNA (lncRNAs) and microRNAs (miRNA), play central roles in progenitor B-cell acute lymphoblastic leukemia (B-ALL) pathogenesis. Furthermore, due to their central roles in cellular homeostasis and their potential as biomarkers, the study of ncRNAs continues to provide new insight into the molecular mechanisms of B-ALL. This article reviews the ncRNA signatures reported for all B-ALL subtypes, focusing on technological developments in transcriptome profiling and recently discovered examples of ncRNAs with biologic and therapeutic relevance in B-ALL.

Epigenetic Control Of Cell Cycle-Promoting Genes By Ikaros and Casein Kinase II In B-Cell Acute Lymphoblastic Leukemia

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 3734-3734
Author(s):  
Sinisa Dovat ◽  
Chunhua Song ◽  
Xiaokang Pan ◽  
Yali Ding ◽  
Chandrika S. Gowda ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Acute Lymphoblastic Leukemia ◽  
B Cell ◽  
Target Genes ◽  
Lymphoblastic Leukemia ◽  
Regulatory Elements ◽  
Preclinical Model ◽  
Cycle Arrest ◽  
Cell Acute Lymphoblastic Leukemia

Abstract IKZF1 (Ikaros) encodes a kruppel-like zinc finger protein that is essential for normal hematopoiesis and acts as a tumor suppressor in acute lymphoblastic leukemia (ALL). The deletion and/or mutation of Ikaros is associated with the development of human T-cell and B-cell acute lymphoblastic leukemia (B-ALL) with poor outcome. In vivo, Ikaros binds DNA and regulates gene expression by chromatin remodeling. Since there is a paucity of known genes that are regulated by Ikaros, the molecular mechanisms through which Ikaros exerts its tumor suppressor function remain unknown. Here we describe studies that identify the targets and mechanisms of Ikaros-mediated epigenetic regulation in human B-ALL. We used chromatin immunoprecipitation coupled with next generation sequencing (ChIP-seq) to identify target genes that are bound by Ikaros in vivo in human B-ALL, and to define epigenetic patterns associated with Ikaros binding. ChIP-seq revealed a large set of Ikaros target genes that contain a characteristic Ikaros binding motif. The largest group of genes that are direct Ikaros targets included genes that are essential for cell cycle progression. These included CDC2, CDC7, CDK2 and CDK6 genes whose deregulation is associated with malignant transformation. The strong binding of ikaros to the promoters of cell cycle-promoting genes was confirmed by quantitative immunoprecipitation in primary leukemia cells. To establish whether Ikaros directly regulates transcription of the cell cycle-promoting genes, their expression was measured in B-ALL cells that were transduced with either a retroviral vector that contains Ikaros, or a control vector. Target gene expression was monitored by qRT-PCR. Ikaros strongly repressed transcription of the cell cycle-promoting genes, which resulted in cell cycle arrest. Global epigenetic profiling using ChIP-seq suggested that Ikaros represses cell cycle-promoting genes by inducing epigenetic changes that are consistent with repressive chromatin. High-resolution epigenetic profiling of the upstream regulatory elements of the cell cycle-promoting genes targeted by Ikaros showed that increased Ikaros expression results in the formation of heterochromatin, which is characterized by the presence of the H3K9me3 histone modification and associated transcriptional repression. Functional analysis revealed that phosphorylation of Ikaros by the oncogenic protein. Casein kinase II (CK2), impairs its function as a transcriptional repressor of the cell cycle-regulating genes. Inhibition of CK2 by specific inhibitors enhances Ikaros-mediated repression of the cell cycle-regulating genes resulting in cessation of cellular proliferation and cell cycle arrest in vitro and in vivo in a B-cell ALL preclinical model. This was associated with increased Ikaros binding and the formation of heterochromatin at upstream regulatory elements of the cell cycle-promoting genes. Our results provide evidence that Ikaros functions as a repressor of cell cycle-promoting genes in B-ALL by directly binding their promoters and inducing the formation of heterochromatin with characteristic H3K9me3 histone modifications Ikaros repressor function is negatively regulated by CK2 kinase in B-cell ALL. Inhibition of CK2 enhances Ikaros mediated-repression of cell cycle-promoting genes resulting in an anti-leukemia effect in a preclinical model of B-cell ALL. Presented data identified the mechanism of action of CK2 inhibitors and demonstrated their efficacy in B-cell ALL preclinical model. Results support the use of CK2 inhibitors in Phase I clinical trial. Supported by National Institutes of Health R01 HL095120 and a St. Baldrick’s Foundation Career Development Award (to S.D.). Disclosures: No relevant conflicts of interest to declare.

scholarly journals CXCR4 Blockade By BL-8040 in T Cell Acute Lymphoblastic Leukemia Decreases Mitochondrial Mass and Induces Non-Apoptotic Cell Death

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 2745-2745
Author(s):  
Jun Xia ◽  
Stephanie Sun ◽  
Matthew RM Jotte ◽  
Geoffrey L. Uy ◽  
Osnat Bohana-Kashtan ◽  
...  
Keyword(s):  
Cell Death ◽  
Apoptotic Cell ◽  
Lymphoblastic Leukemia ◽  
Apoptotic Cell Death ◽  
Employment Equity ◽  
Cxcr4 Signaling ◽  
Cell Acute Lymphoblastic Leukemia ◽  
Equity Ownership

T cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematologic malignancy that accounts for 10-15% of pediatric and 25% of adult ALL cases. CXCL12 is a CXC chemokine that is constitutively expressed at high levels in the bone marrow. CXCR4 is the major receptor for CXCL12 and is by far the most highly expressed chemokine receptor on T-ALL cells. Two groups recently showed that genetic loss of CXCR4 signaling in murine or human T-ALL cells markedly suppressed their growth in vivo. We previously reported that BL-8040, a potent new CXCR4 antagonist with sustained receptor occupancy, is active as monotherapy against T-ALL in mice. Indeed, a 2-week course of daily BL-8040 resulted in a median reduction in tumor burden of 32.1-fold (range 6.8 to 176) across 5 different T-ALL xenografts. Preliminary data from a clinical trial of BL-8040 plus nelarabine for relapsed T-ALL also suggest therapeutic activity, with a complete remission rate observed in 4/8 patients (50%), which compares favorably to published response rates of approximately 30% with single agent nelarabine. Here, we explore molecular mechanisms by which CXCR4 blockade induces T-ALL death. NOD-scid IL2Rgammanull (NSG) mice were injected with P12-Ichikawa cells, a T-ALL cell line modified to express click beetle red luciferase and GFP. Following T-ALL engraftment, mice were treated with a single dose of BL-8040, and then leukemic cells in the bone marrow harvested 24-48 hours later. Treatment with BL-8040 resulted in a marked suppression of Akt and Erk1/2 phosphorylation, suggesting that signaling through CXCR4 is the major source of PI3 kinase pathway activation in T-ALL cells. Surprisingly, treatment with BL-8040 did not affect cellular proliferation, as measured by Ki67/FxCycle Violet staining or by EdU labeling. Moreover, no increase in apoptosis, as measured by annexin V or activated caspase 3 expression, was observed. These data suggest that CXCR4 blockade induces a non-apoptotic cell death. To explore this possibility further, we performed transcriptome sequencing on T-ALL cells recovered from mice 24 hours after 1 dose of BL-8040. A total of 151 differentially expressed genes (FDR of < 0.05% and ≥ 2-fold change) were identified. Gene set enrichment analysis was strongly positive for alterations in oxidative phosphorylation, ribosome biogenesis, and carbohydrate metabolism. Ribosome function was assessed using O-propargyl-puromycin (OPP), which monitors global protein translation. No difference in global protein synthesis in T-ALL cells was observed after CXCR4 blockade in vivo. T-ALL cells are dependent on glutamine as a source of carbon, and PI3 kinase signaling positively regulates glutaminolysis. Thus, we hypothesized that CXCR4 blockade may induce T-ALL cell death by reducing glutamine metabolism. However, treatment of T-ALL cells in vitro with BL-8040 did not alter the cellular levels of glutamine or glutamate, as measured using a commercial bioluminescent assay. Confirmatory metabolic tracing studies using 13C-labeled glutamine and glucose are in progress. Finally, to explore the reduction in oxidative phosphorylation, we examined mitochondria function using Mitotracker Green. Treatment of T-ALL cells in vitro with BL-8040 for 24-48 hours induced a significant decrease in mitochondria number, suggesting induction of mitophagy. Collectively, these data suggest that T-ALL cells are addicted to CXCR4 signaling in vivo. CXCR4 blockade with BL-8040 induces a non-apoptotic cell death that is characterized by a loss of mitochondria. Disclosures Uy: Astellas: Consultancy; Pfizer: Consultancy; Curis: Consultancy; GlycoMimetics: Consultancy. Bohana-Kashtan:BiolineRx: Employment, Equity Ownership. Sorani:BiolineRx: Employment, Equity Ownership. Vainstein:BiolineRx: Employment, Equity Ownership.

scholarly journals BCR-ABL1 gene rearrangement as a subclonal change in ETV6-RUNX1–positive B-cell acute lymphoblastic leukemia

2016 ◽  
Vol 1 (2) ◽  
pp. 132-138 ◽  
Author(s):  
Karen A. Dun ◽  
Rob Vanhaeften ◽  
Tracey J. Batt ◽  
Louise A. Riley ◽  
Giuseppe Diano ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Tyrosine Kinase ◽  
B Cell ◽  
Tyrosine Kinase Inhibitors ◽  
Gene Rearrangement ◽  
Kinase Inhibitors ◽  
Lymphoblastic Leukemia ◽  
Rare Occurrence ◽  
Key Points ◽  
Cell Acute Lymphoblastic Leukemia

Key Points BCR-ABL1 rearrangement as a subclonal change in ETV6-RUNX1–positive B-ALL is a rare occurrence not previously reported. The prognosis of this rare subclonal change has not been determined, yet inclusion of tyrosine kinase inhibitors in treatment is ubiquitous.

Sign in / Sign up

Export Citation Format

Share Document