Temporal multiomic modeling reveals a B-cell receptor proliferative program in chronic lymphocytic leukemia

AbstractB-cell receptor (BCR) signaling is crucial for the pathophysiology of most mature B-cell lymphomas/leukemias and has emerged as a therapeutic target whose effectiveness remains limited by the occurrence of mutations. Therefore, deciphering the cellular program activated downstream this pathway has become of paramount importance for the development of innovative therapies. Using an original ex vivo model of BCR-induced proliferation of chronic lymphocytic leukemia cells, we generated 108 temporal transcriptional and proteomic profiles from 1 h up to 4 days after BCR activation. This dataset revealed a structured temporal response composed of 13,065 transcripts and 4027 proteins, comprising a leukemic proliferative signature consisting of 430 genes and 374 proteins. Mathematical modeling of this complex cellular response further highlighted a transcriptional network driven by 14 early genes linked to proteins involved in cell proliferation. This group includes expected genes (EGR1/2, NF-kB) and genes involved in NF-kB signaling modulation (TANK, ROHF) and immune evasion (KMO, IL4I1) that have not yet been associated with leukemic cells proliferation. Our study unveils the BCR-activated proliferative genetic program in primary leukemic cells. This approach combining temporal measurements with modeling allows identifying new putative targets for innovative therapy of lymphoid malignancies and also cancers dependent on ligand–receptor interactions.

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Targeting Early Events of B-Cell Receptor Signaling in Chronic Lymphocytic Leukemia: Suppressed Syk and PLCγ2 Activities Predict Apoptotic Response of Leukemic Cells to Dasatinib

Blood ◽

10.1182/blood.v112.11.5023.5023 ◽

2008 ◽

Vol 112 (11) ◽

pp. 5023-5023

Author(s):

Y. Lynn Wang ◽

Zibo Song ◽

Pin Lu ◽

John P. Leonard ◽

Morton Coleman ◽

...

Keyword(s):

B Cells ◽

Chronic Lymphocytic Leukemia ◽

B Cell ◽

Kinase Inhibitor ◽

Ex Vivo ◽

Cell Receptor ◽

B Cell Receptor ◽

Lymphocytic Leukemia ◽

Leukemic Cells ◽

Bcr Signaling

Abstract B cell receptor (BCR) signaling plays an essential role in the pathogenesis of chronic lymphocytic leukemia. In a subset of patients with a poor clinical outcome, BCR ligation leads to increased cell metabolism and cell survival (Cancer Research66, 7158–66, 2006). Based on these findings, we tested whether targeting BCR signaling with dasatinib, an inhibitor of Src kinase, would interfere with the signaling cascade and cause death of CLL B cells. CLL leukemic cells were isolated from 34 patients and were incubated with or without dasatinib at a low dose of 128 nM. Among 34 cases, viability of leukemic cells was reduced by 2% to 90%, with an average of ~50% reduction on day 4 of ex vivo culture. Further study showed that CLL B cells undergo death by apoptosis via the intrinsic pathway which involves the generation of reactive oxygen species. Analysis of the Src family kinases showed that phosphorylation of Src, Lyn and Hck was inhibited by dasatinib not only in those cases that responded to dasatinib with apoptosis, but also in those that did not respond well (<20% apoptosis). Further analysis revealed that suppressed activity of two downstream molecules, Syk and PLC Statistical analysis showed a significant correlation between CLL dasatinib response and their IgVH mutation and ZAP70 status. Cases with worse prognoses by these criteria have a better response to the kinase inhibitor. Lastly, we have also found that ZAP70 positive cases showed a greater degree of PLC

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Transcriptional Modulation by Idelalisib Synergizes with Bendamustine in Chronic Lymphocytic Leukemia

Cancers ◽

10.3390/cancers11101519 ◽

2019 ◽

Vol 11 (10) ◽

pp. 1519 ◽

Cited By ~ 3

Author(s):

Kost ◽

Saleh ◽

Mejia ◽

Mostafizar ◽

Bouchard ◽

...

Keyword(s):

Chronic Lymphocytic Leukemia ◽

B Cell ◽

Ex Vivo ◽

Phosphatidyl Inositol ◽

Cell Receptor ◽

B Cell Receptor ◽

Lymphocytic Leukemia ◽

Interleukin 4 ◽

Cross Resistance ◽

Cellular Effects

: The phosphatidyl-inositol 3 kinase (PI3K) δ inhibitor, idelalisib (IDE), is a potent inhibitor of the B-cell receptor pathway and a novel and highly effective agent for the treatment of chronic lymphocytic leukemia (CLL). We evaluated the activities of IDE in comparison to bendamusine (BEN), a commonly used alkylating agent, in primary CLL cells ex vivo. In contrast to BEN, IDE was cytotoxic to cells from extensively-treated patients, including those with a deletion (del)17p. Cross-resistance was not observed between BEN and IDE, confirming their different modes of cytotoxicity. Marked synergy was seen between BEN and IDE, even in cases that were resistant to BEN or IDE individually, and those with deletion (del) 17p. CD40L/interleukin 4 (IL4) co-treatment mimicking the CLL microenvironment increased resistance to IDE, but synergy was retained. PI3Kδ-deficient murine splenic B cells were more resistant to IDE and showed reduced synergy with BEN, thus confirming the importance of functional PI3Kδ protein. Although IDE was observed to induce γH2AX, IDE did not enhance activation of the DNA damage response nor DNA repair activity. Interestingly, IDE decreased global RNA synthesis and was antagonistic with 5,6-Dichlorobenzimidazole 1-b-D-ribofuranoside (DRB), an inhibitor of transcription. These findings add to the increasingly complex cellular effects of IDE, and B cell receptor (BCR) inhibitors in general, in CLL.

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Leukemic B Cells from Patients with Chronic Lymphocytic Leukemia Show Anomalous Lyn Tyrosine Kinase which Contributes to Defective Apoptosis.

Blood ◽

10.1182/blood.v104.11.1917.1917 ◽

2004 ◽

Vol 104 (11) ◽

pp. 1917-1917

Author(s):

Livio Trentin ◽

Anna Maria Brunati ◽

Antonella Contri ◽

Anna Cabrelle ◽

Marta Miorin ◽

...

Keyword(s):

Chronic Lymphocytic Leukemia ◽

Tyrosine Kinase ◽

Cyclosporin A ◽

B Cell ◽

B Lymphocytes ◽

Protein Level ◽

Cell Receptor ◽

B Cell Receptor ◽

Lymphocytic Leukemia ◽

Leukemic Cells

Abstract B-cell chronic lymphocytic leukemia (B-CLL) is the most common leukemia in adults and is characterized by the accumulation of mature B lymphocytes in the G0/G1 phase of the cell cycle, expressing B-cell related (i.e. CD19, surface immunoglobulins) and unrelated molecules (CD5 and CD23). The signal transduction pathways underlying the abnormalities of these leukemic cells are poorly understood and no data are available on deregulated cell signalling in B-CLL. Since Lyn activation plays a pivotal role in the signaling cascade triggered by BCR engagement, we investigated whether this kinase may be involved in CLL pathogenesis. In this study, we investigated freshly isolated and purified malignant B cells obtained from 40 CLL patients and we observed that the Src-kinase Lyn, the switch molecule coupling B-cell-receptor to downstream signaling, displays anomalous properties. Western blot and confocal analyses demonstrated that Lyn is overexpressed at the protein level in leukemic cells as compared to normal B-lymphocytes with a substantial aliquot of the kinase anomalously present in the cytosol of leukemic cells. While in normal B lymphocytes Lyn activation is triggered by B-cell-receptor engagement with anti-IgM antibodies, in freshly isolated leukemic cells this kinase is constitutively active and accounts for high basal protein tyrosine-phosphorylation and low responsiveness to IgM-ligation. To address the question of whether the upregulation of Lyn protein and activity plays a role in the defective apoptosis of leukemic cells, we investigated the relationship between Lyn and the cell survival of malignant lymphocytes in the presence of either dexamethazone and cyclosporin A, which are known to induce apoptosis of human lymphocytes, or PP2 and SU6656, which are selective inhibitors of Lyn. When leukemic cells were cultured in the presence of cyclosporin A or dexamethazone, a marked increase in apoptosis was observed as compared to cells cultured in medium alone, and this effect correlated with a great decrease in both basal activity and protein level of Lyn. The exposure of the leukemic cells to PP2 and SU6656 caused both the inhibition of the overexpressed Lyn activity and marked cell apoptosis. These findings suggest a direct correlation between high basal Lyn activity and defects in the induction of apoptosis in leukemic cells. They also support a critical role for Lyn in B-CLL pathogenesis and identify this tyrosine kinase as a potential therapeutic target for drugs capable of inducing apoptosis in B-CLL leukemic cells.

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Targeted inhibition of eIF4A suppresses B-cell receptor-induced translation and expression of MYC and MCL1 in chronic lymphocytic leukemia cells

Cellular and Molecular Life Sciences ◽

10.1007/s00018-021-03910-x ◽

2021 ◽

Author(s):

Sarah Wilmore ◽

Karly-Rai Rogers-Broadway ◽

Joe Taylor ◽

Elizabeth Lemm ◽

Rachel Fell ◽

...

Keyword(s):

B Cells ◽

Chronic Lymphocytic Leukemia ◽

B Cell ◽

Mrna Translation ◽

Cell Receptor ◽

Memory B Cells ◽

B Cell Receptor ◽

Lymphocytic Leukemia ◽

Mrna Levels ◽

Mrna Stabilization

AbstractSignaling via the B-cell receptor (BCR) is a key driver and therapeutic target in chronic lymphocytic leukemia (CLL). BCR stimulation of CLL cells induces expression of eIF4A, an initiation factor important for translation of multiple oncoproteins, and reduces expression of PDCD4, a natural inhibitor of eIF4A, suggesting that eIF4A may be a critical nexus controlling protein expression downstream of the BCR in these cells. We, therefore, investigated the effect of eIF4A inhibitors (eIF4Ai) on BCR-induced responses. We demonstrated that eIF4Ai (silvestrol and rocaglamide A) reduced anti-IgM-induced global mRNA translation in CLL cells and also inhibited accumulation of MYC and MCL1, key drivers of proliferation and survival, respectively, without effects on upstream signaling responses (ERK1/2 and AKT phosphorylation). Analysis of normal naïve and non-switched memory B cells, likely counterparts of the two main subsets of CLL, demonstrated that basal RNA translation was higher in memory B cells, but was similarly increased and susceptible to eIF4Ai-mediated inhibition in both. We probed the fate of MYC mRNA in eIF4Ai-treated CLL cells and found that eIF4Ai caused a profound accumulation of MYC mRNA in anti-IgM treated cells. This was mediated by MYC mRNA stabilization and was not observed for MCL1 mRNA. Following drug wash-out, MYC mRNA levels declined but without substantial MYC protein accumulation, indicating that stabilized MYC mRNA remained blocked from translation. In conclusion, BCR-induced regulation of eIF4A may be a critical signal-dependent nexus for therapeutic attack in CLL and other B-cell malignancies, especially those dependent on MYC and/or MCL1.

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Chronic lymphocytic leukemia: revelations from the B-cell receptor

Blood ◽

10.1182/blood-2003-12-4312 ◽

2004 ◽

Vol 103 (12) ◽

pp. 4389-4395 ◽

Cited By ~ 286

Author(s):

Freda K. Stevenson ◽

Federico Caligaris-Cappio

Keyword(s):

Chronic Lymphocytic Leukemia ◽

B Cell ◽

Cell Receptor ◽

B Cell Receptor ◽

Lymphocytic Leukemia ◽

Cell Of Origin ◽

Regulatory B Cell ◽

V Genes

Abstract The finding that chronic lymphocytic leukemia (CLL) consists of 2 clinical subsets, distinguished by the incidence of somatic mutations in the immunoglobulin (Ig) variable region (V) genes, has clearly linked prognosis to biology. Antigen encounter by the cell of origin is indicated in both subsets by selective but distinct expression of V genes, with evidence for continuing stimulation after transformation. The key to distinctive tumor behavior likely relates to the differential ability of the B-cell receptor (BCR) to respond. Both subsets may be undergoing low-level signaling in vivo, although analysis of blood cells limits knowledge of critical events in the tissue microenvironment. Analysis of signal competence in vitro reveals that unmutated CLL generally continues to respond, whereas mutated CLL is anergized. Differential responsiveness may reflect the increased ability of post-germinal center B cells to be triggered by antigen, leading to long-term anergy. This could minimize cell division in mutated CLL and account for prognostic differences. Unifying features of CLL include low responsiveness, expression of CD25, and production of immunosuppressive cytokines. These properties are reminiscent of regulatory T cells and suggest that the cell of origin of CLL might be a regulatory B cell. Continuing regulatory activity, mediated via autoantigen, could suppress Ig production and lead to disease-associated hypogammaglobulinemia. (Blood. 2004;103:4389-4395)

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B-cell receptor signaling in chronic lymphocytic leukemia leans on Lyn

Leukemia & Lymphoma ◽

10.3109/10428194.2012.754097 ◽

2013 ◽

Vol 54 (6) ◽

pp. 1125-1126 ◽

Cited By ~ 2

Author(s):

Yair Herishanu ◽

Aaron Polliack

Keyword(s):

Chronic Lymphocytic Leukemia ◽

B Cell ◽

Cell Receptor ◽

B Cell Receptor ◽

Lymphocytic Leukemia ◽

Receptor Signaling ◽

B Cell Receptor Signaling ◽

Cell Receptor Signaling

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Silenced B-cell receptor response to autoantigen in a poor-prognostic subset of chronic lymphocytic leukemia

Haematologica ◽

10.3324/haematol.2014.106054 ◽

2014 ◽

Vol 99 (11) ◽

pp. 1722-1730 ◽

Cited By ~ 7

Author(s):

A.-C. Bergh ◽

C. Evaldsson ◽

L. B. Pedersen ◽

C. Geisler ◽

K. Stamatopoulos ◽

...

Keyword(s):

Chronic Lymphocytic Leukemia ◽

B Cell ◽

Cell Receptor ◽

B Cell Receptor ◽

Lymphocytic Leukemia ◽

Receptor Response

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Role of NFAT in Chronic Lymphocytic Leukemia and Other B-Cell Malignancies

Frontiers in Oncology ◽

10.3389/fonc.2021.651057 ◽

2021 ◽

Vol 11 ◽

Author(s):

Ilenia Sana ◽

Maria Elena Mantione ◽

Piera Angelillo ◽

Marta Muzio

Keyword(s):

Chronic Lymphocytic Leukemia ◽

B Cell ◽

Therapeutic Potential ◽

Cell Receptor ◽

Distinct Pattern ◽

B Cell Receptor ◽

Lymphocytic Leukemia ◽

B Cell Malignancies ◽

Activated T Cells ◽

Inflammatory Microenvironment

In recent years significant progress has been made in the clinical management of chronic lymphocytic leukemia (CLL) as well as other B-cell malignancies; targeting proximal B-cell receptor signaling molecules such as Bruton Tyrosine Kinase (BTK) and Phosphoinositide 3-kinase (PI3Kδ) has emerged as a successful treatment strategy. Unfortunately, a proportion of patients are still not cured with available therapeutic options, thus efforts devoted to studying and identifying new potential druggable targets are warranted. B-cell receptor stimulation triggers a complex cascade of signaling events that eventually drives the activation of downstream transcription factors including Nuclear Factor of Activated T cells (NFAT). In this review, we summarize the literature on the expression and function of NFAT family members in CLL where NFAT is not only overexpressed but also constitutively activated; NFAT controls B-cell anergy and targeting this molecule using specific inhibitors impacts on CLL cell viability. Next, we extend our analysis on other mature B-cell lymphomas where a distinct pattern of expression and activation of NFAT is reported. We discuss the therapeutic potential of strategies aimed at targeting NFAT in B-cell malignancies not overlooking the fact that NFAT may play additional roles regulating the inflammatory microenvironment.

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