CXCR4 hyperactivation cooperates with TCL1 in CLL development and aggressiveness

AbstractAberrant CXCR4 activity has been implicated in lymphoma pathogenesis, disease progression, and resistance to therapies. Using a mouse model with a gain-of-function CXCR4 mutation (CXCR4C1013G) that hyperactivates CXCR4 signaling, we identified CXCR4 as a crucial activator of multiple key oncogenic pathways. CXCR4 hyperactivation resulted in an expansion of transitional B1 lymphocytes, which represent the precursors of chronic lymphocytic leukemia (CLL). Indeed, CXCR4 hyperactivation led to a significant acceleration of disease onset and a more aggressive phenotype in the murine Eµ-TCL1 CLL model. Hyperactivated CXCR4 signaling cooperated with TCL1 to cause a distinct oncogenic transcriptional program in B cells, characterized by PLK1/FOXM1-associated pathways. In accordance, Eµ-TCL1;CXCR4C1013G B cells enriched a transcriptional signature from patients with Richter’s syndrome, an aggressive transformation of CLL. Notably, MYC activation in aggressive lymphoma was associated with increased CXCR4 expression. In line with this finding, additional hyperactive CXCR4 signaling in the Eµ-Myc mouse, a model of aggressive B-cell cancer, did not impact survival. In summary, we here identify CXCR4 hyperactivation as a co-driver of an aggressive lymphoma phenotype.

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Analysis with antiidiotype antibody of a patient with chronic lymphocytic leukemia and a large cell lymphoma (Richter's syndrome)

Blood ◽

10.1182/blood.v70.1.45.45 ◽

1987 ◽

Vol 70 (1) ◽

pp. 45-50 ◽

Cited By ~ 30

Author(s):

LF Bertoli ◽

H Kubagawa ◽

GV Borzillo ◽

M Mayumi ◽

JT Prchal ◽

...

Keyword(s):

B Cells ◽

Chronic Lymphocytic Leukemia ◽

Plasma Cells ◽

Cell Lymphoma ◽

Large Cell ◽

Lymphocytic Leukemia ◽

Immunoglobulin Gene ◽

Anaplastic Lymphoma ◽

Large Cell Lymphoma ◽

Richter’S Syndrome

Abstract A murine monoclonal antibody made against an idiotypic determinant (Id) of surface IgM/IgD lambda molecules on chronic lymphocytic leukemia (CLL) cells of a 71-year-old woman was used for clonal analysis by two- color immunofluorescence. The anti-Id antibody identified IgM+/IgD+/lambda+ B cells as the predominant cell type of her CLL clone. In addition, substantial proportions of the IgG and IgA B cells and most of the IgM plasma cells in her bone marrow and blood were Id+. Six years after diagnosis, the patient died of respiratory failure due to infiltration of lungs by malignant cells. Autopsy revealed a dramatic change in the tumor cell morphology. The lungs, hilar nodes, and liver were infiltrated by a diffuse large cell lymphoma admixed with the leukemic cells. By immunohistologic staining these anaplastic lymphoma cells were IgM+/IgD-/lambda+ B cells expressing the same Id noted earlier on the CLL cells. The immunoglobulin gene rearrangement pattern on Southern blot analysis was also the same in leukemic blood cells and in the tissues involved by the lymphoma. Thus, the combination of antiidiotype and immunoglobulin gene analyses in this patient with Richter's syndrome revealed that a CLL clone, seemingly “frozen” in differentiation, was actually undergoing isotype switching, differentiation into plasma cells, and evolution into a rapidly growing and fetal lymphoma.

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Identical Clonal Origin in Three Cases of Hodgkin’s Lymphoma Variant of Richter’s Syndrome Associated with EBV Infection.

Blood ◽

10.1182/blood.v104.11.3119.3119 ◽

2004 ◽

Vol 104 (11) ◽

pp. 3119-3119

Author(s):

Pierre Feugier ◽

Nathalie Monhoven ◽

Luc Legres ◽

Louis Saout ◽

Laurent Martin ◽

...

Keyword(s):

Single Cell ◽

Hodgkin’S Lymphoma ◽

Lymphocytic Leukemia ◽

Hodgkin's Lymphoma ◽

Aggressive Lymphoma ◽

Cardiac Complications ◽

Chain Gene ◽

Ebv Infection ◽

Clonal Origin ◽

Richter’S Syndrome

Abstract Occurrence of an aggressive lymphoma in patients with chronic lymphocytic leukemia (CLL), clinically referred to as Richter’s syndrome, occasionally manifests as a lymphoproliferation resembling Hodgkin’s lymphoma (HL) and often containing the Epstein-Barr virus (EBV). Only a limited number of HL variants have been subject to informative analysis regarding EBV status and their clonal relation-ship to the CLL, with evidence of an identical clonal origin in some cases and of clonally unrelated neoplasms in most cases. We reported a clinical, pathological and molecular analysis of three new cases of Richter’s syndrome with HL features. The three patients (two males, one female), aged of 67, 70 and 74, have 4, 8 and 15-year history of CLL, respectively. At HL, clinical stages (CS) were IIIAa, IIBb and IIIBb, at HL diagnosis. All the three patients responded to HL specific chemotherapy but two of them died from infectious and cardiac complications and the last one died from HL relapse. Histologically the lesions were composite comprising areas of CLL, areas of CLL with Hodgkin Reed-Sternberg (HRS) cells and areas of HL. The HRS cells were CD45RC−/CD20−/CD79a−/CD30+/CD15+/J-chain-(all cases), Oct-2+ (case 2) and Pax-5+ (cases1 and 3). In all cases, HRS cells were positive for EBV with a latence 2 profile. Using single cell laser microdissection and PEP-PCR amplification of the immunoglobulin heavy chain gene (IgH) FR3A region of each cell type, we demonstrated a same clonal origin for the CLL cells and the HRS cells in all cases. These data were confirmed in two cases by Genescan reamplification with fluorescent FR3A primer and sequenced with BigDye Terminator chemistry. Clinicopathological presentation was similar to classical HL but persistent EBV infection is probably related to an underlined immunosuppression due to CLL and chemotherapy. Seven other cases have been reported in the literature using similar single cell techniques, four of them, EBV +, were clonally unrelated while the three other one, EBV−, were of same clonal origin. At the opposite and although our three cases are EBV+, we show that the HRS cells and CLL cells belong to the same clonal population. Our data do not support the hypothesis that EBV+ HL in CLL patients occurs as unrelated secondary neoplasm in immunocompromised patients.

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The Hsp90 Inhibitor 17-DMAG Increases SOCS3 and Regulates Cytokine Production, Migration and Cell Death in Chronic Lymphocytic Leukemia

Blood ◽

10.1182/blood.v120.21.1362.1362 ◽

2012 ◽

Vol 120 (21) ◽

pp. 1362-1362

Author(s):

Erin K Hertlein ◽

Timothy L. Chen ◽

Rainer Claus ◽

Christoph Plass ◽

Amy Lehman ◽

...

Keyword(s):

Gene Expression ◽

B Cells ◽

Gene Silencing ◽

Cell Survival ◽

Cytokine Production ◽

Transcriptional Silencing ◽

Hsp90 Inhibitor ◽

Lymphocytic Leukemia ◽

Cytokine Signaling ◽

Cxcr4 Signaling

Abstract Abstract 1362 Epigenetic or transcriptional silencing of important tumor suppressors has been described to contribute to cell survival and tumorigenesis in chronic lymphocytic leukemia (CLL). We investigated gene silencing in CLL using gene expression microarray analysis, and found that over 2000 genes are repressed more than 2-fold in CLL B cells compared to normal B cells, including genes involved in cell signaling and survival (Jun/Fos signaling, ATF family, cyclin dependent kinases and suppressors of cytokine signaling). In particular, the suppressor of cytokine signaling protein 3 (SOCS3) was decreased 60-fold in CLL B cells compared to peripheral blood B cells from normal donors. Despite this profound effect, few therapeutic approaches have focused on reversing this gene silencing in CLL. NF-κB has been shown to transcriptionally silence gene expression in several types of cancer, and our group has previously identified a similar role for this transcription factor in CLL. In addition, we have previously shown that the Hsp90 inhibitor 17-DMAG targets NF-κB signaling to induce apoptosis in CLL through transcriptional down-regulation of oncogenes such as MCL1 and BCL2. However, our microarray analysis revealed that treatment with 17-DMAG also leads to the re-expression of nearly 200 genes silenced in CLL compared to normal B cells. One of the genes significantly silenced in CLL and re-expressed by 17-DMAG is SOCS3. This increase in SOCS3 was evident as early as 8 hours following treatment with 17-DMAG, and peaking at 16–24 hours just prior to 17-DMAG induced cell death (up-regulated 5.6 fold at 8 hours, 59.8 fold at 16 hours, and 25.7 fold at 24 hours). The magnitude of induction in CLL cells was significantly greater than SOCS3 induction in peripheral blood B cells from normal donors, which correlates with a decreased apoptotic response of normal B cells to 17-DMAG (25.7 fold up-regulated in CLL versus 3.5 fold in normal B cells). While little is known about its regulation or functional impact in CLL, SOCS3 has been shown to be methylated in solid tumors as well as myeloid leukemia, leading to aberrant cytokine production and cell survival. While it is known that promoter hypermethylation and subsequent gene silencing contributes to CLL disease progression, we found that there was no significant methylation of the SOCS3 promoter in CLL compared to normal B lymphocytes, indicating an alternative mechanism of SOCS3 silencing in CLL. In order to further characterize the downstream effect of SOCS3 regulation, we investigated the pathways known to be regulated by this protein, specifically IL-6 and CXCR4 signaling. We found that 17-DMAG prevents phosphorylation of STAT3 induced by IL-6 stimulation, which leads to decreased production of pro-survival cytokines including negative feedback by decreasing IL-6 itself. While STAT3 is a known Hsp90 client protein, the effect on phosphorylation of STAT3 was evident before a decrease in the total protein was observed, indicating a distinct effect on the signaling pathway independent of Hsp90's role as a protein chaperone. SOCS3 has also been shown to prevent phosphorylation of focal adhesion kinase (FAK) and therefore block both integrin and CXCR4 signaling pathways. We found that 17-DMAG prevents constitutive phosphorylation of FAK in primary CLL cells, and subsequently reduces AKT phosphorylation following recombinant SDF-1 stimulation. In order to determine if 17-DMAG inhibits migration of CLL cells towards recombinant SDF-1 as well as the direct signaling through the CXCR4 receptor, we performed transwell migration assays and found that 17-DMAG significantly inhibits migration towards both recombinant SDF-1 and CXCL13 (migration towards CXCL12, 12.4% with Vehicle vs. 8.9% with 17-DMAG, p=0.0061, towards CXCL13, 12.4% with Vehicle vs. 6.1% with 17-DMAG, p<0.0001). Similar results were obtained by over-expression of SOCS3 in a CLL B cell line, suggesting that 17-DMAG inhibits migration through an increase in SOCS3. Based on these results, we suggest that 17-DMAG reverses gene silencing in CLL, and through re-expression SOCS3 inhibits the migration and signaling associated with SDF-1/CXCR4, an important factor in the tumor microenvironment that contributes to CLL cell survival. Therefore Hsp90 inhibitors represent a novel approach to target transcriptional silencing in CLL and other B cell lymphoproliferative disorders. Disclosures: No relevant conflicts of interest to declare.

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Origin of the Hodgkin/Reed-Sternberg Cells in Chronic Lymphocytic Leukemia With “Hodgkin's Transformation”

Blood ◽

10.1182/blood.v91.5.1757 ◽

1998 ◽

Vol 91 (5) ◽

pp. 1757-1761 ◽

Cited By ~ 99

Author(s):

Toshiyuki Ohno ◽

Bassam N. Smir ◽

Dennis D. Weisenburger ◽

Randy D. Gascoyne ◽

Steven D. Hinrichs ◽

...

Keyword(s):

B Cells ◽

Chronic Lymphocytic Leukemia ◽

Pathological Process ◽

Lymphocytic Leukemia ◽

Nucleotide Sequencing ◽

Chain Gene ◽

Large Tumor ◽

Clonal Relationship ◽

Characteristic Features ◽

Richter’S Syndrome

Abstract A lymphoma with the characteristic features of Hodgkin's disease (HD) occasionally develops in patients with B-cell chronic lymphocytic leukemia (CLL), and has been called Richter's syndrome with HD features. In such cases, large tumor cells have the morphological and immunophenotypic features of classical Hodgkin and Reed-Sternberg (H-RS) cells. However, it is not known whether the H-RS cells arise from transformation of the underlying CLL cells or from a different pathological process. We report herein a study of the clonal relationship between the CLL cells and the H-RS cells in three cases of Richter's syndrome with HD features by using a single cell assay. We isolated single CLL cells and H-RS cells from immunostained tissue sections by micromanipulation. The immunoglobulin heavy chain gene (IgH) complementarity determining region (CDR) III of each cell was amplified by the polymerase chain reaction (PCR). The products were then compared by gel electrophoresis and nucleotide sequencing. The IgH CDRIII sequences from the H-RS cells were identical to those from the CLL cells in two cases. In one case, the clonal relationship between the two types of cells could not be determined because PCR products could not be obtained from any of the H-RS cells. This study shows that the H-RS cells and the CLL cells belong to the same clonal population in some cases of Richter's syndrome with HD features. Furthermore, our findings indicate that mature B cells can undergo transformation to cells with the features of H-RS cells, in association with a cellular background typical of HD. This study also supports recent findings suggesting that the H-RS cells in classical HD are derived from transformed B cells.

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Origin of the Hodgkin/Reed-Sternberg Cells in Chronic Lymphocytic Leukemia With “Hodgkin's Transformation”

Blood ◽

10.1182/blood.v91.5.1757.1757_1757_1761 ◽

1998 ◽

Vol 91 (5) ◽

pp. 1757-1761

Author(s):

Toshiyuki Ohno ◽

Bassam N. Smir ◽

Dennis D. Weisenburger ◽

Randy D. Gascoyne ◽

Steven D. Hinrichs ◽

...

Keyword(s):

B Cells ◽

Chronic Lymphocytic Leukemia ◽

Pathological Process ◽

Lymphocytic Leukemia ◽

Nucleotide Sequencing ◽

Chain Gene ◽

Large Tumor ◽

Clonal Relationship ◽

Characteristic Features ◽

Richter’S Syndrome

A lymphoma with the characteristic features of Hodgkin's disease (HD) occasionally develops in patients with B-cell chronic lymphocytic leukemia (CLL), and has been called Richter's syndrome with HD features. In such cases, large tumor cells have the morphological and immunophenotypic features of classical Hodgkin and Reed-Sternberg (H-RS) cells. However, it is not known whether the H-RS cells arise from transformation of the underlying CLL cells or from a different pathological process. We report herein a study of the clonal relationship between the CLL cells and the H-RS cells in three cases of Richter's syndrome with HD features by using a single cell assay. We isolated single CLL cells and H-RS cells from immunostained tissue sections by micromanipulation. The immunoglobulin heavy chain gene (IgH) complementarity determining region (CDR) III of each cell was amplified by the polymerase chain reaction (PCR). The products were then compared by gel electrophoresis and nucleotide sequencing. The IgH CDRIII sequences from the H-RS cells were identical to those from the CLL cells in two cases. In one case, the clonal relationship between the two types of cells could not be determined because PCR products could not be obtained from any of the H-RS cells. This study shows that the H-RS cells and the CLL cells belong to the same clonal population in some cases of Richter's syndrome with HD features. Furthermore, our findings indicate that mature B cells can undergo transformation to cells with the features of H-RS cells, in association with a cellular background typical of HD. This study also supports recent findings suggesting that the H-RS cells in classical HD are derived from transformed B cells.

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Analysis with antiidiotype antibody of a patient with chronic lymphocytic leukemia and a large cell lymphoma (Richter's syndrome)

Blood ◽

10.1182/blood.v70.1.45.bloodjournal70145 ◽

1987 ◽

Vol 70 (1) ◽

pp. 45-50

Author(s):

LF Bertoli ◽

H Kubagawa ◽

GV Borzillo ◽

M Mayumi ◽

JT Prchal ◽

...

Keyword(s):

B Cells ◽

Chronic Lymphocytic Leukemia ◽

Plasma Cells ◽

Cell Lymphoma ◽

Large Cell ◽

Lymphocytic Leukemia ◽

Immunoglobulin Gene ◽

Anaplastic Lymphoma ◽

Large Cell Lymphoma ◽

Richter’S Syndrome

A murine monoclonal antibody made against an idiotypic determinant (Id) of surface IgM/IgD lambda molecules on chronic lymphocytic leukemia (CLL) cells of a 71-year-old woman was used for clonal analysis by two- color immunofluorescence. The anti-Id antibody identified IgM+/IgD+/lambda+ B cells as the predominant cell type of her CLL clone. In addition, substantial proportions of the IgG and IgA B cells and most of the IgM plasma cells in her bone marrow and blood were Id+. Six years after diagnosis, the patient died of respiratory failure due to infiltration of lungs by malignant cells. Autopsy revealed a dramatic change in the tumor cell morphology. The lungs, hilar nodes, and liver were infiltrated by a diffuse large cell lymphoma admixed with the leukemic cells. By immunohistologic staining these anaplastic lymphoma cells were IgM+/IgD-/lambda+ B cells expressing the same Id noted earlier on the CLL cells. The immunoglobulin gene rearrangement pattern on Southern blot analysis was also the same in leukemic blood cells and in the tissues involved by the lymphoma. Thus, the combination of antiidiotype and immunoglobulin gene analyses in this patient with Richter's syndrome revealed that a CLL clone, seemingly “frozen” in differentiation, was actually undergoing isotype switching, differentiation into plasma cells, and evolution into a rapidly growing and fetal lymphoma.

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Methylome-based cell-of-origin modeling (Methyl-COOM) identifies aberrant expression of immune regulatory molecules in CLL

10.1101/2020.02.04.933937 ◽

2020 ◽

Author(s):

Justyna A. Wierzbinska ◽

Reka Toth ◽

Naveed Ishaque ◽

Karsten Rippe ◽

Jan-Philipp Mallm ◽

...

Keyword(s):

B Cells ◽

B Cell ◽

Cell Biology ◽

Disease Onset ◽

Lymphocytic Leukemia ◽

B Cell Differentiation ◽

Cell Of Origin ◽

Aberrant Expression ◽

Aberrant Transcript ◽

Epigenetic Patterns

ABSTRACTBackgroundIn cancer, normal epigenetic patterns are disturbed and contribute to gene expression changes, disease onset and progression. The cancer epigenome is composed of the epigenetic patterns present in the tumor-initiating cell at the time of transformation, and the tumor-specific epigenetic alterations that are acquired during tumor initiation and progression. The precise dissection of these two components of the tumor epigenome will facilitate a better understanding of the biological mechanisms underlying malignant transformation. Chronic lymphocytic leukemia (CLL) originates from differentiating B cells, which undergo extensive epigenetic programming. This poses the challenge to precisely determine the epigenomic ground-state of the cell-of-origin in order to identify CLL-specific epigenetic aberrations.MethodsWe developed a linear regression model, methylome-based cell-of-origin modeling (Methyl-COOM), to map the cell-of-origin for individual CLL patients based on the continuum of epigenomic changes during normal B cell differentiation.ResultsMethyl-COOM accurately maps the cell-of-origin of CLL and identifies CLL-specific aberrant DNA methylation events that are not confounded by physiologic epigenetic B cell programming. Furthermore, Methyl-COOM unmasks abnormal action of transcription factors, altered super-enhancer activities, and aberrant transcript expression in CLL. Among the aberrantly regulated transcripts were many genes that have previously been implicated in T cell biology. Flow cytometry analysis of these markers confirmed their aberrant expression on malignant B cells at the protein level.ConclusionsMethyl-COOM analysis of CLL identified disease-specific aberrant gene regulation. The aberrantly expressed genes identified in this study might play a role in immune-evasion in CLL and might serve as novel targets for immunotherapy approaches. In summary, we propose a novel framework for in silico modeling of reference DNA methylomes and for the identification of cancer-specific epigenetic changes, a concept that can be broadly applied to other human malignancies.

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Blinatumomab for Patients with Richter's Syndrome: A Multicenter Phase 2 Trial from the Filo Group

Blood ◽

10.1182/blood-2021-147467 ◽

2021 ◽

Vol 138 (Supplement 1) ◽

pp. 3570-3570

Author(s):

Romain Guieze ◽

Loic Ysebaert ◽

Damien Roos-Weil ◽

Lysiane Molina ◽

Luc Mathieu Fornecker ◽

...

Keyword(s):

T Cell ◽

T Cell Activation ◽

Research Funding ◽

Lymphoblastic Leukemia ◽

B Cell Lymphoma ◽

Lymphocytic Leukemia ◽

Aggressive Lymphoma ◽

Phase 2 ◽

Grade 3 ◽

Richter’S Syndrome

Abstract Background Richter syndrome (RS) refers to the onset of aggressive lymphoma, mostly diffuse large B cell lymphoma (DLBCL), in patients with chronic lymphocytic leukemia (CLL). The outcome of RS patients is usually very poor with both low response rates to chemoimmunotherapy and short survival. While BCR and BCL2 inhibitors have transformed the management of CLL patients, these drugs do not prevent the onset of RS. Modulating anti-tumor immunity has recently been suggested as a promising approach in RS (Ding, 2017). Blinatumomab is a bi-specific T-cell engaging antibody construct that transiently links CD3-positive T cells to CD19-positive B-cells, inducing T-cell activation and subsequent lysis of tumor cells. It has been approved for the treatment of patients with relapsed or refractory B-ALL and has also been evaluated in the setting of persisting minimal residual disease. Recently, blinatumomab (stepwise dosing 9-28-112 μg/d) has been evaluated in patients with relapsed or refractory DLBCL and demonstrated promising results (ORR 43%) with acceptable safety (Viardot, 2016). We hypothesized that blinatumomab would improve response in RS patients failing to achieve CR after initial debulking with R-CHOP. Methods We report here the first results of a phase 2 multicenter study investigating the efficacy and safety of blinatumomab after R-CHOP debulking therapy for patients with untreated RS of DLBCL histology (NCT03931642). The patients with persisting (PR, SD) or progressive disease (PD) after 2 cycles of R-CHOP were eligible to receive an 8-week course of blinatumomab induction. Blinatumomab was administered at a stepwise dose of 9 μg/d in the first week, 28 μg/d in the second week, and 112 μg/d thereafter. The primary endpoint was CR rate according to the revised Lugano criteria after the 8-week induction course of blinatumomab. An additional 4-week consolidation cycle was optional. Allo-HSCT was further allowed for eligible patients. Results A total of 34 patients out of 41 has already been enrolled in the trial to date. Median age was 66 years (range, 38-82) and sex ratio M/F was 23/12. CLL features at baseline were as follows: 57% had 17p deletion and 67% TP53 mutations. Sixty-five percent had complex karyotype and 79% unmutated IGHV status. Median number of prior therapeutic lines for CLL was 2 (range, 0-7): 19 (54%) patients previously received chemo-immunotherapy, 23 (66%) patients were exposed to ibrutinib and 11 (31%) to venetoclax. As of the data cut-off of June 1st, 2021, the blinatumomab induction course has been completed for 18 patients. Ten patients did not receive blinatumomab for the following reasons: 7 patients achieved CR after R-CHOP, 2 patients died because of febrile neutropenia after R-CHOP and 1 patient presented severe pneumonia after R-CHOP. Three patients are still on R-CHOP and 3 others on blinatumomab to date. Regarding toxicity during blinatumomab, data are available for the 18 patients having completed the blinatumomab induction to date. All patients had at least one grade 1 adverse event (AE), 10 had grade ≥3 AE. The most common AE (> 1 case), regardless of relationship to blinatumomab, were fever (4 patients), CRS (2 patients), sepsis (2 patients), vein thrombosis (2 patients), anemia (4 patients), neutropenia (3 patients), lymphopenia (5 patients), thrombocytopenia (3 patients) and hyperglycemia (5 patients). In terms of neurologic events, 5 (28%) experienced neurotoxicity (all recovered) including grade 3 encephalopathy, grade 4 confusion, grade 3 anxiety, grade 1 myoclonus, grade 2 ataxia, grade 1 sleep disorder and grade 1 ICANS (each in 1 pt). Blinatumomab was temporarly stopped in 3 patients and permanently in 2. In terms of efficacy, after R-CHOP debulking therapy (n=31 evaluable patients), 7 patients achieved CR, 6 patients were in PR, 7 patients were stable and 11 patients were progressive. At evaluation after the blinatumomab induction (n=18 evaluable patients), 4 (22.2%) patients achieved CR, 4 (22.2%) patients PR, 2 (11.1%) patients were stable and the remaining 8 (44.5%) were progressive. Considering the whole strategy (including R-CHOP debulking) (n=28), 15 (54%) patients achieved overall response including 11 (39%) CR. Conclusions Our preliminary data suggest that blinatumomab suggests encouraging anti-tumor activity and acceptable toxicity in patients with RS. Disclosures Ysebaert: Abbvie, AstraZeneca, Janssen, Roche: Other: Advisory Board, Research Funding. Ferrant: AstraZeneca: Honoraria; Janssen: Other: Travel, Accommodations, Expenses; AbbVie: Honoraria, Other: Travel, Accommodations, Expenses. de Guibert: Janssen: Consultancy, Honoraria; AbbVie: Consultancy, Honoraria; Gilead: Consultancy, Honoraria. Laribi: Astellas Phama, Inc.: Other: Personal Fees; AstraZeneca: Other: Personal Fees; Novartis: Other: Personal Fees, Research Funding; Le Mans Hospital: Research Funding; IQONE: Other: Personal Fees; Jansen: Research Funding; BeiGene: Other: Personal Fees; Takeda: Other: Personal Fees, Research Funding; AbbVie: Other: Personal Fees, Research Funding. Feugier: Abbvie: Consultancy, Honoraria; Gilead: Consultancy, Honoraria; Amgen: Honoraria; Astrazeneca: Consultancy, Honoraria; Janssen: Consultancy, Honoraria. OffLabel Disclosure: blinatumomab is approved for acute lymphoblastic leukemia. The aim of this phase 2 study is to evaluated it in patients with Richter's syndrome.

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Association of CXCR4 with IgM and IgD BCR Isotypes: Role in B Cell Malignancies

Blood ◽

10.1182/blood-2018-99-116585 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 1852-1852

Author(s):

Eva Gentner ◽

Andrea Nicola Mazzarello ◽

Martin Becker ◽

Antonella Nicolò ◽

Valerio Renna ◽

...

Keyword(s):

B Cells ◽

B Cell ◽

Cytoplasmic Tail ◽

B Cell Lymphoma ◽

Lymphocytic Leukemia ◽

In Vitro Assays ◽

B Cell Malignancies ◽

Cxcr4 Signaling

Abstract B cell malignancies including chronic lymphocytic leukemia (CLL) and diffuse large B cell lymphoma (DLBCL) are age-associated diseases driven by clonal B cell proliferation. Signaling through B cell antigen receptor (BCRs) is dysregulated in these diseases. In addition to BCRs, chemokine receptors, such as CXCR4 and CXCR5, are used to predict clinical course. The chemokine receptor CXCR4 is expressed at different developmental stages of B cells, serving different homeostatic functions including migration. We previously reported that the cross-talk of CXCR4 and the BCR isotype IgD is supporting survival and activation of mature B cells in mice. In this process, the B cell marker and co-activator CD19 plays a pivotal role. Nevertheless, interaction of BCR with CXCR4 has not been analyzed in detail in B cell malignancies. In this study, we further elucidated the CXCR4 signaling in mouse as well as human B cell subsets including immature and mature B cells. Consistent with murine B cells, CXCR4 signaling in human B cells from healthy donors remains tightly linked to surface IgD-BCR expression, although CXCR4 is highly expressed in human IgG positive memory B cell compartment. Furthermore, proximity of CXCR4 and IgD in human mature B cells is reminiscent of that of mouse B cells. In contrast, IgD:CXCR4 proximity is skewed towards IgM:CXCR4 in CLL cells. In in vitro assays, unmutated (U)-CLL cells migrate better compared to mutated (M)-CLL. Nevertheless, our analyses reveal a frequent association of IgM:CXCR4 in M-CLL. Taking together our murine and human data, we propose that IgD:CXCR4 association is crucial for CXCR4 signaling in both CLL and healthy B cells. Apart from CXCR4, mutations within the immunoreceptor tyrosine-based activation motif (ITAM) residues of CD79a and CD79b are frequently associated with B cell malignancies including DLBCL. Knowing the potential role of BCR and its isotypes in CXCR4 induced signaling, we further analyzed the role of CD79a and CD79b. Here, we took advantage of transgenic mice, whose CD79a and CD79b cytoplasmic tails carrying ITAM motifs can be inducibly deleted. Our analysis reveals the indispensable role of the CD79b cytoplasmic tail, whose loss of function causes complete impairment of CXCR4 induced signaling in murine B cells. In contrast, loss of CD79a cytoplasmic tail partially blocks CXCR4 induced signaling, which could be rescued by CD19 co-stimulation. Extending our murine results, we established an in vitro read-out system to test the role of ITAM mutants derived from DLBCLs, as well as DLBCL-derived isotypes for analyzing their impact on CXCR4 signaling. Taking our findings together, IgD:CXCR4 association is crucial for CXCR4 signaling in CLL and healthy B cells. An increased association of IgM:CXCR4 in M-CLL compared to U-CLL suggests the necessity of IgD:CXCR4 for functional CXCR4 signaling. Furthermore, CD79b is crucial for CXCR4 induced signaling in mature B cells and loss of CD79b function abrogates CXCR4 signaling in mature B cells. Disclosures Chiorazzi: AR Pharma: Equity Ownership; Janssen, Inc: Consultancy.

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HS1 has a central role in the trafficking and homing of leukemic B cells

Blood ◽

10.1182/blood-2009-12-258814 ◽

2010 ◽

Vol 116 (18) ◽

pp. 3537-3546 ◽

Cited By ~ 44

Author(s):

Cristina Scielzo ◽

Maria T. S. Bertilaccio ◽

Giorgia Simonetti ◽

Antonis Dagklis ◽

Elisa ten Hacken ◽

...

Keyword(s):

B Cells ◽

B Lymphocytes ◽

Disease Onset ◽

Lymphocytic Leukemia ◽

In Vivo Models ◽

Cytoskeletal Remodeling ◽

Cytoskeleton Remodeling ◽

Deficient Mice

Abstract The function of the intracellular protein hematopoietic cell–specific Lyn substrate-1 (HS1) in B lymphocytes is poorly defined. To investigate its role in migration, trafficking, and homing of leukemic B lymphocytes we have used B cells from HS1−/− mice, the HS1-silenced human chronic lymphocytic leukemia (CLL) MEC1 cell line and primary leukemic B cells from patients with CLL. We have used both in vitro and in vivo models and found that the lack of expression of HS1 causes several important functional effects. In vitro, we observed an impaired cytoskeletal remodeling that resulted in diminished cell migration, abnormal cell adhesion, and increased homotypic aggregation. In vivo, immunodeficient Rag2−/−γc−/− mice injected with HS1-silenced CLL B cells showed a decreased organ infiltration with the notable exception of the bone marrow (BM). The leukemic-prone Eμ-TCL1 transgenic mice crossed with HS1-deficient mice were compared with Eμ-TCL1 mice and showed an earlier disease onset and a reduced survival. These findings show that HS1 is a central regulator of cytoskeleton remodeling that controls lymphocyte trafficking and homing and significantly influences the tissue invasion and infiltration in CLL.

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