The microenvironment differentially impairs passive and active immunotherapy in chronic lymphocytic leukaemia – CXCR4 antagonists as potential adjuvants for monoclonal antibodies

Buchner, Maike; Brantner, Philipp; Stickel, Natalie; Prinz, Gabriele; Burger, Meike; Bär, Constance; Dierks, Christine; Pfeifer, Dietmar; Ott, Ariane; Mertelsmann, Roland; Gribben, John G.; Veelken, Hendrik; Zirlik, Katja

doi:10.1111/j.1365-2141.2010.08316.x

Cited by 30 publications

(27 citation statements)

References 61 publications

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“…Consistent with previous reports, 50 coculture of Raji cells with M2 10B4 BM stromal cells increased their resistance to rituximab treatment ( Figure 5C). To determine whether the protective effect of M2 10B4 cells could be reversed by blockade of the CXCR4 receptor, Raji cells were treated with vehicle or rituximab (10 g/mL) in the presence or absence of CXCR4 pepducins.…”

Section: Cxcr4 Pepducins Reverse the Protective Effect Of Stroma And supporting

confidence: 81%

“…All patient CLL B cells tested had high CXCR4 expression levels (data not shown). Both PZ-218 and PZ-210 significantly inhibited cell migration/chemotaxis to CXCL12 with comparable IC 50 Chemotaxis of vehicle-treated cells to CXCL12 was set at 100%, and random chemotaxis (chemotaxis of cells in the absence of CXCL12) was set at 0%. Data are presented as the means Ϯ SEM of triplicate experiments with n ϭ 3 for each experiment.…”

Section: Resultsmentioning

confidence: 99%

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Targeting CXCR4 with cell-penetrating pepducins in lymphoma and lymphocytic leukemia

O’Callaghan

Lee

Nguyen

et al. 2012

Blood

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The chemokine receptor CXCR4, which normally regulates stromal stem cell interactions in the bone marrow, is highly expressed on a variety of malignant hematologic cells, including lymphoma and lymphocytic leukemias. A new treatment concept has arisen wherein CXCR4 may be an effective therapeutic target as an adjunct to treatment of hematologic neoplasms with chemo-and immunotherapy. In the present study, we developed pepducins, cell-penetrating lipopeptide antagonists of CXCR4, to interdict CXCL12-CXCR4 transmembrane signaling to intracellular G-proteins. We demonstrate that pepducins targeting the first (i1) or third (i3) intracellular loops of CXCR4 completely abrogate CXCL12-mediated cell migration of lymphocytic leukemias and lymphomas. Stromal-cell coculture protects lymphoma cells from apoptosis in response to treatment with the CD20-targeted Ab rituximab. However, combination treatment with CXCR4 pepducins and rituximab significantly increases the apoptotic effect of rituximab. Furthermore, treatment of mice bearing disseminated lymphoma xenografts with pepducins alone or in combination with rituximab significantly increased their survival. These data demonstrate that CXCL12-CXCR4 signaling can be effectively inhibited by cell-penetrating pepducins, which represents a potential new treatment strategy for lymphoid malignancies. (Blood. 2012;119(7):1717-1725) IntroductionHematologic malignancies account for almost 10% of new cancer cases in the United States each year. 1 The last decade has seen the introduction of rituximab, a humanized mAb directed against the CD20 Ag, as a treatment option for B-cell leukemia and lymphomas, and combination chemotherapy with rituximab is now standard treatment for aggressive non-Hodgkin lymphoma (NHL). 2 However, because approximately 60% of patients with aggressive NHL are not cured, new biologic therapies and targets are urgently needed to further improve overall survival.The chemokine G-protein-coupled receptor (GPCR) CXCR4 and its ligand, CXCL12 (also called stromal cell-derived factor-1␣ [SDF-1]), regulate a diverse array of cellular processes, including leukocyte trafficking, B-cell lymphopoiesis, and bone marrow myelopoiesis 3 ; survival and proliferation of hematopoietic stem cells (HSCs) 4 ; and homing of HSCs to the BM. Under normal physiologic conditions, HSCs and hematopoietic progenitor cells (HPCs) are predominantly present in the BM, where they give rise to the mature cells of the hematopoietic system that are released into the blood circulation. 5 CXCL12 is constitutively secreted at high levels by BM stromal cells, 6 and it is this chemokine gradient that retains HSCs and HPCs in the BM and regulates homing of CXCR4-expressing cells. 7 The small-molecule antagonist plerixafor (AMD3100), which targets the CXCR4/CXCL12-SDF1 signaling axis, is an effective clinical tool with which to enhance mobilization of HSCs to the peripheral blood for subsequent autologous transplantation, 8,9 and has recently been approved for use in combination with G-CSF as a stem c...

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Section: Cxcr4 Pepducins Reverse the Protective Effect Of Stroma And supporting

confidence: 81%

Section: Resultsmentioning

confidence: 99%

Targeting CXCR4 with cell-penetrating pepducins in lymphoma and lymphocytic leukemia

O’Callaghan

Lee

Nguyen

et al. 2012

Blood

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“…39,40 This is because microenvironmental signals, including interactions between CLL cells and CD40L-expressing T cells, stromal and nurse-like cells are known to increase the apoptotic threshold of CLL cells. 41,42 CD40-CD40L interactions have been shown to augment the antigen-presenting capacity of CLL cells due to increased surface expression of adhesion molecules (CD54) and co-stimulatory molecules such as CD80, CD86 and CD70 leading to CLL cell activation and protection from fludarabine-mediated killing.…”

Section: © F E R R a T A S T O R T I F O U N D A T I O Nmentioning

confidence: 99%

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confidence: 99%

Blinatumomab induces autologous T-cell killing of chronic lymphocytic leukemia cells

et al. 2013

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© F e r r a t a S t o r t i F o u n d a t i o n 2 0 1 3of CLL activation and pro-survival signals. Given these data, this approach represents a novel and promising therapeutic strategy for the treatment of CLL and, potentially, other lymphoid malignancies. Methods Blood samples from patients with chronic lymphocytic leukemiaTwenty-eight CLL patients (aged 53-83 years) were studied with full ethical approval. All blood samples from patients were obtained with informed consent (see Online Supplementary Methods). The majority of the assays were performed on fresh samples from treatment-naïve patients, except those for which the results are shown in Figure 1: in these assays, samples from patients who had received standard chemotherapy treatment were also tested. Antibodies and flow cytometryA full list of the antibodies used can be found in the Online Supplementary Methods. Blinatumomab (MT103, AMG103) was provided by Amgen Inc. (Munich, Germany). T cells were identified using CD4 and CD8 markers, while CLL cells were CD5 + CD4-CD8 -(>99% CD19 + ). Cell culturesPeripheral blood mononuclear cells (PBMC) were incubated in RPMI 1640 supplemented with 5% AB serum (AB media) for 3-7 days. Blinatumomab was added to PBMC cultures at a concentration of 10 or 100 ng/mL. Human T-cell activator CD3/CD28 dynabeads (Invitrogen) were added at 1x10 5 beads/1x10 6 PBMC as a positive control for T-cell activation. For absolute counts of T cells and CLL cells, an anti-CD3 antibody (Invitrogen) at a dose of 27.7 or 277 ng/mL was used as a positive control (same molar concentration as 10 or 100 ng/mL blinatumomab). Intracellular Ki67 stainingPBMC were labeled with antibodies against CD4, CD8, CCR7 and CD45RA, fixed and permeabilized (Fix and Perm with 1% NP40, Caltag-Medsystems, Buckingham, UK) before staining with a Ki67-FITC antibody for flow cytometric analysis. Cytokine secretion assayCytokines in tissue culture supernatants were measured using a Human Th1/Th2 11plex RTU Flowcytomix kit (eBioscience) for simultaneous detection of 11 cytokines. Intracellular cytokine staining and the determination of CD107 and granzyme B expressionPBMC were cultured (3 days) with 10 ng/mL blinatumomab, before treatment with Golgi plug and Golgi stop (BD biosciences), and incubation with an anti-CD107 antibody (5 h at 37°C). Intracellular expression of granzyme B or interferon (IFN)-g and tumor necrosis factor (TNF)-α in CD4 or CD8 T cells was measured by flow cytometry. Absolute counts of T cells and chronic lymphocytic leukemia cellsPBMC samples that had or had not been treated with blinatumomab (7 days) were surface-stained with antibodies against annexin V, CD5, CD8 and CD4. Absolute counts were determined by flow cytometry using Cytocount beads (Dako, Stockport, UK). Flow cytometry-based cytotoxicity assayMouse fibroblast cells transfected with CD40L [TL(CD40L)] and non-transfected cells (NTL) were cultured as previously described. 21 For co-cultures, irradiated (80 Gy) NTL or TL(CD40L) cells were seeded into a 48-well plate, and allow...

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“…[1][2][3][4][5][6] The combined use of ibrutinib with anti-CD20 antibodies [7][8][9] or other monoclonal antibodies (mAbs) has been suggested for the treatment of patients with CLL because they use different mechanisms for antileukemia activity. Additionally, we and others have previously shown that microenvironmental interactions protect CLL and lymphoma cells from rituximab-induced cytotoxicity [10][11][12] and chemotherapyinduced apoptosis. 11,13 The ibrutinib-induced lymphocytosis suggests that a combinatorial therapy with mAbs might overcome adhesionmediated antibody resistance and synergize with anti-CD20 mAbs.…”

Section: Introductionmentioning

confidence: 99%

Ibrutinib inhibits CD20 upregulation on CLL B cells mediated by the CXCR4/SDF-1 axis

Pavlasová

Borský

Šeda

et al. 2016

Blood

100

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Key Points• Microenvironmental interactions upregulate CD20 expression in CLL cells through the CXCR4/SDF-1 axis.• Ibrutinib treatment causes downregulation of CD20 in CLL cells.Agents targeting B-cell receptor (BCR) signaling-associated kinases such as Bruton tyrosine kinase (BTK) or phosphatidylinositol 3-kinase can induce mobilization of neoplastic B cells from the lymphoid tissues into the blood, which makes them potentially ideal to combine with anti-CD20 monoclonal antibodies (such as rituximab, obinutuzumab, or ofatumumab) for treatment of B-cell lymphomas and chronic lymphocytic leukemia (CLL). Here we show that interactions between leukemia cells and stromal cells (HS-5) upregulate CD20 on CLL cells and that administering ibrutinib downmodulates CD20 (MS4A1) expression in vivo. We observed that CLL cells that have recently exited the lymph node microenvironment and moved into the peripheral blood (CXCR4 dim CD5 bright subpopulation) have higher cell surface levels of CD20 than the cells circulating in the bloodstream for a longer time (CXCR4 bright CD5 dim cells). We found that CD20 is directly upregulated by CXCR4 ligand stromal cell-derived factor 1 (SDF-1a, CXCL12) produced by stromal cells, and BTK-inhibitor ibrutinib and CXCR4-inhibitor plerixafor block SDF-1a-mediated CD20 upregulation. Ibrutinib also downmodulated Mcl1 levels in CLL cells in vivo and in coculture with stromal cells. Overall, our study provides a first detailed mechanistic explanation of CD20 expression regulation in the context of chemokine signaling and microenvironmental interactions, which may have important implications for microenvironment-targeting therapies. (Blood. 2016;128(12):1609-1613

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The microenvironment differentially impairs passive and active immunotherapy in chronic lymphocytic leukaemia – CXCR4 antagonists as potential adjuvants for monoclonal antibodies

Cited by 30 publications

References 61 publications

Targeting CXCR4 with cell-penetrating pepducins in lymphoma and lymphocytic leukemia

Targeting CXCR4 with cell-penetrating pepducins in lymphoma and lymphocytic leukemia

Blinatumomab induces autologous T-cell killing of chronic lymphocytic leukemia cells

Ibrutinib inhibits CD20 upregulation on CLL B cells mediated by the CXCR4/SDF-1 axis

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