scholarly journals Identifying Immune Resistance Mechanisms to CD33/CD3 BiTE® Antibody Construct (AMG 330) Mediated Cytotoxicity

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3677-3677
Author(s):  
Christina Krupka ◽  
Thomas Köhnke ◽  
Peter Kufer ◽  
Roman Kischel ◽  
Felix S. Lichtenegger ◽  
...  
Keyword(s):  
T Cell ◽  
Cell Lines ◽  
T Cell Activation ◽  
Immune Checkpoint ◽  
Cell Activation ◽  
Constitutive Expression ◽  
Resistance Mechanisms ◽  
Adaptive Resistance ◽  
Immune Checkpoint Molecules ◽  
Immune Resistance

Abstract In our previous work, we showed elimination of primary acute myeloid leukemia (AML) cells by CD33/CD3 BiTE® antibody construct (AMG 330) mediated cytotoxicity (Krupka et al, Blood 2014). The goal of the present study was to identify innate and adaptive resistance mechanisms to AMG 330 mediated lysis of AML cells. Immune checkpoint molecules have been shown to be a highly relevant escape mechanism of malignant cells to evade innate and adaptive immunity. Previously, it was shown that AML cells upregulate the expression of inhibitory ligands in response to proinflammatory cytokines (Krönig et al, 2014). As AMG 330 mediated T-cell activation induces high levels of the proinflammatory cytokines IFNγ and TNFα, we assessed the constitutive and inducible expression profile of different immune checkpoint molecules on AML cell lines and primary AML cells, including PD-L1, HVEM, ILT3 and SLAMF7 by flow cytometry. No constitutive expression was observed for PD-L1 at time of primary diagnosis in 83.7% of the cases (103/123). In contrast, constitutive expression of HVEM and ILT3 was detected in 73.7% (42/57) and 91.9% (68/74) of patient samples, respectively. Adaptive resistance was evaluated by incubating AML cell lines and primary AML samples with IFNγ and TNFα. We observed an upregulation of PD-L1 and SLAMF7 on AML cell lines and on primary AML patient samples whereas HVEM and ILT3 did not show a significant change in expression level. To test the functional relevance of the immune checkpoint molecules upon AMG 330 mediated lysis, we used an ex vivo long term culture system that enabled us to analyse the dynamic process of receptor-ligand interaction over time. Blockade of the PD-1/PD-L1 interaction resulted in a significantly increase in AMG 330 mediated lysis of primary AML cells (n=9, p=0.03). Currently, blockade of the inducible molecule SLAMF7 in AMG 330 mediated cytotoxicity is being tested. Blocking of HVEM or ILT3 did not result in a significant increase in T cell activation and concomitant lysis of AML cells suggesting a less relevant role of HVEM and ILT3 in resistance to AMG 330 mediated cytotoxicity. The latter might also be influenced by the cytokine microenvironment which favours immune resistance of AML cells. Using a bead based multiplex assay we screened the bone marrow (BM) plasma from 16 AML patients and 3 healthy donors (HD) for the presence of 33 cytokines. The cytokine profile differed between AML patients and healthy donors (HDs). The plasma levels of IL-8, IP-10 and CXCL-16 were higher in the AML samples compared to those of HDs (p=0.0041, 0.0248 and 0.0289, respectively). In contrast, EGF, FLT3-ligand, RANTES and IL-4 were significantly lower in AML samples compared to HDs (p=0.0227, 0.0145, 0.0041 and 0.0041, respectively). However, we did observe a high inter-patient variability of cytokine composition in AML. To explore the functional relevance of the BM plasma on AMG 330 mediated cytotoxicity, cocultures of AML cell lines and HD T cells were set up using different sources of plasma including fetal calf serum (FCS) and patient derived BM plasma. Interestingly, AMG 330 mediated cytotoxicity was significantly reduced using patient derived BM plasma (n=5) compared to cultures containing FCS (n=4) (mean % lysis FCS 97.4 vs PT 70.6). This was accompanied by a considerable impairment in T-cell proliferation (mean % proliferation FKS 44.7% vs PT 26.6%). Currently, we are investigating which soluble factors are responsible for the immunosuppressive effects and if we can increase lysis efficacy and T-cell proliferation through specific blocking of them. In summary we have identified possible resistance mechanisms of AML cells to AMG 330 mediated cytotoxicity. Dynamic receptor-ligand interactions between target and effector cells as well as soluble factors contribute to AMG 330 mediated lysis of primary AML cells. We hypothesize that AMG 330 mediated cytotoxicity can be augmented through combinatorial approaches including PD-1 blockade. The significance of our findings will first be validated in an in vivo mouse model and prospectively translated into human studies. Disclosures Krupka: AMGEN Research (Munich): Research Funding. Kufer:AMGEN Research (Munich): Employment, Equity Ownership. Kischel:AMGEN Research (Munich): Employment, Equity Ownership. Subklewe:AMGEN Research (Munich): Research Funding.

scholarly journals Novel immune checkpoint targets: moving beyond PD-1 and CTLA-4

2019 ◽  
Vol 18 (1) ◽  
Author(s):  
Shuang Qin ◽  
Linping Xu ◽  
Ming Yi ◽  
Shengnan Yu ◽  
Kongming Wu ◽  
...  
Keyword(s):  
Cell Death ◽  
T Cell ◽  
Programmed Cell Death ◽  
T Cell Activation ◽  
Immune Checkpoint ◽  
Cell Activation ◽  
De Novo ◽  
Checkpoint Inhibitors ◽  
Immune Checkpoints ◽  
Immune Checkpoint Molecules

Abstract The emergence of immune checkpoint inhibitors (ICIs), mainly including anti-programmed cell death protein 1/programmed cell death ligand 1 (PD-1/PD-L1) and anti-cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) monoclonal antibodies (mAbs), has shaped therapeutic landscape of some type of cancers. Despite some ICIs have manifested compelling clinical effectiveness in certain tumor types, the majority of patients still showed de novo or adaptive resistance. At present, the overall efficiency of immune checkpoint therapy remains unsatisfactory. Exploring additional immune checkpoint molecules is a hot research topic. Recent studies have identified several new immune checkpoint targets, like lymphocyte activation gene-3 (LAG-3), T cell immunoglobulin and mucin-domain containing-3 (TIM-3), T cell immunoglobulin and ITIM domain (TIGIT), V-domain Ig suppressor of T cell activation (VISTA), and so on. The investigations about these molecules have generated promising results in preclinical studies and/or clinical trials. In this review, we discussed the structure and expression of these newly-characterized immune checkpoints molecules, presented the current progress and understanding of them. Moreover, we summarized the clinical data pertinent to these recent immune checkpoint molecules as well as their application prospects.

scholarly journals CMTM6-Deficient Monocytes in ANCA-Associated Vasculitis Fail to Present the Immune Checkpoint PD-L1

2021 ◽  
Vol 12 ◽  
Author(s):  
Markus Zeisbrich ◽  
Nina Chevalier ◽  
Bettina Sehnert ◽  
Marta Rizzi ◽  
Nils Venhoff ◽  
...  
Keyword(s):  
T Cell ◽  
T Cell Activation ◽  
Immune Checkpoint ◽  
Cell Activation ◽  
Surface Expression ◽  
Molecular Defect ◽  
Lysosomal Degradation ◽  
Lysosomal Function ◽  
Immune Checkpoint Molecules

ObjectivesANCA-associated vasculitides (AAV) affect small- and medium-sized blood vessels. In active disease, vessel wall infiltrates are mainly composed of monocytes and macrophages. Immune checkpoint molecules are crucial for the maintenance of self-tolerance and the prevention of autoimmune diseases. After checkpoint inhibitor therapy, the development of autoimmune vasculitis has been observed. However, defects of immune checkpoint molecules in AAV patients have not been identified yet.MethodsMonocytes and monocyte-derived macrophages from AAV patients and healthy age-matched controls were tested for surface expression of immunoinhibitory checkpoint programmed cell death ligand-1 (PD-L1). Using in vitro co-culture approaches, the effect of monocyte PD-L1 expression on CD4+ T cell activation and proliferation was tested.ResultsMonocytes from AAV patients displayed lower PD-L1 expression and a defective PD-L1 presentation upon activation, an effect that was correlated with disease activity. Lower PD-L1 expression was due to increased lysosomal degradation of PD-L1 in AAV monocytes. We identified a reduced expression of CMTM6, a protein protecting PD-L1 from lysosomal breakdown, as the underlying molecular defect. PD-L1low AAV monocytes showed increased stimulatory capacity and induced T cell activation and proliferation. Inhibiting lysosomal function corrected this phenotype by increasing PD-L1, thus normalizing the pro-stimulatory behavior of AAV monocytes.ConclusionsThis study identifies a defect of the immunoinhibitory checkpoint PD-L1 in monocytes from patients with AAV. Low expression of CMTM6 results in enhanced lysosomal degradation of PD-L1, thus providing insufficient negative signaling to T cells. Correcting this defect by targeting lysosomal function may represent a novel strategy to treat AAV.

EP.WE.805From bench to bedside; A rationale for Immunotherapy in the adjuvant setting for Oesophageal cancer

2021 ◽  
Vol 108 (Supplement_7) ◽  
Author(s):  
Noel Donlon ◽  
Maria Davern ◽  
Andrew Sheppard ◽  
John Reynolds ◽  
Joanne Lysaght
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Immune Checkpoint ◽  
Cell Activation ◽  
Single Agent ◽  
T Cell Subsets ◽  
Effector Memory ◽  
Post Surgery ◽  
Post Operative Period

Abstract Background Immunotherapy is being intensively investigated for its utilisation in the curative setting as a single agent and in the multimodal setting, however, the most appropriate time to incorporate ICIs remains unknown. Our study profiles systemic anti-tumour immunity perioperatively to provide a rationale for adjuvant immunotherapy. Methods Systemic immunity was immunophenotyped pre and post-oesophagectomy on days 0, 1, 3, 7 and week 6 by flow cytometry (n = 14). The frequency of circulating lymphocytes, T cells, cytotoxic and helper T lymphocytes was profiled longitudinally including the proportion of T cell subsets in circulation. This study also profiled immune checkpoint expression on circulating T cells including: PD-1, CTLA-4, TIGIT, TIM-3, LAG-3, PD-L1 and PD-L2. Markers of immunogenicity (calreticulin, HMGB1 and MIC-A/B) were also assessed. Results The frequency of circulating CD27 + T cells increases sequentially in the immediate post-operative period peaking on day 7 in OAC patients. (p < 0.01) There is a sequential decrease in the percentage of effector memory and central memory T cells in circulation and an increase in the percentage of naïve T cells in peripheral circulation of OAC patients in the immediate post-operative period. The expression of CTLA-4 on the surface of circulating CD4 + T cells decreases 6 weeks post-operatively in OAC patients. Conclusions We observed increased T cell activation and immune checkpoints immediately post-surgery with returns to baseline by week 6. These results suggest that immune checkpoint inhibitors such as anti-PD-1 may be beneficial immediately post-surgery to maintain T cell activation and prevent exhaustion of this increased population of activated T cells observed immediately post-surgery.

Abstract 1272: Myeloid leukemia cell lines suppress T cell activation and proliferation.

2013 ◽  
Author(s):  
Kazushi Tanimoto ◽  
Sawa Ito ◽  
Samantha Miner ◽  
Pawel Muranski ◽  
Nancy F. Hensel ◽  
...  
Keyword(s):  
T Cell ◽  
Cell Lines ◽  
T Cell Activation ◽  
Myeloid Leukemia ◽  
Cell Activation ◽  
Leukemia Cell ◽  
Leukemia Cell Lines ◽  
Myeloid Leukemia Cell

scholarly journals Functional Interactions Between the Thrombin Receptor and the T-Cell Antigen Receptor in Human T-Cell Lines

Blood ◽  
1997 ◽  
Vol 90 (5) ◽  
pp. 1893-1901 ◽  
Author(s):  
David E. Joyce ◽  
Yan Chen ◽  
Rochelle A. Erger ◽  
Gary A. Koretzky ◽  
Steven R. Lentz
Keyword(s):  
T Cell ◽  
Cell Lines ◽  
T Cell Activation ◽  
Cell Activation ◽  
Jurkat Cells ◽  
Thrombin Receptor ◽  
Cell Antigen Receptor ◽  
Cell Antigen ◽  
Human T Cell ◽  
T Cell Lines

Abstract The proteolytically activated thrombin receptor (TR) is expressed by T lymphocytes, which suggests that thrombin may modulate T-cell activation at sites of hemostatic stress. We examined the relationship between TR function and T-cell activation in the Jurkat human T-cell line and in T-cell lines with defined defects in T-cell antigen receptor (TCR) function. Stimulation with thrombin or the synthetic TR peptide SFLLRN produced intracellular Ca2+ transients in Jurkat cells. As the concentration of TR agonist was increased, peak Ca2+ mobilization increased, but influx of extracellular Ca2+ decreased. TR signaling was enhanced in a TCR-negative Jurkat line and in T-cell lines deficient in the tyrosine kinase lck or the tyrosine phosphatase CD45, both of which are essential for normal TCR function. TCR cross-linking with anti-CD3 IgM desensitized TR signaling in Jurkat cells, but not in CD45-deficient cells. A proteinase-activated receptor (PAR-2)–specific agonist peptide, SLIGKV, produced small Ca2+ transients in both MEG-01 human megakaryocytic cells and Jurkat cells, but was less potent than the TR-specific agonist TFRIFD in both cell types. Like TR signaling, PAR-2 signaling was enhanced in TCR-negative or lck-deficient Jurkat clones. These findings provide evidence for functional cross-talk between proteolytically activated receptors and the TCR.

The Role of Leukemia Derived B7-H1 in Tumor-T-Cell Interactions in Humans.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 4566-4566
Author(s):  
Matthias Krusch ◽  
Sabine Wintterle ◽  
Lieping Chen ◽  
Lothar Kanz ◽  
Heinz Wiendl ◽  
...  
Keyword(s):  
T Cell ◽  
Cell Lines ◽  
T Cell Activation ◽  
Cell Activation ◽  
Cytokine Production ◽  
Leukemia Cells ◽  
Human Leukemia ◽  
Activation Markers ◽  
Murine Leukemia

Abstract Objective: Expression of the B7-homologue B7-H1 (PD1-Ligand) has been proposed to enable tumor cells to evade immune surveillance. Recently, B7-H1 on murine leukemia cells was reported to mediate resistance to cytolytic T-cell destruction. In this study we investigated the expression and functional role of the B7-homologue B7-H1 in human leukemia. Patients and Methods: Leukemia cells from 20 patients and 9 human leukemia cell lines were investigated for B7-H1 expression by flow cytometry. Functional relevance of B7-H1 for tumor-immune interactions was assessed by coculture experiments using purified, alloreactive CD4 and CD8 T-cells in the presence of a neutralizing anti-B7-H1 antibody. Results: Significant B7-H1 expression levels on leukemia cells were detected in 13 of 20 patients and in 8 of 9 cell lines. In contrast to various other tumor entities and the data reported from a murine leukemia system we did not observe any significant inhibitory effect of leukemia-derived B7-H1 on CD4 and CD8 cytokine production (IFN-g, IL-2) or expression of T-cell activation markers (ICOS, CD69). In the presence of a neutralizing B7-H1 antibody (mAb 5H1) no significant changes in T cell IFN-g or IL-2 production were observed. Conclusions: Our data demonstrate that leukemia-derived B7-H1 seems to have no direct influence on T-cell activation and cytokine production in humans. Further experiments are warranted to delineate factors and characterize yet unidentified B7-H1 receptor(s) that determine inhibitory and stimulatory functions of B7-H1 in human leukemia.

scholarly journals Fucoidan-Supplemented Diet Potentiates Immune Checkpoint Blockage by Enhancing Antitumor Immunity

2021 ◽  
Vol 9 ◽  
Author(s):  
Juan Yang ◽  
Xianzhi Yang ◽  
Wenfeng Pan ◽  
Mingshuo Wang ◽  
Yuxiong Lu ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cancer Progression ◽  
T Cell Activation ◽  
Immune Checkpoint ◽  
Cell Activation ◽  
Synergistic Effects ◽  
Immune Checkpoint Blockade ◽  
Stat Pathway

Immune checkpoint blockade (ICB) therapies such as PD-1 antibodies have produced significant clinical responses in treating a variety of human malignancies, yet only a subset of cancer patients benefit from such therapy. To improve the ICB efficacy, combinations with additional therapeutics were under intensive investigation. Recently, special dietary compositions that can lower the cancer risk or inhibit cancer progression have drawn significant attention, although few were reported to show synergistic effects with ICB therapies. Interestingly, Fucoidan is naturally derived from edible brown algae and exhibits antitumor and immunomodulatory activities. Here we discover that fucoidan-supplemented diet significantly improves the antitumor activities of PD-1 antibodies in vivo. Specifically, fucoidan as a dietary ingredient strongly inhibits tumor growth when co-administrated with PD-1 antibodies, which effects can be further strengthened when fucoidan is applied before PD-1 treatments. Immune analysis revealed that fucoidan consistently promotes the activation of tumor-infiltrating CD8+ T cells, which support the evident synergies with ICB therapies. RNAseq analysis suggested that the JAK-STAT pathway is critical for fucoidan to enhance the effector function of CD8+ T cells, which could be otherwise attenuated by disruption of the T-cell receptor (TCR)/CD3 complex on the cell surface. Mechanistically, fucoidan interacts with this complex and augments TCR-mediated signaling that cooperate with the JAK-STAT pathway to stimulate T cell activation. Taken together, we demonstrated that fucoidan is a promising dietary supplement combined with ICB therapies to treat malignancies, and dissected an underappreciated mechanism for fucoidan-elicited immunomodulatory effects in cancer.

scholarly journals The Tumor Microenvironment in SCC: Mechanisms and Therapeutic Opportunities

2021 ◽  
Vol 9 ◽  
Author(s):  
Nádia Ghinelli Amôr ◽  
Paulo Sérgio da Silva Santos ◽  
Ana Paula Campanelli
Keyword(s):  
Cell Death ◽  
Monoclonal Antibodies ◽  
Tumor Microenvironment ◽  
Programmed Cell Death ◽  
T Cell Activation ◽  
Immune Checkpoint ◽  
Cell Activation ◽  
Cytotoxic T Lymphocyte ◽  
Checkpoint Inhibitors ◽  
Immune Checkpoint Molecules

Squamous cell carcinoma (SCC) is the second most common skin cancer worldwide and, despite the relatively easy visualization of the tumor in the clinic, a sizeable number of SCC patients are diagnosed at advanced stages with local invasion and distant metastatic lesions. In the last decade, immunotherapy has emerged as the fourth pillar in cancer therapy via the targeting of immune checkpoint molecules such as programmed cell-death protein-1 (PD-1), programmed cell death ligand-1 (PD-L1), and cytotoxic T-lymphocyte-associated protein 4 (CTLA-4). FDA-approved monoclonal antibodies directed against these immune targets have provide survival benefit in a growing list of cancer types. Currently, there are two immunotherapy drugs available for cutaneous SCC: cemiplimab and pembrolizumab; both monoclonal antibodies (mAb) that block PD-1 thereby promoting T-cell activation and/or function. However, the success rate of these checkpoint inhibitors currently remains around 50%, which means that half of the patients with advanced SCC experience no benefit from this treatment. This review will highlight the mechanisms by which the immune checkpoint molecules regulate the tumor microenvironment (TME), as well as the ongoing clinical trials that are employing single or combinatory therapeutic approaches for SCC immunotherapy. We also discuss the regulation of additional pathways that might promote superior therapeutic efficacy, and consequently provide increased survival for those patients that do not benefit from the current checkpoint inhibitor therapies.

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