scholarly journals Light-based tuning of ligand half-life supports kinetic proofreading model of T cell activation

2018 ◽  
Author(s):  
Doug Tischer ◽  
Orion D. Weiner
Keyword(s):  
T Cells ◽  
T Cell ◽  
Ligand Binding ◽  
T Cell Activation ◽  
Half Life ◽  
Cell Activation ◽  
Receptor Occupancy ◽  
Biophysical Parameters ◽  
Ligand Interaction ◽  
Kinetic Proofreading

AbstractT cells are thought to discriminate stimulatory versus non-stimulatory ligands by converting small changes in ligand binding half-life to large changes in cell activation. Such a kinetic proofreading model has been difficult to test directly, as existing methods of altering ligand binding half-life also change other potentially important biophysical parameters, most notably the stability of the receptor-ligand interaction under load. Here we develop an optogenetic approach to specifically tune the binding half-life of a light-responsive ligand to a chimeric antigen receptor without changing other binding parameters. By simultaneously manipulating binding half-life while controlling for receptor occupancy, we find that signaling is strongly gated by ligand binding half-life. Our results provide direct evidence of kinetic proofreading in ligand discrimination by T cells.One Sentence SummaryDirect control of ligand binding half-life with light shows that lifetime, not occupancy, dominates T cell activation.

scholarly journals Optogenetic control shows that kinetic proofreading regulates the activity of the T cell receptor

2018 ◽  
Author(s):  
O. Sascha Yousefi ◽  
Matthias Günther ◽  
Maximilian Hörner ◽  
Julia Chalupsky ◽  
Maximilian Wess ◽  
...  
Keyword(s):  
T Cell ◽  
Ligand Binding ◽  
T Cell Receptor ◽  
T Cell Activation ◽  
Half Life ◽  
Cell Activation ◽  
Cell Receptor ◽  
Tcr Signaling ◽  
Experimental Systems ◽  
Kinetic Proofreading

AbstractThe pivotal task of the immune system is to distinguish between self and foreign antigens. The kinetic proofreading model (KPR) proposes that T cells discriminate self from foreign ligands by the different ligand binding half-lives to the T cell receptor (TCR). It is challenging to test KPR as the available experimental systems fall short of only altering the binding half-lives and keeping other parameters of the ligand-TCR interaction unchanged. We engineered an optogenetic system using the plant photoreceptor phytochrome B to selectively control the dynamics of ligand binding to the TCR by light. Combining experiments with mathematical modeling we find that the ligand-TCR interaction half-life is the decisive factor for activating downstream TCR signaling, substantiating the KPR hypothesis.One Sentence SummaryThe half-life of the ligand-T cell receptor complex determines T cell activation.

scholarly journals Light-based tuning of ligand half-life supports kinetic proofreading model of T cell signaling

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Doug K Tischer ◽  
Orion David Weiner
Keyword(s):  
T Cell ◽  
Ligand Binding ◽  
Cell Signaling ◽  
Half Life ◽  
Mechanical Stability ◽  
T Cell Signaling ◽  
Biophysical Parameters ◽  
Ligand Interaction ◽  
The Individual ◽  
Kinetic Proofreading

T cells are thought to discriminate self from foreign peptides by converting small differences in ligand binding half-life into large changes in cell signaling. Such a kinetic proofreading model has been difficult to test directly, as existing methods of altering ligand binding half-life also change other potentially important biophysical parameters, most notably the mechanical stability of the receptor-ligand interaction. Here we develop an optogenetic approach to specifically tune the binding half-life of a chimeric antigen receptor without changing other binding parameters and provide direct evidence of kinetic proofreading in T cell signaling. This half-life discrimination is executed in the proximal signaling pathway, downstream of ZAP70 recruitment and upstream of diacylglycerol accumulation. Our methods represent a general tool for temporal and spatial control of T cell signaling and extend the reach of optogenetics to probe pathways where the individual molecular kinetics, rather than the ensemble average, gates downstream signaling.

Preclinical evaluation of AMG 160, a next-generation bispecific T cell engager (BiTE) targeting the prostate-specific membrane antigen PSMA for metastatic castration-resistant prostate cancer (mCRPC).

2019 ◽  
Vol 37 (7_suppl) ◽  
pp. 301-301 ◽  
Author(s):  
Julie Bailis ◽  
Petra Deegen ◽  
Oliver Thomas ◽  
Pamela Bogner ◽  
Joachim Wahl ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Cells ◽  
T Cell Activation ◽  
Half Life ◽  
Cell Activation ◽  
Phase 1 ◽  
Xenograft Model ◽  
Cytokine Release ◽  
Single Chain

301 Background: mCRPC is a disease of high unmet medical need, especially for patients who fail novel hormonal therapies and chemotherapy. BiTE molecules provide an off the shelf therapy that activates a patient’s own immune system and redirects T cells to kill tumor cells. The BiTE mechanism of action is distinct from other immunotherapies and may unlock immune response in mCRPC. PSMA is a compelling BiTE target that is highly expressed on PCa compared to normal tissue and has increased expression in mCRPC. Methods: AMG 160 is a fully human, half-life extended (HLE) BiTE that targets PSMA on tumor cells and CD3 on T cells. AMG 160 comprises two tandem single chain variable fragments fused to an Fc domain. Results: AMG 160 binds human and non-human primate (NHP) PSMA and CD3, leading to T cell activation and proliferation and cytokine production. AMG 160 redirects T cells to kill PSMA-positive cancer cell lines in vitro, including those with low PSMA levels or androgen-independent signaling. Weekly dosing of AMG 160 induces significant antitumor activity in established PCa xenograft model. The pharmacokinetics (PK) and pharmacodynamics of AMG 160 were tested in NHP. AMG 160 treatment led to BiTE target engagement in vivo, including transient T cell activation and cytokine release in blood, and mixed cellular infiltrates in multiple organs known to express PSMA. AMG 160 treatment was well tolerated. Cytokine release associated with the first dose could be attenuated using a step dose regimen. The half-life of AMG 160 in NHP was about one week. Based on allometric scaling, the PK profile of AMG 160 may be projected to enable dosing every other week in humans. Conclusions: AMG 160 is a potent HLE BiTE with specificity for PSMA-positive tumor cells. A Phase 1 study is planned to evaluate the safety and efficacy of AMG 160 in patients with mCRPC.

scholarly journals Preclinical and Nonclinical Characterization of HPN217: A Tri-Specific T Cell Activating Construct (TriTAC) Targeting B Cell Maturation Antigen (BCMA) for the Treatment of Multiple Myeloma

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 3225-3225 ◽  
Author(s):  
Che-Leung Law ◽  
Wade Aaron ◽  
Rick Austin ◽  
Manasi Barath ◽  
Evan Callihan ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Cell ◽  
Tumor Cells ◽  
T Cell Activation ◽  
Half Life ◽  
Cell Activation ◽  
The Us ◽  
Dose Dependent

Abstract About 31,000 new cases of multiple myeloma (MM) will be diagnosed in the US in 2018. In addition to chemotherapeutic agents, several targeted therapies utilizing distinct mechanisms of action, e.g., proteasome inhibitors (bortezomib, carfilzomib, ixazomib), HDAC inhibitors (panobinostat), Cullin-RING E3 ubiquitin ligase activators (thalidomide, lenalidomide, pomalidomide), and antibodies (daratumumab, elotuzumab) have become available for treating MM. However, MM remains an incurable disease. Patients who relapse after or are refractory to standard of care treatments generally have poor prognosis. In 2018, close to 13,000 patients will die of the disease in the US. Targeting the B cell maturation antigen (BCMA), a BAFF/BLyS and APRIL receptor, for treating MM patients can provide a new treatment approach complementary to existing therapies. CAR-T therapies and an antibody-drug conjugate targeting BCMA have demonstrated early clinical success in the treatment of relapsed refractory MM (RRMM). HPN217 is a tri-specific T cell activating construct (TriTAC) consisting of three binding domains: an N-terminal single domain antibody (sdAb) that binds to human BCMA, a middle sdAb that binds to human serum albumin (HSA), and a C-terminal single chain Fv (scFv) that binds to CD3ε of the T cell receptor (TCR) complex. HPN217 is a highly stable single polypeptide of ~ 53 kDa expressed by CHO cells. Simultaneous engagement of BCMA on a target MM cell and CD3 on a T cell results in T cell activation, functional differentiation and the eventual lysis of the target MM cell. Engineering of an HSA binding domain into HPN217 represents a unique strategy in extending serum half-life, giving the TriTAC molecule a small molecular size and flexibility. This approach is different from Fc-engineering applied in other CD3-based bispecific T cell engaging molecules. The KD of HPN217 binding to recombinant human BCMA, HSA, and recombinant human CD3ε was determined to be 5.5 nM, 6 nM, and 17 nM, respectively, as measured by biolayer interferometry. Flow cytometric analysis on a panel of T cells from normal donors and BCMA positive and BCMA negative tumor cell lines confirmed binding of HPN217 to its native targets expressed on cell surface. The in vitro pharmacological activity of HPN217 was evaluated by T cell-dependent cellular cytotoxicity (TDCC) assays. In co-cultures of T cells from normal human or cynomolgus monkey donors, target tumor cells, and HSA, HPN217 mediated dose-dependent and BCMA-dependent cytotoxicity with EC50 values ranging from 0.05 to 0.7 nM. Killing was dependent on expression of BCMA on target tumor cells. Concomitant with target tumor cell killing, HPN217 also mediated dose-dependent upregulation of CD25 and CD69 on T cells in the TDCC co-cultures when BCMA positive tumor cells were presence. Consistent with the mechanism of action of CD3-based T cell engaging molecules, T cell derived cytokines, e.g., TNFα and IFNγ, were detected. Similar T cell activation could be observed using human or cynomolgus monkey whole blood as a source of T cells. Nonclinical in vivo properties of HPN217 were evaluated in xenograft models and a single dose pharmacokinetic (PK) study in cynomolgus monkeys. HPN217 mediated dose-dependent growth suppression against the RPMI-8226 MM model and Jeko-1 mantle cell lymphoma model expressing relatively low levels of 5,600 and 2,200 copies of BCMA per cell, respectively. In the PK study, a single dose of HPN217 at 0.01, 0.1, or 1 mg/kg was given to cynomolgus monkeys. HPN217 exhibited linear PK behavior over this dose range. Serum half-life was in the range of 64 to 85 hours. Serum half-life, volume of distribution, and clearance appeared to be independent of dose. HPN217 was demonstrated to be stable and remained intact up to 3 weeks in vivo as demonstrated by a functional ligand binding assay using recombinant CD3ε and BCMA, respectively, to capture and detect HPN217. Importantly, serum samples collected one week after dosing were as potent as stock HPN217 in MM tumor cell killing in TDCC assays. Collectively, preclinical and nonclinical characterization suggests that HPN217 is an efficacious novel therapeutic candidate that can provide a convenient dosing schedule for patients. A first-in-human phase 1 clinical trial is planned to evaluate HPN217 in RRMM. Disclosures Law: Harpoon Therapeutics: Employment. Aaron:Harpoon Therapeutics: Employment. Austin:Harpoon Therapeutics: Employment. Barath:Harpoon Therapeutics: Employment. Callihan:Harpoon Therapeutics: Employment. Evans:Harpoon Therapeutics: Employment. Gamez Guerrero:Harpoon Therapeutics: Employment. Hemmati:Harpoon Therapeutics: Employment. Jones:Harpoon Therapeutics: Employment. Kwant:Harpoon Therapeutics: Employment. Lao:Harpoon Therapeutics: Employment. Lemon:Harpoon Therapeutics: Employment. Patnaik:Harpoon Therapeutics: Employment. Sexton:Harpoon Therapeutics: Employment. Wesche:Harpoon Therapeutics: Employment. Xiao:Harpoon Therapeutics: Employment. Yu:Harpoon Therapeutics: Employment. Yu:Harpoon Therapeutics: Employment.

scholarly journals Probiotic supplementation reduces inflammatory profiles but does not prevent oral immune perturbations during SIV infection

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Rhianna Jones ◽  
Kyle Kroll ◽  
Courtney Broedlow ◽  
Luca Schifanella ◽  
Scott Smith ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cd4 T Cells ◽  
Cell Activation ◽  
Rhesus Macaques ◽  
Oral Microbiome ◽  
Probiotic Supplementation ◽  
Siv Infection ◽  
Anti Inflammatory

AbstractHIV/SIV infections lead to massive loss of mucosal CD4 + T cells and breakdown of the epithelial mucosa resulting in severe microbial dysbiosis and chronic immune activation that ultimately drive disease progression. Moreover, disruption of one of the most understudied mucosal environments, the oral cavity, during HIV-induced immunosuppression results in significant microbial and neoplastic co-morbidities and contributes to and predicts distal disease complications. In this study we evaluated the effects of oral probiotic supplementation (PBX), which can stimulate and augment inflammatory or anti-inflammatory pathways, on early SIV infection of rhesus macaques. Our study revealed that similar to the GI mucosae, oral CD4 + T cells were rapidly depleted, and as one of the first comprehensive analyses of the oral microflora in SIV infection, we also observed significant modulation among two genera, Porphyromonas and Actinobacillus, early after infection. Interestingly, although PBX therapy did not substantially protect against oral dysbiosis or ameliorate cell loss, it did somewhat dampen inflammation and T cell activation. Collectively, these data provide one of the most comprehensive evaluations of SIV-induced changes in oral microbiome and CD4 + T cell populations, and also suggest that oral PBX may have some anti-inflammatory properties in lentivirus infections.

scholarly journals Controlling T cells spreading, mechanics and activation by micropatterning

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Anaïs Sadoun ◽  
Martine Biarnes-Pelicot ◽  
Laura Ghesquiere-Dierickx ◽  
Ambroise Wu ◽  
Olivier Théodoly ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Young Modulus ◽  
Careful Examination ◽  
Adhesion Receptor ◽  
Force Microscopy ◽  
Atomic Force ◽  
Micro Patterns

AbstractWe designed a strategy, based on a careful examination of the activation capabilities of proteins and antibodies used as substrates for adhering T cells, coupled to protein microstamping to control at the same time the position, shape, spreading, mechanics and activation state of T cells. Once adhered on patterns, we examined the capacities of T cells to be activated with soluble anti CD3, in comparison to T cells adhered to a continuously decorated substrate with the same density of ligands. We show that, in our hand, adhering onto an anti CD45 antibody decorated surface was not affecting T cell calcium fluxes, even adhered on variable size micro-patterns. Aside, we analyzed the T cell mechanics, when spread on pattern or not, using Atomic Force Microscopy indentation. By expressing MEGF10 as a non immune adhesion receptor in T cells we measured the very same spreading area on PLL substrates and Young modulus than non modified cells, immobilized on anti CD45 antibodies, while retaining similar activation capabilities using soluble anti CD3 antibodies or through model APC contacts. We propose that our system is a way to test activation or anergy of T cells with defined adhesion and mechanical characteristics, and may allow to dissect fine details of these mechanisms since it allows to observe homogenized populations in standardized T cell activation assays.

scholarly journals Deep immune profiling of MIS-C demonstrates marked but transient immune activation compared to adult and pediatric COVID-19

2021 ◽  
Vol 6 (57) ◽  
pp. eabf7570
Author(s):  
Laura A. Vella ◽  
Josephine R. Giles ◽  
Amy E. Baxter ◽  
Derek A. Oldridge ◽  
Caroline Diorio ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cd8 T Cells ◽  
Immune Activation ◽  
Cell Activation ◽  
Vascular Complications ◽  
Adult Disease ◽  
Vasoactive Medication ◽  
Inflammatory Syndrome

Pediatric COVID-19 following SARS-CoV-2 infection is associated with fewer hospitalizations and often milder disease than in adults. A subset of children, however, present with Multisystem Inflammatory Syndrome in Children (MIS-C) that can lead to vascular complications and shock, but rarely death. The immune features of MIS-C compared to pediatric COVID-19 or adult disease remain poorly understood. We analyzed peripheral blood immune responses in hospitalized SARS-CoV-2 infected pediatric patients (pediatric COVID-19) and patients with MIS-C. MIS-C patients had patterns of T cell-biased lymphopenia and T cell activation similar to severely ill adults, and all patients with MIS-C had SARS-CoV-2 spike-specific antibodies at admission. A distinct feature of MIS-C patients was robust activation of vascular patrolling CX3CR1+ CD8+ T cells that correlated with the use of vasoactive medication. Finally, whereas pediatric COVID-19 patients with acute respiratory distress syndrome (ARDS) had sustained immune activation, MIS-C patients displayed clinical improvement over time, concomitant with decreasing immune activation. Thus, non-MIS-C versus MIS-C SARS-CoV-2 associated illnesses are characterized by divergent immune signatures that are temporally distinct from one another and implicate CD8+ T cells in the clinical presentation and trajectory of MIS-C.

scholarly journals CCL19 enhances CD8+ T-cell responses and accelerates HBV clearance

2021 ◽  
Author(s):  
Yan Yan ◽  
Wei Zhao ◽  
Wei Liu ◽  
Yan Li ◽  
Xu Wang ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Crucial Role ◽  
Cell Activation ◽  
Hbv Infection ◽  
Host Immune System ◽  
Tnf Α ◽  
Ifn Γ ◽  
High Level

Abstract Background Chemokine (C–C motif) ligand 19 (CCL19) is a leukocyte chemoattractant that plays a crucial role in cell trafficking and leukocyte activation. Dysfunctional CD8+ T cells play a crucial role in persistent HBV infection. However, whether HBV can be cleared by CCL19-activated immunity remains unclear. Methods We assessed the effects of CCL19 on the activation of PBMCs in patients with HBV infection. We also examined how CCL19 influences HBV clearance and modulates HBV-responsive T cells in a mouse model of chronic hepatitis B (CHB). In addition, C–C chemokine-receptor type 7 (CCR7) knockdown mice were used to elucidate the underlying mechanism of CCL19/CCR7 axis-induced immune activation. Results From in vitro experiments, we found that CCL19 enhanced the frequencies of Ag-responsive IFN-γ+ CD8+ T cells from patients by approximately twofold, while CCR7 knockdown (LV-shCCR7) and LY294002 partially suppressed IFN-γ secretion. In mice, CCL19 overexpression led to rapid clearance of intrahepatic HBV likely through increased intrahepatic CD8+ T-cell proportion, decreased frequency of PD-1+ CD8+ T cells in blood and compromised suppression of hepatic APCs, with lymphocytes producing a significantly high level of Ag-responsive TNF-α and IFN-γ from CD8+ T cells. In both CCL19 over expressing and CCR7 knockdown (AAV-shCCR7) CHB mice, the frequency of CD8+ T-cell activation-induced cell death (AICD) increased, and a high level of Ag-responsive TNF-α and low levels of CD8+ regulatory T (Treg) cells were observed. Conclusions Findings in this study provide insights into how CCL19/CCR7 axis modulates the host immune system, which may promote the development of immunotherapeutic strategies for HBV treatment by overcoming T-cell tolerance.

scholarly journals Immunomodulatory effects of different intravenous immunoglobulin preparations in chronic lymphocytic leukemia

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Ana Colado ◽  
Esteban Enrique Elías ◽  
Valeria Judith Sarapura Martínez ◽  
Gregorio Cordini ◽  
Pablo Morande ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
T Cell Activation ◽  
Cell Activation ◽  
Lymphocytic Leukemia ◽  
Immunomodulatory Effects ◽  
Immunoglobulin Preparations

AbstractHypogammaglobulinemia is the most frequently observed immune defect in chronic lymphocytic leukemia (CLL). Although CLL patients usually have low serum levels of all isotypes (IgG, IgM and IgA), standard immunoglobulin (Ig) preparations for replacement therapy administrated to these patients contain more than 95% of IgG. Pentaglobin is an Ig preparation of intravenous application (IVIg) enriched with IgM and IgA (IVIgGMA), with the potential benefit to restore the Ig levels of all isotypes. Because IVIg preparations at high doses have well-documented anti-inflammatory and immunomodulatory effects, we aimed to evaluate the capacity of Pentaglobin and a standard IVIg preparation to affect leukemic and T cells from CLL patients. In contrast to standard IVIg, we found that IVIgGMA did not modify T cell activation and had a lower inhibitory effect on T cell proliferation. Regarding the activation of leukemic B cells through BCR, it was similarly reduced by both IVIgGMA and IVIgG. None of these IVIg preparations modified spontaneous apoptosis of T or leukemic B cells. However, the addition of IVIgGMA on in vitro cultures decreased the apoptosis of T cells induced by the BCL-2 inhibitor, venetoclax. Importantly, IVIgGMA did not impair venetoclax-induced apoptosis of leukemic B cells. Overall, our results add new data on the effects of different preparations of IVIg in CLL, and show that the IgM/IgA enriched preparation not only affects relevant mechanisms involved in CLL pathogenesis but also has a particular profile of immunomodulatory effects on T cells that deserves further investigation.

Sign in / Sign up

Export Citation Format

Share Document