Generation and molecular recognition of melanoma-associated antigen-specific human γδ T cells

2018 ◽  
Vol 3 (30) ◽  
pp. eaav4036 ◽  
Author(s):  
Patricia M. Benveniste ◽  
Sobhan Roy ◽  
Munehide Nakatsugawa ◽  
Edward L. Y. Chen ◽  
Linh Nguyen ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell Receptors ◽  
Γδ T Cells ◽  
Antigen Recognition ◽  
Functional Responses ◽  
In Vitro System ◽  
Histocompatibility Complex ◽  
Human T Cells ◽  
Nonclassical Mhc

Antigen recognition by T cells bearing αβ T cell receptors (TCRs) is restricted by major histocompatibility complex (MHC). However, how antigens are recognized by T cells bearing γδ TCRs remains unclear. Although γδ T cells can recognize nonclassical MHC, it is generally thought that recognition of antigens is not MHC restricted. Here, we took advantage of an in vitro system to generate antigen-specific human T cells and show that melanoma-associated antigens, MART-1 and gp100, can be recognized by γδ T cells in an MHC-restricted fashion. Cloning and transferring of MART-1–specific γδ TCRs restored the specific recognition of the initial antigen MHC/peptide reactivity and conferred antigen-specific functional responses. A crystal structure of a MART-1–specific γδ TCR, together with MHC/peptide, revealed distinctive but similar docking properties to those previously reported for αβ TCRs, recognizing MART-1 on HLA-A*0201. Our work shows that antigen-specific and MHC-restricted γδ T cells can be generated in vitro and that MART-1–specific γδ T cells can also be found and cloned from the naïve repertoire. These findings reveal that classical MHC-restricted human γδ TCRs exist in the periphery and have the potential to be used in developing of new TCR-based immunotherapeutic approaches.

scholarly journals Can Glycosylation Mask the Detection of MHC Expressing p53 Peptides by T Cell Receptors?

Biomolecules ◽  
2021 ◽  
Vol 11 (7) ◽  
pp. 1056
Author(s):  
Thanh Binh Nguyen ◽  
David P. Lane ◽  
Chandra S. Verma
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Receptors ◽  
Human Leukocyte ◽  
Cell Receptors ◽  
Leukocyte Antigen ◽  
Endogenous Peptides ◽  
Histocompatibility Complex ◽  
Normal Individuals

Proteins of the major histocompatibility complex (MHC) class I, or human leukocyte antigen (HLA) in humans interact with endogenous peptides and present them to T cell receptors (TCR), which in turn tune the immune system to recognize and discriminate between self and foreign (non-self) peptides. Of especial importance are peptides derived from tumor-associated antigens. T cells recognizing these peptides are found in cancer patients, but not in cancer-free individuals. What stimulates this recognition, which is vital for the success of checkpoint based therapy? A peptide derived from the protein p53 (residues 161–169 or p161) was reported to show this behavior. T cells recognizing this unmodified peptide could be further stimulated in vitro to create effective cancer killing CTLs (cytotoxic T lymphocytes). We hypothesize that the underlying difference may arise from post-translational glycosylation of p161 in normal individuals, likely masking it against recognition by TCR. Defects in glycosylation in cancer cells may allow the presentation of the native peptide. We investigate the structural consequences of such peptide glycosylation by investigating the associated structural dynamics.

A class of γδ T cell receptors recognize the underside of the antigen-presenting molecule MR1

Science ◽  
2019 ◽  
Vol 366 (6472) ◽  
pp. 1522-1527 ◽  
Author(s):  
Jérôme Le Nours ◽  
Nicholas A. Gherardin ◽  
Sri H. Ramarathinam ◽  
Wael Awad ◽  
Florian Wiede ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Receptors ◽  
Γδ T Cells ◽  
Mhc I ◽  
Cell Receptors ◽  
Histocompatibility Complex ◽  
Antigen Complex ◽  
Binding Cleft ◽  
Antigen Presenting

T cell receptors (TCRs) recognize antigens presented by major histocompatibility complex (MHC) and MHC class I–like molecules. We describe a diverse population of human γδ T cells isolated from peripheral blood and tissues that exhibit autoreactivity to the monomorphic MHC-related protein 1 (MR1). The crystal structure of a γδTCR–MR1–antigen complex starkly contrasts with all other TCR–MHC and TCR–MHC-I-like complex structures. Namely, the γδTCR binds underneath the MR1 antigen-binding cleft, where contacts are dominated by the MR1 α3 domain. A similar pattern of reactivity was observed for diverse MR1-restricted γδTCRs from multiple individuals. Accordingly, we simultaneously report MR1 as a ligand for human γδ T cells and redefine the parameters for TCR recognition.

scholarly journals Cryopreservation of Whole Tumor Biopsies from Rectal Cancer Patients Enable Phenotypic and In Vitro Functional Evaluation of Tumor-Infiltrating T Cells

Cancers ◽  
2021 ◽  
Vol 13 (10) ◽  
pp. 2428
Author(s):  
Frank Liang ◽  
Azar Rezapour ◽  
Peter Falk ◽  
Eva Angenete ◽  
Ulf Yrlid
Keyword(s):  
Rectal Cancer ◽  
T Cells ◽  
Γδ T Cells ◽  
T Cell Subsets ◽  
Functional Evaluation ◽  
Mait Cells ◽  
Ifn Γ ◽  
Tumor Biopsies

TILs comprise functionally distinct conventional and unconventional T cell subsets and their role in responses to CRC treatments is poorly understood. We explored recovery of viable TILs from cryopreserved tumor biopsies of (chemo)-radiated patients with rectal cancer to establish a platform for retrospective TIL analyses of frozen tumors from pre-selected study cohorts. Frequencies of TIL subsets and their capacity to mount IFN-γ responses in cell suspensions of fresh vs. cryopreserved portions of the same tumor biopsies were determined for platform validation. The percentages and proportions of CD4+ TILs and CD8+ cytotoxic T lymphocytes (CTLs) among total TILs were not affected by cryopreservation. While recovery of unconventional γδ T cells and mucosal-associated invariant T cells (MAIT cells) was stable after cryopreservation, the regulatory T cells (Tregs) were reduced, but in sufficient yields for quantification. IFN-γ production by in vitro-stimulated CD4+ TILs, CTLs, γδ T cells, and MAIT cells were proportionally similar in fresh and cryopreserved tumor portions, albeit the latter displayed lower levels. Thus, the proposed platform intended for TIL analyses on cryopreserved tumor biobank biopsies holds promises for studies linking the quantity and quality of TIL subsets with specific clinical outcome after CRC treatment.

627 The DLL3-targeted half-life extended bispecific T cell engager (HLE BiTE®) immune-oncology therapy AMG 757 has potent antitumor activity in neuroendocrine cancer

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A663-A663
Author(s):  
Keegan Cooke ◽  
Juan Estrada ◽  
Jinghui Zhan ◽  
Jonathan Werner ◽  
Fei Lee ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Growth ◽  
Mouse Models ◽  
T Cell Activation ◽  
Phase 1 ◽  
Phase 1 Study ◽  
Human T Cells

BackgroundNeuroendocrine tumors (NET), including small cell lung cancer (SCLC), have poor prognosis and limited therapeutic options. AMG 757 is an HLE BiTE® immune therapy designed to redirect T cell cytotoxicity to NET cells by binding to Delta-like ligand 3 (DLL3) expressed on the tumor cell surface and CD3 on T cells.MethodsWe evaluated activity of AMG 757 in NET cells in vitro and in mouse models of neuroendocrine cancer in vivo. In vitro, co-cultures of NET cells and human T cells were treated with AMG 757 in a concentration range and T cell activation, cytokine production, and tumor cell killing were assessed. In vivo, AMG 757 antitumor efficacy was evaluated in xenograft NET and in orthotopic models designed to mimic primary and metastatic SCLC lesions. NSG mice bearing established NET were administered human T cells and then treated once weekly with AMG 757 or control HLE BiTE molecule; tumor growth inhibition was assessed. Pharmacodynamic effects of AMG 757 in tumors were also evaluated in SCLC models following a single administration of human T cells and AMG 757 or control HLE BiTE molecule.ResultsAMG 757 induced T cell activation, cytokine production, and potent T cell redirected killing of DLL3-expressing SCLC, neuroendocrine prostate cancer, and other DLL3-expressing NET cell lines in vitro. AMG 757-mediated redirected lysis was specific for DLL3-expressing cells. In patient-derived xenograft and orthotopic models of SCLC, single-dose AMG 757 effectively engaged human T cells administered systemically, leading to a significant increase in the number of human CD4+ and CD8+ T cells in primary and metastatic tumor lesions. Weekly administration of AMG 757 induced significant tumor growth inhibition of SCLC (figure 1) and other NET, including complete regression of established tumors and clearance of metastatic lesions. These findings warranted evaluation of AMG 757 (NCT03319940); the phase 1 study includes dose exploration (monotherapy and in combination with pembrolizumab) and dose expansion (monotherapy) in patients with SCLC (figure 2). A study of AMG 757 in patients with neuroendocrine prostate cancer is under development based on emerging data from the ongoing phase 1 study.Abstract 627 Figure 1AMG 757 Significantly reduced tumor growth in orthotopic SCLC mouse modelsAbstract 627 Figure 2AMG 757 Phase 1 study designConclusionsAMG 757 engages and activates T cells to kill DLL3-expressing SCLC and other NET cells in vitro and induces significant antitumor activity against established xenograft tumors in mouse models. These preclinical data support evaluation of AMG 757 in clinical studies of patients with NET.Ethics ApprovalAll in vivo work was conducted under IACUC-approved protocol #2009-00046.

γδ T cells for immune therapy of patients with lymphoid malignancies

Blood ◽  
2003 ◽  
Vol 102 (1) ◽  
pp. 200-206 ◽  
Author(s):  
Martin Wilhelm ◽  
Volker Kunzmann ◽  
Susanne Eckstein ◽  
Peter Reimer ◽  
Florian Weissinger ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Antitumor Activity ◽  
Low Dose ◽  
Γδ T Cells ◽  
Low Grade ◽  
Treatment Schedule ◽  
Γδ T Cell

Abstract There is increasing evidence that γδ T cells have potent innate antitumor activity. We described previously that synthetic aminobisphosphonates are potent γδ T cell stimulatory compounds that induce cytokine secretion (ie, interferon γ [IFN-γ]) and cell-mediated cytotoxicity against lymphoma and myeloma cell lines in vitro. To evaluate the antitumor activity of γδ T cells in vivo, we initiated a pilot study of low-dose interleukin 2 (IL-2) in combination with pamidronate in 19 patients with relapsed/refractory low-grade non-Hodgkin lymphoma (NHL) or multiple myeloma (MM). The objectives of this trial were to determine toxicity, the most effective dose for in vivo activation/proliferation of γδ T cells, and antilymphoma efficacy of the combination of pamidronate and IL-2. The first 10 patients (cohort A) who entered the study received 90 mg pamidronate intravenously on day 1 followed by increasing dose levels of continuous 24-hour intravenous (IV) infusions of IL-2 (0.25 to 3 × 106 IU/m2) from day 3 to day 8. Even at the highest IL-2 dose level in vivo, γδ T-cell activation/proliferation and response to treatment were disappointing with only 1 patient achieving stable disease. Therefore, the next 9 patients were selected by positive in vitro proliferation of γδ T cells in response to pamidronate/IL-2 and received a modified treatment schedule (6-hour bolus IV IL-2 infusions from day 1-6). In this patient group (cohort B), significant in vivo activation/proliferation of γδ T cells was observed in 5 patients (55%), and objective responses (PR) were achieved in 3 patients (33%). Only patients with significant in vivo proliferation of γδ T cells responded to treatment, indicating that γδ T cells might contribute to this antilymphoma effect. Overall, administration of pamidronate and low-dose IL-2 was well tolerated. In conclusion, this clinical trial demonstrates, for the first time, that γδ T-cell–mediated immunotherapy is feasible and can induce objective tumor responses. (Blood. 2003;102:200-206)

107 Effects of IL-2 and IL-15 on the proliferative and antitumor capacities of allogeneic CD20 CAR-engineered γδ T cells in a 3D B cell lymphoma spheroid assay

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A119-A119
Author(s):  
Lu Bai ◽  
Kevin Nishimoto ◽  
Mustafa Turkoz ◽  
Marissa Herrman ◽  
Jason Romero ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Cytokine Production ◽  
Γδ T Cells ◽  
Γδ T Cell ◽  
Car T

BackgroundAutologous chimeric antigen receptor (CAR) T cells have been shown to be efficacious for the treatment of B cell malignancies; however, widespread adoption and application of CAR T cell products still face a number of challenges. To overcome these challenges, Adicet Bio is developing an allogeneic γδ T cell-based CAR T cell platform, which capitalizes on the intrinsic abilities of Vδ1 γδ T cells to recognize and kill transformed cells in an MHC-unrestricted manner, to migrate to epithelial tissues, and to function in hypoxic conditions. To gain a better understanding of the requirements for optimal intratumoral CAR Vδ1 γδ T cell activation, proliferation, and differentiation, we developed a three-dimensional (3D) tumor spheroid assay, in which tumor cells acquire the structural organization of a solid tumor and establish a microenvironment that has oxygen and nutrient gradients. Moreover, through the addition of cytokines and/or tumor stromal cell types, the spheroid microenvironment can be modified to reflect hot or cold tumors. Here, we report on the use of a 3D CD20+ Raji lymphoma spheroid assay to evaluate the effects of IL-2 and IL-15, positive regulators of T cell homeostasis and differentiation, on the proliferative and antitumor capacities of CD20 CAR Vδ1 γδ T cells.MethodsMolecular, phenotypic, and functional profiling were performed to characterize the in vitro dynamics of the intraspheroid CD20 CAR Vδ1 γδ T cell response to target antigen in the presence of IL-2, IL-15, or no added cytokine.ResultsWhen compared to no added cytokine, the addition of IL-2 or IL-15 enhanced CD20 CAR Vδ1 γδ T cell activation, proliferation, survival, and cytokine production in a dose-dependent manner but were only able to alter the kinetics of Raji cell killing at low effector to target ratios. Notably, differential gene expression analysis using NanoString nCounter® Technology confirmed the positive effects of IL-2 or IL-15 on CAR-activated Vδ1 γδ T cells as evidenced by the upregulation of genes involved in activation, cell cycle, mitochondrial biogenesis, cytotoxicity, and cytokine production.ConclusionsTogether, these results not only show that the addition of IL-2 or IL-15 can potentiate CD20 CAR Vδ1 γδ T cell activation, proliferation, survival, and differentiation into antitumor effectors but also highlight the utility of the 3D spheroid assay as a high throughput in vitro method for assessing and predicting CAR Vδ1 γδ T cell activation, proliferation, survival, and differentiation in hot and cold tumors.

scholarly journals Efficient and Non-genotoxic RNA-Based Engineering of Human T Cells Using Tumor-Specific T Cell Receptors With Minimal TCR Mispairing

2018 ◽  
Vol 9 ◽  
Author(s):  
Diana Campillo-Davo ◽  
Fumihiro Fujiki ◽  
Johan M. J. Van den Bergh ◽  
Hans De Reu ◽  
Evelien L. J. M. Smits ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Receptors ◽  
Cell Receptors ◽  
Human T Cells
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