scholarly journals Synthetic mycobacterial diacyl trehaloses reveal differential recognition by human T cell receptors and the C-type lectin Mincle

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Josephine F. Reijneveld ◽  
Mira Holzheimer ◽  
David C. Young ◽  
Kattya Lopez ◽  
Sara Suliman ◽  
...  
Keyword(s):  
T Cells ◽  
Fatty Acid ◽  
Immune System ◽  
T Cell ◽  
Cross Reactivity ◽  
Lipid Binding ◽  
Human Immune System ◽  
Human T Cell ◽  
Human T Cells ◽  
Pure Compounds

AbstractThe cell wall of Mycobacterium tuberculosis is composed of diverse glycolipids which potentially interact with the human immune system. To overcome difficulties in obtaining pure compounds from bacterial extracts, we recently synthesized three forms of mycobacterial diacyltrehalose (DAT) that differ in their fatty acid composition, DAT1, DAT2, and DAT3. To study the potential recognition of DATs by human T cells, we treated the lipid-binding antigen presenting molecule CD1b with synthetic DATs and looked for T cells that bound the complex. DAT1- and DAT2-treated CD1b tetramers were recognized by T cells, but DAT3-treated CD1b tetramers were not. A T cell line derived using CD1b-DAT2 tetramers showed that there is no cross-reactivity between DATs in an IFN-γ release assay, suggesting that the chemical structure of the fatty acid at the 3-position determines recognition by T cells. In contrast with the lack of recognition of DAT3 by human T cells, DAT3, but not DAT1 or DAT2, activates Mincle. Thus, we show that the mycobacterial lipid DAT can be both an antigen for T cells and an agonist for the innate Mincle receptor, and that small chemical differences determine recognition by different parts of the immune system.

scholarly journals Enhanced differentiation of functional human T cells in NSGW41 mice with tissue-specific expression of human interleukin-7

2020 ◽  
Author(s):  
Emilie Coppin ◽  
Bala Sai Sundarasetty ◽  
Susann Rahmig ◽  
Jonas Blume ◽  
Nikita A. Verheyden ◽  
...  
Keyword(s):  
T Cells ◽  
Cell Differentiation ◽  
T Cell ◽  
Regulatory Elements ◽  
T Cell Differentiation ◽  
Human Immune System ◽  
Specific Expression ◽  
Hematopoietic Stem ◽  
Human T Cell ◽  
Human T Cells

AbstractHumanized mouse models have become increasingly valuable tools to study human hematopoiesis and infectious diseases. However, human T cell differentiation remains inefficient. We generated mice expressing human interleukin (IL-7), a critical growth and survival factor for T cells, under the control of murine IL-7 regulatory elements. After transfer of human cord blood-derived hematopoietic stem and progenitor cells, transgenic mice on the NSGW41 background, termed NSGW41hIL7, showed elevated and prolonged human cellularity in the thymus while maintaining physiological ratios of thymocyte subsets. As a consequence, numbers of functional human T cells in the periphery were increased without evidence for pathological lymphoproliferation or aberrant expansion of effector or memory-like T cells. We conclude that the novel NSGW41hIL7 strain represents an optimized mouse model for humanization to better understand human T cell differentiation in vivo and to generate a human immune system with a better approximation of human lymphocyte ratios.

scholarly journals Diffuse Large B Cell Pdtx in Humanized Mice Are Valuable Models to Study Host-Lymphoma Interactions and Immune-Modulating Agents

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 2406-2406
Author(s):  
Giorgia Zanetti ◽  
Giuseppina Astone ◽  
Luca Cappelli ◽  
William Chiu ◽  
Maria Teresa Cacciapuoti ◽  
...  
Keyword(s):  
T Cells ◽  
Immune System ◽  
T Cell ◽  
B Cell ◽  
Research Funding ◽  
Ex Vivo ◽  
Human Immune System ◽  
Human T Cells ◽  
Human Immune

Abstract Introduction: Immunotherapy is a promising therapeutic intervention for cancer treatment. Activation of the immune system via checkpoint blockade has been shown to produce antitumor responses in patients with both solid and hematological tumors. However, many patients do not respond to checkpoint inhibitors, and additional therapies are needed to treat these patients. Testing immunotherapies requires a functional human immune system; thus, it is difficult to evaluate their effectiveness using conventional experimental models. For this reason, establishing in vivo models that closely reproduce not only human tumors, but also their interactions with the human immune system, has become mandatory. Methods: We developed a humanized mouse model and combined it with a patient-derived tumor xenograft (PDTX). Humanized mice (HuMice) were generated by transplantation of cord blood or mobilized peripheral blood CD34+ hematopoietic stem and progenitor cells into preconditioned immunodeficient mice. We compared human engraftment in 3 different mouse strains: NSG (NOD.Cg-Prkdc scidIl2rg tm1Wjl/SzJ), NSGS (NOD.Cg-Prkdc scidIl2rg tm1Wjl Tg(CMV-IL3,CSF2,KITLG)1Eav/MloySzJ) and NBSGW (NOD.Cg-Kit W-41J Tyr + Prkdc scid Il2rg tm1Wjl/ThomJ). Immune cell profiling and distribution was performed using flow cytometry and immunohistochemistry. The B cell receptor (BCR) repertoire was evaluated using an RNA-based NGS assay. To evaluate the maturation and functionality of T cells developing in HuMice we performed proliferation, degranulation and intracellular cytokine staining. Results: Two months after CD34+ cell transplantation, we observed high levels of human hematopoietic chimerism in all the 3 strains. NSGS mice supported high-level chimerism as early as 1 month after transplantation, with more than 25% of human CD45+ cells in the blood. In all mice the majority of human circulating leukocytes were CD19+ B cells. An early appearance of CD3+ human T cells was detected in NSGS mice as compared to the other strains. Notably, the T cell expansion correlated with a decrease in relative B cell abundance while the myeloid cell contribution to the graft remained steady. We documented the differentiation of CD4+ and CD8+ human T cells at a 2:1 ratio. The characterization of the T cell subsets revealed that the majority was represented by CD45RA-CCR7- effector memory cells in both the spleen and the blood of HuMice. Nevertheless, recipient mice did not exhibit overt signs of graft-versus-host disease. We also evaluated the cytotoxic potential of T cells isolated from the spleen of HuMice: ex vivo peptide antigen (i.e. EBV) presentation let to generation of effective and specific cytotoxic T-cells. After assessing a functional human immune system reconstitution in HuMice, we challenged them in vivo with low-passage tumor fragments from a diffuse large B cell lymphoma (DLBCL) PDTX. All tumor implants were successfully engrafted in both HuMice and non-humanized controls. Remarkably, all the 3 HuMice strains showed a significant reduction in the tumor volume and/or eradication compared to matched non-humanized controls. Flow cytometry analysis of the peripheral blood of humanized PDTX revealed that the tumor engraftment elicited a significant expansion of CD3+ T cells and cytotoxic CD8+ lymphocytes. Moreover, tumors developing in HuMice exhibited intermediate to high levels of tumor infiltrating T lymphocytes commingling with the neoplastic B cells, as determined by immunohistochemistry. Large areas of necrosis were often observed in PDTX of HuMice. Infiltrating CD3+ cells were TIGIT, PD-1 and Lag-3 positive, and did not efficiently proliferate ex vivo: all features consistent with an exhaustion phenotype. PDTX of HuMice often displayed larger areas of necrosis. Conclusions: Collectively, our data demonstrate that a robust reconstitution can be achieved in different strains of immunocompromised mice and that HuMice elicit effective anti-lymphoma responses. PDTX HuMice represent a powerful platform to study host-tumor interactions, and to test novel immune-based strategies (CAR-T, bifunctional Abs) and new pharmacological approaches to counteract T-cell exhaustion. Figure 1 Figure 1. Disclosures Scandura: Abbvie: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Constellation: Research Funding; MPN-RF (Foundation): Research Funding; CR&T (Foudation): Research Funding; European Leukemia net: Honoraria, Other: travel fees . Roth: Janssen: Consultancy; Merck: Consultancy.

scholarly journals Enhanced differentiation of functional human T cells in NSGW41 mice with tissue-specific expression of human interleukin-7

Leukemia ◽  
2021 ◽  
Author(s):  
Emilie Coppin ◽  
Bala Sai Sundarasetty ◽  
Susann Rahmig ◽  
Jonas Blume ◽  
Nikita A. Verheyden ◽  
...  
Keyword(s):  
T Cells ◽  
Cell Differentiation ◽  
T Cell ◽  
Regulatory Elements ◽  
T Cell Differentiation ◽  
Human Immune System ◽  
Hematopoietic Stem ◽  
Interleukin 7 ◽  
Human T Cell ◽  
Human T Cells

AbstractHumanized mouse models have become increasingly valuable tools to study human hematopoiesis and infectious diseases. However, human T-cell differentiation remains inefficient. We generated mice expressing human interleukin-7 (IL-7), a critical growth and survival factor for T cells, under the control of murine IL-7 regulatory elements. After transfer of human cord blood-derived hematopoietic stem and progenitor cells, transgenic mice on the NSGW41 background, termed NSGW41hIL7, showed elevated and prolonged human cellularity in the thymus while maintaining physiological ratios of thymocyte subsets. As a consequence, numbers of functional human T cells in the periphery were increased without evidence for pathological lymphoproliferation or aberrant expansion of effector or memory-like T cells. We conclude that the novel NSGW41hIL7 strain represents an optimized mouse model for humanization to better understand human T-cell differentiation in vivo and to generate a human immune system with a better approximation of human lymphocyte ratios.

Transient accumulation of human mature thymocytes and regulatory T cells with CD28 superagonist in “human immune system” Rag2-/-γc-/- mice

Blood ◽  
2006 ◽  
Vol 108 (1) ◽  
pp. 238-245 ◽  
Author(s):  
Nicolas Legrand ◽  
Tom Cupedo ◽  
Anja U. van Lent ◽  
Menno J. Ebeli ◽  
Kees Weijer ◽  
...  
Keyword(s):  
T Cells ◽  
Immune System ◽  
T Cell ◽  
Opportunistic Infections ◽  
Human Immune System ◽  
Independent Manner ◽  
Human T Cell ◽  
Human Immune

Efficient and quick reconstitution of T-cell compartments in lymphopenic patients is of great importance to prevent opportunistic infections, but remains difficult to achieve. Human T-cell proliferation in a T-cell-receptor (TCR)-independent manner is possible in vitro with superagonist anti-CD28 antibodies, and such molecules are therefore promising therapeutic tools. Here, we investigated the in vivo effects of superagonist anti-CD28 treatment on human developing and mature T cells, in the recently developed model of “human immune system” BALB/c Rag2-/-γc-/- mice. Our results show that superagonist anti-CD28 treatment transiently induces a 7-fold increase in thymocyte numbers and up to 18-fold accumulation of mature thymocytes. The increased thymic production lead to transient accumulation of mature T cells in the periphery at the peak of treatment effect (day 6). In addition, long-term peripheral T-cell depletion was induced. Furthermore, the concomitant selective expansion and accumulation of suppressive CD4+CD25+FoxP3+ T cells was induced in a transient manner. Superagonist anti-CD28 therapy could therefore be of clinical interest in humans, both for beneficial effect on thymic T-cell production as well as regulatory T-cell accumulation. (Blood. 2006;108:238-245)

721 AT1412, a patient-derived CD9 antibody in preclinical development promoting tumor immune infiltration and inducing tumor rejection

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A763-A763
Author(s):  
Remko Schotte ◽  
Julien Villaudy ◽  
Martijn Kedde ◽  
Wouter Pos ◽  
Daniel Go ◽  
...  
Keyword(s):  
T Cells ◽  
Immune System ◽  
T Cell ◽  
Tumor Cells ◽  
Tumor Rejection ◽  
Human Immune System ◽  
Preclinical Development ◽  
High Affinity ◽  
Human Immune ◽  
A375 Melanoma

BackgroundAdaptive immunity to cancer cells forms a crucial part of cancer immunotherapy. Recently, the importance of tumor B-cell signatures were shown to correlate with melanoma survival. We investigated whether tumor-targeting antibodies could be isolated from a patient that cured (now 13 years tumor-free) metastatic melanoma following adoptive transfer of ex vivo expanded autologous T cells.MethodsPatient‘s peripheral blood B cells were isolated and tested for the presence of tumor-reactive B cells using AIMM’s immmortalisation technology. Antibody AT1412 was identified by virtue of its differential binding to melanoma cells as compared to healthy melanocytes. AT1412 binds the tetraspanin CD9, a broadly expressed protein involved in multiple cellular activities in cancer and induces ADCC and ADCP by effector cells.ResultsSpontaneous immune rejection of tumors was observed in human immune system (HIS) mouse models implanted with CD9 genetically-disrupted A375 melanoma (A375-CD9KO) tumor cells, while A375wt cells were not cleared. Most notably, no tumor rejection of A375-CD9KO tumors was observed in NSG mice, indicating that blockade of CD9 makes tumor cells susceptible to immune rejection.CD9 has been described to regulate integrin signaling, e.g. LFA-1, VLA-4, VCAM-1 and ICAM-1. AT1412 was shown to modulate CD9 function by enhancing adhesion and transmigration of T cells to endothelial (HUVEC) cells. AT1412 was most potently enhancing transendothelial T-cell migration, in contrast to a high affinity version of AT1412 or other high affinity anti-CD9 reference antibodies (e.g. ALB6). Enhanced immune cell infiltration is also observed in immunodeficient mice harbouring a human immune system (HIS). AT1412 strongly enhanced CD8 T-cell and macrophage infiltration resulting in tumor rejection (A375 melanoma). PD-1 checkpoint blockade is further sustaining this effect. In a second melanoma model carrying a PD-1 resistant and highly aggressive tumor (SK-MEL5) AT1412 together with nivolumab was inducing full tumor rejection, while either one of the antibodies alone did not.ConclusionsThe safety of AT1412 has been assessed in preclinical development and is well tolerated up to 10 mg/kg (highest dose tested) by non human primates. AT1412 demonstrated a half-life of 8.5 days, supporting 2–3 weekly administration in humans. Besides transient thrombocytopenia no other pathological deviations were observed. No effect on coagulation parameters, bruising or bleeding were observed macro- or microscopically. The thrombocytopenia is reversible, and its recovery accelerated in those animals developing anti-drug antibodies. First in Human clinical study is planned to start early 2021.Ethics ApprovalStudy protocols were approved by the Medical Ethical Committee of the Leiden University Medical Center (Leiden, Netherlands).ConsentBlood was obtained after written informed consent by the patient.

Generation of primary antigen-specific human cytotoxic T lymphocytes in human/mouse radiation chimera

Blood ◽  
1996 ◽  
Vol 88 (2) ◽  
pp. 721-730 ◽  
Author(s):  
H Segall ◽  
I Lubin ◽  
H Marcus ◽  
A Canaan ◽  
Y Reisner
Keyword(s):  
T Cells ◽  
T Cell ◽  
Human Lymphocytes ◽  
Scid Mice ◽  
Human Antibody ◽  
Adoptive Therapy ◽  
Activation Markers ◽  
Human T Cell ◽  
Human T Cells

Severe combined immunodeficient (SCID) mice are increasingly used as hosts for the adoptive transfer of human lymphocytes. Human antibody responses can be obtained in these xenogeneic chimeras, but information about the functionality of the human T cells in SCID mice is limited and controversial. Studies using human peripheral blood lymphocytes (PBL) injected intraperitoneally (IP) into SCID mice (hu-PBL-SCID mice) have shown that human T cells from these chimeras are anergic and have a defective signaling via the T-cell receptor. In addition, their antigenic repertoire is limited to xenoreactive clones. In the present study, we tested the functionality of human T cell in a recently described chimeric model. In this system, BALB/c mice are conditioned by irradiation and then transplanted with SCID bone marrow, followed by IP injection of human PBL. Our experiments demonstrated that human T cells, recovered from these hu-PBL-BALB mice within 1 month posttransplant, proliferated and expressed activation markers upon stimulation with anti-CD3 monoclonal antibody. A vigorous antiallogeneic human cytotoxic T-lymphocyte (CTL) response could be generated in these mice by immunizing them with irradiated allogeneic cells. Moreover, anti-human immunodeficiency virus type 1 (HIV-1) Net- specific human CTLs could be generated in vivo from naive lymphocytes by immunization of mouse-human chimeras with a recombinant vaccinia-nef virus. This model may be used to evaluate potential immunomodulatory drugs or cytokines, and could provide a relevant model for testing HIV vaccines, for production of antiviral T-cell clones for adoptive therapy, and for studying human T-cell responses in vivo.

scholarly journals Single-cell transcriptomics of human T cells reveals tissue and activation signatures in health and disease

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Peter A. Szabo ◽  
Hanna Mendes Levitin ◽  
Michelle Miron ◽  
Mark E. Snyder ◽  
Takashi Senda ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Single Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Interferon Response ◽  
Functional Responses ◽  
Multiple Tumor ◽  
Human T Cell ◽  
Human T Cells

Abstract Human T cells coordinate adaptive immunity in diverse anatomic compartments through production of cytokines and effector molecules, but it is unclear how tissue site influences T cell persistence and function. Here, we use single cell RNA-sequencing (scRNA-seq) to define the heterogeneity of human T cells isolated from lungs, lymph nodes, bone marrow and blood, and their functional responses following stimulation. Through analysis of >50,000 resting and activated T cells, we reveal tissue T cell signatures in mucosal and lymphoid sites, and lineage-specific activation states across all sites including distinct effector states for CD8+ T cells and an interferon-response state for CD4+ T cells. Comparing scRNA-seq profiles of tumor-associated T cells to our dataset reveals predominant activated CD8+ compared to CD4+ T cell states within multiple tumor types. Our results therefore establish a high dimensional reference map of human T cell activation in health for analyzing T cells in disease.

scholarly journals Public and private human T-cell clones respond differentially to HCMV antigen when boosted by CD3 copotentiation

2020 ◽  
Vol 4 (21) ◽  
pp. 5343-5356
Author(s):  
Laura R. E. Becher ◽  
Wendy K. Nevala ◽  
Shari Lee Sutor ◽  
Megan Abergel ◽  
Michele M. Hoffmann ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Clonal Expansion ◽  
Antigen Receptors ◽  
Fab Fragment ◽  
Public And Private ◽  
T Cell Clones ◽  
Human T Cell ◽  
Cell Clones ◽  
Human T Cells

Abstract Human cytomegalovirus (HCMV) induces long-lasting T-cell immune responses that control but do not clear infection. Typical responses involve private T-cell clones, expressing T-cell antigen receptors (TCRs) unique to a person, and public T-cell clones with identical TCRs active in different people. Here, we report the development of a pretherapeutic immunostimulation modality against HCMV for human T cells, CD3 copotentiation, and the clonal analysis of its effects in recall assays at single-cell resolution. CD3 copotentiation of human T cells required identification of an intrinsically inert anti-CD3 Fab fragment that conditionally augmented signaling only when TCR was coengaged with antigen. When applied in recall assays, CD3 copotentiation enhanced the expansion of both public and private T-cell clones responding to autologous HLA-A2(+) antigen-presenting cells and immunodominant NLVPMVATV (NLV) peptide from HCMV pp65 protein. Interestingly, public vs private TCR expression was associated with distinct clonal expansion signatures in response to recall stimulus. This implied that besides possible differences in their generation and selection in an immune response, public and private T cells may respond differently to pharmacoimmunomodulation. Furthermore, a third clonal expansion profile was observed upon CD3 copotentiation of T-cell clones from HLA-A2(−) donors and 1 HLA-A2(+) presumed-uninfected donor, where NLV was of low intrinsic potency. We conclude that human T-cell copotentiation can increase the expansion of different classes of T-cell clones responding to recall antigens of different strengths, and this may be exploitable for therapeutic development against chronic, persistent infections such as HCMV.

scholarly journals Broadly targeted human cytomegalovirus-specific CD4+ and CD8+ T cells dominate the memory compartments of exposed subjects

2005 ◽  
Vol 202 (5) ◽  
pp. 673-685 ◽  
Author(s):  
Andrew W. Sylwester ◽  
Bridget L. Mitchell ◽  
John B. Edgar ◽  
Cara Taormina ◽  
Christian Pelte ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Human Cytomegalovirus ◽  
Cd8 T Cells ◽  
Viral Infections ◽  
Infectious Agent ◽  
Cross Reactivity ◽  
T Cell Response ◽  
Open Reading Frames ◽  
Human T Cell

Human cytomegalovirus (HCMV) infections of immunocompetent hosts are characterized by a dynamic, life-long interaction in which host immune responses, particularly of T cells, restrain viral replication and prevent disease but do not eliminate the virus or preclude transmission. Because HCMV is among the largest and most complex of known viruses, the T cell resources committed to maintaining this balance have never been characterized completely. Here, using cytokine flow cytometry and 13,687 overlapping 15mer peptides comprising 213 HCMV open reading frames (ORFs), we found that 151 HCMV ORFs were immunogenic for CD4+ and/or CD8+ T cells, and that ORF immunogenicity was influenced only modestly by ORF expression kinetics and function. We further documented that total HCMV-specific T cell responses in seropositive subjects were enormous, comprising on average ∼10% of both the CD4+ and CD8+ memory compartments in blood, whereas cross-reactive recognition of HCMV proteins in seronegative individuals was limited to CD8+ T cells and was rare. These data provide the first glimpse of the total human T cell response to a complex infectious agent and will provide insight into the rules governing immunodominance and cross-reactivity in complex viral infections of humans.

scholarly journals Inhibition or activation of human T cell receptor transfectants is controlled by defined, soluble antigen arrays.

1992 ◽  
Vol 176 (5) ◽  
pp. 1421-1430 ◽  
Author(s):  
D E Symer ◽  
R Z Dintzis ◽  
D J Diamond ◽  
H M Dintzis
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Receptor ◽  
Interleukin 2 ◽  
Cell Receptor ◽  
Jurkat Cells ◽  
Soluble Antigen ◽  
Human T Cell ◽  
Human T Cells ◽  
Binding Avidity

We present evidence that direct T cell receptor (TCR) occupancy by antigen can either activate or inhibit T cells, depending upon whether or not a threshold number of local TCRs are crosslinked by multivalent arrays of the antigen. Variants of Jurkat cells were previously transfected with TCR alpha and beta chains that bind fluorescein, yielding FL-TCR+ human T cells. The transfectants are activated upon binding soluble multivalent antigen arrays at concentrations well below those required for monovalent interactions. This activation, measured by calcium fluxes and interleukin 2 (IL-2) production, indicates the superior binding avidity of multivalent ligands. Smaller, less multivalent arrays do not activate the cells, but antagonize larger arrays, demonstrating that antigen can bind TCR as either agonist or antagonist. The balance between activation and inhibition depends upon antigen array size, ligand valence, and concentration, indicating that a threshold extent of receptor crosslinking, and not individual perturbations of single TCR, is required for activation by antigen. Approximately 100 stimulatory arrays specifically bind per FL-TCR+ cell at concentrations where IL-2 production is half-maximal.

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