Biology of porcine T lymphocytes

2006 ◽  
Vol 7 (1-2) ◽  
pp. 81-96 ◽  
Author(s):  
Wasin Charerntantanakul ◽  
James A. Roth
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Lymphocytes ◽  
Cell Subset ◽  
Γδ T Cells ◽  
T Cell Responses ◽  
T Cell Subsets ◽  
Γδ T Cell ◽  
T Cell Subset ◽  
Cell Responses

The present review concentrates on the biological aspects of porcine T lymphocytes. Their ontogeny, subpopulations, localization and trafficking, and responses to pathogens are reviewed. The development of porcine T cells begins in the liver during the first trimester of fetal life and continues in the thymus from the second trimester until after birth. Porcine T cells are divided into two lineages, based on their possession of the [@@@]\rmalpha [@@@]β or γδ T-cell receptor. Porcine [@@@]\rmalpha [@@@]β T cells recognize antigens in a major histocompatibility complex (MHC)-restricted manner, whereas the γδ T cells recognize antigens in a MHC non-restricted fashion. The CD4+CD8−and CD4+CD8loT cell subsets of [@@@]\rmalpha [@@@]β T cells recognize antigens presented in MHC class II molecules, while the CD4−CD8+T cell subset recognizes antigens presented in MHC class I molecules. Porcine [@@@]\rmalpha [@@@]β T cells localize mainly in lymphoid tissues, whereas γδ T cells predominate in the blood and intestinal epithelium of pigs. Porcine CD8+[@@@]\rmalpha [@@@]β T cells are a prominent T-cell subset during antiviral responses, while porcine CD4+[@@@]\rmalpha [@@@]β T cell responses predominantly occur in bacterial and parasitic infections. Porcine γδ T cell responses have been reported in only a few infections. Porcine T cell responses are suppressed by some viruses and bacteria. The mechanisms of T cell suppression are not entirely known but reportedly include the killing of T cells, the inhibition of T cell activation and proliferation, the inhibition of antiviral cytokine production, and the induction of immunosuppressive cytokines.

scholarly journals Characterization of Adaptive-like γδ T Cells in Ugandan Infants during Primary Cytomegalovirus Infection

Viruses ◽  
2021 ◽  
Vol 13 (10) ◽  
pp. 1987
Author(s):  
Jessica Tuengel ◽  
Sanya Ranchal ◽  
Alexandra Maslova ◽  
Gurpreet Aulakh ◽  
Maria Papadopoulou ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Γδ T Cells ◽  
T Cell Responses ◽  
Cell Receptor ◽  
T Cell Subsets ◽  
Γδ T Cell ◽  
Cmv Infection ◽  
Cell Responses ◽  
Age Dependent

Gamma-delta (γδ) T cells are unconventional T cells that help control cytomegalovirus (CMV) infection in adults. γδ T cells develop early in gestation, and a fetal public γδ T cell receptor (TCR) clonotype is detected in congenital CMV infections. However, age-dependent γδ T cell responses to primary CMV infection are not well-understood. Flow cytometry and TCR sequencing was used to comprehensively characterize γδ T cell responses to CMV infection in a cohort of 32 infants followed prospectively from birth. Peripheral blood γδ T cell frequencies increased during infancy, and were higher among CMV-infected infants relative to uninfected. Clustering analyses revealed associations between CMV infection and activation marker expression on adaptive-like Vδ1 and Vδ3, but not innate-like Vγ9Vδ2 γδ T cell subsets. Frequencies of NKG2C+CD57+ γδ T cells were temporally associated with the quantity of CMV shed in saliva by infants with primary infection. The public γδ TCR clonotype was only detected in CMV-infected infants <120 days old and at lower frequencies than previously described in fetal infections. Our findings support the notion that CMV infection drives age-dependent expansions of specific γδ T cell populations, and provide insight for novel strategies to prevent CMV transmission and disease.

scholarly journals β2 Integrins differentially regulate γδ T cell subset thymic development and peripheral maintenance

2020 ◽  
Vol 117 (36) ◽  
pp. 22367-22377
Author(s):  
Claire L. McIntyre ◽  
Leticia Monin ◽  
Jesse C. Rop ◽  
Thomas D. Otto ◽  
Carl S. Goodyear ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Ex Vivo ◽  
Cell Subset ◽  
Γδ T Cells ◽  
T Cell Subsets ◽  
Γδ T Cell ◽  
Thymic Development ◽  
T Cell Subset

The γδ T cells reside predominantly at barrier sites and play essential roles in immune protection against infection and cancer. Despite recent advances in the development of γδ T cell immunotherapy, our understanding of the basic biology of these cells, including how their numbers are regulated in vivo, remains poor. This is particularly true for tissue-resident γδ T cells. We have identified the β2family of integrins as regulators of γδ T cells. β2-integrin–deficient mice displayed a striking increase in numbers of IL-17–producing Vγ6Vδ1+γδ T cells in the lungs, uterus, and circulation. Thymic development of this population was normal. However, single-cell RNA sequencing revealed the enrichment of genes associated with T cell survival and proliferation specifically in β2-integrin–deficient IL-17+cells compared to their wild-type counterparts. Indeed, β2-integrin–deficient Vγ6+cells from the lungs showed reduced apoptosis ex vivo, suggesting that increased survival contributes to the accumulation of these cells in β2-integrin–deficient tissues. Furthermore, our data revealed an unexpected role for β2integrins in promoting the thymic development of the IFNγ-producing CD27+Vγ4+γδ T cell subset. Together, our data reveal that β2integrins are important regulators of γδ T cell homeostasis, inhibiting the survival of IL-17–producing Vγ6Vδ1+cells and promoting the thymic development of the IFNγ-producing Vγ4+subset. Our study introduces unprecedented mechanisms of control for γδ T cell subsets.

scholarly journals Reconstitution of T Cell Subsets Following Allogeneic Hematopoietic Cell Transplantation

Cancers ◽  
2020 ◽  
Vol 12 (7) ◽  
pp. 1974 ◽  
Author(s):  
Linde Dekker ◽  
Coco de Koning ◽  
Caroline Lindemans ◽  
Stefan Nierkens
Keyword(s):  
Overall Survival ◽  
T Cells ◽  
T Cell ◽  
Cell Transplantation ◽  
Hematopoietic Cell Transplantation ◽  
Cell Subset ◽  
Γδ T Cells ◽  
Hematopoietic Cell ◽  
T Cell Subsets ◽  
T Cell Subset

Allogeneic (allo) hematopoietic cell transplantation (HCT) is the only curative treatment option for patients suffering from chemotherapy-refractory or relapsed hematological malignancies. The occurrence of morbidity and mortality after allo-HCT is still high. This is partly correlated with the immunological recovery of the T cell subsets, of which the dynamics and relations to complications are still poorly understood. Detailed information on T cell subset recovery is crucial to provide tools for better prediction and modulation of adverse events. Here, we review the current knowledge regarding CD4+ and CD8+ T cells, γδ T cells, iNKT cells, Treg cells, MAIT cells and naive and memory T cell reconstitution, as well as their relations to outcome, considering different cell sources and immunosuppressive therapies. We conclude that the T cell subsets reconstitute in different ways and are associated with distinct adverse and beneficial events; however, adequate reconstitution of all the subsets is associated with better overall survival. Although the exact mechanisms involved in the reconstitution of each T cell subset and their associations with allo-HCT outcome need to be further elucidated, the data and suggestions presented here point towards the development of individualized approaches to improve their reconstitution. This includes the modulation of immunotherapeutic interventions based on more detailed immune monitoring, aiming to improve overall survival changes.

Characterization of a Unique γδ T-Cell Subset as a Specific Marker of Cytomegalovirus Infection Severity

2020 ◽  
Author(s):  
Hannah Kaminski ◽  
Coline Ménard ◽  
Bouchra El Hayani ◽  
And-Nan Adjibabi ◽  
Gabriel Marsères ◽  
...  
Keyword(s):  
Immune Response ◽  
T Cells ◽  
T Cell ◽  
Cell Subset ◽  
Γδ T Cells ◽  
Specific Marker ◽  
Dependent Manner ◽  
Γδ T Cell ◽  
T Cell Subset ◽  
Cmv Disease

Abstract Cytomegalovirus (CMV) is a major infectious cause of death and disease after transplantation. We have previously demonstrated that the tissue-associated adaptive Vδ2neg γδ T cells are key effectors responding to CMV and associated with recovery, contrasting with their innatelike circulating counterparts, the Vγ9posVδ2pos T cells that respond to phosphoantigens but not to CMV. A third Vγ9negVδ2pos subgroup with adaptive functions has been described in adults. In the current study, we demonstrate that these Vγ9negVδ2pos T cells are also components of the CMV immune response while presenting with distinct characteristics from Vδ2neg γδ T cells. In a cohort of kidney transplant recipients, CMV seropositivity was the unique clinical parameter associated with Vγ9negVδ2pos T-cell expansion and differentiation. Extensive phenotyping demonstrated their substantial cytotoxic potential and activation during acute CMV primary infection or reinfection. In vitro, Vγ9negVδ2pos T cells responded specifically to CMV-infected cells in a T-cell receptor–dependent manner and through strong interferon γ production. Finally, Vγ9negVδ2pos T cells were the only γδ T-cell subset in which expansion was tightly correlated with the severity of CMV disease. To conclude, our results identify a new player in the immune response against CMV and open interesting clinical perspectives for using Vγ9negVδ2pos T cells as an immune marker for CMV disease severity in immunocompromised patients.

scholarly journals Characterization of local and circulating bovine γδ T cell responses to respiratory BCG vaccination

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Mariana Guerra-Maupome ◽  
Jodi L. McGill
Keyword(s):  
T Cells ◽  
T Cell ◽  
Γδ T Cells ◽  
T Cell Responses ◽  
Memory Cell ◽  
Effector Memory ◽  
Γδ T Cell ◽  
Respiratory Mucosa ◽  
Bcg Vaccination ◽  
Cell Responses

Abstract The Mycobacterium bovis Bacillus Calmette-Guerin (BCG) vaccine is administered parenterally to infants and young children to prevent tuberculosis (TB) infection. However, the protection induced by BCG is highly variable and the vaccine does not prevent pulmonary TB, the most common form of the illness. Until improved TB vaccines are available, it is crucial to use BCG in a manner which ensures optimal vaccine performance. Immunization directly to the respiratory mucosa has been shown to promote greater protection from TB in animal models. γδ T cells play a major role in host defense at mucosal sites and are known to respond robustly to mycobacterial infection. Their positioning in the respiratory mucosa ensures their engagement in the response to aerosolized TB vaccination. However, our understanding of the effect of respiratory BCG vaccination on γδ T cell responses in the lung is unknown. In this study, we used a calf model to investigate the immunogenicity of aerosol BCG vaccination, and the phenotypic profile of peripheral and mucosal γδ T cells responding to vaccination. We observed robust local and systemic M. bovis-specific IFN-γ and IL-17 production by both γδ and CD4 T cells. Importantly, BCG vaccination induced effector and memory cell differentiation of γδ T cells in both the lower airways and peripheral blood, with accumulation of a large proportion of effector memory γδ T cells in both compartments. Our results demonstrate the potential of the neonatal calf model to evaluate TB vaccine candidates that are to be administered via the respiratory tract, and suggest that aerosol immunization is a promising strategy for engaging γδ T cells in vaccine-induced immunity against TB.

scholarly journals Regulation of Human CD4+ αβ T-Cell-Receptor-Positive (TCR+) and γδ TCR+ T-Cell Responses to Mycobacterium tuberculosis by Interleukin-10 and Transforming Growth Factor β

1999 ◽  
Vol 67 (12) ◽  
pp. 6461-6472 ◽  
Author(s):  
Roxana E. Rojas ◽  
Kithiganahalli N. Balaji ◽  
Ahila Subramanian ◽  
W. Henry Boom
Keyword(s):  
T Cells ◽  
Mycobacterium Tuberculosis ◽  
T Cell ◽  
Growth Factor ◽  
Transforming Growth Factor ◽  
Γδ T Cells ◽  
T Cell Responses ◽  
Transforming Growth Factor Β ◽  
T Cell Subsets ◽  
Cell Responses

ABSTRACT Mycobacterium tuberculosis is the etiologic agent of human tuberculosis and is estimated to infect one-third of the world's population. Control of M. tuberculosis requires T cells and macrophages. T-cell function is modulated by the cytokine environment, which in mycobacterial infection is a balance of proinflammatory (interleukin-1 [IL-1], IL-6, IL-8, IL-12, and tumor necrosis factor alpha) and inhibitory (IL-10 and transforming growth factor β [TGF-β]) cytokines. IL-10 and TGF-β are produced by M. tuberculosis-infected macrophages. The effect of IL-10 and TGF-β on M. tuberculosis-reactive human CD4+and γδ T cells, the two major human T-cell subsets activated byM. tuberculosis, was investigated. Both IL-10 and TGF-β inhibited proliferation and gamma interferon production by CD4+ and γδ T cells. IL-10 was a more potent inhibitor than TGF-β for both T-cell subsets. Combinations of IL-10 and TGF-β did not result in additive or synergistic inhibition. IL-10 inhibited γδ and CD4+ T cells directly and inhibited monocyte antigen-presenting cell (APC) function for CD4+ T cells and, to a lesser extent, for γδ T cells. TGF-β inhibited both CD4+ and γδ T cells directly and had little effect on APC function for γδ and CD4+ T cells. IL-10 down-regulated major histocompatibility complex (MHC) class I, MHC class II, CD40, B7-1, and B7-2 expression on M. tuberculosis-infected monocytes to a greater extent than TGF-β. Neither cytokine affected the uptake of M. tuberculosis by monocytes. Thus, IL-10 and TGF-β both inhibited CD4+ and γδ T cells but differed in the mechanism used to inhibit T-cell responses to M. tuberculosis.

Oligoclonal Expansion of Vδ1+ γδT Cells in Cyclosporine-Responsive Acquired Pure Red Cell Aplasia.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 3845-3845
Author(s):  
Makoto Hirokawa ◽  
Naohito Fujishima ◽  
Yuhta Oyama ◽  
Masumi Fujishima ◽  
Takahiro Horiuchi ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Lymphocytes ◽  
Cell Subset ◽  
Γδ T Cells ◽  
Pure Red Cell Aplasia ◽  
Red Cell ◽  
T Cell Subset ◽  
Pcr Products ◽  
Oligoclonal Expansion

Abstract Background: Acquired pure red cell aplasia (PRCA) is an autoimmune disease in which autoaggressive T lymphocytes selectively suppress erythropoiesis, sometimes associated with clonal T cell disorders. However, little is known about the structure of the T-cell receptors (TCR) used by T lymphocytes involved in the development of acquired PRCA. Previously, a patient with granular lymphocyte-proliferative disorder with an expansion of Vδ1+ γδ T cells has been reported to be associated with cyclosporine-resistant acquired PRCA (Handgretinger et al. NEJM1999;340:278–284). We hypothesized that autoreactive γδ T lymphocytes might also be involved in the pathogenesis of cyclosporine-responsive PRCA, and thus examined whether the oligoclonal expansion would be detected in cyclosporine-responsive acquired PRCA patients. Methods: Five PRCA patients who achieved remission after oral cyclosporine therapy were included in this study, and informed consent was obtained before drawing blood samples. Blood mononuclear cells were stained with monoclonal antibodies specific for Vδ1, Vδ2 or Vδ3, and were analyzed by flow cytometry. Clonality of the Vδ1+ T cell subset was determined by complementarity-determining region 3 (CDR3) size spectratyping analysis of TCR Vδ1-Cδ PCR products. Junctional sequence was determined by DNA sequencing with an ABI377 automated DNA sequencer. Results: All five patients showed an increase of the Vδ1+ T cell subset in circulating blood (Table 1), although the major subset of human γδ T cells in healthy individuals is the Vδ2 (data not shown). Oligoclonal expansion was detected by CDR3 size spectratyping analysis and were confirmed by sequencing of the TCRVδ1-Cδ PCR products in all five patients (Table 2). No apparent consensus sequence among patients was observed in the CDR3 region of clonally expanded Vδ1+ T cells. Conclusions: Oligclonally expanded Vδ1+ T cells may be responsible for the pathogenesis of cyclosporine-responsive acquired PRCA. Biological function of these clonally expanding Vδ1+ T cells, natural ligands of the TCRs and requirements of antigen-presenting molecules remain to be elucidated. Table 1. γδT cell subsets in cyclosporine-responsive acquired PRCA patients UPN01 UPN02 UPN03 UPN04 UPN05 Pan- γδ 9.65 16.53 18.95 22.89 6.89 δ 1 V 6.62 18.37 13.34 40.08 2.33 δ 2 V 0.66 0.20 1.12 0.15 0.72 δ 3 V 0.40 0.18 4.95 0.11 4.06 Table 2. Oligoclonal expansions of Vδ1+ T cells and the CDR3 sequences Vδ 1 NDN Jδ Colony frequency T-cell clones appearing more than twice in each sample are presented. UPN01 FCALG PLGFLPRWGIRG WDTRQMFF 5/17 FCALG GTSSYWGHYT DKLIFGKG 5/17 FCALGE LGQGGYPPVR TAQLFFGKG 4/17 UPN02 FCALGE RAPHWGIRRI TDKLIFGKG 9/23 FCALGE KRTYSPP LTAQLFFGKG 6/23 UPN03 FCALGE LSLPHSLVGPGGSH TDKLIFGKG 5/23 FCALG YGPRVGYPWGRI IFGKG 4/23 FCALG RQPSYGWWGISFPPY TDKLIFGKG 3/23 FCALG GAYVIWGK TDKLIFGKG 3/23 UPN04 FCALGE REYWGMY TDKLIFGKG 6/16 FCALG KRGFPWPYWGIRA TDKLIFGKG 3/23 UPN05 FCALG HSGLFGVQY TDKLIFGKG 5/24 FCALG DPSYRRWGIRLY TDKLIFGKG 3/24

scholarly journals γδ T Cells in Immunity Induced by Mycobacterium bovis Bacillus Calmette-Guérin Vaccination

2004 ◽  
Vol 72 (3) ◽  
pp. 1504-1511 ◽  
Author(s):  
Jinhee Lee ◽  
Keumhwa Choi ◽  
Michael R. Olin ◽  
Sang-Nae Cho ◽  
Thomas W. Molitor
Keyword(s):  
T Cells ◽  
T Cell ◽  
Mycobacterium Bovis ◽  
Γδ T Cells ◽  
T Cell Responses ◽  
Γδ T Cell ◽  
Bcg Vaccination ◽  
Cell Responses ◽  
Ifn Γ ◽  
Time Point

ABSTRACT Mycobacterium bovis bacillus Calmette-Guérin (BCG) vaccination is efficacious for newborns or adults with no previous exposure to environmental mycobacteria. To determine the relative contribution and the nature of γδ T-cell receptor-positive T cells in newborns, compared to CD4+ T cells, in immunity induced by M. bovis BCG vaccination, 4-week-old specific-pathogen-free pigs were vaccinated with M. bovis BCG and monitored by following the γδ T-cell immune responses. A flow cytometry-based proliferation assay and intracellular staining for gamma interferon (IFN-γ) were used to examine γδ T-cell responses. Pigs were found to mount Th1-like responses to M. bovis BCG vaccination as determined by immunoproliferation and IFN-γ production. The γδ T-cell lymphoproliferation and IFN-γ production to stimulation with mycobacterial antigens were significantly enhanced by M. bovis BCG vaccination. The relative number of proliferating γδ T cells after stimulating peripheral blood mononuclear cells with Mycobacterium tuberculosis H37Rv culture filtrate protein was higher than that of CD4+ T cells at an early time point after M. bovis BCG vaccination, but CD4+ T cells were found to be more abundant at a later time point. Although the γδ T-cell responses were dependent on the presence of CD4+ T cells for the cytokine interleukin-2, the enhanced γδ T cells were due to the intrinsic changes of γδ T cells caused by M. bovis BCG vaccination rather than being due solely to help from CD4+ T cells. Our study shows that γδ T cells from pigs at early ages are functionally enhanced by M. bovis BCG vaccination and suggests an important role for this T-cell subset in acquired immunity conferred by M. bovis BCG vaccination.

scholarly journals Selective Recruitment of T-Cell Subsets to the Udder during Staphylococcal and Streptococcal Mastitis: Analysis of Lymphocyte Subsets and Adhesion Molecule Expression

1999 ◽  
Vol 67 (12) ◽  
pp. 6293-6302 ◽  
Author(s):  
Jindrich Soltys ◽  
Mark T. Quinn
Keyword(s):  
T Cells ◽  
T Cell ◽  
Lymphocyte Subsets ◽  
Inflammatory Responses ◽  
Cell Subset ◽  
Γδ T Cells ◽  
T Cell Subsets ◽  
Γδ T Cell ◽  
Cell Numbers ◽  
Milk Samples

ABSTRACT During bacterial infection of the bovine mammary gland, large numbers of leukocytes migrate into the udder, resulting in the establishment of a host response against the pathogen. Currently, the specific leukocyte populations mediating this immune response are not well defined. In the studies described here, we analyzed blood and milk from healthy cows and cows with naturally occurring mastitis to determine if distinct αβ and γδ T-lymphocyte subsets were involved in the response of the udder to a mastitis pathogen and if the type of mastitis pathogen influenced the subset composition of these responding leukocytes. Although blood samples from cows with confirmed staphylococcal and streptococcal mastitis were characterized by increased numbers of γδ T cells, the most dramatic changes in leukocyte distributions occurred in milk samples from these cows, with a 75% increase in αβ T-cell levels and a 100% increase in γδ T-cell levels relative to the levels in milk samples from healthy animals. Interestingly, the increase in αβ T-cell numbers observed in milk from cows with staphylococcal mastitis was primarily due to increased numbers of CD4+ T cells, while the increase in αβ T-cell numbers observed in cows with streptococcal mastitis was due to a parallel increase in both CD4+ and CD8+ T-cell numbers. The increased numbers of γδ T cells in milk from cows with staphylococcal and streptococcal mastitis were due to a selective recruitment of a distinct γδ T-cell subset (GD3.1+), while no change in the numbers of GD197+ γδ T cells was observed. We also analyzed adhesion protein expression on blood and milk leukocytes and found that, in comparison to the situation for healthy cows, L-selectin was down-regulated and CD18 was up-regulated on leukocytes from cows with mastitis. Thus, shedding of L-selectin and up-regulation of CD18 by neutrophils may provide a sensitive indicator of early inflammatory responses during bovine mastitis. Overall, these studies suggest that distinct αβ and γδ T-cell subsets are involved in the host defense of the udder against mastitis infection and that selective recruitment of these T-cell subsets depends on the infectious agent involved.

scholarly journals 413 GEN-009, a personalized neoantigen vaccine, elicits robust immune responses in individuals with advanced or metastatic solid tumors

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A438-A438
Author(s):  
Mara Shainheit ◽  
Devin Champagne ◽  
Gabriella Santone ◽  
Syukri Shukor ◽  
Ece Bicak ◽  
...  
Keyword(s):  
Clinical Trial ◽  
T Cells ◽  
T Cell ◽  
Immune Responses ◽  
Solid Tumors ◽  
Ex Vivo ◽  
T Cell Responses ◽  
Checkpoint Blockade ◽  
T Cell Subsets ◽  
Cell Responses

BackgroundATLASTM is a cell-based bioassay that utilizes a cancer patient‘s own monocyte-derived dendritic cells and CD4+ and CD8+ T cells to screen their mutanome and identify neoantigens that elicit robust anti-tumor T cell responses, as well as, deleterious InhibigensTM.1 GEN-009, a personalized vaccine comprised of 4–20 ATLAS-identified neoantigens combined with Hiltonol®, harnesses the power of neoantigen-specific T cells to treat individuals with solid tumors. The safety and efficacy of GEN-009 is being assessed in a phase 1/2a clinical trial (NCT03633110).MethodsA cohort of 15 adults with solid tumors were enrolled in the study. During the screening period, patients received standard of care PD-1-based immunotherapies appropriate for their tumor type. Subsequently, patients were immunized with GEN-009 with additional doses administered at 3, 6, 12, and 24 weeks. Peripheral blood mononuclear cells (PBMCs) were collected at baseline, pre-vaccination (D1), as well as 29, 50, 92, and 176 days post first dose. Vaccine-induced immunogenicity and persistence were assessed by quantifying neoantigen-specific T cell responses in ex vivo and in vitro stimulation dual-analyte fluorospot assays. Polyfunctionality of neoantigen-specific T cells was evaluated by intracellular cytokine staining. Additionally, potential correlations between the ATLAS-identified profile and vaccine-induced immunogenicity were assessed.ResultsGEN-009 augmented T cell responses in 100% of evaluated patients, attributable to vaccine and not checkpoint blockade. Furthermore, neoantigen-induced secretion of IFNγ and/or TNFα by PBMCs, CD4+, and CD8+ T cells was observed in all patients. Responses were primarily from polyfunctional TEM cells and detectable in both CD4+ and CD8+ T cell subsets. Some patients had evidence of epitope spreading. Unique response patterns were observed for each patient with no apparent relationship between tumor types and time to emergence, magnitude or persistence of response. Ex vivo vaccine-induced immune responses were observed as early as 1 month, and in some cases, persisted for 176 days. Clinical efficacy possibly attributable to GEN-009 was observed in several patients, but no correlation has yet been identified with neoantigen number or magnitude of immune response.ConclusionsATLAS empirically identifies stimulatory neoantigens using the patient‘s own immune cells. GEN-009, which is comprised of personalized, ATLAS-identified neoantigens, elicits early, long-lasting and polyfunctional neoantigen-specific CD4+ and CD8+ T cell responses in individuals with advanced cancer. Several patients achieved clinical responses that were possibly attributable to vaccine; efforts are underway to explore T cell correlates of protection. These data support that GEN-009, in combination with checkpoint blockade, represents a unique approach to treat solid tumors.AcknowledgementsWe are grateful to the patients and their families who consented to participate in the GEN-009-101 clinical trial.Trial RegistrationNCT03633110Ethics ApprovalThis study was approved by Western Institutional Review Board, approval number 1-1078861-1. All subjects contributing samples provided signed individual informed consent.ReferenceDeVault V, Starobinets H, Adhikari S, Singh S, Rinaldi S, Classon B, Flechtner J, Lam H. Inhibigens, personal neoantigens that drive suppressive T cell responses, abrogate protection of therapeutic anti-tumor vaccines. J. Immunol 2020; 204(1 Supplement):91.15.

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