Cross-Reactivity to Mutated Viral Immune Targets Can Influence CD8+ T Cell Functionality: An Alternative Viral Adaptation Strategy

Loss of T cell immunogenicity due to mutations in virally encoded epitopes is a well-described adaptation strategy to limit host anti-viral immunity. Another described, but less understood, adaptation strategy involves the selection of mutations within epitopes that retain immune recognition, suggesting a benefit for the virus despite continued immune pressure (termed non-classical adaptation). To understand this adaptation strategy, we utilized a single cell transcriptomic approach to identify features of the HIV-specific CD8+ T cell responses targeting non-adapted (NAE) and adapted (AE) forms of epitopes containing a non-classical adaptation. T cell receptor (TCR) repertoire and transcriptome were obtained from antigen-specific CD8+ T cells of chronic (n=7) and acute (n=4) HIV-infected subjects identified by either HLA class I tetramers or upregulation of activation markers following peptide stimulation. CD8+ T cells were predominantly dual tetramer+, confirming a large proportion of cross-reactive TCR clonotypes capable of recognizing the NAE and AE form. However, single-reactive CD8+ T cells were identified in acute HIV-infected subjects only, providing the potential for the selection of T cell clones over time. The transcriptomic profile of CD8+ T cells was dependent on the autologous virus: subjects whose virus encoded the NAE form of the epitope (and who transitioned to the AE form at a later timepoint) exhibited an ‘effective’ immune response, as indicated by expression of transcripts associated with polyfunctionality, cytotoxicity and apoptosis (largely driven by the genes GZMB, IFNɣ, CCL3, CCL4 and CCL5). These data suggest that viral adaptation at a single amino acid residue can provide an alternative strategy for viral survival by modulating the transcriptome of CD8+ T cells and potentially selecting for less effective T cell clones from the acute to chronic phase.

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Transfer of Diabetes in the NOD-scid Mouse by CD4 T-Cell Clones: Differential Requirement for CD8 T-Cells

Diabetes ◽

10.2337/diab.45.3.328 ◽

1996 ◽

Vol 45 (3) ◽

pp. 328-336 ◽

Cited By ~ 61

Author(s):

J. D. Peterson ◽

K. Haskins

Keyword(s):

T Cells ◽

T Cell ◽

Cd8 T Cells ◽

Scid Mouse ◽

Cd4 T Cell ◽

T Cell Clones ◽

Cell Clones

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Macrophage colony-stimulating factor (CSF-1) gene expression in human T- lymphocyte clones

Blood ◽

10.1182/blood.v77.4.780.780 ◽

1991 ◽

Vol 77 (4) ◽

pp. 780-786 ◽

Cited By ~ 16

Author(s):

MM Hallet ◽

V Praloran ◽

H Vie ◽

MA Peyrat ◽

G Wong ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Phenotypic Diversity ◽

Calcium Ionophore ◽

Colony Stimulating Factor ◽

T Cell Clones ◽

Cell Clones ◽

Macrophage Colony Stimulating Factor ◽

Macrophage Colony ◽

Stimulating Factor

Abstract Macrophage colony stimulating factor (CSF-1) is one of several cytokines that control the differentiation, survival, and proliferation of monocytes and macrophages. A set of 11 human T-cell clones, chosen for their phenotypic diversity, were tested for their ability to express CSF-1 mRNA. After 5 hours of stimulation with phorbol myristate acetate (PMA) + calcium ionophore (Cal), all T-cell clones expressed a major 4-kb transcript, a less abundant 2-kb transcript, and several other minor species. This pattern of expression is typical for CSF-1 mRNAs. Furthermore, of the two alloreactive T-cell clones analyzed, only one showed a definitive message for CSF-1 on specific antigenic stimulation, but with delayed kinetics and less efficiency. Both conditions of stimulation induced the release of CSF-1 protein by T cells in the culture medium. Together, these findings demonstrate for the first time that normal T cells are able to produce CSF-1, previous reports being limited to two cases of tumoral cells of the T-cell lineage.

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Selective Depletion of Alloreactive T Cells and Study of Anti-Tumor Activity of Specific T Cell Clones in Patients with Leukemia and Renal Carcinoma

Clinical Immunology ◽

10.1016/j.clim.2007.03.482 ◽

2007 ◽

Vol 123 ◽

pp. S106-S107

Author(s):

Eva Matejkova ◽

Zuzana Hrotekova ◽

Drahomira Kyjovska ◽

Jaroslav Michalek ◽

Petra Vidlakova

Keyword(s):

T Cells ◽

T Cell ◽

Renal Carcinoma ◽

T Cell Clones ◽

Alloreactive T Cells ◽

Cell Clones ◽

Selective Depletion ◽

Anti Tumor Activity

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Evaluation of Various Strategies to Generate NPM1mut-Specific T Cell Clones

Blood ◽

10.1182/blood.v128.22.5718.5718 ◽

2016 ◽

Vol 128 (22) ◽

pp. 5718-5718 ◽

Cited By ~ 1

Author(s):

Elke Ruecker-Braun ◽

Falk Heidenreich ◽

Cornelia S Link ◽

Maria Schmiedgen ◽

Rebekka Wehner ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Checkpoint Inhibitors ◽

Single Cells ◽

Molecular Genetic ◽

Specific Antigen ◽

Positive Control ◽

Low Frequencies ◽

T Cell Clones ◽

Cell Clones

Abstract Mutated nucleophosmin (NPM1) was identified as a promising leukemia-specific antigen for cytotoxic T lymphocytes (CTL). NPM1 is a multifunctional nucleocytoplasmic shuttling phosphoprotein. In AML patients with normal cytogenetics NPM1 mutations are the most frequent molecular genetic abnormalities, accounting for up to 60% of the patients. The peptide (AIQDLCLAV) derived from the mutated NPM1 (NPM1mut) has been described to elicit a CTL response restricted to HLA-A*02:01. We observed that NPM1mut multimer+ T cells were very rare in peripheral blood. The limitation of the multimer technology is the absence of a positive control; nevertheless it is an attractive tool to generate antigen positive T cell clones. The goal was to compare strategies for the generation of NPM1mut multimer+ T cell clones systematically. For this purpose we analyzed blood samples from two patients with AML after transplantation and six different healthy donors. We explored different strategies to isolate HLA-A*02:01 restricted NPM1mut multimer+ T single cells. The first strategy was to isolate multimer+ T cells directly from the blood without any supplements by single cell sorting. The second strategy was to sort multimer+ T cells which were previously CD8+ enriched supplementing the media either with or without IL-21. Published by Yongqing et al.IL-21 enhances the generation of human antigen-specific CD8+ T cells. A further strategy was to previously enrich CD14+ cells for the generation of autologous monocyte-derived dendritic cells (MoDCs). The co-cultivation of MoDCs loaded with the NPM1mut peptide and CD8+ cells were performed either with or without IL-21, as well. We expanded the last strategy by a second round of NPM1mut-specific stimulation. So far it was not possible to generate NPM1mut-specific T cell clones based on the advanced strategies and consistently there is no data published on NPM1mut multimer+ T cell clones. This fact raises the question why NPM1mut specific clones display such low frequencies. We want to point out that although we varied the strategies and we used eight different donors the isolation of NPM1mut-specific T cells restricted to HLA-A*02:01 apparently is challenging. Greater efforts, e.g. a larger number of donors or the use of immunological checkpoint inhibitors during cell culture are needed. Disclosures Thiede: AgenDix: Employment, Other: Ownership. Schetelig:Sanofi: Honoraria.

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An Ex Vivo Study of T Lymphocytes Recovered from the Lungs of I/St Mice Infected with and Susceptible toMycobacterium tuberculosis

Infection and Immunity ◽

10.1128/iai.66.10.4981-4988.1998 ◽

1998 ◽

Vol 66 (10) ◽

pp. 4981-4988 ◽

Cited By ~ 33

Author(s):

Irina Lyadova ◽

Vladimir Yeremeev ◽

Konstantin Majorov ◽

Boris Nikonenko ◽

Sergei Khaidukov ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Ex Vivo ◽

Antimycobacterial Activity ◽

Enzymatic Digestion ◽

Interleukin 4 ◽

T Cell Clones ◽

Cell Clones ◽

Ifn Γ

ABSTRACT I/St mice, previously characterized as susceptible toMycobacterium tuberculosis H37Rv, were given 103 or 105 CFU intravenously. At two time points postinoculation, the cell suspensions that resulted from enzymatic digestion of lungs were enumerated and further characterized phenotypically and functionally. Regarding the T-cell populations recovered at 2 and 5 weeks postinfection, two main results were obtained: (i) the population of CD44− CD45RB+cells disappeared within 2 weeks postinfection, while the number of CD44+ CD45RB−/low cells slowly increased between weeks 2 and 5; (ii) when cocultured with irradiated syngeneic splenocytes, these lung T cells proliferated in the presence of H37Rv sonicate. Using H37Rv sonicate and irradiated syngeneic splenocytes to reactivate lung T cells, we selected five CD3+CD4+ CD8− T-cell clones. In addition to the H37Rv sonicate, the five clones react to both a short-term culture filtrate and an affinity-purified 15- to 18-kDa mycobacterial molecule as assessed by the proliferative assay. However, there was a clear difference between T-cell clones with respect to cytokine (gamma interferon [IFN-γ] and interleukin-4 [IL-4] and IL-10) profiles: besides one Th1-like (IFN-γ+ IL-4−) clone and one Th0-like (IFN-γ+ IL-4+IL-10+) clone, three clones produced predominantly IL-10, with only marginal or no IL-4 and IFN-γ responses. Inhibition of mycobacterial growth by macrophages in the presence of T cells was studied in a coculture in vitro system. It was found that the capacity to enhance antimycobacterial activity of macrophages fully correlated with INF-γ production by individual T-cell clones following genetically restricted recognition of infected macrophages. The possible functional significance of cytokine diversity among T-cell clones is discussed.

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Increased antitumor efficacy of PD-1-deficient melanoma-specific human lymphocytes

Journal for ImmunoTherapy of Cancer ◽

10.1136/jitc-2019-000311 ◽

2020 ◽

Vol 8 (1) ◽

pp. e000311 ◽

Cited By ~ 6

Author(s):

Lucine Marotte ◽

Sylvain Simon ◽

Virginie Vignard ◽

Emilie Dupre ◽

Malika Gantier ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Solid Tumors ◽

Adoptive Transfer ◽

Adoptive Cell Transfer ◽

Effector Memory ◽

High Avidity ◽

Cell Transfer ◽

T Cell Clones ◽

Cell Clones

BackgroundGenome editing offers unique perspectives for optimizing the functional properties of T cells for adoptive cell transfer purposes. So far,PDCD1editing has been successfully tested mainly in chimeric antigen receptor T (CAR-T) cells and human primary T cells. Nonetheless, for patients with solid tumors, the adoptive transfer of effector memory T cells specific for tumor antigens remains a relevant option, and the use of high avidity T cells deficient for programmed cell death-1 (PD-1) expression is susceptible to improve the therapeutic benefit of these treatments.MethodsHere we used the transfection of CAS9/sgRNA ribonucleoproteic complexes to editPDCD1gene in human effector memory CD8+T cells specific for the melanoma antigen Melan-A. We cloned edited T cell populations and validatedPDCD1editing through sequencing and cytometry in each T cell clone, together with T-cell receptor (TCR) chain’s sequencing. We also performed whole transcriptomic analyses on wild-type (WT) and edited T cell clones. Finally, we documented in vitro and in vivo through adoptive transfer in NOD scid gamma (NSG) mice, the antitumor properties of WT and PD-1KO T cell clones, expressing the same TCR.ResultsHere we demonstrated the feasibility to editPDCD1gene in human effector memory melanoma-specific T lymphocytes. We showed that PD-1 expression was dramatically reduced or totally absent onPDCD1-edited T cell clones. Extensive characterization of a panel of T cell clones expressing the same TCR and exhibiting similar functional avidity demonstrated superior antitumor reactivity against a PD-L1 expressing melanoma cell line. Transcriptomic analysis revealed a downregulation of genes involved in proliferation and DNA replication in PD-1-deficient T cell clones, whereas genes involved in metabolism and cell signaling were upregulated. Finally, we documented the superior ability of PD-1-deficient T cells to significantly delay the growth of a PD-L1 expressing human melanoma tumor in an NSG mouse model.ConclusionThe use of such lymphocytes for adoptive cell transfer purposes, associated with other approaches modulating the tumor microenvironment, would be a promising alternative to improve immunotherapy efficacy in solid tumors.

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Diminished Production of Interleukin 2 and Gamma-Interferon by Cloned «T» Cells from Patients with the Acquired Immunodeficiency Syndrome (AIDS)

Tumori Journal ◽

10.1177/030089168707300311 ◽

1987 ◽

Vol 73 (3) ◽

pp. 273-278 ◽

Cited By ~ 1

Author(s):

Enrico Maggi ◽

Donatella Macchia ◽

Paola Parronchi ◽

Domenico Milo ◽

Sergio Romagnani

Keyword(s):

T Cells ◽

T Cell ◽

Interleukin 2 ◽

Immune Deficiency Syndrome ◽

Gamma Interferon ◽

Aids Patients ◽

T Cell Clones ◽

Cell Clones ◽

The Mean ◽

Mean Concentration

A total of 76 T-cell clones established from peripheral blood (PB) of 2 patients with the acquired immune deficiency syndrome (AIDS) and of 141 T-cell clones established from PB of 3 normal donors were compared for their ability to produce interleukin 2 (IL-2) and gamma-interferon (γ-IFN). Twenty-seven clones from AIDS patients and 85 clones from controls expressed the CD4 phenotype, whereas 49 clones from AIDS patients and 56 clones from controls expressed the CD8 phenotype. There were no significant differences in the proportions of IL-2-producing CD4 T-cell clones established from PB of patients with AIDS and controls, but the mean concentration of IL-2 produced by CD4 clones from AIDS patients was significantly lower than that produced by CD4 clones from controls. Both the proportion of γ-IFN-producing CD4 clones and the mean concentration of γ-IFN produced by CD4 clones were significantly lower in AIDS patients than in controls. In contrast, there were no differences between AIDS patients and normal individuals in the proportion of IL-2- or Y-IFN-producing CD8 clones, or in the mean concentration of IL-2 and v-IFN produced by CD8 clones. These data suggest that the reduced ability of PB T-cells from patients with AIDS to produce IL-2 and v-IFN is not simply due to altered proportions or numbers of T-cell sub-populations, but also reflects intrinsic abnormalities of individual CD4 T lymphocytes.

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Emergence of CD52-, phosphatidylinositolglycan-anchor-deficient T lymphocytes after in vivo application of Campath-1H for refractory B- cell non-Hodgkin lymphoma

Blood ◽

10.1182/blood.v86.4.1487.bloodjournal8641487 ◽

1995 ◽

Vol 86 (4) ◽

pp. 1487-1492 ◽

Cited By ~ 71

Author(s):

B Hertenstein ◽

B Wagner ◽

D Bunjes ◽

C Duncker ◽

A Raghavachar ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

B Cell ◽

B Cell Lymphoma ◽

Molecular Characteristics ◽

T Cell Clones ◽

Non Hodgkin Lymphoma ◽

Cell Clones ◽

Immune Attack

CD52 is a phosphatidylinositolglycan (PIG)-anchored glycoprotein (PIG- AP) expressed on normal T and B lymphocytes, monocytes, and the majority of B-cell non-Hodgkin lymphomas. We observed the emergence of CD52- T cells in 3 patients after intravenous treatment with the humanized anti-CD52 monoclonal antibody Campath-1H for refractory B- cell lymphoma and could identify the underlaying mechanism. In addition to the absence of CD52, the PIG-AP CD48 and CD59 were not detectable on the CD52- T cells in 2 patients. PIG-AP-deficient T-cell clones from both patients were established. Analysis of the mRNA of the PIG-A gene showed an abnormal size in the T-cell clones from 1 of these patients, suggesting that a mutation in the PIG-A gene was the cause of the expression defect of PIG-AP. An escape from an immune attack directed against PIG-AP+ hematopoiesis has been hypothesized as the cause of the occurrence of PIG-AP-deficient cells in paroxysmal nocturnal hemoglobinuria (PNH) and aplastic anemia. Our results support the hypothesis that an attack against the PIG-AP CD52 might lead to the expansion of a PIG-anchor-deficient cell population with the phenotypic and molecular characteristics of PNH cells.

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

Blood Advances ◽

10.1182/bloodadvances.2020002255 ◽

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.

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