Mapping of Specific and Promiscuous HLA-DR-Restricted T-Cell Epitopes on the Plasmodium falciparum 27-Kilodalton Sexual Stage-Specific Antigen

ABSTRACT We have characterized HLA-DR-restricted T-cell epitopes on the 27-kDa protein (Pfg27), a sexual stage-specific antigen, of the human malaria parasite Plasmodium falciparum in subjects with a history of malaria. Pfg27, expressed early in the sexual stages, is recognized by monoclonal antibodies capable of reducing the infectivity of gametocytes in mosquitoes. By using 16 Pfg27-specific CD4+-T-cell clones derived from three donors, seven different T-cell epitopes were identified. Among them, P11 (amino acids 191 to 210 of the Pfg27 sequence, IDVVDSYIIKPIPALPVTPD) was found to contain a previously described binding motif for multiple HLA-DR allotypes. Indeed, P11 was found to be promiscuous in that it could be recognized by T cells in the context of at least five different HLA-DR molecules. The cytokine profile of the clones was mixed. Seven of nine T-cell clones exhibited a Th0-like cytokine profile, producing high levels of gamma interferon (IFN-γ) and interleukin-4 (IL-4) upon stimulation with specific peptides and mitogens. The other two clones had a Th1-like cytokine profile with high expression of IFN-γ and no IL-4. Identification of a promiscuous epitope in Pfg27 could play a significant role in the design of a subunit vaccine for suppressing malaria transmission.

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Thyrotropin-Receptor and Thyroid Peroxidase-Specific T Cell Clones and Their Cytokine Profile in Autoimmune Thyroid Disease1

The Journal of Clinical Endocrinology & Metabolism ◽

10.1210/jcem.82.11.4336 ◽

1997 ◽

Vol 82 (11) ◽

pp. 3655-3663

Author(s):

Maria Elena Fisfalen ◽

Ellen M. Palmer ◽

Gijs A. van Seventer ◽

Keyoumars Soltani ◽

Yoshikuni Sawai ◽

...

Keyword(s):

Amino Acid ◽

T Cell ◽

Thyroid Tissue ◽

Cytokine Profile ◽

Thyroid Peroxidase ◽

Amino Acid Residues ◽

T Cell Epitopes ◽

T Cell Clones ◽

Cell Clones ◽

Ifn Γ

We studied the cytokine profile and the immune responses to thyroid antigens of specific T cell clones (TCC) isolated from patients with Hashimoto’s thyroiditis (HT) and Graves’ disease (GD). Antigen-specific TCC were reactive to thyroid peroxidase (TPO), thyroglobulin (Tg) or human recombinant TSH-receptor extracellular domain (TSH-R), and/or their respective peptides. Of the 43 clones derived from HT patients, 65% were reactive to TPO, and 59% of the 32 clones derived from GD patients were reactive to TSH-R. TPO epitopes 100–119 and 625–644 were recognized by 75% of HT-derived clones, whereas TSH-R epitopes 158–176, 207–222, and 343–362/357–376 were recognized by 85% of GD-derived TCC. The TCC were classified according to their cytokine profile into T helper cell (Th)0 [secreting interleukin (IL)-4, IL-5, interferon (IFN)-γ], Th1 (secreting IFN-γ) and Th2 (secreting IL-4 and/or IL-5). Tumor necrosis factor-β and IL-10 were produced by all subsets. The specific TCC were predominantly Th1-like cells in HT, and were Th0- and Th1-like cells in GD. Fifty three percent of Th0 clones were derived from GD patients and were reactive to TSH-R, whereas 50% of Th1 clones were derived from HT patients and were reactive to TPO or Tg. Most Th2 clones (82%) were reactive to TPO and were established from peripheral blood. All these clones produced IL-5, and 64% produced IL-4 and IL-10. Interestingly, IFN-γ was highly produced by TPO- or Tg-specific clones established from HT thyroid tissue. These results confirm at the clonal level our previous studies regarding T cell epitopes on TPO and TSH-R molecules and support the concept that immunodominant T cell epitopes are located on amino acid residues 100–119 and 625–644 of TPO in HT and amino acid residues 158–176, 207–222 and 343–362/357–376 of TSH-R in GD. Our studies also demonstrate that thyroid-specific T cells can be classified into Th0, Th1, and Th2 subsets. TPO- or Tg-specific clones with Th1 phenotype appear to be involved in the pathogenesis of HT, mediating thyroid tissue destruction, whereas TSH-R clones with Th0 phenotype may induce thyroid-stimulating autoantibodies in GD.

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Accumulation of Human Heat Shock Protein 60-Reactive T Cells in the Gingival Tissues of Periodontitis Patients

Infection and Immunity ◽

10.1128/iai.70.5.2492-2501.2002 ◽

2002 ◽

Vol 70 (5) ◽

pp. 2492-2501 ◽

Cited By ~ 74

Author(s):

Kazuhisa Yamazaki ◽

Yutaka Ohsawa ◽

Koichi Tabeta ◽

Harue Ito ◽

Kaoru Ueki ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Heat Shock ◽

Heat Shock Protein ◽

Peripheral Blood ◽

Cytokine Profile ◽

T Cell Clones ◽

Cell Clones ◽

Ifn Γ ◽

Human Hsp60

ABSTRACT Heat shock protein 60s (hsp60) are remarkably immunogenic, and both T-cell and antibody responses to hsp60 have been reported in various inflammatory conditions. To clarify the role of hsp60 in T-cell responses in periodontitis, we examined the proliferative response of peripheral blood mononuclear cells (PBMC), as well as the cytokine profile and T-cell clonality, for periodontitis patients and controls following stimulation with recombinant human hsp60 and Porphyromonas gingivalis GroEL. To confirm the infiltration of hsp60-reactive T-cell clones into periodontitis lesions, nucleotide sequences within complementarity-determining region 3 of the T-cell receptor (TCR) β-chain were compared between hsp60-reactive peripheral blood T cells and periodontitis lesion-infiltrating T cells. Periodontitis patients demonstrated significantly higher proliferative responses of PBMC to human hsp60, but not to P. gingivalis GroEL, than control subjects. The response was inhibited by anti-major histocompatibility complex class II antibodies. Analysis of the nucleotide sequences of the TCR demonstrated that human hsp60-reactive T-cell clones and periodontitis lesion-infiltrating T cells have the same receptors, suggesting that hsp60-reactive T cells accumulate in periodontitis lesions. Analysis of the cytokine profile demonstrated that hsp60-reactive PBMC produced significant levels of gamma interferon (IFN-γ) in periodontitis patients, whereas P. gingivalis GroEL did not induce any skewing toward a type1 or type2 cytokine profile. In control subjects no significant expression of IFN-γ or interleukin 4 was induced. These results suggest that periodontitis patients have human hsp60-reactive T cells with a type 1 cytokine profile in their peripheral blood T-cell pools.

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Immunodominant Epitopes in Babesia bovis Rhoptry-Associated Protein 1 That Elicit Memory CD4+-T-Lymphocyte Responses in B. bovis-Immune Individuals Are Located in the Amino-Terminal Domain

Infection and Immunity ◽

10.1128/iai.70.4.2039-2048.2002 ◽

2002 ◽

Vol 70 (4) ◽

pp. 2039-2048 ◽

Cited By ~ 17

Author(s):

Junzo Norimine ◽

Carlos E. Suarez ◽

Terry F. McElwain ◽

Monica Florin-Christensen ◽

Wendy C. Brown

Keyword(s):

T Cell ◽

Cell Lines ◽

Vaccine Development ◽

Specific Antigen ◽

Babesia Bovis ◽

T Cell Epitopes ◽

T Cell Clones ◽

Amino Terminal ◽

Cell Clones ◽

T Cell Lines

ABSTRACT Babesia bovis rhoptry-associated protein 1 (RAP-1), which confers partial protection against B. bovis challenge, is recognized by antibodies and T lymphocytes from cattle that have recovered from infection and are immune to subsequent challenge. RAP-1 is a 60-kDa protein with an N-terminal (NT) region that contains four cysteine residues conserved among all Babesia RAP-1 family members and a C-terminal (CT) region that contains multiple, degenerate, tandem 23-amino-acid (aa) repeats. To define the location of CD4+-T-cell epitopes for vaccine development using a recombinant protein or minigene construct, a series of truncated recombinant RAP-1 proteins and peptides were tested for stimulation of T-cell lines derived from B. bovis-immune cattle. CD4+-T-cell lines from three B. bovis-immune cattle with different DRB3 haplotypes responded to the NT region of RAP-1, whereas T cells from only one animal responded weakly to the CT region. T-cell lines from the three individuals recognized two to six NT-region peptides spanning aa 134 to 316 and representing at least four dominant epitopes. Using RAP-1-specific CD4+-T-cell clones, two NT-region epitopes, EYLVNKVLYMATMNYKT (aa 187 to 203) and EAPWYKRWIKKFR (aa 295 to 307), and one CT-region repeat epitope, FREAPQATKHFL, which is present twice at aa positions 391 to 402 and 414 to 425, were identified. Several peptides representing degenerate repeats of the agonist CT-region peptide FREAPQATKHFL neither stimulated responses of T-cell clones specific for this peptide nor inhibited responses to the agonist peptide. Upon stimulation with specific antigen, T-cell clones specific for NT or CT epitopes produced gamma interferon. The presence of T-helper-cell epitopes in the NT domain of RAP-1, which is highly conserved among otherwise antigenically different strains of B. bovis, supports the inclusion of this region in vaccine constructs to be tested in cattle.

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691 Identification of shared tumor epitopes from endogenous retroviruses inducing high avidity cytotoxic T cells for cancer immunotherapy

Journal for ImmunoTherapy of Cancer ◽

10.1136/jitc-2021-sitc2021.691 ◽

2021 ◽

Vol 9 (Suppl 3) ◽

pp. A719-A719

Author(s):

Paola Bonaventura ◽

Vincent Alcazer ◽

Virginie Mutez ◽

Laurie Tonon ◽

Juliette Martin ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Tumor Cells ◽

Cd8 T Cells ◽

Endogenous Retroviruses ◽

High Avidity ◽

T Cell Epitopes ◽

T Cell Clones ◽

Cell Clones ◽

Ifn Γ

BackgroundHuman endogenous retroviruses (HERVs) are aberrantly expressed by tumor cells and may represent a source of T cell epitopesMethodsUsing TCGA pancancer RNAseq data (n=8,893 samples), we developed a bioinformatics-based method to select cancer-specific HERVs associated with a cytotoxic T cell response (“cyt-HERVs”) and identify shared T cell epitope candidates. T cells were primed with selected short and long peptide candidates from HLA-A2+ healthy donors. Peptide-specific dextramers were used to sort and expand specific CD8+ T cell clones and determine their TCR sequences and avidity. Cytotoxicity was assessed against HERV-expressing tumor cell lines and patient-derived organoids using Incucyte and Nanolive technologies (Flowchart, figure 1).ResultsIn a pancancer analysis, we identified 57 HML-2/HERV-K HLA-A*0201 epitope candidates from 27 distinct open reading frames. Six shared HLA-A2 strong binders 9-mer peptides, present on multiple HERVs located on different chromosomes, and with translational evidence found in mass spectrometry public datasets, were selected and synthetized. In vitro HLA binding assay confirmed peptide-HLA affinity. Priming assays showed the presence of specific CD8+ T cells leading to polyfunctional IFN-γ+ TNF-α+ T cell responses with upregulation of the degranulation marker CD107A upon co-culture with peptide-pulsed T2 cells. Synthetic long peptides containing the epitopes were used to confirm the correct processing by antigen-presenting cells. The functionality of the sorted T cell clones was confirmed using an Elispot assay (GrzB+ IFN-γ+). Their sequenced TCRs were predicted to stably interact with their respective MHC-peptide complexes in a 3D model. This was confirmed by measurement of the functional avidity, which was in the same order as CMV-specific T cell clones. HERV-specific CD8+ T cells induced specific cell death of HLA-A2+ cancer cell lines, associated with IFN-g production, in a HLA-A2 restricted manner. Finally, pre-existing HERV-specific CD8+ T cells were identified using dextramers among tumor infiltrating lymphocytes (TILs) from cancer patients. HERV-specific T cells co-cultured with patient derived organoids showed signs of activation with lysis of the organoid.ConclusionsOur bioinformatics-based approach allowed us to identify shared HERV-derived CD8+ T cell epitopes specifically expressed by tumor cells and inducing high avidity T cell clones able to kill tumor cells in a class I-restricted manner. The detection of TILs recognizing HERV peptides suggests natural presentation of these epitopes in the tumors. These HERV-derived epitopes may thus represent relevant targets for the development of new cancer vaccines or T cell-based therapies, especially in tumors with low mutational burden.Abstract 691 Figure 1Graphical flowchart of HERV antigen validation. Graphical representation of the flowchart used to identify and validate specific CD8+ T cells for shared tumor epitopes from endogenous retroviruses http://dx.doi.org/10.1136/jitc-2021-SITC2021.691

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Elimination of a Human T-cell Region in Staphylokinase by T-cell Screening and Computer Modeling

Thrombosis and Haemostasis ◽

10.1055/s-0037-1613064 ◽

2002 ◽

Vol 87 (04) ◽

pp. 666-673 ◽

Cited By ~ 35

Author(s):

Stéphane Plaisance ◽

Kristel Vanderlick ◽

Petra Vandervoort ◽

Kathleen Brepoels ◽

Désiré Collen ◽

...

Keyword(s):

Immune Response ◽

Amino Acid ◽

T Cell ◽

Computer Modeling ◽

T Cell Epitopes ◽

Cell Reactivity ◽

T Cell Clones ◽

Human T Cell ◽

Hla Dr ◽

Cell Clones

SummaryStaphylokinase is a potent highly fibrin-selective thrombolytic agent, but it induces a humoral immune response in most treated patients. Staphylokinase-specific T-lymphocytes can be found in normal healthy individuals, from whom a large panel of staphylokinasespecific T-cells were cloned. The staphylokinase amino acid sequence 71-87 was widely recognized, as it induced proliferation of T-cell clones isolated from 90% of the donors. Computer modeling of this area, threaded as 11-mer peptides within the peptide-binding groove of the major HLA-DR alleles, indicated two putative partially overlapping binding sequences. The region-(71-87)-specific T-cell clones recognized either one or the other minimal peptide, confirming that both sequences could be functional T-cell epitopes. Furthermore, to guide the mutagenesis to eliminate T-cell reactivity, the contribution of each residue to the HLA-DR-anchoring and T-cell receptor exposure was evaluated for both binding motifs. Computer calculations combined with functional assays resulted in the design of staphylokinasevariants, including 2 to 4 amino acid substitutions in the region 71-87. These variants were no longer recognized by the region-(71-87)specific T-cell clones, and importantly no new staphylokinase-variantspecific cellular immune response could be measured.

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T-cell clones from early-stage cutaneous T-cell lymphoma show no polarized Th-1 or Th-2 cytokine profile

Archives of Dermatological Research ◽

10.1007/pl00007454 ◽

2000 ◽

Vol 292 (1) ◽

pp. 1-8 ◽

Cited By ~ 14

Author(s):

S. Harwix ◽

K. Zachmann ◽

C. Neumann

Keyword(s):

T Cell ◽

Cell Lymphoma ◽

Early Stage ◽

Cytokine Profile ◽

T Cell Lymphoma ◽

Cutaneous T Cell Lymphoma ◽

T Cell Clones ◽

Cell Clones ◽

Th 1

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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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