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

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

Alloreactivity of Virus Specific T-Cells.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3249-3249
Author(s):  
Avital L. Amir ◽  
Lloyd J.A. D’Orsogna ◽  
Marleen M. van Loenen ◽  
Dave L. Roelen ◽  
Ilias I.N. Doxiadis ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Lines ◽  
Memory T Cells ◽  
Target Cells ◽  
T Cell Clones ◽  
Hla Alleles ◽  
Alloreactive T Cells ◽  
Cell Clones ◽  
T Cell Lines

Abstract Graft versus host disease (GVHD) in allogeneic stem cell transplantation (SCT) and graft rejection is caused by alloreactive T-cells. Alloreactivity can be exerted by naïve as well as by memory T-cells. Persistent latent viral infections, like those with herpes viruses, have a profound impact on the repertoire of memory T-cells. This implies that virus specific memory T-cells are also potentially alloreactive. Previously it has been shown that virus specific T-cell clones can cross react against allo-HLA. We investigated the frequency of alloreactivity mediated by virus specific T-cells. Mixed lymphocyte reactions, previously used to determine precursor frequencies of alloreactive T-cells, give an underestimation of the total frequency of alloreactive T-cells, due to limited number of allo-HLA alleles tested in this system. Therefore, in this study multiple CD8+ virus specific T-cells lines and clones were tested for alloreactivity against almost all frequent HLA class I and II alleles. From different healthy individuals we derived CD8+ virus specific T-cell lines, specific for Epstein Barr virus (EBV), Cytomegalovirus (CMV), Varicella Zoster virus (VZV) and Influenza virus (Flu) which were restricted to different HLA molecules. The generation of the T-cell lines and clones was performed by bulk sorting and single cell sorting, based on staining with viral peptide/MHC complex specific tetramers. The viral specificity of the expanded lines and clones was confirmed by tetramer staining and cytotoxicity and cytokine production assays. Polyclonality of the T-cell lines and monoclonality of the T-cell clones was confirmed by TCR Vβ analysis. Next, the T-cell lines and clones were screened for alloreactivity by testing against a panel of 29 different EBV transformed LCLs, together covering almost all frequent HLA class I and II molecules. 90% of tested virus specific T-cell lines and 40% of virus specific T-cell clones were found to be alloreactive, recognizing at least one of the allo-HLA alleles. For several lines and clones the specific recognized allo-HLA molecule was further identified using a panel of HLA typed target cells in combination with HLA specific blocking antibodies. Additionally, single HLA antigen expressing cell lines were used as target cells. Thus far we found EBV EBNA3A specific, HLA-A3 restricted T-cell clones to recognize HLA-A31. A CMV pp50 specific, HLA-A1 restricted T-cell line recognized HLA-A68. One VZV IE62 specific, HLA-A2 restricted clone showed recognition of HLA-B57, while another clone with the same specificity but with a different TCR Vβ recognized HLA-B55. An EBV BMLF specific, HLA-A2 restricted T-cell line showed recognition of HLA-A11. Finally an EBV BRLF specific, HLA-A3 restricted clone recognized HLA-A2. Our results show that a high percentage of virus specific T-cells can exert alloreactivity against allo- HLA molecules. Previously it was assumed that virus specific T-cells are not alloreactive against foreign HLA, allowing safe application of virus specific T-cell lines derived from HLA disparate donors in patients without the risk of inducing GVHD. Our data indicate that applying virus specific T-cell lines over HLA barriers does give a significant risk of GVHD and suggest that lines should be tested for alloreactivity against patient specific HLA alleles prior to application. A substantial part of the memory T-cell pool consists of virus specific T-cells, which are dominated by a limited repertoire of virus specific T-cell clones, present in high frequencies. Thus, virus specific T-cells recognizing allo-HLA alleles may also play an essential role in graft rejection.

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

Blood ◽  
1991 ◽  
Vol 77 (4) ◽  
pp. 780-786 ◽  
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.

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

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 5718-5718 ◽  
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.

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

1998 ◽  
Vol 66 (10) ◽  
pp. 4981-4988 ◽  
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.

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

2020 ◽  
Vol 8 (1) ◽  
pp. e000311 ◽  
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.

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

1987 ◽  
Vol 73 (3) ◽  
pp. 273-278 ◽  
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.

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

Blood ◽  
1995 ◽  
Vol 86 (4) ◽  
pp. 1487-1492 ◽  
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.

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 Diversification of the antigen-specific T cell receptor repertoire after varicella zoster vaccination

2016 ◽  
Vol 8 (332) ◽  
pp. 332ra46-332ra46 ◽  
Author(s):  
Qian Qi ◽  
Mary M. Cavanagh ◽  
Sabine Le Saux ◽  
Hong NamKoong ◽  
Chulwoo Kim ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Receptor ◽  
Chronic Infection ◽  
Cell Receptor ◽  
Control Mechanisms ◽  
Cell Repertoire ◽  
T Cell Clones ◽  
Varicella Zoster ◽  
Cell Clones

Diversity and size of the antigen-specific T cell receptor (TCR) repertoire are two critical determinants for successful control of chronic infection. Varicella zoster virus (VZV) that establishes latency during childhood can escape control mechanisms, in particular with increasing age. We examined the TCR diversity of VZV-reactive CD4 T cells in individuals older than 50 years by studying three identical twin pairs and three unrelated individuals before and after vaccination with live attenuated VZV. Although all individuals had a small number of dominant T cell clones, the breadth of the VZV-specific repertoire differed markedly. A genetic influence was seen for the sharing of individual TCR sequences from antigen-reactive cells but not for repertoire richness or the selection of dominant clones. VZV vaccination favored the expansion of infrequent VZV antigen–reactive TCRs, including those from naïve T cells with lesser boosting of dominant T cell clones. Thus, vaccination does not reinforce the in vivo selection that occurred during chronic infection but leads to a diversification of the VZV-reactive T cell repertoire. However, a single-booster immunization seems insufficient to establish new clonal dominance. Our results suggest that repertoire analysis of antigen-specific TCRs can be an important readout to assess whether a vaccination was able to generate memory cells in clonal sizes that are necessary for immune protection.

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