Detailed Analysis of CD8+ T Cell Immunity and Identification of a Novel Minor Histocompatibility Antigen Contributing to Graft-Versus- Leukemia Reactivity.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3250-3250
Author(s):  
Marieke Griffioen ◽  
M. Willy Honders ◽  
Edith D. van der Meijden ◽  
Roelof Willemze ◽  
J.H. Frederik Falkenburg
Keyword(s):  
T Cells ◽  
T Cell ◽  
Detailed Analysis ◽  
Cd8 T Cells ◽  
Cell Clone ◽  
Hematopoietic Cells ◽  
Cd8 T Cell ◽  
T Cell Clones ◽  
Cell Clones ◽  
T Cell Clone

Abstract Donor lymphocyte infusion (DLI) can be an effective cellular immunotherapy for patients with hematological malignancies after HLA-matched allogeneic stem cell transplantation (alloSCT). The effect of DLI is mediated by donor derived T cells recognizing minor histocompatibility antigens (mHags) on malignant cells of the recipient. These donor originated T cells may also induce Graft-versus-Host Disease (GvHD) when directed against mHags with broad expression on non-malignant tissues of the patient. In this study, we performed a detailed analysis and characterization of mHags recognized by CD8+ T cells contributing to Graft-versus-Leukemia (GvL) reactivity in a patient treated with DLI for relapsed chronic myeloid leukemia (CML) more than one year after HLA-matched alloSCT. The GvL effect in this patient was accompanied with only mild GvHD of the skin. To investigate the specificity of the CD8+ T cell response induced in this patient, activated (HLA-DR+) CD8+ T cells were single cell sorted from a bone marrow sample obtained five weeks after DLI by flowcytometry. A number of isolated CD8+ T cell clones were shown to be specific for mHags, as determined by differential recognition of patient and donor EBV-transformed B cells (EBV-LCL) in IFN-g ELISA and 51Crrelease assays. By screening a panel of third party EBV-LCL sharing one or more HLA class I restriction molecules with the patient, CD8+ T cell clones directed against 7 different mHags were identified, including the known hematopoiesis restricted mHags HA-1 and HA-2, which are presented in HLA-A*0201. Of the 5 remaining specificities, 4 mHags were presented in HLA-B*4001 (B60) and one mHag in HLA-B*0801. To determine the tissue distribution patterns in more detail, we tested recognition of selected non-malignant hematopoietic cells (monocytes, B cells, T cells), malignant CD34+ CML precursor cells, and skin-derived fibroblasts. One HLA-B*4001-restricted T cell clone (clone ZRZ16) failed to recognize all primary hematopoietic cells and skin fibroblasts. The four remaining T cell clones were all capable of recognizing and lysing (specific subsets of) non-malignant hematopoietic cells and malignant CD34+ CML precursor cells. Fibroblast recognition could be demonstrated for two of these four T cell clones. Since clone ZRZ16 failed to recognize all hematopoietic and non-hematopoietic cells, except for EBV-LCL, we tested whether this clone selectively recognizes antigen presenting cells (APC), which are known to be required for efficient induction of immune responses in vivo. Clone ZRZ16 showed indeed strong recognition of monocyte-derived dendritic cells as well as in vitro differentiated CD34+ CML cells with APC phenotype. To identify the mHag recognized by the CD8+ T cell clone, we screened a cDNA expression library constructed from EBV-LCL from the patient. One single cDNA was isolated as the target for B*4001 restricted CD8+ T cell clone ZRZ16. The epitope recognized by this clone was derived from the 3’ untranslated region (UTR) of a cDNA encoding thyroid hormone receptor interactor 10 (TRIP10). The peptide epitope was translated in a reading frame different from the TRIP10 protein and comprises three single nucleotide polymorphisms, which were all different between patient and donor. Two of the three SNPs were shown to be important for recognition by clone ZRZ16. Despite ubiquitous tissue expression of the TRIP10 gene as determined by public microarray analysis, CD8+ T cells specific for the newly-identified LB-TRIP10-1EPC mHag selectively recognized APC and failed to recognize CD34+ CML precursor cells, suggesting a predominant role in the initiation, but not effector, phase of the anti-tumor response. In conclusion, our data show a detailed analysis of mHag specific CD8+ T cell immunity induced in a patient successfully treated with DLI for relapsed CML and provide evidence for differential involvement of HLA class I restricted mHags in the onset and execution of GvL reactivity.

scholarly journals TCR-MHC Interaction Strength Defines Trafficking and Resident Memory Status of CD8 T cells in the Brain

2018 ◽  
Author(s):  
Anna Sanecka ◽  
Nagisa Yoshida ◽  
Elizabeth Motunrayo Kolawole ◽  
Harshil Patel ◽  
Brian D. Evavold ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Chronic Infection ◽  
Cd8 T Cell ◽  
T Cell Clones ◽  
Cell Clones ◽  
T Cell Clone ◽  
Wide Range ◽  
The Brain

AbstractT cell receptor-Major histocompatibility complex (TCR-MHC) affinities span a wide range in a polyclonal T cell response, yet it is undefined how affinity shapes long-term properties of CD8 T cells during chronic infection with persistent antigen. Here, we investigate how the affinity of the TCR-MHC interaction shapes the phenotype of memory CD8 T cells in the chronically Toxoplasma gondii-infected brain. We employed CD8 T cells from three lines of transnuclear (TN) mice that harbour in their endogenous loci different T cell receptors specific for the same Toxoplasma antigenic epitope ROP7. The three TN CD8 T cell clones span a wide range of affinities to MHCI-ROP7. These three CD8 T cell clones have a distinct and fixed hierarchy in terms of effector function in response to the antigen measured as proliferation capacity, trafficking, T cell maintenance and memory formation. In particular, the T cell clone of lowest affinity does not home to the brain. The two higher affinity T cell clones show differences in establishing resident memory populations (CD103+) in the brain with the higher affinity clone persisting longer in the host during chronic infection. Transcriptional profiling of naïve and activated ROP7-specific CD8 T cells revealed that Klf2 encoding a transcription factor that is known to be a negative marker for T cell trafficking is upregulated in the activated lowest affinity ROP7 clone. Our data thus suggest that TCR-MHC affinity dictates memory CD8 T cell fate at the site of infection.

scholarly journals IL4I1 Accelerates the Expansion of Effector CD8+ T Cells at the Expense of Memory Precursors by Increasing the Threshold of T-Cell Activation

2020 ◽  
Vol 11 ◽  
Author(s):  
Marie-Line Puiffe ◽  
Aurélie Dupont ◽  
Nouhoum Sako ◽  
Jérôme Gatineau ◽  
José L. Cohen ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Activation ◽  
Cd8 T Cells ◽  
Cell Activation ◽  
Cell Clone ◽  
Cd8 T Cell ◽  
Lymphocytic Choriomeningitis ◽  
T Cell Clone

IL4I1 is an immunoregulatory enzyme that inhibits CD8 T-cell proliferation in vitro and in the tumoral context. Here, we dissected the effect of IL4I1 on CD8 T-cell priming by studying the differentiation of a transgenic CD8 T-cell clone and the endogenous repertoire in a mouse model of acute lymphocytic choriomeningitis virus (LCMV) infection. Unexpectedly, we show that IL4I1 accelerates the expansion of functional effector CD8 T cells during the first several days after infection and increases the average affinity of the elicited repertoire, supporting more efficient LCMV clearance in WT mice than IL4I1-deficient mice. Conversely, IL4I1 restrains the differentiation of CD8 T-cells into long-lived memory precursors and favors the memory response to the most immunodominant peptides. IL4I1 expression does not affect the phenotype or antigen-presenting functions of dendritic cells (DCs), but directly reduces the stability of T-DC immune synapses in vitro, thus dampening T-cell activation. Overall, our results support a model in which IL4I1 increases the threshold of T-cell activation, indirectly promoting the priming of high-affinity clones while limiting memory T-cell differentiation.

Diversity of HLA Class I and Class II Restricted Minor Histocompatibility Antigens in Graft-Versus-Leukemia Reactivity.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4084-4084
Author(s):  
Marieke Griffioen ◽  
M. Willy Honders ◽  
Anita N. Stumpf ◽  
Edith D. van der Meijden ◽  
Cornelis A.M. van Bergen ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Hematopoietic Cells ◽  
Cd4 T Cell ◽  
Cell Recognition ◽  
T Cell Clones ◽  
T Cell Recognition ◽  
Cell Clones ◽  
T Cell Clone ◽  
Ifn Γ

Abstract Abstract 4084 Poster Board III-1019 Donor lymphocyte infusion (DLI) can be an effective cellular immunotherapy for patients with hematological malignancies after HLA-matched allogeneic stem cell transplantation (alloSCT). The effect of DLI is mediated by donor derived T-cells recognizing minor histocompatibility antigens (mHags) encoded by single nucleotide polymorphisms (SNPs) on malignant cells of the recipient. Donor T-cells may also induce Graft-versus-Host Disease (GvHD) when directed against mHags with broad expression on non-malignant tissues. The aim of this study was to investigate the specificity and diversity of mHags recognized by T-cells in Graft-versus-Leukemia (GvL) reactivity. Activated (HLA-DR+) CD8+ and CD4+ T-cell clones were isolated from a patient successfully treated with DLI for relapsed chronic myeloid leukemia (CML) more than one year after HLA-matched alloSCT. GvL reactivity in this patient was accompanied with mild GvHD of the skin. Isolated T-cell clones were shown to recognize 13 different mHags. CD8+ T-cell clones were specific for HA-1 and HA-2 in HLA-A*0201, one unknown mHag in B*0801 and 4 unknown mHags in B*4001. CD4+ T-cell clones were specific for one unknown mHag in HLA-DQ and 5 unknown mHags in DR. By screening plasmid (class I) and bacteria (class II) cDNA libraries, we identified a mHag in HLA-DQ encoded by the PI4K2B gene (Griffioen et al., PNAS 2008), 4 mHags in HLA-DR encoded by the PTK2B, MR-1, LY75 and MTHFD1 genes (Stumpf et al., Blood 2009) and a mHag in B*4001 encoded by the TRIP10 gene. For the 3 T cell clones recognizing unknown mHags in B*4001, we performed Whole Genome Assocation scanning (WGAs). A panel of 60 EBV-LCL was retrovirally-transduced with B*4001 and tested for T-cell recognition. In parallel, genomic DNA was isolated and more than one million single nucleotide polymorphisms (SNPs) were determined by the Illumina beadchip array. Statistical analysis revealed significant association between T-cell recognition of EBV-LCL and the presence of coding SNPs in the SON DNA-binding protein and SWAP-70 genes. To get more insight into the role and potential use of the mHags in GvL reactivity and GvHD, all T-cell clones were analyzed in detail for reactivity against hematopoietic and non-hematopoietic cells. Hematopoietic cells included peripheral blood cells (monocytes, B-cells and T-cells), professional antigen presenting cells (APC) and leukemic cells (CML, ALL and AML). All CD8+ T-cell clones recognized (subsets of) peripheral blood cells as well as CML cells, except for the T-cell clone for TRIP10. Recognition of (subsets of) peripheral blood cells was also observed for all CD4+ T-cell clones, but CML cells were differentially recognized. CML cells were strongly recognized by the T-cell clones for MTHFD1 and the unknown mHag in HLA-DR, whereas no or low reactivity was observed for all other CD4+ T-cell clones. All CD8+ and CD4+ T-cell clones strongly recognized professional APC, including monocyte-derived dendritic cells and in vitro differentiated CML cells with APC phenotype. All T-cell clones were also capable of recognizing AML and ALL, except for the T-cell clone for TRIP10, which showed restricted recognition of AML-M4 and -M5 of monocytic origin. As non-hematopoietic cells, patient-derived fibroblasts were cultured with and without IFN-γ and tested for T-cell recognition. In the absence of IFN-γ, all T-cell clones failed to recognize fibroblasts, except for the T-cell clone for the unknown mHag in B*0801. After treatment with IFN-γ, additional reactivity was observed for the T-cell clones for SON DNA-binding protein and the unknown mHag in B*4001. Our data showed the specificity and diversity of mHags recognized by T-cells induced in a patient successfully treated with DLI for relapsed CML. The T-cell response was directed against 13 different mHags, of which 10 mHags in HLA class I and class II have now been identified by different techniques. Detailed analysis of T-cell recognition of hematopoietic and non-hematopoietic cells provides evidence that the mHags played different roles in the onset and execution of GvL and GvHD. Moreover, only one of the 10 identified mHags was expressed on fibroblasts after treatment with IFN-γ, indicating the characterization of mHags with potential relevance for T-cell based immunotherapy. Disclosures: No relevant conflicts of interest to declare.

HLA Class II Upregulation During An Ongoing Viral Infection Can Lead to HLA-DP Directed Graft-Versus-Host Disease After HLA-DPB1 Mismatched CD4+ Donor Lymphocyte Infusion

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 3062-3062 ◽  
Author(s):  
Sanja Stevanovic ◽  
Cornelis A.M. van Bergen ◽  
Simone A.P. van Luxemburg-Heijs ◽  
Jessica C. Harskamp ◽  
C.J.M. Halkes ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Hematopoietic Cells ◽  
Class Ii ◽  
Hla Class Ii ◽  
T Cell Response ◽  
Cd4 T Cell ◽  
T Cell Clones ◽  
Cell Clones

Abstract Abstract 3062 T cell depletion of the graft in allogeneic hematopoietic stem cell transplantation (alloSCT) prevents the occurrence of severe acute Graft-versus-Host Disease (GvHD), but also impairs post-transplant anti-tumor and anti-viral immunity. Early intervention with donor lymphocyte infusion (DLI) after alloSCT may prevent relapse of the malignancy and improve immune reconstitution, but can be associated with reintroduction of GvHD. Since under non-inflammatory conditions HLA class II molecules are predominantly expressed on hematopoietic cells, DLI consisting of only CD4+ T cells can selectively target residual patient (pt) HLA class II + hematopoietic cells without inducing severe GvHD. However, recently in two pts with acute myeloid leukemia we observed severe GvHD after prophylactic CD4+ DLI following a 10/10 HLA allele matched, but HLA-DPB1 mismatched unrelated donor alloSCT. Both pts received a T cell depleted SCT after a non-myeloablative conditioning regimen, resulting in mixed chimerism (>97 % donor) at 3 months after alloSCT, and no GvHD. A single infusion of 0.5*106 purified CD4+ T cells/kg was administered 3.5 months after alloSCT, resulting in a decreasing pt chimerism coinciding with grade 1 skin GvHD, followed by grade 3–4 colonic GvHD 3–8 weeks later. Both pts were successfully treated with immune suppression and are in complete remission (CR) more than one year later. During the clinical immune responses high percentages of activated CD4+ (30–74 %) and CD8+ T cells (9–56 %) were demonstrated in peripheral blood (PB). Using cell sorting, we clonally isolated 777 and 289 CD4+, and 204 and 34 CD8+ T cell clones from pts 1 and 2, respectively, and tested these clones for recognition of multiple pt and donor derived target cells using IFNg ELISA. None of the CD8+ clones were alloreactive. In contrast, 3 and 8 % of the CD4+ T cell clones from pts 1 and 2, respectively, recognized various pt hematopoietic cells, but not donor cells, indicating alloreactivity. Retroviral transduction of donor EBV-LCL with pt HLA-DPB1 alleles identified specific recognition of the mismatched alleles for 2 and 7 % of all CD4+ T cell clones isolated, respectively. The remaining alloreactive CD4+ T cell clones showed a hematopoiesis-restricted minor histocompatibility antigen recognition pattern, since they failed to recognize pt skin fibroblasts pretreated with IFNg to upregulate HLA class II expression. In contrast, the majority of HLA-DPB1 specific CD4+ T cell clones recognized pt IFNg treated skin fibroblasts, indicating a direct role as mediators of GvHD after HLA-DPB1 mismatched CD4+ DLI. Since both pts were in CR, but mixed chimeric at the time of CD4+ DLI, we hypothesized that residual pt HLA-DP+ hematopoietic cells after alloSCT may have served as antigen presenting cells (APC) to induce the HLA-DPB1 specific CD4+ T cell response. Lineage specific chimerism analysis of PB samples prior to CD4+ DLI showed complete donor chimerism in the B cell and myeloid compartments, whereas predominantly pt chimerism (89–100% pt) was demonstrated in the T cell compartment. Flowcytometric analysis showed that 5–25 % of the pt CD4+ and CD8+ T cells were activated and expressed HLA-DP. CMV tetramer analysis demonstrated that 31 % of CD8+ T cells from pt 1 and 10 % from pt 2 were CMV specific, which had expanded as a consequence of CMV reactivation. We hypothesize that the HLA-DPB1 specific CD4+ T cell response has been induced by upregulated HLA-DP expression on activated pt T cells due to preexisting CMV infection, and/or by residual pt derived skin-resident APC, resulting in limited skin GvHD. We demonstrated CMV infection in a colon biopsy at the time of colonic GvHD, suggesting that local production of cytokines by pt derived CMV specific T cells may have upregulated HLA class II expression on non-hematopoietic cells and enhanced the HLA-DPB1 specific CD4+ T cell response, resulting in exacerbation of GvHD. In conclusion, we show in two pts that GvHD after prophylactic CD4+ DLI administered early after HLA-DPB1 mismatched T cell depleted alloSCT was caused by alloreactive CD4+ T cells directed against pt mismatched HLA-DPB1 alleles. Our results suggest that the presence of active viral infections inducing immune responses by residual pt T cells at the time of prophylactic HLA class II mismatched CD4+ DLI increases the likelihood of development of GvHD by influencing HLA class II expression on pt hematopoietic and non-hematopoietic cells. Disclosures: No relevant conflicts of interest to declare.

ROR1 Specific T Cell Clones from Healthy Individuals Show Common T Cell Receptor Motifs

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 3364-3364
Author(s):  
Falk Heidenreich ◽  
Elke Ruecker-Braun ◽  
Juliane S. Stickel ◽  
Anne Eugster ◽  
Denise Kühn ◽  
...  
Keyword(s):  
Amino Acids ◽  
T Cells ◽  
T Cell ◽  
Mhc Class I ◽  
T Cell Receptor ◽  
Cd8 T Cells ◽  
Cd8 T Cell ◽  
Cell Receptor ◽  
T Cell Clones ◽  
Cell Clones

Abstract Background Immunotherapy for CLL with new antibodies or T-cells with modified TCR relies on attractive targets. ROR1 is such a promising target since it is highly overexpressed in CLL. Chimeric antigen receptor engineered T cells and antibodies directed against the extracellular part of ROR1 have already been developed and tested in vitro or in animal models, but still there is no MHC-class I presented peptide serving as target structure for CD8+ T cells (with or without a genetically modified T cell receptor) available. Aim The aim of this study was (1) to identify an immunogenic MHC-class I presented ROR1 peptide, (2) to generate respective ROR1 peptide specific CD8+ T cell clones, and (3) to analyze the nucleotide sequence of the CDR3 region of the expressed alpha and beta T cell receptor chain. Results In mass spectrometric-based analyses of the HLA-ligandome a HLA-B*07 presented ROR1 peptide was identified in primary CLL cells of two patients. Six T cell clones specific for this particular ROR1-peptide were generated from single CD8+ T cells from 2 healthy individuals with 3 T cell clones generated from each donor. Functionality and specificity of those T cell clones were tested in cytotoxicity assays. All 6 dextramer+ CD8+ T cell clones lysed peptide loaded and HLA-B*07+ transduced K562 cells (kindly provided by Lorenz Jahn, [Jahn et al., Blood, 2015 Feb 5;125(6):949-58]). Two selected clones (XD8 and XB6) were tested for their cytotoxic potential against 2 ROR1+ HLA-B*07+ tumor cell lines (with the ROR1 peptide identified by mass spectrometry for both of them) and against 2 primary CLL cell samples. Tested clones showed a significant lysis of the respective target cells. CDR3 regions of the alpha and beta T cell receptor chain were sequenced on a single cell level. The CDR3 alpha region from each of the 3 ROR1 specific T cell clones from donor A showed some similarities to T cell clones derived from donor B (Table 1). Conclusion For the first time a MHC-class I presented ROR1 peptide antigen is reported. ROR1 positive CLL cells can be targeted by specific HLA-B*07 restricted CTLs. Respective CD8+ T cell clones with anti-leukemic activity from 2 donors share some amino acid motifs of the CDR3 alpha and beta regions. In conclusion, this information provides the possibility of generating ROR1 specific CD8+ T cells with genetically modified T cell receptors for immunotherapy and for tracking those cells after administration with next generation sequencing in peripheral blood samples of patients. Furthermore, data suggest the existence of public TCR motifs in leukemia antigen specific CTLs, which needs to be proven in follow-up experiments with larger cohorts of donors and patients. Finally, the presented strategy to identify leukemia specific peptide antigens for CD8+ T cells might be an attractive method for similar projects. Table 1 Amino acid sequences of CDR3 alpha and beta regions of the TCR of ROR1 specific CD8+ T cell clones. When comparing two clones, matching amino acids are depicted in red. The aromatic amino acids phenylalanine (F) and tyrosine (Y) are shown in blue when situated at the same position. Gaps inserted during the sequence alignment process are indicated by a hyphen '-'. Table 1. Amino acid sequences of CDR3 alpha and beta regions of the TCR of ROR1 specific CD8+ T cell clones. When comparing two clones, matching amino acids are depicted in red. The aromatic amino acids phenylalanine (F) and tyrosine (Y) are shown in blue when situated at the same position. Gaps inserted during the sequence alignment process are indicated by a hyphen '-'. Disclosures Middeke: Sanofi: Honoraria. Schetelig:Sanofi: Honoraria.

scholarly journals Association of Donor T Cell Repertoire in Host Lymphoid and Target Organs and Gvhd Development

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 4525-4525
Author(s):  
Yongxia Wu ◽  
Jianing Fu ◽  
Anusara Daenthanasanmak ◽  
Hung D Nguyen ◽  
Mohammed Hanief Sofi ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Clone ◽  
T Cell Repertoire ◽  
Cell Repertoire ◽  
T Cell Clones ◽  
Cell Clones ◽  
Target Organs ◽  
T Cell Clone ◽  
Donor T Cells

Abstract The diversity and composition of T cell receptor (TCR) repertoire, which is the result of V, D and J gene recombination in TCR gene locus, has been found to impact immune responses in autoimmune and infectious diseases. The correlation of T-cell repertoire with the pathogenesis and outcome of graft-versus-host disease (GVHD) remain undefined. Here, by utilizing high-throughput sequencing of the gene encoding the TCRβ-chain, we comprehensively analyzed the profile of T-cell repertoire in host lymphoid and GVHD target organs after bone marrow transplantation (BMT). To understand whether T-cell repertoire is affected by different strength of alloantigen stimulation, we transferred same donor T cells derived from C57BL/6 (B6) mice into irradiated BALB/c (MHC-fully mismatched), B6D2F1 (MHC-haploidentical), BALB.b (MHC-matched ) and B6 recipients (syngeneic). Fourteen days later, T cells were isolated from recipient peripheral blood, spleen, peripheral lymphoid nodes (pLN), mesenteric lymphoid nodes (mLN), liver, lung, gut and skin for TCR sequencing. Clonality of donor T cells, which is inversely associated with TCR diversity, was significantly increased in either syngeneic or allogeneic recipients when compared with naïve donor T-cells, consistent with the concept that TCR diversity is reduced after T-cell activation and expansion. Increased TCR clonality was observed in lymphoid organs of allogeneic compared with syngeneic recipients, confirming that donor T cells were further activated in allogeneic recipients. However, decreased TCR clonality was observed in GVHD target organs of allogeneic compared with syngeneic recipients, suggesting that only limited donor T-cell clones were able to migrate in target organs in syngeneic compared to allogeneic recipients. The frequency of top clones in total productive rearrangements was increased in GVHD target organs especially liver of allogenic than syngeneic receipts. Interestingly, the frequency of top clones was positively associated with MHC disparity between donor and host, ranging from low to high in syngeneic, MHC-matched, haploidentical, and fully-mismatched recipients, respectively. To understand the extent to which TCR rearrangement is shared among different organs after BMT, we analyzed the overlap of TCR clones across different organs in the same recipients. T-cell clones were highly overlapping across organs, especially among GVHD target organs, in the same recipients after allogeneic BMT, although much lower overlapping in recipients after syngeneic BMT. The results suggest that alloantigen stimulation selectively activate certain T-cell clones and enrich antigen specific clones. On the other hand, much fewer shared clones were found among different recipients within the same group, regardless of MHC-disparity between donor and host. These results suggest that specific T-cell clones activated and expanded by alloantigens stimulation were highly different in individual recipients even with the same MHC-disparity between donor and host. Interestingly, the levels of clone overlapping were different in distinct organs among individual recipients. The level of T-cell clone overlapping was found high in liver of individual recipients regardless of the strength of alloantigen stimulation. The level of T cell clone overlapping was relatively high in pLNs and skin of the recipients after haploidentical BMT; whereas the level of T cell clone overlapping was relatively high in mLNs and gut of the recipients after MHC-matched BMT. These results suggest that skin may be a dominant target in haploidentical BMT and gut as a dominant target in MHC-matched BMT; whereas liver is a common target organ regardless. In conclusion, the current study establishes the association between MHC disparity, T-cell activation, and GVHD development in the level of donor T-cell repertoire. While TCR repertoire of donor T cells in peripheral blood or lymph nodes likely is representative in any individual recipient/patient, it is nearly impossible to identify T-cell clones that are pathogenic and shared among groups of recipients/patients even with the same MHC-disparity between donor and host. Disclosures No relevant conflicts of interest to declare.

scholarly journals Functional Characterization and Modulation of Cytokine Production by CD8+ T Cells from Human Immunodeficiency Virus-Infected Individuals

Blood ◽  
1997 ◽  
Vol 89 (10) ◽  
pp. 3672-3681 ◽  
Author(s):  
Enrico Maggi ◽  
Roberto Manetti ◽  
Francesco Annunziato ◽  
Lorenzo Cosmi ◽  
Maria Grazia Giudizi ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Cd8 T Cell ◽  
T Cell Clones ◽  
Cell Clones ◽  
Immunodeficiency Virus ◽  
Ifn Γ ◽  
Hiv Seropositive

CD8+ T-cell clones were generated from peripheral blood mononuclear cells (PBMC) of three human immunodeficiency virus (HIV)-seronegative individuals and six HIV-seropositive individuals and assessed for their cytokine secretion profile, cytolytic potential, and chemokine production. While the great majority of CD8+ T-cell clones generated from HIV-seronegative individuals produced interferon (IFN)-γ, but not interleukin-4 (IL-4), that is a type 1 cytotoxic (Tc1) profile, high numbers of CD8+ T-cell clones generated from HIV-seropositive individuals produced IL-4 in addition to IFN-γ or IL-4 alone, thus showing a type 0 cytotoxic (Tc0)- or a type 2 cytotoxic (Tc2) profile, respectively. Tc0/Tc2 cells displayed lower cytolytic activity than Tc1 cells, including a reduced ability to lyse autologous targets pulsed with HIV or HIV peptides. By contrast, the production of chemokines RANTES and macrophage inflammatory protein-1α was comparable in Tc1, Tc0, and Tc2 clones irrespective of whether they were derived from HIV-seronegative or HIV-seropositive individuals. When CD8+ T-cell clones were generated from PBMC cultures of HIV-seronegative individuals conditioned with IL-4 plus an anti–IL-12 antibody (Ab), a shift towards the Tc0/Tc2-like profile was observed. Conversely, the addition to PBMC cultures of IL-12 plus an anti – IL-4 Ab shifted the differentiation of CD8+ T cells from HIV-infected individuals towards the Tc1-like profile, whereas IL-12 or anti–IL-4 Ab alone had a lower Tc1-promoting effect. Irradiated PBMC from HIV-infected individuals, used as feeder cells, shifted the differentiation of CD8+ T cells from a healthy HIV-seronegative individual towards the Tc0/Tc2-like profile. On the other hand, a shift towards the Tc1-like profile was noted in CD8+ T-cell clones generated from the skin specimens of two HIV-seropositive patients with Kaposi's sarcoma, successfully treated with IFN-α, in comparison to CD8+ clones generated from the same skin areas before treatment. The IFN-α–induced Tc1 shift could be prevented by the incubation of skin-infiltrating CD8+ T cells with IL-4 before cloning. Taken together, these data indicate that both defective production of IL-12 and abnormal IL-4 production in bulk PBMC populations of HIV-infected individuals may contribute to the development of high numbers of CD8+ T-cell clones showing a Tc0/Tc2-like phenotype and reduced cytolytic potential against HIV itself. They also suggest that the cytokine profile of CD8+ T-cell clones can be modulated by cytokines (or anticytokine Ab) both in vitro and in vivo.

Generating of Human CD4+ and CD8+ T Lymphocytes toward NY-ESO-1 by T Cell Receptor (TCR) Modification and Transduction for Adoptive TCR Gene-Transfer

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3533-3533
Author(s):  
Holger Krönig ◽  
Kathrin Hofer ◽  
Daniel Sommermeyer ◽  
Christian Peschel ◽  
Wolfgang Uckert ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Gene Transfer ◽  
T Cell Receptor ◽  
Cell Clone ◽  
Cd8 T Cell ◽  
Cell Receptor ◽  
Cell Clones ◽  
T Cell Clone ◽  
T Cell Transfer

Abstract The Cancer Testis (CT) antigen NY-ESO-1 is one of the most immunogenic cancer antigens eliciting strong humoral and cellular immune responses in tumor patients and therefore it is a promising candidate antigen for successful adoptive T cell transfer. The aim of our studies is the transfer of autologous T cells re-directed towards CT antigens by T cell receptor (TCR) gene transfer. The first precondition for genetic transfer of CT-Ag-specific TCRs is the availability of tumor-reactive CD4+ and CD8+ T cell clones that express a CT-Ag-specific TCR. Therefore, we generated the autologous CD8+ T cell clone ThP2 through stimulating HLA-A2.1− PBMCs with autologous HLA-A2+DCs loaded with synthetic NY-ESO-1157–165. After two restimulations, FACS-sorting and cloning, the T cell line specifically recognized the NY-ESO-1157–165 peptide and also specifically lysed NY-ESO-1157–165 expressing tumor cells. In addition, we generated NY-ESO-1 specific T helper1 clones from HLA-DR1+ and HLA-DR4+ healthy donors by stimulation of CD4+ T cells with autologous dendritic cells (DC) pulsed with the NY-ESO-187–111 peptide. The specificity of CD4+ T helper cell clones was determined by proliferation assays and IFN gamma ELISPOT through screening with the NY-ESO-187–111 peptide. By limiting dilution of the NYESO- 1-specific T cell populations we succeeded to isolate CD4+ T cell clones, which recognized NY-ESO-1-pulsed target cells and DCs pulsed with NY-ESO-1 protein. The second precondition for TCR gene transfer is the availability of efficient vector systems. Using vectors based upon mouse myelo-proliferative sarcoma virus (MPSV), it was possible to achieve a high transgene expression in the TCR-transduced T cells. Therefore, we cloned the TCR of the HL-A2-restricted NY-ESO-1-specific CTL clone ThP2 in the retroviral vector and documented the correct expression of the TCR-chains using peptide/HLA-multimers following retroviral transduction of peripheral PBMCs. Moreover, the NY-ESO-1 specific lysis of HLA-A2+ NY-ESO-1+ tumor cell lines after transduction in primary T cells was as well effective as the primary T cell clone. Because the expression of naive transgenic T cell receptors in recipient human T cells is often insufficient to achieve highly reactive T cell bulks we modified the TCR of the ThP2 CTL clone by, murinisation, codon optimalization or by introducing cysteins into the constant regions. Afterwards we compared the expression efficiency of the three different modifications on naive T cells by tetramer-staining. We were able to show that codon optimalization leads to an increase in the expression levels of the transgenic TCRs in human CD8+ T cells. The next step is the development of T cell transfer regiments, which are based on class-II-restricted TCR-transduced T cells.

Demonstration of Frequent Occurrence of Clonal T Cells in the Peripheral Blood But Not in the Skin of Patients With Small Plaque Parapsoriasis

Blood ◽  
1999 ◽  
Vol 94 (4) ◽  
pp. 1409-1417 ◽  
Author(s):  
J. Marcus Muche ◽  
Ansgar Lukowsky ◽  
Jürgen Heim ◽  
Markus Friedrich ◽  
Heike Audring ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Peripheral Blood ◽  
Cell Clone ◽  
Small Plaque ◽  
Blood Samples ◽  
T Cell Clones ◽  
Cell Clones ◽  
T Cell Clone

Clinical, immunohistological, and molecular biological data suggest the chronic dermatosis small plaque parapsoriasis (SPP) to be a precursor of mycosis fungoides (MF). However, most data are contradictory and confusing due to inexact definition of SPP. Recently, clonal T cells were detected in skin and blood samples of early MF. Because demonstration of identical T-cell clones in skin and blood of SPP patients would indicate a close relationship of SPP to MF, we investigated the clonality of skin and blood specimens from 14 well-defined SPP patients. By a polymerase chain reaction (PCR) amplifying T-cell receptor γ rearrangements and subsequent high-resolution electrophoresis, clonal T cells were detected in 9 of 14 initial and 32 of 49 follow-up blood samples, but in 0 of 14 initial skin specimens. Even a clone-specific PCR showing the persistence of the initial blood T-cell clone in 20 of 20 follow-up samples, failed to detect the T-cell clone in the skin. In 2 patients, the clonal T cells were shown to be CD4+. For the first time, the majority of SPP patients was shown to carry a T-cell clone in the peripheral blood. Although a relation between circulating clonal T cells and SPP cannot directly be proven by the applied techniques, our results indicate blood T-cell clonality to be a characteristic feature of SPP and CTCL because analysis of multiple controls and clinical workup of our SPP patients excluded other factors simulating or causing a clonal T-cell proliferation. A sufficient cutaneous antitumor response but also an extracutaneous origin of the T-cell clones might explain the failure to detect skin infiltrating clonal T cells.

scholarly journals Regional and clonal T cell dynamics at single cell resolution in immune checkpoint blockade

2021 ◽  
Author(s):  
Joy A. Pai ◽  
Andrew Chow ◽  
Jennifer Sauter ◽  
Marissa Mattar ◽  
Hira Rizvi ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Immune Checkpoint ◽  
Cd8 T Cell ◽  
Immune Checkpoint Blockade ◽  
Rna Seq ◽  
Cell Dynamics ◽  
T Cell Clones ◽  
Cell Clones

Paired T cell receptor and RNA single cell sequencing (scTCR/RNA-seq) has allowed for enhanced resolution of clonal T cell dynamics in cancer. Here, we report a scTCR/RNA-seq dataset of 162,062 single T cells from 31 tissue regions, including tumor, adjacent normal tissues, and lymph nodes (LN), from three patients who underwent resections for progressing lung cancers after immune checkpoint blockade (ICB). We found marked regional heterogeneity in tumor persistence that was associated with heterogeneity in CD4 and CD8 T cell phenotypes; regions with persistent cancer cells were enriched for follicular helper CD4 T cells (TFH), regulatory T cells (Treg), and exhausted CD8 T cells. Clonal analysis demonstrated that highly-expanded T cell clones were predominantly of the CD8 subtype, were ubiquitously present across all sampled regions, found in the peripheral circulation, and expressed gene signatures of 'large' and 'dual-expanded' clones that have been predictive of response to ICB. Longitudinal tracking of CD8 T cell clones in the peripheral blood revealed that the persistence of ubiquitous CD8 T cell clones, as well as phenotypically distinct clones with tumor-reactive features, correlated with systemic tumor control. Finally, tracking CD8 T cell clones across tissues revealed the presence of TCF-1+ precursor exhausted CD8 T cells in tumor draining LNs that were clonally linked to expanded exhausted CD8 T cells in tumors. Altogether, this comprehensive scTCR/RNA-seq dataset with regional, longitudinal, and clonal resolution provides fundamental insights into the tissue distribution, persistence, and differentiation trajectories of ICB-responsive T cells that underlie clinical responses to ICB.

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