scholarly journals Differential lysis of tumors by polyclonal T cell lines and T cell clones specific for hTERT

2007 ◽  
Vol 6 (12) ◽  
pp. 1991-1996 ◽  
Author(s):  
Dih-Yih Chen ◽  
Barbara A. Vance ◽  
Lara B. S. Thompson ◽  
Susan M. Domchek ◽  
Robert H. Vonderheide
Keyword(s):  
T Cell ◽  
Cell Lines ◽  
T Cell Clones ◽  
Cell Clones ◽  
Differential Lysis ◽  
T Cell Lines

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.

A Novel Strategy for Generation of Human Tumor-Specific T Cell Clones for Adoptive Transfer.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 3713-3713
Author(s):  
Seung-Tae Lee ◽  
Shujuan Liu ◽  
Pariya Sukhumalchandra ◽  
Jeffrey Molldrem ◽  
Patrick Hwu ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Tumor Cells ◽  
Cell Lines ◽  
Cd4 T Cells ◽  
Cd4 T Cell ◽  
Autologous Tumor ◽  
T Cell Clones ◽  
Cell Clones ◽  
T Cell Lines

Abstract Adoptive T-cell therapy using donor lymphocyte infusions is a promising approach for treating hematological malignancies. But, efficacy is limited by the induction of graft-versus-host disease. Transfer of tumor-specific T-cell clones could enhance the graft-versus-tumor effect and eliminate graft-versus-host disease. However, isolating antigen-specific T-cell clones by the traditional limiting dilution approach is a time-consuming and laborious process. Here, we describe a novel strategy for rapidly cloning tumor-specific T cells. Lymphoma-specific T-cell lines were generated from two follicular lymphoma patients by repeated in vitro stimulation of lymphocytes isolated from tumor or blood with autologous soluble CD40 ligand-activated tumor cells. After four in vitro stimulations at 10-day intervals in the presence of IL-2 and IL-15, T-cell lines were found to be predominantly CD4+ T cells and produced significant amounts of TNF-a, GM-CSF, and IFN-γ in response to autologous tumor cells. The tumor reactivity was MHC class II restricted suggesting that it was mediated by CD4+ T cells. Staining with a TCR Vb antibody panel, a set of monoclonal antibodies against 24 human TCR Vb families, revealed that certain Vb families were overrepresented in each CD4+ T-cell line. In patient 1, 51% of CD4+ T cells were Vb1 positive, and in patient 2, 27% of CD4+ T cells were Vb8 positive. To clone lymphoma-specific T cells, CD4+ T-cell lines were labeled with CFSE and stimulated with autologous tumor cells. After 9 days of in vitro expansion in the presence of IL-2 and IL-15, CD4+ T-cell lines were stained with an anti-human CD4-APC monoclonal antibody and an anti-human TCR Vb-PE monoclonal antibody for each CD4+ T-cell line. Proliferating Vb1 cells from patient 1 and Vb8 cells from patient 2 were identified by their reduction in CFSE staining, and CD4+TCRV b +CFSEdim cells were sorted by flow cytometer. Monoclonality of the sorted cells was confirmed by PCR using a panel of optimized primers specific for 24 TCR Vb families, by TCR Vb spectratype analysis, and finally, by sequencing the TCR Vb gene used by each T-cell clone. Sorted tumor-specific T-cell clones could be expanded to large numbers using a 14-day rapid expansion protocol with allofeeder PBMCs, and confirmed to retain specificity against autologous tumor cells in a cytokine induction assay. This approach was also successfully used to isolate melanoma-specific CD8+ T-cell clones from two patients. We conclude that this approach is highly reproducible, rapid, and efficient for generating antigen-specific T-cell clones for adoptive T-cell therapy against human malignancies in the autologous or allogeneic setting.

HLA-DPB1 Mismatching Results in the Generation of a Full Repertoire of HLA-DPB1 Specific T Cell Responses Showing Immunogenicity of All HLA-DPB1 Alleles

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3504-3504
Author(s):  
Caroline E. Rutten ◽  
Simone A.P. van Luxemburg-Heijs ◽  
Edith D. van der Meijden ◽  
Marieke Griffioen ◽  
Roelof Willemze ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Lines ◽  
Hela Cells ◽  
Cd4 T Cells ◽  
Cross Reactivity ◽  
T Cell Clones ◽  
Cell Clones ◽  
T Cell Lines

Abstract In unrelated donor hematopoietic stem cell transplantation (URD-SCT) patients are preferably transplanted with stem cells from a fully HLA matched donor, usually defined as identical for HLA-class I, -DR and -DQ. Since HLA-DPB1 is often not taken into consideration in donor selection, 80–90% of URD-SCTs are mismatched for HLA-DPB1. The role of HLA-DPB1 as transplantation antigen has been unclear, since clinical reports on the impact of matching for HLA-DPB1 on transplant outcome showed conflicting results. HLA-DPB1 mismatching has been associated with an increased risk of graft versus host disease (GVHD). However, we recently demonstrated that HLA-DPB1 specific T cells can mediate a potent graft versus leukemia effect without inducing GVHD. It has been suggested that the controversial effects of matching for HLA-DPB1 in URD-SCT could partly be explained by the assumption that not all HLA-DPB1 differences are immunogenic. This theory was based on the cross-reactive recognition of two HLA-DPB1* 09 specific T cell clones that recognized other HLA-DPB1 alleles sharing amino acids (aa) in position 8–11 of HLA-DPB1 (Zino et al, blood 2004). It was hypothesized that there would be no induction of T cell responses between individuals expressing HLA-DPB1 molecules sharing this aa sequence. This was translated into a classification of permissive and non-permissive HLA-DPB1 mismatches in order to allow a broader donor selection. To investigate whether cross-reactive recognition of other HLA-DPB1 molecules by our previously generated HLA-DPB1*02 or *03 specific CD4+ T cell clones depended on the presence of specific aa sequences we tested recognition of a panel of 14 EBV-LCL expressing 9 different HLA-DPB1 molecules. All HLA-DPB1*02 as well as all *03 specific T cell clones showed cross-reactivity with other HLA-DPB1 alleles and each T cell clone exhibited its own pattern of cross-reactivity. Two HLA-DPB1*0201 specific T cell clones with different TCR-Vβ showed also recognition of EBV-LCL expressing HLA-DPB1*1001 and *1701 or HLA-DPB1*1001, *0901 and *1601 respectively. Five HLA-DPB1*03 reactive T cells clones with different TCR-Vβ showed differential cross-recognition of EBV-LCL expressing HLA-DPB1*0101, *0601, *1101, *1301 and *1401. To identify immunogenic differences the aa sequences of the HLA-DPB1 molecules recognized by the various T cell clones were compared. The HLA-DPB1 molecules recognized by the HLA-DPB1*02 specific T cell clones shared an aa substitution at position 69 compared to the responder cell. However, HLA-DPB1*0601,*0901 and *1901 with the same substitution were not recognized by both T cell clones. This phenomenon was also observed for the HLA-DPB1*03 specific T cell clones, indicating that the cross-reactive recognition of HLA-DPB1 could not be predicted by aa sequences. Next, we analyzed the immunogenicity of various HLA-DPB1 alleles in different stimulator/responder combinations to verify the classification of permissive and non-permissive mismatches. We developed a model to generate allo-HLA-DP responses by transducing HLA-class II negative HELA cells with various HLA-DP molecules and used these cells to stimulate purified CD4+ T cells from HLA-DPB1 homozygous donors. HELA cells transduced with HLA-DPB1*0101, *0201, *0301, *0401, *0402, *0501, *0601, *0901, *1101, *1301, *1401 or *1701 were used as stimulator cells. Responder CD4+ T cells were typed HLA-DPB1* 0201, *0301, *0401 or *0402. 14 days after stimulation, CD4+ T cells were tested for recognition of the stimulator cells and of HELA cells transduced with the responder HLA-DPB1 molecule as a negative control. For these 4 responders, stimulation with 12 different HLA-DP transduced HELA cell lines resulted in specific IFN-γ production in response to the stimulator cells in 47 out of 48 stimulations. 28 CD4+ T cell lines also showed cross-reactive recognition of HELA cells transduced with at least one other HLA-DPB1 molecule. In conclusion, we showed that cross-reactive recognition of various HLA-DPB1 molecules by HLA-DPB1 specific T cells is a common observation. However, we demonstrated that cross-reactivity between HLA-DPB1 molecules by allo-HLA-DPB1 specific T cells does not exclude the generation of immune response between individuals expressing these HLA-DPB1 molecules. By generating multiple allo-HLA-DP specific T cell lines, we showed that all HLA-DPB1 mismatch combinations are immunogenic.

scholarly journals Alloreactive cloned T cell lines. I. Interactions between cloned amplifier and cytolytic T cell lines.

1980 ◽  
Vol 151 (4) ◽  
pp. 876-895 ◽  
Author(s):  
A L Glasebrook ◽  
F W Fitch
Keyword(s):  
T Cell ◽  
Cell Lines ◽  
Cytolytic Activity ◽  
Surface Proteins ◽  
Target Cells ◽  
Spleen Cells ◽  
T Cell Clones ◽  
Cell Clones ◽  
T Cell Lines ◽  
Cytolytic Cell

Several T cell clones have been derived by limiting dilution of secondary mixed leukocyte culture cells stimulated by H-2- and M locus (Mls)-disparate spleen cells. When examined for the expression of cytolytic activity and the ability to proliferate, these cell clones can be classified into two major categories. One type of cell is noncytolytic; when cultured with irradiated spleen cells, such clones proliferate in response to Mls determinants. Some, but not all, of these clones express Lyt-1 alloantigens. The other type of cell is cytolytic; these clones do not proliferate when cultured with irradiated allogeneic spleen cells unless supernatant fluid (SF) is added. These cytolytic clones express Lyt-2 alloantigens. Some cytolytic clones are specific for H-2Kd and others for H-2Dd alloantigens. Still other cytolytic cell clones exhibit cross-reactive lysis of different H-2-bearing tumor and Con A blast target cells. Noncytolytic T cell clones, when stimulated by Mls antigens, were examined for their ability to promote proliferation of cytolytic T cell clones. All of the noncytolytic cell clones tested were able to promote proliferation of cytolytic cell clones with the concomitant expression of cytolytic activity directed toward the original stimulating alloantigen (H-2d). Amplification of cytolytic activity was dependent upon stimulation of the noncytolytic amplifier T cell clones by Mls antigens. Specific alloantigen (signal 1), however, was not required for proliferation of the cytolytic cell clones; the amplifying signal (signal 2), delivered by the amplifier cell clones, was sufficient alone to promote proliferation of the cytolytic cell clones. Whereas proliferation of the amplifier cells was radiosensitive, the generation of the soluble amplifying signal was radioresistant. Amplification of cytolytic activity was observed when either amplifier cells were physically separated from responding cytolytic cells in Marbrook cultures or when cytolytic cells were cultured with SF collected from amplifier cell cultures. The amplifying factors were neither antigen specific nor strain specific and could be produced by Lyt-1- cells. The availability of cloned T cell lines that retain specific biologic function offers unique opportunities to characterize cell surface proteins and cell-cell interactions.

T Cell Lines and T Cell Clones Bearing Cross-Reactive Idiotype

1982 ◽  
pp. 397-404
Author(s):  
Jacques Thèze ◽  
Masao Kimoto ◽  
Marie-Lise Gougeon ◽  
Jean-Louis Moreau ◽  
Gerard Somme ◽  
...  
Keyword(s):  
T Cell ◽  
Cell Lines ◽  
T Cell Clones ◽  
Cell Clones ◽  
T Cell Lines

scholarly journals 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

2002 ◽  
Vol 70 (4) ◽  
pp. 2039-2048 ◽  
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.

scholarly journals Natural killer cell stimulatory factor (interleukin 12 [IL-12]) induces T helper type 1 (Th1)-specific immune responses and inhibits the development of IL-4-producing Th cells.

1993 ◽  
Vol 177 (4) ◽  
pp. 1199-1204 ◽  
Author(s):  
R Manetti ◽  
P Parronchi ◽  
M G Giudizi ◽  
M P Piccinni ◽  
E Maggi ◽  
...  
Keyword(s):  
T Cell ◽  
Cell Lines ◽  
Cd4 T Cell ◽  
Interleukin 12 ◽  
T Helper ◽  
Ifn Gamma ◽  
T Cell Clones ◽  
Cell Clones ◽  
T Cell Lines ◽  
Helper Type

The effects exerted on the in vitro development of antigen-specific T cell lines and T cell clones by addition or neutralization of interleukin 12 (IL-12) in lymphocyte bulk culture were examined. T cell lines specific for Dermatophagoides pteronyssinus group I (Der p I) derived in the presence of IL-12 exhibited reduced ability to produce IL-4 and increased ability to produce interferon gamma (IFN-gamma), and developed into Der p I-specific CD4+ T cell clones showing a T helper type 0 (Th0)- or Th1-, instead of Th2-, like cytokine profile. In contrast, purified protein derivative (PPD)-specific T cell lines derived in the presence of anti-IL-12 antibody exhibited an increased ability to produce IL-4 and developed into PPD-specific CD4+ T cell clones showing a Th0-, instead of Th1-, like profile. The influence of IL-12 on the cytokine secretion profile of Der p I-specific T cell lines was not prevented by addition to lymphocyte bulk cultures of anti-IFN-gamma antibody, but could be at least partially inhibited by the removal from bulk cultures of CD16+ cells. Thus, IL-12 and CD16+ cells appear to have inhibitory effects on the development of IL-4-producing cells and to play an inductive role in promoting Th1-like responses.

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