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Functional Reconstitution of Class II MHC-Restricted T Cell Immunity Mediated by Retroviral Transfer of the ß TCR Complex¹

Keishi Fujio^*,, Yoshikata Misaki, Keigo Setoguchi, Sumiyo Morita^*, Kimito Kawahata, Ikunoshin Kato, Tetsuya Nosaka^*, Kazuhiko Yamamoto and Toshio Kitamura^2,*

^* Department of Hematopoietic Factors, Institute of Medical Science, University of Tokyo, Tokyo, Japan; Department of Allergy and Rheumatology, Graduate School of Medicine, University of Tokyo, Tokyo, Japan; and Biotechnology Research Laboratories, Takara Shuzo, Shiga, Japan

	Abstract

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Transfer of the ß TCR genes into T lymphocytes will provide a means to enhance Ag-specific immunity by increasing the frequency of tumor- or pathogen-specific T lymphocytes. We generated an efficient ß TCR gene transfer system using two independent monocistronic retrovirus vectors harboring either of the class II MHC-restricted or ß TCR genes specific for chicken OVA. The system enabled us to express the clonotypic TCR in 44% of the CD4⁺ T cells. The transduced cells showed a remarkable response to OVA_323–339 peptide in the in vitro culture system, and the response to the Ag was comparable with those of the T lymphocytes derived from transgenic mice harboring OVA-specific TCR. Adoptive transfer of the TCR-transduced cells in mice induced the Ag-specific delayed-type hypersensitivity in response to OVA_323–339 challenge. These results indicate that ß TCR gene transfer into peripheral T lymphocytes can reconstitute Ag-specific immunity. We here propose that this method provides a basis for a new approach to manipulation of immune reactions and immunotherapy.

	Introduction

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Antigen-specific immune response plays a key role in immunity to pathogens or neoplasms (1, 2, 3). T cells bearing virus- or tumor-specific TCR have been used to establish the concept of adoptive immunotherapy, a possible choice for therapy. CTLs have been used in many Ag-specific immunotherapeutical experiments. Adoptive transfer of tumor-specific CTLs causes regression of murine tumors (4), mediates a partial response in some melanoma patients (5), and contributes to the resolution of EBV-associated lymphoproliferative disease in bone marrow transplant recipients (6). On one hand, Ag-specific CD4⁺ helper T lymphocytes are also important for tumor immunity in activating peripheral CTLs via the CD40-CD40 ligand interaction (7); adoptive transfer of either subset of tumor-specific helper T lymphocytes, Th1 or Th2, leads to tumor eradication in mice (8). However, the number of tumor- or pathogen-specific T lymphocytes is usually limited, even under conditions of vast Ag loading or vaccination (9, 10). Reconstitution of functional specific TCRs by the transfer of ß TCR genes into T cell hybridomas and human Jurkat cells was reported previously (11, 12, 13, 14). If we could transfer the ß TCR genes into T lymphocytes, we would be able to increase the frequency of tumor- or pathogen-specific T lymphocytes in the body by infusing the Ag-specific T cells after in vitro expansion. To this end, Pogulis and Pease used a bicistronic retroviral vector harboring the - and ß-chain of TCR genes in an attempt to express TCR on a hybridoma (15); however, cell sorting was needed to detect the transferred TCR complex as the transduction efficiency was low. In addition, this model experiment was performed using a cell line, which could not be applied for practical therapy. Therefore, it is required to establish a system that provides efficient expression of both of the TCR chains on nontransformed lymphocytes. Using an efficient retrovirus system, we here report functional reconstitution of TCR in CD4⁺ T cells that is verified by both in vitro and in vivo assays.

	Materials and Methods

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Animals

BALB/c mice were obtained from Japan SLC (Shizuoka, Japan). The transgenic (Tg)³ mice expressing OVA_323–339-specific I-A^d-restricted TCR (DO11.10 TCR) maintained on the BALB/c background were kindly provided by Dr. T. Watanabe (Medical Institute of Bioregulation, Kyushu University, Fukuoka, Japan) (16). All the mice used were female and were used at 7–8 wk of age.

Cytokines, Abs, Ags, and cell separation

Recombinant murine (m) IL-2 was obtained from R&D Systems (Minneapolis, MN). FITC-anti-CD3 mAb, FITC-anti-CD8 mAb, FITC-anti-CD11b mAb, FITC-anti-CD19 mAb, and peridinin-chlorophyll (PerCP)-anti-CD4 mAb were obtained from PharMingen (San Diego, CA). KJ1-26, a clonotype-specific mAb raised against TCR from a T cell hybridoma DO11.10, was kindly provided by Dr. T. Watanabe. Anti-FITC-conjugated microbeads, the MS⁺ separation columns, and the MiniMACS^- separation device were purchased from Miltenyi Biotec (Bergisch Gladbach, Germany) and were used according to the manufacturer’s instructions.

Cell culture

TG40 is a cell-surface TCR-negative, intracytoplasmic CD3-positive mutant of the 21.2.2 mouse T cell line (17, 18). A packaging cell line, PLAT-E, can produce retroviruses with a titer of about 1 x 10⁷/ml, which we have recently developed (19). PLAT-E was cultured in DMEM supplemented with 10% FCS, 2 mM L-glutamine, 1 µg/ml puromycin, and 10 µg/ml blasticidin. Splenocytes were cultured in RPMI 1640 medium containing 10% FCS, 2 mM L-glutamine, 100 U/ml penicillin, 100 µg/ml streptomycin, 0.05 mM 2-ME, 50 ng/ml mIL-2, and 10 µg/ml Con A for 24 h before retroviral infection. Irradiated (3300 rad) syngeneic spleen cells were used as a source of APCs.

Construction of TCR expression vectors

Complementary DNAs for the TCR - and ß-chains were isolated from a cDNA library of DO11.10 TCR Tg splenocytes and were inserted into a retroviral vector pMX (20) to generate pMX-DOTAE and pMX-DOTBE, respectively, or inserted into pMX-puro (21) or pMX-neo retroviral vectors to generate pMX-puro-DOTAE and pMX-neo-DOTBE, respectively (Fig. 1). A retrovirus vector pMX-neo harbors a SV40 early promoter-driven neomycin resistance gene between the multicloning site and the 3' long terminal repeat of the pMX vector. The resulting expression plasmids were transfected to PLAT-E cells, and the supernatants were collected, as described (22).

View larger version (13K): [in this window] [in a new window]   FIGURE 1. Schematic diagrams of the protocol for reconstitution of T cell immunity mediated by retroviral gene transfer. The supernatants were mixed and coinfected three times to splenocytes. The cells coexpressing transduced - and ß-chains (hatched cell) were detected by the anti-clonotypic Ab.

The two plasmids, pMX-puro-DOTAE and pMX-neo-DOTBE, were retrovirally introduced into TG40, as described (22). Splenocytes were incubated with a mixture of both viral supernatants derived from pMX-DOTAE and pMX-DOTBE on nontissue culture 24-well plates coated with recombinant fibronectin fragment CH-296, RetroNectin (TaKaRa, Ohtsu, Japan) (23), according to the manufacturer’s instruction. Infections were repeated at 8 and 16 h after the first round (Fig. 1).

Enrichment of CD4⁺ cells and analysis of surface expression of the TCR complex

CD4⁺ T cells were enriched by negative selection; CD4⁺ T cells were magnetically purified from mock-infected splenocytes (mock-splenocytes), splenocytes coinfected with pMX-DOTAE and -DOTBE (AB-splenocytes), and DO11.10 Tg splenocytes (Tg-splenocytes) using FITC-labeled anti-CD8, -CD11b, and -CD19 mAbs, followed by the anti-FITC-conjugated paramagnetic beads and the MS⁺ separation columns (Miltenyi Biotec). Depletion of CD8-, CD11b-, or CD19-positive cells was achieved by collecting only the first eluted fraction through the column. For detection of the transduced TCR complex, mock-splenocytes, AB-splenocytes, and Tg-splenocytes, as well as CD4⁺ fraction of each culture (mock-CD4⁺ T cells, AB-CD4⁺ T cells, Tg-CD4⁺ T cells), were stained with FITC-anti-CD8 mAb, biotinylated KJ1-26 (b-KJ1-26) plus streptavidin-PE conjugate (SAPE), and PerCP-anti-CD4 mAb. For detection of the CD3 complex, cells were stained with FITC-anti-CD3 mAb, b-KJ1-26 plus SAPE, and PerCP-anti-CD4 mAb. Fluorescence intensity was measured using flow cytometry (FACSCalibur; Becton Dickinson, San Diego, CA) 54 h after the first infection.

Proliferation assay

Splenocytes were cultured at 1 x 10⁴ cells/well, with 1 x 10⁵ cells/well of APCs in 96-well, flat-bottom microtiter plates in a volume of 100 µl of complete medium 48 h after the first infection. After 24 h of culture, the cells were pulse-labeled with 1 µCi of [³H]thymidine/well (NEN Life Science Products, Boston, MA) for 12 h. After the labeling, the cells were harvested using Printed Filtermat A (Wallac, Turuku, Finland) with a Micro96 Harvester (Skatron, Sterling, VA), and the incorporation of [³H]thymidine was determined by MicroBeta (Pharmacia, Piscataway, NJ) according to the manufacturer’s instruction.

Local adoptive transfer (LAT) assays

The LAT assay was used to investigate in vivo functions of TCR-transduced CD4⁺ lymphocytes (24). Mock-, AB-, or Tg-CD4⁺ T cells (1 x 10⁶) were mixed with APCs (1 x 10⁷) in 20 µl of PBS and were injected with or without OVA_323–339 (3 µM) into the right and left hind footpads of the mice, respectively, 60 h after the infection started. Thickness of the footpads was measured after 20 h, and swelling was expressed as (changes of right footpad thickness) - (changes of left footpad thickness) and was measured using a Digimatic Caliper (Mitsutoyo, Kanagawa, Japan). In each experiment, eight mice were used for injection of mock- and AB-transfected CD4⁺ T cells, and four mice were used for injection of Tg-transfected CD4⁺ T cells.

	Results

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Surface expression of the TCR complex derived from DO11.10 hybridoma on the TG40 cell line

The TG40 cells transduced with TCR and selected for 7 days in the presence of 1 µg/ml puromycin and 1 mg/ml G418 (Life Technologies, Grand Island, NY) (AB-TG40) and parental TG40 cells were stained with FITC-conjugated anti-CD3 mAb and b-KJ1-26 plus SAPE. Most AB-TG40 cells were found to express both CD3 and the clonotypic TCR (Fig. 2), indicating that retrovirally transferred TCR - and ß-chains were paired and coexpressed with CD3 on the cell surface.

View larger version (22K): [in this window] [in a new window]   FIGURE 2. FACS analysis of the expression of clonotypic TCR complex on the cell surface of transduced TG40 cells.

In an attempt to investigate functional properties of the reconstituted TCR, we next transferred TCR genes into splenocytes. The expression and function of the transduced TCR in the splenocytes were compared with those of control cells derived from DO11.10 Tg mice. In each well of a 24-well plate, 4 x 10⁶ splenocytes were infected, and an average of 2 x 10⁶ blastic cells were harvested after 48 h. Expression levels of the clonotypic TCR on the surface of the splenocytes were determined by FACS analysis. A gate was set on the lymphocyte population, and T cells accounted for over 98% of the gated cells (data not shown). Clonotype-positive cells were detected in 18.5% of AB-splenocytes and 64.4% in Tg-splenocytes, and these clonotype-positive cells were mostly CD4⁺ cells (Fig. 3A). To assay the efficiency and function of the transduced class II-restricted TCR on CD4⁺CD8^- cells, CD4-positive splenocytes were enriched by MACS beads as described in Materials and Methods. The enriched cells consisted of a large number of CD4⁺CD8^- cells, a few CD4^-CD8^- cells, and rare CD4^-CD8⁺ cells. In AB-CD4⁺ T cells, the clonotype positivity in CD4⁺CD8^- cells reached 44.0% (Fig. 3B), and the expression level of clonotypic TCR reached approximately half that of Tg-CD4⁺ T cells, as assessed based on mean fluorescent intensities in histogram analysis. No significant right-shift of CD3 staining level was evident in the CD4⁺ clonotype-positive population, as compared with the mock-CD4⁺ population (Fig. 3C).

View larger version (42K): [in this window] [in a new window]   FIGURE 3. FACS analysis of the expression of clonotypic TCR on the cell surface of transduced mouse T lymphocytes. About 98% of the cells in the gate used were positive for anti-CD3 (data not shown). A, Anti-CD4 and KJ1-26 staining of mock-, AB-, and Tg-splenocytes. B, Anti-CD4 and KJ1-26 staining of mock-, AB-, and Tg-CD4+ T cells. Depletion of CD8+ cells was confirmed. Approximately 40% of AB-CD4+ T cells were positive for clonotype TCR. The mean staining level of AB-CD4+ T cells reached approximately half that of Tg-CD4+ T cells in the histogram analysis. C, Anti-CD3 staining of the clonotype-positive population, mock-CD4+ T cells, and Tg-CD4+ T cells. No evident right-shift was observed in the clonotype-positive population. Three independent experiments yielded similar results, and a representative experiment is shown.

The specific reactivity of AB-CD4⁺ T cells to OVA_323–339 was examined in in vitro culture. While AB-CD4⁺ T cells showed a remarkable dose-dependent response to OVA_323–339, no such response was detected in mock-CD4⁺ T cells (Fig. 4). Maximum response of AB-CD4⁺ T cells was comparable to that of Tg-CD4⁺ T cells.

View larger version (15K): [in this window] [in a new window]   FIGURE 4. Reactivity of mock-, AB-, and Tg-CD4+ T cells to OVA323–339 peptide. Incorporated cpm of [3H]thymidine at the indicated concentrations is shown. Data represent the mean ± SD of three independent experiments.

Definite swelling was noted in the right hind footpad after 20 h in the case of AB-CD4⁺ T cell-injected mice, but not in the mock-CD4⁺ T cell-injected mice. Footpad swellings of mice injected with AB-CD4⁺ T cells were comparable to those of mice injected with Tg-CD4 cells (Fig. 5).

View larger version (11K): [in this window] [in a new window]   FIGURE 5. Footpad swelling in the LAT assay. Mock-, AB-, or Tg-CD4+ T cells (1 x 106) were mixed with APCs (1 x 107). The mixture (20 µl) with and without OVA323–339 peptide were injected into the right and left hind footpads, respectively. After 20 h, thickness of the footpads was measured, and swelling expressed as (changes of right footpad thickness) - (changes of left footpad thickness). Left lane, Mock-CD4+ T cells. Middle lane, AB-CD4+ T cells. Right lane, Tg-CD4+ T cells. Three independent experiments yielded similar results, and a representative experiment is shown. In each experiment, eight mice were used for injection of mock- and AB-CD4+ T cells, and four mice were used for injection of Tg-CD4+ T cells.

	Discussion

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For cotransfection of TCR- and -ß genes, either a single bicistronic vector containing both genes tandemly or two independent monocistronic vectors can be used. We chose to use two independent monocistronic vectors because of the higher titers of the produced viruses and the stable expression compared with that by bicistronic vectors (25). As a packaging cell line, we used an efficient packaging cell line, PLAT-E, which we recently established (19). PLAT-E uses the EF1 promoter in combination with the Kozak’s consensus sequence to efficiently drive expression of gag-pol and env gene of the Moloney Murine Leukemia Virus (MoMLV), and PLAT-E produces high-titer retroviruses with an average titer of 1 x 10⁷/ml.

Because retroviral vectors based on MoMLV can infect only dividing cells, most of the infected cells cultured in the presence of IL-2 and Con A are expected to be T lymphocytes. Moreover, macrophages or B lymphocytes cannot express the TCR complex because the CD3 complex, which is required for surface expression of ß TCR chains (26, 27), is expressed exclusively on the T lineage. In addition, results from Tg mice expressing a class I or II MHC-restricted ß TCR indicate that T lymphocytes managed to express an appropriate combination of ß TCR and coreceptor molecule for signaling, i.e., class I-restricted receptors with CD8 and class II-restricted receptors with CD4 (28). Therefore, we assume that the exogenously expressed class-II MHC-restricted TCRs would transduce signals only in CD4⁺CD8^- but not in CD4^-CD8⁺ cells.

It is important to investigate whether the transduced TCR could be expressed on cell surface of T cells. Although phenotypic allelic exclusion (posttranslational regulation) of TCR V was noted in mature lymphocytes (29, 30), the transduced TCR -chain was expressed in >40% of CD4⁺ T cells at relatively high levels in our experiments. Phenotypic allelic exclusion appears to be maintained by competition for the ß-chain between the -chains (31, 32, 33); hence, efficient clonotype expression may be derived from specific pairing of the transduced -chain with the cotransduced TCR ß-chain, which originally paired in the DO11.10 cell line. Alternatively, the abundance of the transduced - and ß-chains was sufficient to overwhelm endogenous - and ß-chains. The lack of evident up-regulation of the CD3 complex despite of the efficient expression of the transduced TCR suggests that the transduced subunits were expressed in a substitutive rather than in an additive manner to endogenous TCR. While this suggestion is consistent with the exclusion model, the details of exogenous TCR expression remain to be explored.

While we were able to reconstitute the function of the transduced class II-restricted TCR in CD4⁺ T cells, CD8⁺ T cells were also found to express the transduced TCR (Fig. 3B). This result suggests that transducing class I-restricted TCR to CD8⁺ T cells with this method would also lead to reconstitution of the class I-restricted T cell immunity. Manipulation of both class I and II MHC-restricted T cell immunities will pave a road to the investigation of the immune system and to the development of immunotherapy for patients with neoplasms or pathogenic microorganisms.

	Acknowledgments

 
We thank M. Ohara for critical reading of the manuscript, Dr. T. Saito (Department of Molecular Genetics, Chiba University Graduate School of Medicine, Chiba, Japan) for the kind gift of the TG40 cell line, and Dr. T. Watanabe (Kyusyu University) for making available to us the Tg mice and the Ab.

	Footnotes

 
¹ This work was supported in part by a grant-in-aid from the Ministry of Education, Science, Sports, and Culture of Japan, and by a grant-in-aid from the Ministry of Health and Welfare. The Department of Hematopoietic Factors is supported by Chugai Pharmaceutical Company Ltd.

² Address correspondence and reprint requests to Dr. Toshio Kitamura, Department of Hematopoietic Factors, Institute of Medical Science, University of Tokyo, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639, Japan.

³ Abbreviations used in this paper: Tg, transgenic; LAT, local adoptive transfer; MoMLV, Molony murine leukemia virus; m, murine; PerCP, peridinin-chlorophyll; SAPE, streptavidin-PE conjugate; b-KJ1-26, biotinylated Kj1-26.

Received for publication January 18, 2000. Accepted for publication April 11, 2000.

	References

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1. Boon, T., J. C. Cerottini, B. Van den Eynde, P. van der Bruggen, A. Van Pel. 1994. Tumor antigens recognized by T lymphocytes. Annu. Rev. Immunol. 12:337.[Medline]
2. Cox, A. L., J. Skipper, Y. Chen, R. A. Henderson, T. L. Darrow, J. Shabanowitz, V. H. Engelhard, D. F. Hunt, Jr C. L. Slingluff. 1994. Identification of a peptide recognized by five melanoma-specific human cytotoxic T cell lines. Science 264:716.[Abstract/Free Full Text]
3. Finn, O. J.. 1993. Tumor-rejection antigens recognized by T lymphocytes. Curr. Opin. Immunol. 5:701.[Medline]
4. Melief, C. J., W. M. Kast. 1995. T-cell immunotherapy of tumors by adoptive transfer of cytotoxic T lymphocytes and by vaccination with minimal essential epitopes. Immunol. Rev. 145:167.[Medline]
5. Ettinghausen, S. E., S. A. Rosenberg. 1995. Immunotherapy and gene therapy of cancer. Adv. Surg. 28:223.[Medline]
6. Rooney, C. M., C. A. Smith, C. Y. Ng, S. Loftin, C. Li, R. A. Krance, M. K. Brenner, H. E Heslop. 1995. Use of gene-modified virus-specific T lymphocytes to control Epstein- Barr-virus-related lymphoproliferation. Lancet 345:9.[Medline]
7. Schoenberger, S. P., R. E. Toes, E. I. van der Voort, R. Offringa, C. J. Melief. 1998. T-cell help for cytotoxic T lymphocytes is mediated by CD40-CD40L interactions. Nature 393:480.[Medline]
8. Nishimura, T., K. Iwakabe, M. Sekimoto, Y. Ohmi, T. Yahata, M. Nakui, T. Sato, S. Habu, H. Tashiro, M. Sato, A. Ohta. 1999. Distinct role of antigen-specific T helper type 1 (Th1) and Th2 cells in tumor eradication in vivo. J. Exp. Med. 190:617.[Abstract/Free Full Text]
9. Staveley-O’Carroll, K., E. Sotomayor, J. Montgomery, I. Borrello, L. Hwang, S. Fein, D. Pardoll, H. Levitsky. 1998. Induction of antigen-specific T cell anergy: an early event in the course of tumor progression. Proc. Natl. Acad. Sci. USA 95:1178.[Abstract/Free Full Text]
10. Aichele, P., K. Brduscha-Riem, R. M. Zinkernagel, H. Hengartner, H. Pircher. 1995. T cell priming versus T cell tolerance induced by synthetic peptides. J. Exp. Med. 182:261.[Abstract/Free Full Text]
11. Saito, T., A. Weiss, J. Miller, M. A. Norcross, R. N. Germain. 1987. Specific antigen-Ia activation of transfected human T cells expressing murine Ti ß-human T3 receptor complexes. Nature 325:125.[Medline]
12. Gabert, J., C. Langlet, R. Zamoyska, J. R. Parnes, A. M. Schmitt-Verhulst, B. Malissen. 1987. Reconstitution of MHC class I specificity by transfer of the T cell receptor and Lyt-2 genes. Cell 50:545.[Medline]
13. Saito, T., R. N. Germain. 1987. Predictable acquisition of a new MHC recognition specificity following expression of a transfected T-cell receptor ß-chain gene. Nature 329:256.[Medline]
14. Kuo, C. L., L. Hood. 1987. Antigen/major histcompatibility complex-specific activation of murine T cells transfected with functionally rearranged T-cell receptor genes. Proc. Natl. Acad. Sci. USA 84:7614.[Abstract/Free Full Text]
15. Pogulis, R. J., L. R. Pease. 1998. A retroviral vector that directs simultaneous expression of and ß T cell receptor genes. Hum. Gene Ther. 9:2299.[Medline]
16. Murphy, K. M., A. B. Heimberger, D. Y. Loh. 1990. Induction by antigen of intrathymic apoptosis of CD4⁺CD8⁺TCR^lo thymocytes in vivo. Science 250:1720.[Abstract/Free Full Text]
17. Sussman, J. J., T. Saito, E. M. Shevach, R. N. Germain, J. D. Ashwell. 1988. Thy-1- and Ly-6-mediated lymphokine production and growth inhibition of a T cell hybridoma require co-expression of the T cell antigen receptor complex. J. Immunol. 140:2520.[Abstract]
18. Zumla, A., C. Marguerie, A. So, W. M. Yokoyama, T. Saito, J. R. Batchelor, R. I. Lechler. 1992. Co-expression of human T cell receptor chains with mouse CD3 on the cell surface of a mouse T cell hybridoma. J. Immunol. Methods 149:69.[Medline]
19. Morita, S., T. Kojima, and T. Kitamura. 2000. Plat-E; an efficient and stable system for transient packaging of retroviruses. Gene Ther. In press.
20. Onishi, M., S. Kinoshita, Y. Morikawa, A. Shibuya, J. Phillips, L. L. Lanier, D. M. Gorman, G. P. Nolan, A. Miyajima, T. Kitamura. 1996. Applications of retrovirus-mediated expression cloning. Exp. Hematol. 24:324.[Medline]
21. Onishi, M., T. Nosaka, K. Misawa, A. L. Mui, D. Gorman, M. McMahon, A. Miyajima, T. Kitamura. 1998. Identification and characterization of a constitutively active STAT5 mutant that promotes cell proliferation. Mol. Cell. Biol. 18:3871.[Abstract/Free Full Text]
22. Kitamura, T., M. Onishi, S. Kinoshita, A. Shibuya, A. Miyajima, G. P. Nolan. 1995. Efficient screening of retroviral cDNA expression libraries. Proc. Natl. Acad. Sci. USA 92:9146.[Abstract/Free Full Text]
23. Hanenberg, H., X. L. Xiao, D. Dilloo, K. Hashino, I. Kato, D. A. Williams. 1996. Colocalization of retrovirus and target cells on specific fibronectin fragments increases genetic transduction of mammalian cells. Nat. Med. 2:876.[Medline]
24. Kawahata, K., Y. Misaki, Y. Komagata, K. Setoguchi, S. Tsunekawa, Y. Yoshikawa, J. Miyazaki, K. Yamamoto. 1999. Altered expression level of a sytemic nuclear autoantigen determines the fate of immune response to self. J. Immunol. 162:6482.[Abstract/Free Full Text]
25. Hildinger, M., B. Fehse, S. Hegewisch-Becker, J. John, Jr W. Rafferty, W. Ostertag, C. Baum. 1998. Dominant selection of hematopoietic progenitor cells with retroviral MDR1 co-expression vectors. Hum. Gene Ther. 9:33.[Medline]
26. Alarcon, B., B. Berkhout, J. Breitmeyer, C. Terhorst. 1988. Assembly of the human T cell receptor-CD3 complex takes place in the endoplasmic reticulum and involves intermediary complexes between the CD3- core and single T cell receptor or ß chains. J. Biol. Chem. 263:2953.[Abstract/Free Full Text]
27. Soudais, C., J. P. de Villartay, F. Le Deist, A. Fischer, B. Lisowska-Grospierre. 1993. Independent mutations of the human CD3- gene resulting in a T cell receptor/CD3 complex immunodeficiency. Nat. Genet. 3:77.[Medline]
28. von Boehmer, H.. 1990. Developmental biology of T cells in T cell-receptor transgenic mice. Annu. Rev. Immunol. 8:531.[Medline]
29. Alam, S. M., I. N. Crispe, N. R. Gascoigne. 1995. Allelic exclusion of mouse T cell receptor chains occurs at the time of thymocyte TCR up-regulation. Immunity 3:449.[Medline]
30. Alam, S. M., N. R. Gascoigne. 1998. Posttranslational regulation of TCR V allelic exclusion during T cell differentiation. J. Immunol. 160:3883.[Abstract/Free Full Text]
31. Malissen, M., J. Trucy, F. Letourneur, N. Rebai, D. E. Dunn, F. W. Fitch, L. Hood, B. Malissen. 1988. A T cell clone expresses two T cell receptor genes but uses one ß heterodimer for allorecognition and self MHC-restricted antigen recognition. Cell 55:49.[Medline]
32. Malissen, M., J. Trucy, E. Jouvin-Marche, P. A. Cazenave, R. Scollay, B. Malissen. 1992. Regulation of TCR and ß gene allelic exclusion during T-cell development. Immunol. Today 13:315.[Medline]
33. Borgulya, P., H. Kishi, Y. Uematsu, H. von Boehmer. 1992. Exclusion and inclusion of and ß T cell receptor alleles. Cell 69:529.[Medline]

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				M. H. M. Heemskerk, R. S. Hagedoorn, M. A. W. G. van der Hoorn, L. T. van der Veken, M. Hoogeboom, M. G. D. Kester, R. Willemze, and J. H. F. Falkenburg Efficiency of T-cell receptor expression in dual-specific T cells is controlled by the intrinsic qualities of the TCR chains within the TCR-CD3 complex Blood, January 1, 2007; 109(1): 235 - 243. [Abstract] [Full Text] [PDF]

				K. Fujio, A. Okamoto, Y. Araki, H. Shoda, H. Tahara, N. H. Tsuno, K. Takahashi, T. Kitamura, and K. Yamamoto Gene Therapy of Arthritis with TCR Isolated from the Inflamed Paw J. Immunol., December 1, 2006; 177(11): 8140 - 8147. [Abstract] [Full Text] [PDF]

				S.-A. Xue, L. Gao, D. Hart, R. Gillmore, W. Qasim, A. Thrasher, J. Apperley, B. Engels, W. Uckert, E. Morris, et al. Elimination of human leukemia cells in NOD/SCID mice by WT1-TCR gene-transduced human T cells Blood, November 1, 2005; 106(9): 3062 - 3067. [Abstract] [Full Text] [PDF]

				T. Tsuji, M. Yasukawa, J. Matsuzaki, T. Ohkuri, K. Chamoto, D. Wakita, T. Azuma, H. Niiya, H. Miyoshi, K. Kuzushima, et al. Generation of tumor-specific, HLA class I-restricted human Th1 and Tc1 cells by cell engineering with tumor peptide-specific T-cell receptor genes Blood, July 15, 2005; 106(2): 470 - 476. [Abstract] [Full Text] [PDF]

				E. C. Morris, A. Tsallios, G. M. Bendle, S.-a. Xue, and H. J. Stauss A critical role of T cell antigen receptor-transduced MHC class I-restricted helper T cells in tumor protection PNAS, May 31, 2005; 102(22): 7934 - 7939. [Abstract] [Full Text] [PDF]

				D. Zhang, K. Fujio, Y. Jiang, J. Zhao, N. Tada, K. Sudo, H. Tsurui, K. Nakamura, K. Yamamoto, H. Nishimura, et al. Dissection of the role of MHC class II A and E genes in autoimmune susceptibility in murine lupus models with intragenic recombination PNAS, September 21, 2004; 101(38): 13838 - 13843. [Abstract] [Full Text] [PDF]

				K. Fujio, A. Okamoto, H. Tahara, M. Abe, Y. Jiang, T. Kitamura, S. Hirose, and K. Yamamoto Nucleosome-Specific Regulatory T Cells Engineered by Triple Gene Transfer Suppress a Systemic Autoimmune Disease J. Immunol., August 1, 2004; 173(3): 2118 - 2125. [Abstract] [Full Text] [PDF]

				M. H.M. Heemskerk, M. Hoogeboom, R. Hagedoorn, M. G.D. Kester, R. Willemze, and J.H. F. Falkenburg Reprogramming of Virus-specific T Cells into Leukemia-reactive T Cells Using T Cell Receptor Gene Transfer J. Exp. Med., April 5, 2004; 199(7): 885 - 894. [Abstract] [Full Text] [PDF]

				K. Chamoto, T. Tsuji, H. Funamoto, A. Kosaka, J. Matsuzaki, T. Sato, H. Abe, K. Fujio, K. Yamamoto, T. Kitamura, et al. Potentiation of Tumor Eradication by Adoptive Immunotherapy with T-cell Receptor Gene-Transduced T-Helper Type 1 Cells Cancer Res., January 1, 2004; 64(1): 386 - 390. [Abstract] [Full Text] [PDF]

				H. Tahara, K. Fujio, Y. Araki, K. Setoguchi, Y. Misaki, T. Kitamura, and K. Yamamoto Reconstitution of CD8+ T Cells by Retroviral Transfer of the TCR {alpha}{beta}-Chain Genes Isolated from a Clonally Expanded P815-Infiltrating Lymphocyte J. Immunol., August 15, 2003; 171(4): 2154 - 2160. [Abstract] [Full Text] [PDF]

				J. J. Roszkowski, D. C. Yu, M. P. Rubinstein, M. D. McKee, D. J. Cole, and M. I. Nishimura CD8-Independent Tumor Cell Recognition Is a Property of the T Cell Receptor and Not the T Cell J. Immunol., March 1, 2003; 170(5): 2582 - 2589. [Abstract] [Full Text] [PDF]

				N. Schaft, R. A. Willemsen, J. de Vries, B. Lankiewicz, B. W. L. Essers, J.-W. Gratama, C. G. Figdor, R. L. H. Bolhuis, R. Debets, and G. J. Adema Peptide Fine Specificity of Anti-Glycoprotein 100 CTL Is Preserved Following Transfer of Engineered TCR{alpha}{beta} Genes Into Primary Human T Lymphocytes J. Immunol., February 15, 2003; 170(4): 2186 - 2194. [Abstract] [Full Text] [PDF]

				M. P. Rubinstein, A. N. Kadima, M. L. Salem, C. L. Nguyen, W. E. Gillanders, M. I. Nishimura, and D. J. Cole Transfer of TCR Genes into Mature T Cells Is Accompanied by the Maintenance of Parental T Cell Avidity J. Immunol., February 1, 2003; 170(3): 1209 - 1217. [Abstract] [Full Text] [PDF]

				T. Ueno, H. Tomiyama, and M. Takiguchi Single T Cell Receptor-Mediated Recognition of an Identical HIV-Derived Peptide Presented by Multiple HLA Class I Molecules J. Immunol., November 1, 2002; 169(9): 4961 - 4969. [Abstract] [Full Text] [PDF]

				L. Yang, X.-F. Qin, D. Baltimore, and L. Van Parijs Generation of functional antigen-specific T cells in defined genetic backgrounds by retrovirus-mediated expression of TCR cDNAs in hematopoietic precursor cells PNAS, April 30, 2002; 99(9): 6204 - 6209. [Abstract] [Full Text] [PDF]

				S.-i. Kagami, H. Nakajima, A. Suto, K. Hirose, K. Suzuki, S. Morita, I. Kato, Y. Saito, T. Kitamura, and I. Iwamoto Stat5a regulates T helper cell differentiation by several distinct mechanisms Blood, April 15, 2001; 97(8): 2358 - 2365. [Abstract] [Full Text] [PDF]

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