In vitro activated γ δ lymphocytes

Drug – bio-affecting and body treating compositions – Whole live micro-organism – cell – or virus containing – Animal or plant cell

Reexamination Certificate

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C424S534000, C435S007100, C435S007240

Reexamination Certificate

active

07078034

ABSTRACT:
Patients who develop increased numbers of γδ+ cytotoxic T lymphocytes 2–6 months after allogeneic bone marrow transplantation are less likely to relapse than those who do not. The γδ+ T cells isolated from blood of patients with increased γδ+ T cells are CD3+CD4−CD8−CD57+, cytolytic to K562 cells, and express the Vδ1 T cell receptor phenotype. Similar γδ+ T cells can be generated in vitro by culture of donor mononuclear cells which are enriched for γδ+ T cells by immunomagnetic depletion of depleted of CD4+ and CD8+ cells. This γδ-enriched cell preparation was cultured on a combination of immobilized pan-δ monoclonal antibody and irradiated recipient B cell leukemia. After four weeks, the cultures were almost exclusively Vδ1+CD3+CD4−CD8− cells that co-expressed activation-associated antigens CD69, CD25, and HLA-DR. Furthermore, they were cytolytic against the primary leukemia obtained from the recipient and lymphoblastic leukemia cell lines, yet had minimal cytotoxicity against normal donor-derived mononuclear cells or myeloid leulemia cell lines. These observations suggest that donor-derived cytotoxic γδ+ T cells can be generated in vitro, and may provide therapeutic potential for prevention of disease relapse.

REFERENCES:
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Henslee-Downey PJ, Abhyankar SH, Parrish RS, Pati AR, Godder KT, Neglia WJ, Goon-Johnson KS, Geier SS, Lee CG, Gee AP. Use of partially mismatched related donors extends access to allogeneic marrow transplant. Blood 89 (10):3864, 1997, discloses the increase in disease relapse upon the use of T cell depletion to prevent graft-versus-host disease.
Horowitz, M.M., Gale, R.P., Sondel, P.M., Goldman, J.M. et al. Graft versus leukemia reactions after bone marrow transplantation. Blood 75, 555, 1990, discloses that graft-versus-leukemia effectors are predominantly T cells that can either recognize allospecific molecules and cell surface molecules.
Sykes, M., Romick, M.L., Sachs, D.H. Interleukin 2 prevents graft-versus-host disease while preserving the graft-versus-leukemia effect of allogeneic T cells. Proc. Nat. Acad. Sci. 1990; 87: 5633, discloses evidence that graft-versus-host and graft-versus-leukemia effects can be separated to some degree.
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Viale M, Ferrini S, Bacigalupo A. TCR positive lymphocytes after allogeneic bone marrow transplantation. Bone Marrow Transplantation 10:249, 1992, discloses an increase in the ration of V 1:V 2 cells in patients with acute graft-versus-host disease.
Yabe M, Yabe H, Hattori K, Hinorhars T, Morimoto T, Kato S, Kusonoki A. Transition of T-cell receptor gamma/delta expressing double negative (CD4/CD8) lymphocytes after allogeneic bone marrow transplantation. Bone Marrow Transplantation 14:741, 1994, discloses increases in the ration of CD4—CD8—gd+ T cells during post-bone marrow transplantation.
Tsuji S, Char D, Bucy RP, Simonsen M, Chen C, Cooper MD. +T-cells are secondary participants in acute graft-versus-host reactions initiated by CD4+ T-cells. European Journal of Immunology 26: 420, 1996, discloses gd+ T cells that are recruited into donor + lesions where they are activated and induced to proliferate.
Cela ME, Holliday MS, Rooney CM, Richardson S, Alexander B, Krance RA, Brenner MK, Heslop HE. + T-lymphocyte regeneration after T-lymphocyte-depleted bone marrow transplantation from mismatched family members or matched unrelated donors. Bone Marrow Transplantation 17:243, 1996, discloses the use of selective or targeted type of T cell delpletion combined with post-transplant immune suppression.
Esslin A, Formby B. Comparison of cytolytic and proliferative activities of human and T-cells from peripheral blood against various human tumor cell lines. Journal of the National Cancer Institute 83:1564, 1994, discloses that in vitro activated gd+ T cells can mediate broadly-based non-MHC restricted cytolytic activity to selected human tumor cell lines.
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Hoffman, T., Theobald, M., Bunjes, D., Weiss, M., Heimpel, H., Heidt, W. Frequency of bone marrow T cells responding to HLA-identical non-leukemic and leukemic stimulator cells. Bone Marrow Transplantation 1993; 12: 1, discloses the generation of leukemia-reactive clones.
Fisch, P., Malkovska, M., Braakman, E., Sturm, E., Bolhuis, R.L., Prieve, A., Sosman, J.A., Lam, V.A., Sondel, P.M. Gamma/delta T cell clones and natural killer cell clones mediate distinct patterns of non-major histocompatibility-restricted cytolysis. J. Exp. Med. 1990; 171: 1567, discloses the identification of potential leukemia-associated antigens.
Kaur, I., Voss, S.D., Gupta, R.S., Schell, K., Fisch, P., Sondel, P.M. Human peripheral gamma/delta T cells recognize hsp60 molecules on Daudi Burkitt's lymphoma cells. J. Immunol. 1993; 150: 2046, discloses gd+ T cells response to heat shock proteins.
Potential antileukemic effect of gamma delta T cells in acute lymphoblastic leukemia. Leukemia 1995: 9: 863, discloses cytotoxic anti-leukemic activity in a patient by gd+ T cells expressing the V 1 form of the T cell receptor.
Lamb LS, Henslee-Downey PJ, Parrish RS, Godder KT, Thompson J, Lee C, Gee AP. Increased frequency of TCR- + T-cells in disease-free survivors following T-cell depleted partially mismatched bone marrow transplantation for leukemia. Journal of Hematotherapy 5:503, 1996, discloses a higher percentage of survival among patients having an increased proportion of gd+ T cells as compared to patients have a normal number of gd+ T cells.
Lamb, L.S., Gee, A., Hazlett, L, Musk, P., Parrish, R.S., O'Hanlon, T.P., Geier, S., Folk, R.S., Harris, W.G., McPherson, K., Lee, C., Henslee-Downey, P.J. Influence of T cell depletion method on circulating gd+ T cell reconstitution and potential role in the graft-versus-leukemia effect Cytotherapy (in press), discloses that enrichment of a graft with gd+ T cells may contribute to later development of increased gd+ T cells.
Anasetti, C., Amos, D., Beatty, P.G., Appelbaum, F.R. et al. Effect of HLA compatibility on engraftment of bone marrow transplants in patients with leukemia or lymphoma. New Engl. J. Med. 1989; 320: 197, discloses that minor histocompatibility differences can initiate graft-versus-host disease.
Beatty, P.G., Clift, R.A., Mickelson, E.M., Nisperos, B.B. Flournoy, N., Martin, P.J., Sanders, J.E., Stewart, P., Buckner, C.D., Storb, R., Thomas, E.D., Hansen, J. Marrow transplantation from related donors other than HLA-identical siblings. New. Engl. J. Med. 1985; 313: 765, discloses that initial attempts to use non-manipulated marrow from mismatched unrelated donors and partially mismatched related donors have resulted

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