Drug – bio-affecting and body treating compositions – Designated organic active ingredient containing – Carbohydrate doai
Reexamination Certificate
1997-05-01
2001-06-19
Clark, Deborah J. (Department: 1633)
Drug, bio-affecting and body treating compositions
Designated organic active ingredient containing
Carbohydrate doai
C424S900000, C800S008000, C800S011000, C435S320100, C435S235100, C435S375000
Reexamination Certificate
active
06248721
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to the evaluation of vaccines. In particular, the present invention provides humanized animal models for the evaluation of vaccines designed to confer immunity against human pathogens, including vaccines directed against the human immunodeficiency virus. The present invention further relates to HIV vaccines. In particular, the present invention provides attenuated replication-competent HIV vaccines and replication-defective HIV vaccines.
BACKGROUND OF THE INVENTION
The development of effective vaccines for the prevention of human immunodeficiency virus (HIV) infection has been hampered by the lack of suitable in vivo HIV-1 infection animal models. Given the complex interaction of HIV with the immune system, there is no substitute for in vivo models for HIV infection and HIV vaccine evaluation. Existing animal models employed for HIV infection and vaccine development include chimpanzees infected with HIV-1, macaques infected with either SIV or HIV-2, cats infected with feline immunodeficiency virus and scid/scid mice reconstituted with human cells and infected with HIV. None of these models are ideal for the evaluation of HIV vaccines. The replication of HIV in chimpanzees is more restricted than in humans; therefore, vaccines shown to be protective in chimpanzees may fail to protect humans as chimpanzees may be more easy to protect from HIV infection. In addition, the number of chimpanzees available for research is small and therefore, it is difficult to obtain statistically significant experimental results, these animals are costly to maintain and once exposed to HIV these animals must be confined for the rest of their lives at a cost of greater than $100,000.00/chimp [Stott and Almond (1995) Nat. Med. 1:295]. While the supply of macaques is greater than that of chimpanzees, these animals are also costly to maintain. Most importantly, macaques cannot be infected with HIV-1 and thus, infection with HIV-2 or SIV must be used as a surrogate model for HIV-1 infection. Scid/scid mice reconstituted with human lymphocytes (hu-PBL-SCID mice), while permitting infection of the human cells with HIV-1, have failed to provide a model in which HIV vaccines and HIV immunity can be evaluated. This failure is due to the fact that most of the human T lymphocytes in the reconstituted scid/scid mice exhibit activated cell phenotypes soon after reconstitution, and almost all (>99%) human T cells exhibit mature memory phenotypes in a state of reversible anergy [Rizza et al. (1996) J. Virol. 70:7958; Tarry-Lehmann and Saxon (1992) J. Exp. Med. 175:503; Tarry-Lehmann et al. (1995) Immunol. Today 16:529]. Therefore, the lack of sufficient numbers of immature naive T cells after reconstitution renders the hu-PBL-SCID model unsuitable for the evaluation of anti-HIV immunity following administration of a HIV vaccine.
The ultimate system for the study of HIV vaccines is, of course, the human. However, given the fact that HIV infection is almost invariably fatal in humans, ethical considerations limit the use of human volunteers in HIV vaccine evaluation. The art needs animal models in which anti-HIV immunity can be assessed to permit the evaluation of HIV vaccines.
SUMMARY
The present invention provides novel humanized animal models that permit the identification of immune-modulating genes and combinations thereof useful for the treatment of human tumors. In addition, the present invention provides methods of treating subjects having a tumor with one or more immune-modulating genes and provides tumor cell vaccines comprising tumor cells modified to express immune-modulating genes. The novel animals of the present invention provide a means of evaluating vaccines, including cancer vaccines and vaccines directed against human pathogens (e.g., HIV, malaria, Leishmania, etc.). In addition, the invention provides HIV vaccines including live attenuated HIV vaccines and HIV DNA vaccines.
Accordingly, the present invention provides an immunodeficient mouse comprising human T lymphocytes expressing the CD45 antigen wherein at least 5% of the human T lymphocytes expressing the CD45 antigen represent immature naive T lymphocytes. The invention is not limited by the nature of the immunodeficient mouse strain employed. In a preferred embodiment, the immunodeficient mouse is a SCID/beige mouse.
In another preferred embodiment, the immunodeficient mouse comprising human T lymphocytes further comprising human tumor cells. The invention is not limited by the nature of the human tumor cells employed. The human tumor cells may be established tumor cells, primary tumors cells or tumor cells (established or primary) modified to express one or more immune-modulating genes, genes encoding cell cycle regulators and genes encoding inducers of apoptosis.
In another embodiment, the present invention provides a SCID/beige mouse comprising human immune cells. The invention is not limited by the nature of the human immune cells, these cells may be human PBLs, splenocytes, cells isolated from lymph nodes and/or peritoneal lavage. In a preferred embodiment, the SCID/beige mouse comprising human immune cells further comprising human tumor cells. The invention is not limited by the nature of the human tumor cells employed. The human tumor cells may be established tumor cells, primary tumors cells or tumor cells (established or primary) modified to express one or more immune-modulating genes, genes encoding cell cycle regulators and genes encoding inducers of apoptosis. In a preferred embodiment, the tumor cells are derived from central nervous system cells, most preferably glioblastoma cells. In another preferred embodiment, the tumor cells are malignant melanoma cells.
The present invention further provides a method comprising: a) providing: i) a SCID/beige mouse; ii) human tumor cells; iii) human peripheral blood lymphocytes; b) introducing a first dose of the tumor cells into said mouse; c) reconstituting the mouse containing said tumor cells with the lymphocytes; and d) monitoring the reconstituted mouse for the growth of the tumor cells. The invention is not limited by the nature of the human tumor cells employed. The human tumor cells may be established tumor cells, primary tumors cells or tumor cells (established or primary) modified to express one or more immune-modulating genes, genes encoding cell cycle regulators and genes encoding inducers of apoptosis. In a preferred embodiment, the tumor cells are derived from central nervous system cells, most preferably glioblastoma cells. In another preferred embodiment, the tumor cells are malignant melanoma cells.
In a preferred embodiment, the method further comprises identifying at least one immune modulating gene (or gene encoding a cell cycle regulator or inducer of apoptosis) whose expression prevents the growth of the introduced tumor cells in the reconstituted mouse. In another preferred embodiment, the method comprises, following the reconstitution, the additional step of vaccinating the reconstituted mouse with a second dose of tumor cells. In a preferred embodiment, the first dose of tumor cells comprises unmodified tumor cells and the second dose of tumor cells comprises irradiated tumor cells. In a particularly preferred embodiment, the irradiated tumor cells express at least one immune-modulating gene (or gene encoding a cell cycle regulator or inducer of apoptosis).
In one embodiment of the methods of the present invention, the tumor cells and the lymphocytes come from the same donor. In another embodiment, the tumor cells and the lymphocytes come from different donors.
The present invention further provides a method comprising: a) providing: i) a SCID/beige mouse; ii) irradiated and unirradiated human tumor cells; iii) human peripheral blood lymphocytes; b) reconstituting said mouse with the lymphocytes; c) vaccinating the mouse with the irradiated tumor cells; d) introducing the unirradiated tumor cells into the vaccinated mouse; and e) monitoring the vaccinated mouse for the growth of the unir
Clark Deborah J.
Saliwanchik Lloyd & Saliwanchik
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