Chemistry: molecular biology and microbiology – Animal cell – per se ; composition thereof; process of... – Method of regulating cell metabolism or physiology
Reexamination Certificate
2000-06-05
2002-11-12
Ketter, James (Department: 1632)
Chemistry: molecular biology and microbiology
Animal cell, per se ; composition thereof; process of...
Method of regulating cell metabolism or physiology
C435S325000, C435S375000, C435S455000, C424S093100, C424S093400, C424S093710
Reexamination Certificate
active
06479286
ABSTRACT:
FIELD OF THE INVENTION
The invention relates to generation of dendritic cells from expanded populations of monocytes. These dendritic cells are potent antigen presenting cells which can mediate a variety of T cell responses. The invention relates to the fields of immunology, molecular biology, and medicine.
BACKGROUND OF THE INVENTION
T cells mediate most forms of cellular immunity, including cell lympholysis, delayed type hypersensitivity (DTH), transplantation rejection, and allograft rejection. An introduction to T cells and cell mediated immunity is found in Paul (1993)
Fundamental Immunology, Third Edition
Raven Press, New York, N.Y. and the references cited therein.
Typical T cells do not respond to free antigenic peptides. Some T cells interact with a specialized set of cell surface proteins (the class I and class II major histocompatibility complexes, or MHC) which present antigens on the surface of cells (T cells also recognize antigens in the context of other molecules). Cytotoxic and helper T cells are induced to proliferate by specialized antigen presenting cells, such as macrophage and dendritic cells, which present antigenic epitopes, such as peptides or carbohydrates, on their cellular surfaces in conjunction with MHC molecules. T cells are induced by these antigen presenting cells to recognize corresponding antigens expressed, e.g., on MHC antigens on the surface of target cells. T cells destroy these target cells, or induce other cells to destroy these target cells.
Certain T cells can recognize the antigen in the form of a polypeptide fragment bound to the MHC class I molecules on target cells, rather than the intact polypeptide itself. The polypeptide is endogenously synthesized by the cell, and a portion of the polypeptide is degraded into small peptide fragments in the cytoplasm. Some of these small peptides translocate into a pre-Golgi compartment and interact with class I heavy chains to facilitate proper folding and association with the subunit &bgr;2 microglobulin. The peptide-MHC class I complex is then routed to the cell surface for expression and potential recognition by specific T cells. Investigations of the crystal structure of the human MHC class I molecule HLA-A2.1 indicate that a peptide binding groove is created by the folding of the &agr;1 and &agr;2 domains of the class I heavy chain (Bjorkman et al., (1987)
Nature
329:506). Falk et al., (1991)
Nature
351:290 have developed an approach to characterize naturally processed peptides bound to class I molecules. Other investigators have successfully achieved direct amino acid sequencing of the more abundant antigenic peptides in various HPLC fractions by conventional automated sequencing of peptides eluted from class I molecules (Jardetzky, el al. (1991)
Nature
353:326 and mass spectrometry Hunt, et al.,
Science
225:1261 (1992). A review of the characterization of naturally processed peptides in MHC Class I is found in Rötzschke and Falk (1991)
Immunol. Today
12:447.
Target T cells recognizing antigenic peptides can be induced to differentiate and proliferate in response, for example, to antigen presenting cells bearing antigenic peptides in the context of MHC class I and class II complexes. There are differences in the antigenic peptides bound to MHC class I and class II molecules, but the two classes of bound peptides share common epitopes within the same protein which enable a T cell activated by an antigen presenting cell to recognize a corresponding epitope in the context of MHC class I or II, or other cell surface molecules. MHC class I molecules on target cells typically bind 9 amino acid antigenic peptides, while corresponding MHC class II-peptide complexes have greater heterogeneity in the size of the bound antigenic peptide.
Dendritic cells are the most potent antigen presenting cells known, being capable of activating T cells, NK cells and other immune cells by presentation of peptides and carbohydrate antigens on the MHC class I and class II molecules on the surface of the cells. An extensive review of the origin, maturation and antigen presenting function of dendritic cells in reviewed in Banchereau and Schmitt (1995)
Dendritic Cells In Fundamental and Clinical Immunology Volume
2, in
Advances in Experimental Medicine and Biology
(Back et al. eds), volume 378 Plenum Press, NY. A short review is found in Cella et al. (1997)
Current Opinion in Immunology
9:10-16, and the references cited therein. See also, Hart (1997) Blood 90:3245 (1997); J. Banchereau and R. M. Steinman (1998)
Nature
392: 245; Schuler et al. (1997)
Int Arch Allergy Immunol
112:317-322; Rescigno et al. (1997)
Journal of Leukocyte Biology
61:415-421, Clark (1997)
J. Exp. Med
. 185(3) 801-803, Sprent (1995)
Current Biology
5(10): 1095-1097; Nair et al. (1995)
International Immunology
7(4):679-688; Caux et al. (1995)
immunology Today
16(1):2-4; Liu et al. (1996)
International Review of Cytology
166:139-179, and O'Doherty et al. (1993)
J.Exp. Med
. 178:1067-1078, and the references cited in each article.
“Immature” and “mature” phenotypic subsets of dendritic cells have been characterized and methods for the isolation and/or generation of DCs have been described, including various conditions for generating “immature” and “mature” DC subsets. See, e.g., Steinman (1991)
Ann Rev Immunol
9:271; Steinman et al. (1993)
Adv Exp Med Biol
329:1; Schuler et al. (1997)
Int Arch Allergy Immunol
112:317; Jaffe (1993)
Pediatric Pathology
13:821. Peters, et al., (1996)
Immunology Today
276:273; Herbst, et al. (1996)
Blood
88:2541. Romani et al. (1996)
J Immunol Methods
196:137. Cella et al. (1997)
Current Opinion Immunology
9: 10. Morse, et al. (1997)
Annals of Surgery
226:6; Santiago-Schwartz, et al. (1992)
J Leuk Biol
52:274; Zhou and Tedder (1996)
Proc Natl Acad Sci USA
. 93:2588; C. Caux, et al. (1996)
J Exp Med
184;695; C. Caux, et al. (1997) Blood 90:14589; Winzler, et al., (1997)
J Exp Med
185:317. Sallusto and Lanzavecchia (1994)
J Exp Med
179:1109.
A general introduction to the use of dendritic cells for immunotherapy is provided by Girolomoni et al. (1997) in
Immunology Today
. In addition to presenting antigens to T-cells and NK cells, dendritic cells stimulate T cell mitogenesis, e.g., by producing the T cell mitogen IL-12. See, e.g., Jonuleit et al. (1997)
Journal of Immunol
. 2610-2614.
Despite the clear value of dendritic cells for immunotherapy, problems remain in using dendritic cells for therapeutic applications. Primarily, dendritic cells are very rare in peripheral blood, making isolation of sufficient numbers of such cells for therapeutic applications impractical. For example, autologous therapies in which dendritic cells are isolated from a patient and loaded with a particular peptide or carbohydrate antigen for T cell activation are impractical in the absence of large numbers of dendritic cells. Accordingly, there exists a need in the art for a method of making dendritic cells which are capable of T cell activation, particularly in the context of autologous therapeutic approaches. This invention solves these and other problems.
SUMMARY OF THE INVENTION
The present invention derives, in part, from the surprising discovery that IL-3 cultured expanded populations of monocytes are suitable for in vitro differentiation into dendritic cells. Thus, the present invention overcomes problems of the prior art by providing an easy method of generating large numbers of dendritic cells, i.e., from cultured monocytes. This, in turn facilitates the use of dendritic cells to generate cell-mediated immune responses.
Accordingly, in one embodiment, the invention provides methods of differentiating monocytes into dendritic cells. In the methods, monocytes are incubated in the presence of IL-3, causing the monocytes to proliferate, yielding an expanded population of monocytes. The expanded population of monocytes is differentiated into dendritic cells, e.g., by culturing the expanded population of cells with GM-CSF and IL-4 (to produce baseline or Type I DCs) and, o
Nelson Edward L.
Strobl Susan L
Ketter James
Li Q Janice
The United States of America as represented by the Secretary of
Townsend and Townsend / and Crew LLP
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