Chemistry: molecular biology and microbiology – Animal cell – per se ; composition thereof; process of... – Fused or hybrid cell – per se
Reexamination Certificate
2000-05-23
2002-05-21
Stucker, Jeffrey (Department: 1648)
Chemistry: molecular biology and microbiology
Animal cell, per se ; composition thereof; process of...
Fused or hybrid cell, per se
C435S325000, C435S326000, C435S328000, C435S329000, C435S337000, C435S339100, C530S387500, C530S388100, C530S388150, C530S388250, C530S388350
Reexamination Certificate
active
06391635
ABSTRACT:
BACKGROUND OF THE INVENTION
The effector molecules of the immune system include a repertoire of circulating immunoglobulins non-attributable to exogenous antigenic induction, variously referred to as “autoantibodies” or “natural antibodies”. The two terms are not synonymous. Thus, for the “self-attacking” antibodies the term “autoantibodies” is customarily applied, while for the “self-protective” antibodies the term “natural antibodies” is used.
A vast majority of natural antibodies react with one or more “self” antigens. Their importance in immune regulation has long been neglected, since tolerance to “self” was thought to be primarily dependent on the deletion of autoreactive clones, rather than on peripheral suppressive mechanisms. Clonal deletion cannot account, however, for the prevalence of natural autoreactivity among healthy individuals. It is now well established that autoreactive repertoires are predominantly selected early in ontogeny and that autoreactive antibodies, B cells, and T cells, are present in healthy individuals, and in virtually all vertebrate species (Lactoix-Desmazes et al., 1998, J. Immunol. Methods, 216:117-137 and references therein).
Natural antibodies are mostly IgM, polyreactive, and are generally encoded by V genes in germline configuration (Casali et al., 1996, Curr. Top. Microbiol. Immunol., 210:167-79 and references therein). They are mainly produced by B-1 cells which account for most of the B cell repertoire in the fetus and neonate, and possibly play a major role in the development of the adult B cell repertoire.
It is still unclear whether precursors of B-1 cells are capable of undergoing an antigen-driven clonal selection process, thereby producing natural antibodies with a high affinity for the selecting antigen. In this respect, it has been clearly established that B-1 cells can express a hypermutation mechanism similar to that of conventional (B-2) cells and that the main structural correlate for antibody polyreactivity and antigen binding in monoreactive antibodies is provided by the somatically generated CDR3 heavy chain (Casali et al., supra).
Although endowed with self-reactivity, natural antibodies also bind exogenous antigens. Exposure to environmental antigens is not necessary for the emergence of natural antibody-producing cell precursor clones to exogenous antigens, as suggested by the significant population of B cells capable of producing antibodies to a variety of bacterial antigens in germ-free animals (Casali et al., supra). Because of their ability to bind a variety of exogenous antigens, including those on bacteria and viruses, natural antibodies play a major role in the primary line of defense against infections.
U.S. Pat. No. 5,872,012 discloses a circulating natural human antibody immunoreactive with an arginine-rich epitope present on human protamine. U.S. Pat. No. 5,606,026 discloses that the arginine-rich epitope is present in the Tat protein of HIV-1 and further discloses a second circulating human natural antibody immunoreactive with a different epitope on the Tat protein. It has been also shown that these Tat-reactive circulating human natural antibodies decrease after HIV infection reaching minimal levels as the patient progresses to AIDS (Rodman et al., 1999, Hum. Immunol., 60:631-639). In addition, a novel circulating human natural antibody immunoreactive with a cryptic epitope present on human lactoferrin is disclosed therein.
As the correlation of the titers of some of the circulating natural antibodies with disease progression has been established in HIV infection, there is a need in the art to develop new treatment strategies based on supplementing the patient's immune system with effective amounts of exogenously produced natural antibodies. An ideal source of such natural antibodies would be monoclonal counterparts of the circulating human natural antibodies that can be produced in large quantities and used for various therapeutic and diagnostic purposes.
SUMMARY OF THE INVENTION
The present invention provides monoclonal forms of human natural antibodies.
In one aspect, the present invention provides a method for producing human hybridoma cells producing monoclonal human natural antibodies comprising the steps of
fusing immortalized or transformed human umbillicord blood cells with mouse: human
heteromyeloma cells,
isolating fused cells,
plating said fused cells under limited dilution conditions, and
recovering said hybridoma cells.
In another aspect, the present invention provides human hybridoma cells producing monoclonal human natural antibodies produced by the method of the present invention.
These and other aspects of the present invention will be apparent to those of ordinary skill in the art in light of the present description, claims and drawings.
REFERENCES:
patent: 4997764 (1991-03-01), Dalla Favera
patent: 5606026 (1997-02-01), Rodman
patent: 5656272 (1997-08-01), Le et al.
Rodman et al. “Epitopes for Natural Antibodies of Human Immunodeficiency Virus (HIV)-Negative (Normal) and HIV-Positive Sera are Coincident with Two Key Functional Sequences of HIV Tat Protein” Proceedings of the National Academy of Sciences of the United States of America, vol. 90, No. 16 (1993), pp. 7719-7723. Q11.N26.*
Jahn et al. “Human hybridomas derived from CD5+ B lymphocytes of patients with chronic lymphocytic leukemia (B-CLL) produce multi-specific nature IGM (kappa) antibodies” Clin. Exp. Immunol., vol. 83(1991), pp. 413-417. RC583.C54.*
Manchester et al., Lactoferrin-Reactive Natural Antibodies, Annals New York Acad. of Sciences, 815:475 (1997).
Lachgar et al., Repair of the in Vitro HIV-1-Induced Immunosuppression and Blockade of the Generation of Functional Suppressive CD8 Cells by Anti-Alpha Interferon and ANTIT-TAT Antibodies, Biomed & Pharmacother. 50:13-18 (1996).
Brocke et al., Treatment of Experimental Encephalomyelitis with a Peptide Analogue of Myelin Basic Protein, Nature 379:343-46 (1996).
Re et al., Effect of Antibody to HIV-1 TAT Protein on Viral Replication in Vitro and Progression of HIV-1 Disease in Vivo, J. Acq. Imm. Def. Syndromes and Human Retrovir. 10:408-416 (1995).
Friedman et al., Predicting Molecular Interactions and Inducible Complementarity: Fragment Docking of Fab-Peptide Complexes, Proteins: Structure, Function and Genetics 20:15-24 (1994).
Coffman et al., Mechanism and Regulation of Immunoglobulin Isotype Switching, Advances in Immuno. 54:229-70 (1993).
Rodman et al., Human Immunodeficiency Virus (HIV) TAT-Reactive Antibodies Present in Normal HIV-Negative Sera and Depleted in HIV-Positive Sera. Identification of the Epitope, J. Exp. Med. 175:1247-53 (1992).
Varela et al., Population Dynamics of Natural Antibodies in Normal and Autoimmune Individuals, Proc. Natl. Acad. Sci. USA 88:5917-21 (1991).
Avrameas, Natural Autoantibodies: from ‘Horror Autotoxicus’ to ‘GNOTHI SEAUTON’, Goday 12:154-160 (1991).
Urlacher et al., IgM Anti-Idiotypes that Block Anti-HLA Antibodies: Naturally Occurring or Immune Antibodies?, Clin. Exp. Immunol. 83:116-120 (1991).
Rodman et al., Identification of a Low-Affinity Subset of Protamine-Reactive IgM Antibodies Prsent in Normal, Deficient in AIDS, SERA: Implications of HIV Latency, Cl. Immun. and Immunopath. 57:430-440 (1990).
Posner et al., The Construction and Use of a Human-Mouse Myeloma Analogue Suitable for the Routine Production of Hybridomas Secreting Human Monoclonal Antibodies, Hybridoma 6:611-625 (1987).
Muñoz et al., New Experimental Criteria for Optimization of Solid-Phase Antigen Concentration and Stability in ELISA, J. Immuno. Methods 94:137-144 (1986).
Rodman et al., Naturally Occurring Antibodies Reactive with Sperm Proteins: Apparent Deficiency in AIDS SERA, Science 228:1211-15 (1985).
Rodman et al., p15, A Nuclear-Associated Protein of Human Sperm: Identification of Four Variants and Their Occurrence in Normal and Abnormal Seminal Cells, Gamete Research 8:129-47 (1983).
Goodman et al., Immunological Identification of Lactoferrin as a Shared Antigen on Radioiodinated Human Sperm Surface and in Radioiodinated Human Semi
Darby & Darby
Institute for Human Genetics and Biochemistry
Stucker Jeffrey
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