Chemistry: natural resins or derivatives; peptides or proteins; – Proteins – i.e. – more than 100 amino acid residues – Blood proteins or globulins – e.g. – proteoglycans – platelet...
Reexamination Certificate
1999-11-29
2001-12-11
Nolan, Patrick J. (Department: 1644)
Chemistry: natural resins or derivatives; peptides or proteins;
Proteins, i.e., more than 100 amino acid residues
Blood proteins or globulins, e.g., proteoglycans, platelet...
C530S387100, C530S387300
Reexamination Certificate
active
06329511
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates generally to the combination of recombinant DNA and monoclonal antibody technologies for developing novel biologics and, more particularly, for example, to the production of non-immunogenic (in humans) immunoglobulins specific for gamma-interferon (&ggr;-IFN) and their uses in vitro and in vivo. The present invention also relates more specifically to humanized monoclonal antibodies against &ggr;-IFN, polynucleotide sequences encoding the antibodies, a method of producing the antibodies, pharmaceutical compositions comprising the antibodies as an active ingredient, and therapeutic agents for suppressing undesired immune responses comprising the antibodies as an active ingredient.
BACKGROUND
The mammalian immune response is mediated by several types of cells that interact specifically with foreign material, i.e., antigens. One of these cell types, B cells, is responsible for the production of antibodies. Another cell type, T cells, include a wide variety of cellular subsets that destroy cells infected with virus or control the in vivo function of both B cells and other hematopoietic cells, including T cells. A third cell type, macrophages, process and present antigens in conjunction with major histocompatibility complex (MHC) proteins to T cells. Communication between these cell types is mediated in a complex manner by lymphokines, such as interleukins 1-6 and &ggr;-IFN (see, generally, Paul, W. E., ed.,
Fundamental Immunology
, 3rd ed., Raven Press, New York (1993), which is incorporated herein in relevant part by reference.)
One important lymphokine is &ggr;-IFN, which is secreted by some T cells. In addition to its anti-viral activity, &ggr;-IFN stimulates natural killer (NK) cells and T helper 1 (Th1) cells, activates macrophages, and stimulates the expression of MHC molecules on the surface of cells (Paul, op. cit., pp. 764-766). Hence &ggr;-IFN generally serves to enhance many aspects of immune function, and is a logical candidate for a therapeutic drug in cases where such enhancement is desired, e.g., in treating cancer. Conversely, in disease states where the immune system is over-active, e.g., autoimmune diseases and organ transplant rejection, antagonists of &ggr;-IFN can be useful to treat the disease by neutralizing the stimulatory effects of &ggr;-IFN.
Mouse monoclonal antibodies that bind to and neutralize &ggr;-IFN have been reported (see, e.g., Van der Meide et al.,
J. Gen. Virol
, 67, 1059 (1986)). Such anti-&ggr;-IFN antibodies have been reported to delay or prevent rejection in vitro and in vivo mouse models of transplants, (Landolfo et al.,
Science
229, 176 (1985) and Rosenberg et al.,
J. Immunol
. 144, 4648 (1990)), both of which are incorporated herein by reference). Treatment of mice prone to develop a syndrome like systemic lupus erythematosus (SLE) with a monoclonal antibody to &ggr;-IFN was reported to delay onset of the disease (Jacob et al.,
J. Exp. Med
. 166, 798 (1987)). An anti-&ggr;-IFN antibody has also been reported to alleviate adjuvant arthritis in rats (Jacob et al.,
J. Immunol
. 142, 1500 (1989))and colitis in mice. (Powrie et al.,
Immunity
1, 553-562 (1994)). Queen et al., WO 92/11018 discuss the mouse AF2 antibody to &ggr;-IFN, certain humanized immunoglobulins, and use of the same for treating inflammatory disease.
The use of non-human monoclonal antibodies such as AF2 has certain drawbacks in human treatment, particularly in repeated therapeutic regimens as explained below. Mouse monoclonal antibodies, for example, have a relatively short circulating half-life in humans, and lack other important immunoglobulin functional characteristics when used in humans.
Perhaps more importantly, murine monoclonal antibodies contain substantial amino acid sequences that will be immunogenic when injected into a human patient. Numerous studies have shown that, after injection of a foreign antibody, the immune response elicited by a patient against the injected antibody can be quite strong, essentially eliminating the antibody's therapeutic utility after an initial treatment. Moreover, if mouse or other antigenic (to humans) monoclonal antibodies are used to treat various human diseases, subsequent treatments with unrelated mouse antibodies may be ineffective or even dangerous in themselves, because of cross-reactivity.
Thus, there is a need for improved forms of humanized immunoglobulins specific for &ggr;-IFN antigen that are substantially non-immunogenic in humans, yet easily and economically produced in a manner suitable for therapeutic formulation and other uses. The present invention fulfills these and other needs.
OBJECTS AND SUMMARY OF THE INVENTION
It is the object of the present invention to provide humanized monoclonal antibodies against &ggr;-IFN; polynucleotide sequences encoding the antibodies; a method for producing the antibodies; a pharmaceutical composition comprising the antibodies as an active ingredient; a therapeutic agent for treating diseases, particularly autoimmune diseases, and for immune system suppression comprising the antibody as an active ingredient; and a method for treating such diseases.
The invention provides humanized immunoglobulins that are humanized versions of the mouse AF2 immunoglobulin. The mouse AF2 immunoglobulin is characterized by a light chain variable region designated SEQ ID No:2 and a heavy chain variable region designated SEQ ID No:4. The humanized immunoglobulins of the invention comprise humanized heavy and light chains. Position 11 of the humanized heavy chain variable region framework is occupied by the amino acid present in the equivalent position of the mouse AF2 heavy chain variable region framework. A preferred humanized immunoglobulin of the invention comprises a humanized light chain variable region designated SEQ ID No:6 and a humanized heavy chain variable region designated SEQ ID No:8.
The humanized immunoglobulins specifically bind to the &ggr;-IFN antigen and neutralize &ggr;-IFN. The humanized immunoglobulins are also capable of blocking the binding of the CDR-donating mouse monoclonal antibody to &ggr;-IFN. —IFN. Preferred humanized immunoglobulins have two pairs of light chain/heavy chain complexes, at least one chain comprising one or more mouse complementarity determining regions (CDRs) functionally joined to human framework region segments. For example, mouse CDRs, with or without additional naturally-associated mouse amino acid residues, can be introduced into human framework regions to produce humanized immunoglobulins capable of binding to the antigen at affinity levels stronger than about 10
7
M
−1
.
The immunoglobulins, including binding fragments and other derivatives thereof, of the present invention can be produced readily by a variety of recombinant DNA techniques, with ultimate expression in transfected cells, preferably immortalized eukaryotic cells, such as myeloma or hybridoma cells. Polynucleotides comprising a first sequence coding for humanized immunoglobulin framework regions and a second sequence coding for the desired immunoglobulin CDRs can be produced synthetically or by combining appropriate cDNA and genomic DNA segments.
The humanized immunoglobulins can be utilized in substantially pure form and can be prepared in a pharmaceutically accepted dosage form, which varies depending on the mode of administration.
REFERENCES:
patent: 5530101 (1996-06-01), Queen et al.
patent: 5585089 (1996-12-01), Queen et al.
patent: 5693761 (1997-12-01), Queen et al.
patent: 5693762 (1997-12-01), Queen et al.
Colman, PM Res. Immunology 145:33-36, 1994.
Landolfi Nicholas F.
Queen Cary L.
Tsurushita Naoya
Vasquez Maximiliano
Ewoldt Gerald R.
Nolan Patrick J.
Protein Design Labs, Inc.
Townsend and Townsend / and Crew LLP
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