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
1999-06-10
2001-06-26
Saunders, David (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...
C424S143100, C424S174100, C435S334000, C436S548000, C530S388150, C530S388220, C530S389100
Reexamination Certificate
active
06252050
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
This invention relates generally to a method for making monoclonal antibodies. The invention further pertains to antibodies obtainable by the method which specifically cross-react with two or more different receptors to which Apo-2 ligand (Apo-2L) can bind.
2. Description of Related Art
Native antibodies are synthesized primarily by specialized lymphocytes called “plasma cells”. Production of a strong antibody response in a host animal is controlled by inducing and regulating the differentiation of B cells into these plasma cells. This differentiation involves virgin B cells (which have a modified antibody as a cell-surface antigen receptor and do not secrete antibodies) becoming activated B cells (which both secrete antibodies and have cell-surface antibodies), then plasma cells (which are highly specialized antibody factories with no surface antigen receptors). This differentiation process is influenced by the presence of antigen and by cellular communication between B cells and helper T cells.
Because of their ability to bind selectively to an antigen of interest, antibodies have been used widely for research, diagnostic and therapeutic applications. The potential uses for antibodies were expanded with the development of monoclonal antibodies. In contrast to polyclonal antiserum, which includes a mixture of antibodies directed against different epitopes, monoclonal antibodies are directed against a single determinant or epitope on the antigen and are homogeneous. Moreover, monoclonal antibodies can be produced in unlimited quantities.
The seminal work by Kohler and Milstein described the first method for obtaining hybridomas that can produce monoclonal antibodies [Kohler and Milstein
Nature
256:495 (1975)]. In this method, an antibody-secreting immune cell, isolated from an immunized mouse, is fused with a myeloma cell, a type of B cell tumor. The resultant hybrid cells (i.e. hybridomas) can be maintained in vitro and continue to secrete antibodies with a defined specificity.
Since murine monoclonal antibodies are derived from mice, their use as therapeutic agents in humans is limited because of the human anti-mouse response that occurs upon administration of the murine antibody to a patient. Accordingly, researchers have engineered non-human antibodies to make them appear more human. Such engineered antibodies are called “chimeric” antibodies; in which a non-human antigen-binding domain is coupled to a human constant domain (Cabilly et al., U.S. Pat. No. 4,816,567). The isotype of the human constant domain may be selected to tailor the chimeric antibody for participation in antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity. In a further effort to resolve the antigen binding functions of antibodies and to minimize the use of heterologous sequences in human antibodies, Winter and colleagues [(Jones et al.,
Nature
321:522-525 (1986); Riechmann et al.,
Nature
332:323-327 (1988); Verhoeyen et al.,
Science
239:1534-1536 (1988)] have substituted rodent complementarity determining region (CDR) residues for the corresponding segments of a human antibody to generate humanized antibodies. As used herein, the term “humanized” antibody is an embodiment of chimeric antibodies wherein substantially less than an intact human variable domain has been substituted by the corresponding sequence from a non-human species. In practice, humanized antibodies are typically human antibodies in which CDR residues and possibly some framework region (FR) residues are substituted by residues from analogous sites in rodent antibodies.
Other groups have developed methods for making fully “human” monoclonal antibodies. Such antibodies may be generated by immortalizing a human cell secreting a specific antibody using an Epstein-Barr virus (EBV) [Steinitz et al.
Nature
269:420-422 (1977)]; or by preparing a human—human hybridoma secreting the monoclonal antibody [Olsson et al.
PNAS
(
USA
) 77:5429-5431 (1980)]. Human antibodies can also be derived from phage-display libraries [Hoogenboom et al.,
J. Mol. Biol
., 227:381 (1991); Marks et al.,
J. Mol. Biol
., 222:581-597 (1992); Vaughan et al.
Nature Biotech
14:309 (1996)].
Alternatively, human antibodies have been made in transgenic laboratory animals, in which human immunoglobulin loci have been introduced into the animal and the endogenous immunoglobulin genes are partially or completely inactivated [Fishwild et al.
Nature Biotech
. 14:845-851 (1996); and Mendez et al.
Nature Genetics
15:146-156 (1997)].
SUMMARY OF THE INVENTION
The present invention, in one aspect, provides a method for making monoclonal antibodies wherein an animal is immunized with two or more different antigens and monoclonal antibodies are made and identified which bind to each antigen. Surprisingly, it was discovered herein that sera titers from animals immunized with a mixture of different antigens were similar to those achieved in animals immunized with a single antigen.
This method is thought to be useful for reducing the number of animals that need to be immunized and sacrificed in order to make two or more monoclonal antibodies with differing antigen-binding specificities.
Moreover, it was discovered that the method was useful for making antibodies that cross-reacted with two or more different antigens. For example, antibodies were made which specifically cross-reacted with two or more different Apo-2L receptors.
Accordingly, the invention provides a method for making antibodies comprising the following steps:
(a) immunizing an animal with two or more different antigens so as to generate polyclonal antibodies against each antigen in the animal;
(b) preparing monoclonal antibodies using immune cells of the immunized animal which produce said polyclonal antibodies; and
(c) screening said monoclonal antibodies to identify one or more monoclonal antibodies that bind to each antigen. In the screening step, one finds at least one monoclonal antibody against at least two different antigens. Preferably, at least one monoclonal antibody is found for each antigen with which the animal was immunized.
Preferably, the animal is immunized with a composition comprising a mixture of the two or more different antigens; and step (b) comprises fusing immune cells from the immunized animal with myeloma cells in order to generate hybridoma cell lines producing the monoclonal antibodies.
In one embodiment, the method further comprises identifying one or more monoclonal antibodies that cross-react with two or more of the different antigens.
The invention further provides a monoclonal antibody that has been made according to the above method (e.g. one that cross-reacts with two or more structurally or functionally related antigens).
The invention also relates to an antibody that specifically cross-reacts with two or more different Apo-2L receptors; e.g. which specifically binds to Apo-2 polypeptide and further specifically cross-reacts with another Apo-2L receptor.
The present application further supplies a monoclonal antibody which has the biological characteristics of a monoclonal antibody selected from the group consisting of 3H1.18.10, 3H3.14.5 and 3D5.1.10.
Moreover, the invention provides hybridoma cell lines that produce any of the monoclonal antibodies disclosed herein.
The invention also relates to isolated nucleic acid comprising DNA encoding an antibody as herein disclosed; a vector comprising the nucleic acid; a host cell comprising the vector; a method of producing an antibody comprising culturing the host cell under conditions wherein the DNA is expressed and, optionally, further comprising recovering the antibody from the host cell culture.
The invention further provides a composition comprising an antibody as described herein and a carrier.
In addition, a method of inducing apoptosis in mammalian cancer cells is provided which comprises exposing mammalian cancer cells to an effective amount of a cross-reactive, agonistic
Ashkenazi Avi J.
Chuntharapai Anan
Kim K. Jin
Genentech Inc.
Lee Wendy M.
Saunders David
LandOfFree
Method for making monoclonal antibodies and cross-reactive... does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Method for making monoclonal antibodies and cross-reactive..., we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Method for making monoclonal antibodies and cross-reactive... will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-2535161