Human Monoclonal Antibodies Against CD20

Abstract

Isolated human monoclonal antibodies which bind to and inhibit human CD20, and related antibody-based compositions and molecules, are disclosed. The human antibodies can be produced by a transfectoma or in a non-human transgenic animal, e.g., a transgenic mouse, capable of producing multiple isotypes of human monoclonal antibodies by undergoing V-D-J recombination and isotype switching. Also disclosed are pharmaceutical compositions comprising the human antibodies, non-human transgenic animals and hybridomas which produce the human antibodies, and therapeutic and diagnostic methods for using the human antibodies.

1 . An isolated human monoclonal antibody which binds to human CD20, comprising a heavy chain with a variable region (V H ) comprising CDRs 1-3 and a light chain with a variable region (V L ) comprising CDRs 1-3, wherein:
 (i) the V H  CDR1 amino acid sequence comprises the sequence of SEQ ID NO: 13; the V H  CDR2 amino acid sequence comprises the sequence of SEQ ID NO: 14; the V H  CDR3 amino acid sequence comprises the sequence of SEQ ID NO: 15; the V L  CDR1 amino acid sequence comprises the sequence of SEQ ID NO: 16; the V L  CDR2 amino acid sequence comprises the sequence of SEQ ID NO: 17; and the V L  CDR3 amino acid sequence comprises the sequence of SEQ ID NO: 18,   (ii) the V H  CDR1 amino acid sequence comprises the sequence of SEQ ID NO: 19; the V H  CDR2 amino acid sequence comprises the sequence of SEQ ID NO: 20; the V H  CDR3 amino acid sequence comprises the sequence of SEQ ID NO: 21; the V L  CDR1 amino acid sequence comprises the sequence of SEQ ID NO: 22; the V L  CDR2 amino acid sequence comprises the sequence of SEQ ID NO: 23 and the V L  CDR3 amino acid sequence comprises the sequence of SEQ ID NO: 25, or   (iii) the V H  CDR1 amino acid sequence comprises the sequence of SEQ ID NO: 26; the V H  CDR2 amino acid sequence comprises the sequence of SEQ ID NO: 27; the V H  CDR3 amino acid sequence comprises the sequence of SEQ ID NO: 28; the V L  CDR1 amino acid sequence comprises the sequence of SEQ ID NO: 29; the V L  CDR2 amino acid sequence comprises the sequence of SEQ ID NO: 30 and the V L  CDR3 amino acid sequence comprises the sequence of SEQ ID NO: 31.   
     
     
         2 . The antibody of  claim 1 , selected from the group consisting of an IgG1, an IgG2, an IgG3, an IgG4, an IgM, an IgA1, an IgA2, a secretory IgA, an IgD, and an IgE antibody. 
     
     
         3 . The antibody of  claim 2 , wherein the antibody is an IgG1 antibody. 
     
     
         4 .- 11 . (canceled) 
     
     
         12 . The antibody of  claim 1  encoded by human heavy chain and human kappa light chain nucleic acids comprising nucleotide sequences as set forth in SEQ ID NO:1 and SEQ ID NO:3, respectively. 
     
     
         13 . The antibody of  claim 1  encoded by human heavy chain and human kappa light chain nucleic acids comprising nucleotide sequences as set forth in SEQ ID NO:5 and SEQ ID NO:7, respectively. 
     
     
         14 . The antibody of  claim 1  encoded by human heavy chain and human kappa light chain nucleic acids comprising nucleotide sequences as set forth in SEQ ID NO:9 and SEQ ID NO:11, respectively. 
     
     
         15 . The antibody of  claim 1  comprising human heavy chain and human kappa light chain variable regions amino acid sequences comprising SEQ ID NO:2 and SEQ ID NO:4, respectively. 
     
     
         16 . (canceled) 
     
     
         17 . The antibody of  claim 1  comprising human heavy chain and human kappa light chain variable regions amino acid sequences comprising SEQ ID NO:6 and SEQ ID NO:8, respectively. 
     
     
         18 . (canceled) 
     
     
         19 . The antibody of  claim 1  comprising human heavy chain and human kappa light chain variable regions amino acid sequences comprising SEQ ID NO:10 and SEQ ID NO:12, respectively. 
     
     
         20 .- 21 . (canceled) 
     
     
         22 . An isolated human monoclonal antibody which binds to an epitope on human CD20 defined by the antibody of  claim 1 . 
     
     
         23 . An isolated human monoclonal antibody which binds to an epitope on CD20,
 which epitope does not comprise or require proline at position 172.   
     
     
         24 .- 43 . (canceled) 
     
     
         44 . The antibody of  claim 1  which is an intact antibody selected from an intact IgG1 antibody, an intact IgG2 antibody, an intact IgG3 antibody, an intact IgG4 antibody, an intact IgM antibody, an intact IgA1 antibody, an intact IgA2 antibody, an intact secretory IgA antibody, an intact IgD antibody, and an intact IgE antibody, wherein the antibody is glycosylated when expressed in a eukaryotic cell. 
     
     
         45 . The antibody of  claim 1  which is an antibody fragment or a single chain antibody. 
     
     
         46 .- 52 . (canceled) 
     
     
         53 . A transfectoma which produces a human monoclonal antibody encoded by nucleic acids comprising a human IgG heavy chain nucleotide sequences as set forth in SEQ ID NOs: 1, 5, or 9 and a human kappa light chain nucleotide sequence as set forth in SEQ ID NOs: 3, 7, or 11. 
     
     
         54 . A transfectoma which produces a human monoclonal antibody having IgG heavy chain and kappa light chain variable regions amino acid sequences comprising SEQ ID NOs:2, 6, or 10, and SEQ ID NOs:4, 8, or 12, respectively. 
     
     
         55 . A eukaryotic or prokaryotic host cell which produces a human monoclonal antibody having heavy chain and light chain variable regions amino acid sequences comprising SEQ ID NOs:2, 6, or 10, and SEQ ID Nos:4, 8, or 12, respectively. 
     
     
         56 .- 61 . (canceled) 
     
     
         62 . A composition comprising the human antibody of  claim 1  and a pharmaceutically acceptable carrier. 
     
     
         63 .- 64 . (canceled) 
     
     
         65 . A composition according to  claim 62  further comprising an additional therapeutic agent. 
     
     
         66 . The antibody according to  claim 1 , further comprising a chelator linker for attaching the antibody to a radioisotope. 
     
     
         67 . An immunoconjugate comprising an antibody according to  claim 1  linked to a cytotoxic agent, a radioisotope, or a drug. 
     
     
         68 . A bispecific molecule comprising an antibody according to  claim 1  wherein the bispecific molecule has binding specificity for a human effector cell. 
     
     
         69 . The bispecific molecule according to  claim 68  wherein the bispecific molecule has binding specificity for a human Fc receptor or for a T cell receptor. 
     
     
         70 .- 72 . (canceled) 
     
     
         73 . A method of treating a disease or disorder involving cells expressing CD20, comprising administering to a subject in need thereof a human antibody according to  claim 1 , in an amount effective to treat the disease. 
     
     
         74 . The method of  claim 73 , wherein the disease or disorder is a B cell lymphoma. 
     
     
         75 . The method of  claim 73 , wherein the disease or disorder is B cell non-Hodgkin's lymphoma. 
     
     
         76 . The method of  claim 73 , wherein the disease or disorder is precursor B cell lymphoblastic leukemia/lymphoma or a mature B cell neoplasm. 
     
     
         77 . The method of  claim 73 , wherein the disease or disorder is follicular lymphoma (FL). 
     
     
         78 . The method of  claim 76 , wherein the disease or disorder is B cell chronic lymphocytic leukemia (CLL)/small lymphocytic lymphoma (SLL). 
     
     
         79 . The method of  claim 73 , wherein the disease or disorder is selected from lymphomatoid granulomatosis, primary effusion lymphoma, intravascular large B cell lymphoma, mediastinal large B cell lymphoma, heavy chain and lymphomas induced by therapy with immunosuppressive agents. 
     
     
         80 . A method of treating an immune disease involving CD20 expressing immune cells, comprising administering to a subject in need thereof the antibody of  claim 1 , in an amount effective to treat the immune disease. 
     
     
         81 . The method of  claim 80 , wherein treatment results in killing of B cells which produce antibodies against autoantigens. 
     
     
         82 . The method of  claim 73 , wherein the disease or disorder is selected from psoriasis, psoriatic arthritis, dermatitis, systemic scleroderma and sclerosis, inflammatory bowel disease (IBD), Crohn's disease, ulcerative colitis, respiratory distress syndrome, meningitis, encephalitis, uveitis, glomerulonephritis, eczema, asthma, atherosclerosis, leukocyte adhesion deficiency, multiple sclerosis, Raynaud's syndrome, Sjögren's syndrome, juvenile onset diabetes, Reiter's disease, Behçet's disease, immune complex nephritis, IgA nephropathy, IgM polyneuropathies, immune-mediated thrombocytopenias, hemolytic anemia, myasthenia gravis, lupus nephritis, systemic lupus erythematosus, rheumatoid arthritis (RA), atopic dermatitis, pemphigus, Graves' disease, Hashimoto's thyroiditis, Wegener's granulomatosis, Omenn's syndrome, chronic renal failure, acute infectious mononucleosis, HIV, and herpes virus associated diseases. 
     
     
         83 . The method of  claim 82 , wherein the disease or disorder is rheumatoid arthritis (RA). 
     
     
         84 . The method of  claim 73 , wherein the disease or disorder is an inflammatory, immune and/or autoimmune disorder selected from ulcerative colitis, Crohn's disease, juvenile onset diabetes, multiple sclerosis, immune-mediated thrombocytopenias, hemolytic anemia, myasthenia gravis, systemic sclerosis, and pemphigus vulgaris. 
     
     
         85 . The method of  claim 73 , wherein the disease or disorder is an inflammatory, immune and/or autoimmune disorder selected from inflammatory bowel disease (IBD), ulcerative colitis, Crohn's disease, and multiple sclerosis. 
     
     
         86 . The method of  claim 73 , further comprising separately administering another therapeutic agent to the subject. 
     
     
         87 . The method of  claim 86 , wherein the other therapeutic agent is a cytotoxic agent or a radiotoxic agent. 
     
     
         88 . The method of  claim 86 , wherein the other therapeutic agent is an immunosuppressant. 
     
     
         89 . The method of  claim 86 , wherein the other therapeutic agent is an immunological modulating agent. 
     
     
         90 . The method of  claim 86 , wherein the other therapeutic agent is selected from doxorubicin, cisplatin, bleomycin, carmustine, chlorambucil, and cyclophosphamide. 
     
     
         91 . The method of  claim 86 , wherein the other therapeutic agent is selected from anti-CD25 antibodies, anti-CD 19 antibodies, anti-CD21 antibodies, anti-CD22 antibodies, anti-CD37 antibodies, anti-CD38 antibodies, antilL6R antibodies, anti-IL8 antibodies, anti-IL15 antibodies, anti-IL15R antibodies, antiCD4 antibodies, anti-CD11a antibodies, anti-alpha-4/beta-1 integrin (VLA4) antibodies, CTLA4-Ig, and anti-C3b(i) antibodies. 
     
     
         92 . An in vitro method for detecting the presence of CD20 antigen, or a cell expressing CD20, in a sample comprising:
 contacting the sample with the antibody of  claim 1  under conditions that allow for formation of a complex between the antibody and CD20; and   detecting the formation of a complex.   
     
     
         93 . A kit for detecting the presence of CD20 antigen, or a cell expressing CD20, in a sample, the kit comprising the antibody of  claim 1 . 
     
     
         94 . An in vivo method for detecting CD20 antigen, or a cell expressing CD20, in an subject comprising:
 administering the antibody of  claim 1  under conditions that allow for formation of a complex between the antibody and CD20; and   detecting the formed complex.   
     
     
         95 .- 96 . (canceled) 
     
     
         97 . An expression vector comprising a nucleotide sequence encoding a heavy chain variable region comprising a nucleotide sequence set forth in SEQ ID NO: 1, 5, or 9, and a light variable region comprising a nucleotide sequence as set forth in SEQ ID NO: 3, 7, or 11. 
     
     
         98 . An expression vector comprising a nucleotide sequence encoding a heavy chain variable region comprising an amino acid sequence as set forth in SEQ ID NO:2, 6, or 10, and a light chain variable region comprising an amino acid sequence as set forth in SEQ ID NO: 4, 8, or 12. 
     
     
         99 . A pharmaceutical composition comprising the expression vector of claim  95  and a pharmaceutically acceptable carrier. 
     
     
         100 . An anti-idiotypic antibody binding to an antibody of  claim 1 . 
     
     
         101 . (canceled) 
     
     
         102 . A method for detecting the level of the antibody of  claim 1  in a sample, comprising obtaining a sample potentially comprising the antibody and contacting the sample with an anti-idiotypic antibody to the antibody. 
     
     
         103 . A method for producing a human monoclonal antibody which binds to human CD20 in a host cell, wherein the method comprises:
 (a) transfecting the host cell with an expression vector comprising human IgG heavy chain and human kappa light chain nucleic acids comprising nucleotide sequences as set forth in: (i) SEQ ID NOs: 1 and 3, respectively, (ii) SEQ ID NOs: 5 and 7, respectively, or (iii) SEQ ID NOs: 9 and 11, respectively;   (b) culturing the host cell in culture medium;   (c) expressing the antibody in the host cell; and   (d) isolating and purifying the expressed antibody from the culture supernatant and/or host cell.   
     
     
         104 . The method according to  claim 103 , wherein the host cell is a eukaryotic or prokaryotic cell. 
     
     
         105 . An antibody produced according to the method of  claim 103 .