Single-chain multivalent binding proteins with effector function

Abstract

Multivalent binding peptides, including bi-specific binding peptides, having immunoglobulin effector function are provided, along with encoding nucleic acids, vectors and host cells as well as methods for making such peptides and methods for using such peptides to treat or prevent a variety of diseases, disorders or conditions, as well as to ameliorate at least one symptom associated with such a disease, disorder or condition.

1 - 81 . (canceled) 
     
     
         82 . A single-chain protein comprising from amino to carboxy terminus:
 (a) a first binding domain derived from an immunoglobulin-like molecule or the variable regions of an immunoglobulin;   (b) a Fc region, wherein said Fc region does not comprise a domain derived from an immunoglobulin C H1  domain;   (c) a linker peptide of at least 5 amino acids; and   (d) a second binding domain derived from an immunoglobulin-like molecule or the variable regions of an immunoglobulin, wherein the first or second binding domain binds CD3.   
     
     
         83 . The protein of  claim 82 , wherein the first and/or second binding domain is a single-chain variable antibody fragment (scFv). 
     
     
         84 . The protein of  claim 82 , wherein the first and/or second binding domain comprises chimeric, humanized, or human immunoglobulin variable regions. 
     
     
         85 . The protein of  claim 82 , wherein the first and/or second binding domain comprises a light chain immunoglobulin variable region (VL1) and a heavy chain immunoglobulin variable region (VH1), wherein said variable regions are positioned in a VH1-VL1 or a VL1-VH1 orientation. 
     
     
         86 . The protein of  claim 82 , wherein the first or second binding domain comprises variable regions derived from the G19-4 antibody. 
     
     
         87 . The protein of  claim 86 , wherein the first or second binding domain comprises the amino acid sequence of SEQ ID NO: 107 or SEQ ID NO: 109. 
     
     
         88 . The protein of  claim 82 , wherein the immunoglobulin-like molecule is a receptor. 
     
     
         89 . The protein of  claim 82 , wherein the other binding domain binds to a target selected from the group consisting of a tumor antigen, a B-cell target, a TNF receptor superfamily member, a Hedgehog family member, a receptor tyrosine kinase, a proteoglycan-related molecule, a TGF-beta superfamily member, a Wnt-related molecule, a receptor ligand, a T-cell target, a Dendritic cell target, an NK cell target, a monocyte/macrophage cell target and an angiogenesis target. 
     
     
         90 . The protein of  claim 89 , wherein the tumor antigen is selected from the group consisting of SQUAMOUS CELL CARCINOMA ANTIGEN 1, SQUAMOUS CELL CARCINOMA ANTIGEN 2, Ovarian carcinoma antigen CA125, MUCIN 1, CTCL tumor antigen se1-1, CTCL tumor antigen se1-4-3, CTCL tumor antigen se20-4, CTCL tumor antigen se20-9, CTCL tumor antigen se33-1, CTCL tumor antigen se37-2, CTCL tumor antigen se57-1, CTCL tumor antigen se89-1, Prostate-specific membrane antigen, 5T4 oncofetal trophoblast glycoprotein, Orf73 Kaposi's sarcoma-associated herpesvirus, MAGE-C1, MAGE-B1 ANTIGEN, MAGE-B2 ANTIGEN, MAGE-2 ANTIGEN, MAGE-4a antigen, MAGE-4b antigen, Colon cancer antigen NY-CO-45, Lung cancer antigen NY-LU-12 variant A, Cancer associated surface antigen, Adenocarcinoma antigen ART1, Paraneoplastic associated brain-testis-cancer antigen, Neuro-oncological ventral antigen 2, Hepatocellular carcinoma antigen gene 520, TUMOR-ASSOCIATED ANTIGEN CO-029, Tumor-associated antigen MAGE-X2, Synovial sarcoma X breakpoint 2, Squamous cell carcinoma antigen recognized by T cell, Serologically defined colon cancer antigen 1, Serologically defined breast cancer antigen NY-BR-15, Serologically defined breast cancer antigen NY-BR-16, Chromogranin A; parathyroid secretory protein 1, DUPAN-2, CA 19-9, CA 72-4, CA 195 and L6. 
     
     
         91 . The protein of  claim 82 , wherein the Fc region further comprises an immunoglobulin hinge region. 
     
     
         92 . The protein of  claim 91 , wherein the hinge region is a hinge region selected from the group consisting of IgG1, IgG2, IgG3, IgG4, IgE, IgA2, synthetic hinge and the hinge-like C H2  domain of IgM. 
     
     
         93 . The protein of  claim 92 , wherein the hinge region is an IgG1 hinge region. 
     
     
         94 . The protein of  claim 93 , wherein the hinge region is a human IgG1 hinge region with a mutation at one or two cysteine residues. 
     
     
         95 . The protein of  claim 82 , wherein the Fc region comprises a domain derived from an immunoglobulin C H2  and C H3  domain. 
     
     
         96 . The protein of  claim 82 , wherein the Fc region comprises a human immunoglobulin hinge region, C H2  domain, and C H3  domain. 
     
     
         97 . The protein of  claim 82 , wherein the linker peptide is between 5 and 45 amino acids in length. 
     
     
         98 . The protein of  claim 82 , wherein the linker peptide is derived from an immunoglobulin hinge region. 
     
     
         99 . The protein of  claim 82 , wherein the linker peptide is derived from a stalk region of a C-type lectin. 
     
     
         100 . The protein of  claim 99 , wherein the C-type lectin is selected from the group consisting of CD69, CD72, CD94, NKG2A, and NKG2D. 
     
     
         101 . The protein of  claim 82 , wherein the linker peptide comprises an amino acid sequence selected from the group consisting of SEQ ID NO:11, 113, 115, 117, 119, 121, 123, 125, 127, 129, 131, 133, 135, 149, 151, 153, 155, 157, 159, 161, 163, 165, 167, 169, 231, 233, 235, 237, 239, 241, 243, 245, 247, 249, 251, 253, 255, 257, 259, 261, 263, 265, 267, 269, 271, 273, 275, 277, 279, 281, 287, 289, 297, 305, 307, 309, 310, 311, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331, 346, 373, 374, 375, 376, 377, 380, or 381. 
     
     
         102 . The protein of  claim 82 , wherein the linker peptide is derived from an interdomain region between the Ig V-like and Ig C-like region of CD2, CD4, CD22, CD33, CD48, CD58, CD66, CD80, CD86, CD150, CD166, or CD244. 
     
     
         103 . The protein of  claim 82 , wherein the protein is capable of forming dimers. 
     
     
         104 . The protein of  claim 82 , wherein the other binding domain binds to a target on a cell of an infectious organism. 
     
     
         105 . The protein of  claim 104 , wherein the infectious organism is a bacterium, mycobacterium, fungus, or parasite. 
     
     
         106 . The protein of  claim 82 , wherein the other binding domain binds to a virus. 
     
     
         107 . A method for treating, preventing, or mitigating an infection associated with an infectious organism or virus in a subject in need thereof comprising administering to the subject a therapeutically effective amount of the protein of  claim 82 , wherein the other binding domain binds to the virus or a target on a cell of the infectious organism. 
     
     
         108 . A nucleic acid encoding the protein of  claim 82 . 
     
     
         109 . A vector comprising the nucleic acid of  claim 108 . 
     
     
         110 . A host cell comprising the vector of  claim 109 . 
     
     
         111 . A composition comprising the protein of  claim 82  and a pharmaceutically acceptable carrier. 
     
     
         112 . A method of inducing damage to a target cell comprising contacting the target cell with the protein of  claim 82 . 
     
     
         113 . A method of treating cancer, an autoimmune disease, or inflammation in a subject in need thereof comprising administering to the subject a therapeutically effective amount of the protein of  claim 82 . 
     
     
         114 . The protein of  claim 82 , wherein the Fc region consists essentially of a human immunoglobulin hinge region, C H2  domain, and C H3  domain.