Multimeric il-15-based molecules

Abstract

The invention features multi-specific protein complexes with one domain comprising IL-15 or a functional variant and a binding domain specific to a disease antigen, immune checkpoint or signaling molecule.

1 . An isolated soluble fusion protein complex comprising at least two soluble proteins, wherein
 a first soluble protein comprises an interleukin-15 (IL-15) polypeptide domain and a second soluble protein comprises a soluble IL-15 receptor alpha sushi-binding domain (IL-15RαSu) fused to an immunoglobulin Fc domain,   wherein one of the first or second soluble protein further comprises a binding domain that specifically binds to a disease antigen, immune checkpoint molecule or immune signaling molecule, and   wherein the IL-15 domain of the first soluble protein binds to the IL-15RαSu domain of the second soluble protein to form a soluble fusion protein complex.   
     
     
         2 . The soluble fusion protein complex of  claim 1 , wherein one of the first or second soluble protein further comprises a second binding domain that specifically binds to a disease antigen, immune checkpoint molecule, or immune signaling molecule. 
     
     
         3 . The soluble fusion protein complex of  claim 1 , wherein the IL-15 polypeptide is an IL-15 variant comprising an N72D mutation (IL-15N72D). 
     
     
         4 . The soluble fusion protein complex of  claim 1 , wherein the binding domain comprises an immunoglobulin light chain variable domain covalently linked to an immunoglobulin heavy chain variable domain by a polypeptide linker sequence. 
     
     
         5 . The soluble fusion protein complex of  claim 1 , wherein the binding domain specifically binds to one or more molecules comprising: programmed death ligand 1 (PD-L1), programmed death 1 (PD-1), cytotoxic T-lymphocyte associated protein 4 (CTLA-4), cluster of differentiation 33 (CD33), cluster of differentiation 47 (CD47), glucocorticoid-induced tumor necrosis factor receptor (TNFR) family related gene (GITR), lymphocyte function-associated antigen 1 (LFA-1), tissue factor (TF), delta-like protein 4 (DLL4), single strand DNA or T-cell immunoglobulin and mucin-domain containing-3 (Tim-3). 
     
     
         6 . The soluble fusion protein complex of  claim 1 , wherein the first soluble protein comprises the amino acid sequence set forth in one of SEQ ID NOs: 2, 6, 10, 18, 20, 24, 28, 32, or 38. 
     
     
         7 . The soluble fusion protein complex of  claim 1 , wherein the second soluble protein comprises the amino acid sequence set forth in one of SEQ ID NOs: 4, 8, 12, 14, 16, 22, 26, 30, 34, 36, 40, 42, 44, 46, 51, 52, 53, or 54. 
     
     
         8 . A soluble fusion protein complex comprising a first soluble fusion protein complex wherein the first soluble protein complex comprises an interleukin-15 (IL-15) polypeptide domain covalently linked to a second soluble fusion protein complex wherein the second soluble protein complex comprises a soluble IL-15 receptor alpha sushi-binding domain (IL-15RαSu). 
     
     
         9 . The soluble fusion protein complex of  claim 8 , wherein the first soluble fusion protein complex is covalently linked to the second soluble fusion protein complex by a disulfide bond linking the Fc domain of the first soluble fusion protein complex to the Fc domain of the second soluble fusion protein complex. 
     
     
         10 . A nucleic acid sequence encoding a first soluble protein, wherein said nucleic acid sequence comprises the sequence set forth in one of SEQ ID NOs: 1, 5, 9, 17, 19, 23, 27, 31 or 37. 
     
     
         11 . The nucleic acid sequence of  claim 10 , wherein the nucleic acid sequence further comprises a promoter, translation initiation signal, and leader sequence operably linked to the sequence encoding the soluble protein. 
     
     
         12 . A nucleic acid sequence encoding a second soluble protein, wherein said nucleic acid sequence comprises the sequence set forth in one of SEQ ID NOs: 3, 7, 11, 13, 15, 21, 25, 29, 33, 35, 39, 41, 43, 45, 47, 48, 49, or 50. 
     
     
         13 . The nucleic acid sequence of  claim 12 , wherein the nucleic acid sequence further comprises a promoter, translation initiation signal, and leader sequence operably linked to the sequence encoding the soluble protein. 
     
     
         14 . A DNA vector comprising the nucleic acid sequences set forth in one of SEQ ID NOs: 1, 5, 9, 17, 19, 23, 27, 31 or 37 and/or nucleic acid sequences set forth in one of SEQ ID NOs: 3, 7, 11, 13, 15, 21, 25, 29, 33, 35, 39, 41, 43, 45, 47, 48, 49, or 50, wherein the nucleic acid sequences further comprise a promoter, translation initiation signal, and leader sequence operably linked to the sequence encoding the soluble protein. 
     
     
         15 . The soluble fusion protein complex of  claim 1 , wherein the disease antigen is associated with neoplasia, infectious disease, or autoimmune disease. 
     
     
         16 . A method for killing a target cell, the method comprising:
 a) contacting a plurality of cells with a soluble fusion protein complex the soluble fusion protein complex comprising at least two soluble proteins, wherein   
       the first soluble protein comprises an interleukin-15 (IL-15) polypeptide domain and the second soluble protein comprises a soluble IL-15 receptor alpha sushi-binding domain (IL-15RαSu) fused to an immunoglobulin Fc domain, 
       wherein one of the first or second soluble protein further comprises a binding domain that specifically binds to a disease antigen, immune checkpoint molecule or immune signaling molecule, and 
       wherein the IL-15 domain of the first soluble protein binds to the IL-15RαSu domain of the second soluble protein to form a soluble fusion protein complex, wherein the plurality of cells further comprises immune cells bearing the IL-15 receptor (IL-15R) chains recognized by the IL-15 domain of the soluble fusion protein complex, or immune cells bearing a disease antigen, checkpoint or signaling molecules recognized by the binding domains of the soluble fusion protein complex, and the target cells;
 b) activating the immune cells via the IL-15R or signaling molecules or via blockade of the checkpoint molecules; and 
 c) killing the target cells by the activated immune cells; or, 
 a1) contacting a plurality of cells with a soluble fusion protein complex, the soluble fusion protein complex comprising at least two soluble proteins, wherein 
 
       the first soluble protein comprises an interleukin-15 (IL-15) polypeptide domain and the second soluble protein comprises a soluble IL-15 receptor alpha sushi-binding domain (IL-15RαSu) fused to an immunoglobulin Fc domain, 
       wherein one of the first or second soluble protein further comprises a binding domain that specifically binds to a disease antigen, immune checkpoint molecule or immune signaling molecule, and 
       wherein the IL-15 domain of the first soluble protein binds to the IL-15RαSu domain of the second soluble protein to form a soluble fusion protein complex, wherein the plurality of cells further comprises immune cells bearing Fc receptor chains recognized by the Fc domain of the soluble fusion protein complex and the target cells bearing an antigen recognized by the antigen-specific binding domain of the soluble fusion protein complex,
 b1) forming a specific binding complex (bridge) between the antigen on the target cells and the Fc receptor chains on the immune cells sufficient to bind and activate the immune cells; and 
 c1) killing the target cells by the bound activated immune cells. 
 
     
     
         17 . (canceled) 
     
     
         18 . The method of  claim 16 , wherein the target cells are tumor cells, infected cells or autoimmune disease associated cells. 
     
     
         19 . The method of  claim 16  or  17 , wherein binding domain comprises an anti-PD-L1 antibody. 
     
     
         20 . (canceled) 
     
     
         21 . A method for treating a neoplasia, infectious disease or autoimmune disease in a subject in need thereof comprising administering to said subject an effective amount of a pharmaceutical composition comprising a soluble fusion protein complex, the soluble fusion protein complex comprising at least two soluble proteins, wherein
 the first soluble protein comprises an interleukin-15 (IL-15) polypeptide domain and the second soluble protein comprises a soluble IL-15 receptor alpha sushi-binding domain (IL-15RαSu) fused to an immunoglobulin Fc domain,   
       wherein one of the first or second soluble protein further comprises a binding domain that specifically binds to a disease antigen, immune checkpoint molecule or immune signaling molecule, and
 wherein the IL-15 domain of the first soluble protein binds to the IL-15RαSu domain of the second soluble protein to form a soluble fusion protein complex, thereby treating said neoplasia, infectious disease or autoimmune disease. 
 
     
     
         22 . The method of  claim 21 , wherein said neoplasia is selected from the group consisting of a glioblastoma, prostate cancer, hematological cancer, B-cell neoplasms, multiple myeloma, B-cell lymphoma, B cell non-Hodgkin lymphoma, Hodgkin's lymphoma, chronic lymphocytic leukemia, acute myeloid leukemia, cutaneous T-cell lymphoma, T-cell lymphoma, a solid tumor, urothelial/bladder carcinoma, melanoma, lung cancer, renal cell carcinoma, breast cancer, gastric and esophageal cancer, prostate cancer, pancreatic cancer, colorectal cancer, ovarian cancer, non-small cell lung carcinoma, and squamous cell head and neck carcinoma. 
     
     
         23 . The method of  claim 21 , wherein said effective amount is between about 1 and 100 μg/kg said fusion protein complex. 
     
     
         24 . The method of  claim 21 , wherein said fusion protein complex is administered at least one time per week. 
     
     
         25 . The method of  claim 21 , wherein said pharmaceutical composition is administered systemically, locally, intravenously, subcutaneously or intratumorally. 
     
     
         26 . The method of  claim 21 , wherein said fusion protein complex induces an immune response in the subject. 
     
     
         27 . The method of  claim 21 , wherein said fusion protein complex increases immune cell proliferation. 
     
     
         28 . The method of  claim 21 , wherein said fusion protein complex stimulates immune cell responses against cells associated with said neoplasia, infectious disease or autoimmune disease. 
     
     
         29 . (canceled) 
     
     
         30 . (canceled) 
     
     
         31 . (canceled) 
     
     
         32 . (canceled)