Interleukin-2 variants and methods of uses thereof

Abstract

The present invention relates to polypeptides which share primary sequence with human IL-2, except for several amino acids that have been mutated. One panel of IL-2 variants comprise mutations with impressive manufacturability that preferentially promotes the proliferation, survival, activation and/or function of immunosuppressive regulatory T cells (Treg: CD4+CD25+FoxP3+) over effector T cells and NK cells. Also includes therapeutic uses of such IL-2 selective agent, used alone, or in combination with immune modulating agents or disease-tissue targeting antibody, protein or peptide to treat Treg cell-deficiency, various autoimmune and inflammatory disorders, organ transplantation and graft-versus-host disease. In another aspect the present invention relates to pharmaceutical compositions comprising the polypeptides disclosed. Finally, the present invention relates to the therapeutic use of the polypeptides and pharmaceutical compositions disclosed due to their selective modulating effect of the immune system on diseases like autoimmune and inflammatory disorders.

1 - 33 . (canceled) 
     
     
         34 . An isolated interleukin-2 (IL-2) variant polypeptide, wherein said IL-2 variant polypeptide comprises the amino acid sequence of SEQ ID NO: 3 having one or more of amino acid residues position L19, D20, L21, Q22, R38, F42, N88, S125 or Q126 substituted with another amino acid, and wherein said IL-2 variant polypeptide has a reduced ability to activate the IL-2Rβγ receptor complex as compared to the polypeptide represented by SEQ ID NO: 3, while retaining the ability to activate the IL-2Rαβγ receptor complex. 
     
     
         35 . The IL-2 variant polypeptide according to  claim 34 , wherein the amino acid substitution is selected from the group consisting of: the substitution of L19D, L19H, L19N, L19R, L19S, L19P and L19Y at position 19, the substitution of D20E, D20I, D20N, D20Q, D20S, D20T and D20Y at position 20, the substitution of L21S, L21R and L21N at position 21, the substitution of Q22N, Q22H, Q22K, Q22Y, Q22I at position 22, the substitution of N88E, N88G, N88T, N88M, N88Q, N88R, and N88I at position 88, the substitution of S125E, S125K, S125H, S125W and S125I at position 125, and the substitution of Q126D, Q126E, Q126H, Q126K, Q126L, Q126M, Q126N and Q126Y at position 126 of SEQ ID NO: 3, or any combination of these substitutions. 
     
     
         36 . The IL-2 variant polypeptide according to  claim 35 , wherein said IL-2 variant polypeptide comprises two amino acid substitutions at amino acid residues position L19 and Q126 of SEQ ID NO: 3; wherein the amino acid substitution is selected from the group consisting of: the substitution of L19D, L19H, L19N, L19R, L19S, L19P and L19Y at position 19, and the substitution of Q126D, Q126E, Q126H, Q126K, Q126L, Q126M, Q126N and Q126Y at position 126 of SEQ ID NO: 3. 
     
     
         37 . The IL-2 variant polypeptide according to  claim 35 , wherein said IL-2 variant polypeptide comprises three amino acid substitutions at amino acid residues position L19, S125 and Q126 of SEQ ID NO: 3; wherein the amino acid substitution is selected from the group consisting of: the substitution of L19D, L19H, L19N, L19R, L19S, L19P and L19Y at position 19, the substitution of S125E, S125K, S125H, S125W and S125I at position 125, and the substitution of Q126D, Q126E, Q126H, Q126K, Q126L, Q126M, Q126N and Q126Y at position 126 of SEQ ID NO: 3. 
     
     
         38 . The IL-2 variant polypeptide according to  claim 34 , wherein the IL-2 variant polypeptide comprises the amino acid sequence is selected from the group consisting of the amino acid sequences set forth in SEQ ID NOS: 4-43, 108-146 and 193-197. 
     
     
         39 . The IL-2 variant polypeptide according to  claim 38 , wherein the IL-2 variant polypeptide comprises the amino acid sequence is selected from the group consisting of the amino acid sequences set forth in the IL-2 variants comprise SEQ ID NOS: 5-14, 26-43, 108-111, 125-146 and 193-197. 
     
     
         40 . An isolated IL-2 variant polypeptide comprising an amino acid sequence that is at least about 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 3 and wherein said IL-2 variant polypeptide has a reduced ability to activate the IL-2Rβγ receptor complex as compared to the polypeptide represented by SEQ ID NO: 3, while retaining the ability to activate the IL-2Rαβγ receptor complex. 
     
     
         41 . An isolated fusion protein comprising 1) an IL-2 variant polypeptide according to  claims 34  and  2 ) a heterologous protein; wherein the fusion protein is in the form of either a dimer or monomer. 
     
     
         42 . The isolated fusion protein according to  claim 41 , wherein said IL-2 variant polypeptide is fused at its N-terminal amino acid to the C-terminal amino acid of the heterologous protein, optionally through a peptide linker. 
     
     
         43 . The isolated fusion protein according to  claim 41 , wherein said IL-2 variant polypeptide is fused at its C-terminal amino acid to the N-terminal amino acid of the heterologous protein, optionally through a peptide linker. 
     
     
         44 . The isolated fusion protein according to  claim 41 , wherein the heterologous protein increases the circulating half-life of the IL-2 variant polypeptide. 
     
     
         45 . The isolated fusion protein according to  claim 41 , wherein the heterologous protein is selected from the group consisting of a full-length null antibody or antibody fragment which provides for half-life extension. 
     
     
         46 . The isolated fusion protein according to  claim 41 , wherein the heterologous protein enhances the expression levels and overall purity of the IL-2 variant polypeptide. 
     
     
         47 . The isolate fusion protein according to  claim 41 , wherein the heterologous protein serves as a marker or tag or targeting moiety. 
     
     
         48 . The isolated fusion protein according to  claim 41 , wherein the heterologous protein is an Fc domain selected from the group consisting of a human IgG1 Fc domain, a human IgG2 Fc domain, a human IgG3 Fc domain, a human IgG4 Fc domain, an IgA Fc domain, an IgD Fc domain, an IgE Fc domain, an IgG Fc domain, and an IgM Fc domain. 
     
     
         49 . The isolated fusion protein according to  claim 41 , wherein the Fc domain is an Fc domain having an amino acid sequence selected from the group consisting of the amino acid sequences set forth in SEQ ID NOS: 44-47 and 212-213. 
     
     
         50 . The isolated fusion protein according to  claim 41 , wherein said IL-2 variant polypeptide is fused to said heterologous protein through a peptide linker, and wherein said peptide linker comprises between 1 and 40 amino acids. 
     
     
         51 . The isolated fusion protein of  claim 41 , wherein the fusion protein comprises the amino acid sequence selected from the group consisting of the amino acid sequences set forth in SEQ ID NOS: 69-107, 147-189, and 198-211. 
     
     
         52 . A pharmaceutical composition comprising an IL-2 variant polypeptide or isolated fusion protein according to any one of claims  1  to  22  in admixture with a pharmaceutically acceptable carrier. 
     
     
         53 . A method of treating a disorder in a subject, wherein the disorder is selected from the group consisting of an autoimmune disease, organ transplantation rejection, or associated graft-versus-host disease (GvHD), an inflammatory disease, rheumatoid arthritis, inflammatory bowel disease, multiple sclerosis, atopic dermatitis, diabetes and immune-related adverse events (irAE), comprising administering to the subject a therapeutically effective amount of the pharmaceutical composition according to claim  23 .