Non-Natural Consensus Albumin Binding Domains

Abstract

Non-natural albumin binding domains, polynucleotides encoding thereof and methods of making and using these domains and polynucleotides are useful in controlling the half-life of therapeutic molecules for patients.

What is claimed: 
     
         1 . An isolated non-natural albumin binding domain comprising an amino acid sequence at least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to the amino acid sequence of SEQ ID NO: 21 and wherein the albumin binding domain has a dissociation constant (K D ) of binding to human albumin between about 50 pM-1 μM using ProteOn™ XPR-36 Protein Interaction Array System (Bio-Rad) using GLC sensor chips at a flow rate of 100 μL/min using PBST (PBS, 0.005% Tween20) as running buffer. 
     
     
         2 . An isolated non-natural albumin binding domain comprising an amino acid of SEQ ID NO: 21 having substitutions at 1, 2, 3, 4, 5, or 6 residues and wherein the albumin binding domain has a dissociation constant (K D ) of binding to human albumin between about 50 pM-1 μM using ProteOn™ XPR-36 Protein Interaction Array System (Bio-Rad) using GLC sensor chips at a flow rate of 100 μL/min using PBST (PBS, 0.005% Tween20) as running buffer. 
     
     
         3 . The isolated albumin binding domain of  claim 2 , wherein the substitutions at 1, 2, 3, 4, 5, or 6 residues occur at one or more of amino acid positions Y21, Y22, L25, K30, T31, E33, G34, A37, L38, E41, I42 and A45 of SEQ ID NO: 21. 
     
     
         4 . The isolated albumin binding domain of  claim 2 , wherein the substitutions at 1, 2, 3, 4, 5, or 6 residues occur at one or more of amino acid positions Y21, Y22, K30, T31, A37, and E41 of SEQ ID NO: 21. 
     
     
         5 . The isolated albumin binding domain of  claim 1 , further comprising an extension of 5 amino acids at its N-terminus. 
     
     
         6 . The isolated albumin binding domain of  claim 5 , wherein the extension of 5 amino acids comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 42, 43 and 45-55. 
     
     
         7 . The isolated albumin binding domain of  claim 5 , wherein the extension of 5 amino acids comprises an amino acid sequence of SEQ ID NO: 43. 
     
     
         8 . The isolated albumin binding domain of  claim 2 , further comprising an extension of 5 amino acids at its N-terminus. 
     
     
         9 . The isolated albumin binding domain of  claim 8 , wherein the extension of 5 amino acids comprises an amino acid sequence of SEQ ID NOs: 42, 43 or 45-55. 
     
     
         10 . The isolated albumin binding domain of  claim 8 , wherein the extension of 5 amino acids comprises an amino acid sequence of SEQ ID NO: 43. 
     
     
         11 . The isolated albumin binding domain of  claim 1 , wherein the albumin binding domain has an off rate constant (Koff) of binding to human albumin at 3.6×10 −5  to 1.1×10 −1  using ProteOn™ XPR-36 Protein Interaction Array System (Bio-Rad) using GLC sensor chips at a flow rate of 100 μl/min using PBST (PBS, 0.005% Tween20) as running buffer. 
     
     
         12 . A fusion protein comprising the albumin binding domain of  claim 1  and a bioactive agent. 
     
     
         13 . The fusion protein of  claim 12 , wherein the bioactive agent is a protein scaffold specifically binding a target molecule. 
     
     
         14 . The fusion protein of  claim 13 , wherein the protein scaffold is based on Tencon25 comprising amino acid residues 1-90 of SEQ ID NO: 39 or variants thereof. 
     
     
         15 . The fusion protein of  claim 13 , wherein the albumin binding protein and the bioactive agent are operably linked using a linker. 
     
     
         16 . The fusion protein of  claim 15 , wherein the linker comprises a Gly-Ser linker. 
     
     
         17 . The fusion protein of  claim 16 , wherein the linker comprises an amino acid sequence of SEQ ID NO: 40. 
     
     
         18 . A pharmaceutical composition comprising the fusion protein of  claim 12  and at least one pharmaceutically acceptable carrier or diluent.