Single-chain cd137-receptor agonist proteins

Abstract

Provided herein are specific CD137 receptor agonist proteins, nucleic acids encoding the same, and methods of treating a subject having a CD137L-associated disease or disorder. The CD137 receptor agonist proteins provided herein comprise three soluble CD137L domains and an Fe fragment. The CD137 receptor agonist proteins are substantially non-aggregating and suitable for therapeutic, diagnostic and/or research applications.

1 . (canceled) 
     
     
         2 . A CD137 receptor agonist protein comprising a single-chain fusion polypeptide comprising:
 (i) a first soluble CD137L domain,   (ii) a first peptide linker,   (iii) a second soluble CD137L domain,   (iv) a second peptide linker, and   (v) a third soluble CD137L domain, and   (vi) an antibody derived Fab- or Fc-fragment, wherein the CD137L domains (i), (iii) and (v) are N- and C-terminally shortened domains consisting of amino acids starting at an amino acid between 86 and 90 and ending at amino acid 240, or 241, or 243 of SEQ ID NO:1.   
     
     
         3 . The CD137 receptor agonist protein of  claim 2 , wherein the Fc fragment of (vi) consists of the amino acid sequence as shown in SEQ ID NO: 13 or 14 or amino acids 1-217 of SEQ ID NO: 13 or 14. 
     
     
         4 . The CD137 receptor agonist protein of  claim 2 , wherein the Fc fragment of (vi) is fused to the C-terminal end of the third CD137L domain (v) via a hinge-linker of any one of SEQ ID NOS: 16 or 19-24. 
     
     
         5 . The CD137 receptor agonist protein of  claim 2 , wherein the Fab fragment of (vi) is fused to the N-terminal end of the first CD137L domain. 
     
     
         6 . The CD137 receptor agonist protein of  claim 5 , wherein antibody Fab fragment of (vi) is fused to the N-terminal end of the first CD137L domain via short G/C rich linkers according to SEQ ID NOS: 2-12. 
     
     
         7 . The CD137 receptor agonist protein of  claim 2 , wherein in at least one of the soluble CD137L domains, the starting amino acid D86 or R88 or Q89 is replaced by a neutral amino acid. 
     
     
         8 . The CD137 receptor agonist protein of  claim 2 , wherein the soluble CD137L domains optionally comprises a mutation at position D86, L87, R88, Q89, D112, V118, A154, A174, A176, A188, T241, or at two or more of said positions. 
     
     
         9 . The CD137 receptor agonist protein of  claim 2 , wherein the first and second peptide linkers (ii) and (iv) independently have a length of 3-8 amino acids. 
     
     
         10 . The CD137 receptor agonist protein of  claim 2 , wherein the first and the second peptide linkers (ii) and (iv) consist of the amino acid sequence according to SEQ ID NO: 2. 
     
     
         11 . The CD137 receptor agonist protein of  claim 2 , which additionally comprises an N-terminal signal peptide domain. 
     
     
         12 . The CD137 receptor agonist protein of  claim 2 , wherein the polypeptide is further post-translationally modified. 
     
     
         13 . The CD137 receptor agonist protein of  claim 4 , wherein the polypeptide is further post-translationally modified. 
     
     
         14 . The CD137 receptor agonist protein of  claim 13 , wherein the post-translational modification comprises modification of the N-terminal glutamine to pyroglutamate. 
     
     
         15 . A nucleic acid molecule encoding the CD137 receptor agonist protein of  claim 2 , in an operative linkage with an expression control sequence. 
     
     
         16 . An expression vector comprising the nucleic acid molecule of  claim 15 . 
     
     
         17 . A cell or a non-human organism transformed or transfected with the nucleic acid molecule of  claim 15 . 
     
     
         18 . A pharmaceutical or diagnostic composition comprising as an active agent the CD137 receptor agonist protein of  claim 2  and one or more pharmaceutically acceptable carriers, diluents, excipients and/or adjuvants.