Soluble gp130 muteins with improved binding activity

Abstract

Described are soluble gp130 polypeptide monomers and dimers, wherein, in a preferred embodiment, at least one of the three amino acid residues Thr 102 GIn 113 or ASn 114 of the N-terminal Ig-like domain of gp130 is mutated to Tyr 102 , Phe 113 or Leu 114 , respectively. These mutations, alone or in combination, specifically enhance binding of gp130 to its ligand complex of interleukin-6 and soluble interleukin-6 receptor, thus increasing the biological activity of the gp130 muteins. In a particularly preferred embodiment, all three mutations are combined in the triple mutein Thr102Tyr/Gln113Phe/Asn114Leu (T102Y/Q113F/N114L). Moreover, a pharmaceutical composition containing said monomers or dimers and various medical uses are described.

1 - 15 . (canceled) 
     
     
         16 . A polynucleotide encoding a polypeptide comprising the entire extracellular part of glycoprotein gp130 or variants or fragments thereof, which is capable of inhibiting the activity of the agonistic complex IL-6/sIL-6R and wherein Thr 102  is substituted by a large neutral amino acid residue and/or Gln 113  is substituted by a hydrophobic and nonpolar amino acid residue and/or Asn 114  is substituted by a hydrophobic and nonpolar amino acid residue. 
     
     
         17 . The polynucleotide of  claim 16 , wherein the polynucleotide sequence is codon optimized for the production of the encoded polypeptide in prokaryotic or eukaryotic host cells or in a cell-free expression system. 
     
     
         18 . An expression vector containing a polynucleotide of  claim 16 . 
     
     
         19 . A host cell or cell-free expression system containing the expression vector of  claim 18 . 
     
     
         20 . A method of producing a polypeptide, comprising culturing a host cell or using a cell-free expression system of  claim 19  and recovering and purifying the polypeptide monomer or dimer from said host cell, the culture medium or the cell-free expression system. 
     
     
         21 . A pharmaceutical composition containing a polynucleotide of  claim 16  or  17 , or an expression vector of  claim 18 . 
     
     
         22 . Use of a polynucleotide of  claim 16  or  17 , or an expression vector of  claim 18  for the preparation of a pharmaceutical composition for the treatment and/or prevention of a disease or disorder where blockage of the agonistic complex IL-6/sIL-6R has a beneficial effect, wherein said disease is bone resorption, hypercalcemia, cachexia, a tumor or other type of cancer, an autoimmune disease, an inflammatory or atopic disease, an infection, sepsis, an endocrinologic disorder or a metabolic or catabolic disease. 
     
     
         23 . An antibody binding to a polypeptide comprising the entire extracellular part of glycoprotein gp130 or variants or fragments thereof, which is capable of inhibiting the activity of the agonistic complex IL-6/sIL-6R and wherein Thr 102  is substituted by a large neutral amino acid residue and/or Gln 113  is substituted by a hydrophobic and nonpolar amino acid residue and/or Asn 114  is substituted by a hydrophobic and nonpolar amino acid residue, which is specific for the mutated peptide motif(s) or the specific linker region of said polypeptide. 
     
     
         24 . A diagnostic method which is based on the detection of the binding of an antibody of  claim 23  to a polypeptide comprising the entire extracellular part of glycoprotein gp130 or variants or fragments thereof, which is capable of inhibiting the activity of the agonistic complex IL-6/sIL-6R and wherein Thr 102  is substituted by a large neutral amino acid residue and/or Gln 113  is substituted by a hydrophobic and nonpolar amino acid residue and/or Asn 114  is substituted by a hydrophobic and nonpolar amino acid residue.