Parathyroid hormone fusion polypeptide

Abstract

The disclosure relates to long acting parathyroid or parathyroid hormone like fusion polypeptides comprising a receptor polypeptide and its use in the treatment of hypoparathyroidism and osteoporosis.

We claim: 
     
         1 . A fusion polypeptide comprising
 a polypeptide comprising an amino acid sequence of a parathyroid hormone or biologically active fragment or analogue thereof,   a polypeptide comprising an amino acid sequence of a growth hormone binding domain of growth hormone receptor or fragment or analogue thereof, wherein said parathyroid hormone or biologically active fragment or analogue thereof is linked either directly or indirectly as a translational fusion to said growth hormone binding domain of growth hormone receptor.   
     
     
         2 . The fusion polypeptide according to  claim 1 , wherein said fusion polypeptide comprises a parathyroid hormone comprising or consisting of:
 the amino acid sequence set forth in SEQ ID NO: 10 or SEQ ID NO: 9; or   a fragment of SEQ ID NO: 10 including amino acids 1-28, 1-29, 1-30, 1-31, 1-32, 1-33, 1-34, or 1-35 of SEQ ID NO: 10, or a fragment of SEQ ID NO: 9 including amino acids 1-28, 1-29, 1-30, 1-31, 1-32, 1-33, 1-34, or 1-35 of SEQ ID NO: 9.   
     
     
         3 . The fusion polypeptide according to  claim 2  wherein said fusion polypeptide comprises a parathyroid hormone comprising or consisting of the amino acid sequence set forth in SEQ ID NO: 54. 
     
     
         4 . The fusion polypeptide according to  claim 1  wherein said fusion polypeptide comprises a parathyroid hormone analogue comprising or consisting of the amino acid sequence of formula (I): 
       
         
           
                 
               
                   (SEQ ID NO: 8) 
                 
                   Xaa01-Val-Xaa03-Glu-Ile-Gln-Leu-Xaa08-His-Xaa10- 
                 
                     
                 
                   Xaa11-Xaa12-Xaa13-Xaa14-Leu-Xaa16-Xaa17-Xaa18-Arg- 
                 
                     
                 
                   Arg-Arg-Xaa22-Phe-Leu-Xaa25-Xaa26-Leu-Ile-Ala-Glu- 
                 
                     
                 
                   Ile-His-Thr-Ala-Glu-Ile 
                 
             
                
                
                
                
                
                
                
                
               
            
           
         
         (I) where Xaa01 is Ser or Ala; Xaa03 is Ser or Ala; Xaa08 is Met or Leu; Xaa10 is Asn, Ala, Val, Asp, Glu, or Gln; Xaa11 is Leu, Ala, Val, Met, Lys, Arg, or Trp; Xaa12 is Gly, Ala, His, or Arg; Xaa13 is Lys, Ala, Leu, Gln, Arg, His, or Trp; Xaa14 is His, Leu, Arg, Phe, Trp, or Ser; Xaa16 is Gln or Asn; Xaa17 is Asp or Ser; Xaa18 is Ala, Leu, Met, Glu, Ser, or Phe; Xaa22 is Ala, Phe, Glu, Ser, Leu, Asn, Trp, or Lys; Xaa25 is His, Arg, Leu, Trp, or Lys; and Xaa26 is Lys, His, Ala, Ser, Asn, or Arg or a fragment thereof including amino acids 1-28, 1-29, 1-30, 1-31, 1-32, 1-33, 1-34, or 1-35 of formula (I), with the proviso that at least one of Xaa18 is not Leu or Met, Xaa22 is not Phe, and Xaa26 is not His. 
       
     
     
         5 . The fusion polypeptide according to  claim 1  wherein said fusion polypeptide comprises:
 a parathyroid hormone and a parathyroid hormone analogue amino acid sequence comprising or consisting of the amino acid sequence set forth in SEQ ID NO: 37; or 
 a parathyroid hormone related protein or biologically active fragment or analogue thereof, comprising the amino acid sequence set forth in SEQ ID NO: 40. 
 
     
     
         6 . The fusion polypeptide according to  claim 1  wherein said fusion polypeptide comprises:
 the extracellular growth hormone binding domain polypeptide of human growth hormone receptor; or 
 an extracellular growth hormone binding domain polypeptide comprising of the amino acid sequence set forth in SEQ ID NO: 5. 
 
     
     
         7 . The fusion polypeptide according to  claim 6  wherein said fusion polypeptide comprises:
 a modified extracellular growth hormone binding domain polypeptide wherein said modification is by addition, deletion or substitution of at least one amino acid residue wherein said modified polypeptide substantially lacks growth hormone binding activity or has reduced growth hormone binding activity; 
 a modification of one or more of the amino acid residues selected from the group consisting of: W169, R43, E44, I103, W104, I105, P106, I164 and D165 as set forth in SEQ ID NO: 5; 
 a deletion or substitution of amino acid residue tryptophan 104 of the amino acid sequence set forth in SEQ ID NO: 5; or 
 a substitution of tryptophan 104 for alanine as set forth in SEQ ID NO: 5. 
 
     
     
         8 . The fusion polypeptide according to  claim 1  wherein said fusion polypeptide comprises or consists of a parathyroid hormone fragment comprising or consisting of the amino acid residues 1-28, 1-29, 1-30, 1-31, 1-32, 1-33, 1-34, or 1-35 of SEQ ID NO: 8. 
     
     
         9 . The fusion polypeptide according to  claim 8  wherein said fusion polypeptide comprises or consists of the amino acid sequence set forth in SEQ ID NO: 54 wherein said parathyroid hormone amino acid sequence is modified by addition, deletion or substitution of at least one amino acid residue and wherein said modified fusion polypeptide retains parathyroid hormone activity. 
     
     
         10 . The fusion polypeptide according to  claim 1  wherein said fusion polypeptide comprises a growth hormone binding domain polypeptide wherein said growth hormone binding domain polypeptide is located at the carboxyl terminal end or amino terminal end of said fusion polypeptide. 
     
     
         11 . The fusion polypeptide according to  claim 1  wherein said fusion polypeptide comprises a pro-peptide and/or a peptide secretion signal. 
     
     
         12 . The fusion polypeptide according to  claim 1  wherein said fusion polypeptide comprises a parathyroid hormone, fragment or analogue thereof and is linked directly or indirectly to said growth hormone binding domain polypeptide of growth hormone receptor by a peptide linker and/or is directly linked to said receptor polypeptide as an in-frame translational fusion. 
     
     
         13 . A nucleic acid molecule that encodes a fusion polypeptide according to  claim 1 . 
     
     
         14 . A vector comprising a nucleic acid molecule according to  claim 13 . 
     
     
         15 . An isolated cell transfected or transformed with a nucleic acid molecule according to  claim 13  or a vector comprising the nucleic acid molecule. 
     
     
         16 . A method for the production of a fusion polypeptide comprising the steps consisting of:
 i) providing a cell transfected or transformed with a nucleic acid molecule encoding the fusion polypeptide of  claim 1  or a vector comprising the nucleic acid molecule and cell culture medium;   ii) culturing said cell; and   iii) isolating the fusion polypeptide from said cell or medium.   
     
     
         17 . A pharmaceutical composition comprising a fusion polypeptide according to  claim 1  including an excipient or carrier. 
     
     
         18 . A method of treating hypoparathyroidism in a subject comprising administering the fusion polypeptide according to  claim 1  to the subject. 
     
     
         19 . A method of treating osteoporosis in a human subject comprising administering the fusion polypeptide according to  claim 1  to the human subject. 
     
     
         20 . The method of  claim 19  wherein said fusion polypeptide comprises the amino acid sequence set forth in SEQ ID NO: 40. 
     
     
         21 . The method of  claim 19  wherein said fusion polypeptide is administered to the subject in combination with calcium carbonate and/or vitamin D supplements such as calcitrol or alfacalcitrol. 
     
     
         22 . The fusion polypeptide according to  claim 1 , wherein said fusion polypeptide comprises or consists of an amino acid sequence set forth in SEQ ID NO: 20, 27, 28, 33 or 34. 
     
     
         23 . The fusion polypeptide according to  claim 1 , wherein said fusion polypeptide comprises or consists of an amino acid sequence set forth in SEQ ID NO: 20. 
     
     
         24 . The fusion polypeptide according to  claim 1 , wherein said fusion polypeptide comprises or consists of an amino acid sequence set forth in SEQ ID NO: 27.