Interferon Analogs

Abstract

The invention relates to the field of medicine. Among others, it relates to biologically active analogs of interferons (IFNs) which show less unwanted side-effects and to the therapeutic uses thereof. Provided is an IFN analog, wherein the moiety mediating binding to its natural receptor is at least functionally disrupted and wherein the analog comprises a signaling moiety capable of mediating intracellular IFN activity, said signaling moiety being provided at its N-terminus, optionally via a linker, with at least one targeting domain capable of binding to a cell surface receptor other than the IFN receptor.

1 . An analog of interferon gamma (IFNγ), wherein the moiety mediating binding to its natural receptor is at least functionally disrupted and wherein the analog comprises a signaling moiety capable of mediating intracellular IFNγ activity, said signaling moiety being provided at its N-terminus, optionally via a linker, with at least one targeting domain capable of binding to a cell surface receptor other than the IFNγ receptor. 
     
     
         2 . The analog according to  claim 1 , wherein the signaling moiety mediating intracellular activity comprises a polybasic nuclear localization signal (NLS) motif. 
     
     
         3 . The analog according to  claim 2 , wherein the polybasic NLS motif comprises the amino acid sequence (R)KRXRS(R), wherein X is any amino acid residue. 
     
     
         4 . The analog according to  claim 1 , wherein the signaling moiety comprises a sequence selected from the group consisting of
 (a) the amino acid sequence   
       
         
           
                 
               
                   (SEQ ID NO: 6) 
                 
                 
                 
               
                     
                   KFEVNNPQVQRQAFNELIRVVHQLLPESSLRKRKRSR; 
                 
             
                
               
            
             
                
               
            
           
         
         (b) the amino acid sequence 
       
       
         
           
                 
               
                   (SEQ ID NO: 8) 
                 
                 
                 
               
                     
                   YSVTDLNVQRKAIHELIQVMAELSPAAKTGKRKRSQ; 
                 
             
                
               
            
             
                
               
            
           
         
         (c) the amino acid sequence 
       
       
         
           
                 
               
                   (SEQ ID NO: 9) 
                 
                 
                 
               
                     
                   AKFEVNNPQIQHKAVNELIRVIHQLSPESSLRKRKRSRC; 
                 
             
                
               
            
             
                
               
            
           
         
         (d) a stretch of at least 10, preferably at least 15, contiguous amino acids of the sequence under (a), (b) or (c); 
         (e) an amino acid sequence showing at least 70%, preferably at least 80%, more preferably at least 90% identity to (a) or (b) or (c) provided that the intracellular signaling activity is maintained; 
         (f) the amino acid sequence under (a) or (b) or (c) wherein at most 10, preferably at most 8, more preferably at most 5 amino acid residues are deleted, added or substituted, provided that the signaling activity, for example nuclear translocation, is maintained; 
         (g) the consensus sequence VxxxxVQRxAxxELlxVxxxLxPxxxxxKRxRS (SEQ ID NO: 10) wherein x is any amino acid residue; 
         (h) the consensus sequence VxxxxxQxxAxxELlxVxxxLxPxxxxxKRKRS (SEQ ID NO: 11) wherein x is any amino acid residue; and 
         (i) the consensus sequence Vxxx[Q/N][F/V/I]Q [R/H][Q/K]A[F/V/I][N/H]ELI[R/Q]Vx[H/A][Q/E]L[L/S]P[E/A][S/A][S/A][L/K]xxKRKRS (SEQ ID NO: 64) wherein x is any amino acid residue; 
       
     
     
         5 . The analog according to  claim 4 , comprising a signaling moiety according to the sequence: 
       
         
           
                 
               
                   (SEQ ID NO: 61) 
                 
                 
               
                   Xaa 1 Xaa 2 Xaa 3 Xaa 4  Val Xaa 5  Xaa 6  Xaa 7  Xaa 8  Xaa 9   
                 
                     
                 
                   Gln Xaa 10  Xaa 11  Ala Xaa 12  Xaa 13  Glu Leu Ile Xaa 14   
                 
                     
                 
                   Val Xaa 15  Xaa 16  Xaa 17  Leu Xaa 18  Pro Xaa 19  Xaa 20   
                 
                     
                 
                   Xaa 21  Xaa 22  Xaa 23  Lys Arg Lys Arg Ser Xaa 24  Xaa 25 , 
                 
             
                
               
            
             
                
                
                
                
                
                
                
               
            
           
         
       
       wherein
 Xaa 1 , Xaa 2 , Xaa 3 , Xaa 4 , Xaa 6 , Xaa 11 , Xaa 13 , Xaa 14 , Xaa 16  Xaa 17 , Xaa 18  Xaa 19  Xaa 20  Xaa 21  Xaa 22  Xaa 23  Xaa 24  and Xaa 25  is any amino acid residue, 
 Xaa 5  is a polar, uncharged residue such as Asn or Thr; 
 Xaa 7  is a non-polar, hydrophobic residue such as Pro or Leu; 
 Xaa 8  is a polar, uncharged residue such as Gln or Asn; 
 Xaa 9  is a non-polar hydrophobic residue such as Val, Ile or Leu; 
 Xaa 10  is a polar, basic residue such as Arg, His or Lys; 
 Xaa 12  is a non-polar hydrophobic residue such as Phe, Val or Ile; 
 Xaa 15  is a non-polar hydrophobic residue such as Val, Ile, Met. 
 
     
     
         6 . The analog according to  claim 1 , wherein the targeting domain can bind to a receptor that is specific for fibroblast and fibroblast-like cells, preferably myofibroblasts, portal fibroblasts, mesangial cells, interstitial fibroblasts, alveolar fibroblasts and/or stromal cells. 
     
     
         7 . The analog according to  claim 1 , wherein the targeting domain can bind to a receptor for an oligosaccharide or glycoprotein, preferably the mannose 6-phosphate/insulin-like growth factor-II receptor (M6P/IGF2R), asialoglycoprotein (ASGP) receptor or the mannose receptor (CD 206). 
     
     
         8 . The analog according to  claim 7 , wherein the targeting domain is an oligosaccharide, preferably mannose(-6-phosphate) or lactose, conjugated to a carrier molecule. 
     
     
         9 . The analog according to  claim 1 , wherein the receptor is selected from the group consisting of the PDGF receptors, collagen type VI receptor, and cytokine receptors including TGFβ receptor, TNFα receptor, Insulin growth factor receptors, VEGF receptors, chemokine receptors and IL1β receptor. 
     
     
         10 . The analog according to  claim 9 , wherein receptor is the PDGF receptor, preferably the PDGFβ receptor. 
     
     
         11 . The analog according to  claim 1 , wherein the targeting domain comprises at least one cyclic peptide portion. 
     
     
         12 . The analog according to  claim 11 , wherein the targeting domain comprises at least one tandem repeat of a cyclic peptide portion, preferably identical cyclic peptide portions. 
     
     
         13 . The analog according to  claim 11 , wherein the two cyclic peptide portions within a tandem repeat are connected via a linker of 2 to 7 amino acids. 
     
     
         14 . The analog according to  claim 13 , wherein the linker is selected from the group consisting of (i) the sequences KGSGG (SEQ ID NO: 37) and KGSGSGG (SEQ ID NO: 38) and (ii) the sequences of the general formula [G n D m ] wherein n+m is from 4 to 7, wherein n≧4 and M an integer between 0 and 3. 
     
     
         15 . The analog according to  claim 1 , wherein the targeting domain comprises at least the amino acid sequence RGD, KPT, SRN, NLI and/or LID. 
     
     
         16 . The analog according to  claim 15 , wherein the targeting domain comprises the amino acid sequence CSRNLIDC (SEQ ID NO: 46)-linker-CSRNLIDCS (SEQ ID NO: 62), wherein the linker is an amino acid sequence of 3 to 7, preferably 4 or 5, amino acid residues. 
     
     
         17 . A conjugate comprising a compound of interest conjugated to a targeting domain, said targeting domain comprising the amino acid sequence X 1 SRNLIDX 2  (SEQ ID NO: 21)-linker-X 3 SRNLIDX 4  (SEQ ID NO: 21), wherein the pair of X 1  and X 2  and the pair of X 3  and X 4  can form a bond, preferably a peptidic bond, such that a bicyclic structure is formed wherein the sequences SRNLID are each part of a ring, and wherein the linker is an amino acid sequence of 2 to 7 amino acid residues. 
     
     
         18 . The analog according to  claim 1 , further comprising at least one non-antigenic polymer. 
     
     
         19 . An isolated nucleic acid sequence encoding an analog according to  claim 1 , wherein the analog is proteinaceous. 
     
     
         20 . An expression vector comprising an isolated nucleic acid sequence according to  claim 19 . 
     
     
         21 . A host cell comprising an isolated nucleic acid sequence according to  claim 18 . 
     
     
         22 . A pharmaceutical composition comprising an analog according to  claim 1  and a pharmaceutically acceptable carrier, adjuvant and/or excipient. 
     
     
         23 . An analog according to  claim 1  for the therapeutic or prophylactic treatment of a disease selected from cancer, viral disease, fibrotic disease, sclerotic disease and chronic or acute inflammatory diseases. 
     
     
         24 . An analog to  claim 23 , for the treatment of a liver disease, preferably a chronic liver disease such as liver cirrhosis. 
     
     
         25 . An analog to  claim 23 , for the treatment of a viral disease, preferably a disease caused by influenza virus or respiratory syncytial virus (RSV). 
     
     
         26 . A method for the therapeutic or prophylactic treatment of a disease selected from cancer, viral disease, fibrotic disease, sclerotic disease and chronic or acute inflammatory diseases, comprising administering to a subject in need thereof an effective dose of an analog according to  claim 1 . 
     
     
         27 . The method according to  claim 26 , wherein the disease is selected from the group consisting of glomerulosclerosis, interstitial fibrosis, lung fibrosis, atherosclerosis, rheumatoid arthritis, Crohns disease, colitis ulcerosa, glomerulonephritis and sepsis. 
     
     
         28 . The method according to  claim 26 , wherein said administering comprises mucosal administration. 
     
     
         29 . The method according to  claim 26 , wherein the disease is a liver disease. 
     
     
         30 . The method according to  claim 29 , wherein the liver disease is a chronic liver disease. 
     
     
         31 . The analog according to  claim 3 , wherein the polybasic NLS motif comprises the amino acid sequence
 (R)KRXRS(R) (SEQ ID NO: 1) wherein X is R, K, S or T.