Peptide for use in the reduction of side effects in the form of immunostimulatory reactions/effects

Abstract

What is described is a peptide for use in the reduction of side-effects in form of immunostimulatory reactions/effects which occur with gene therapy, wherein the peptide is coupled to a nucleic acid molecule used in gene therapy. Furthermore, such a peptide is described for use in the reduction of side-effects in form of immunostimulatory reactions/effects which occur with gene therapy, wherein the peptide is coupled to a nucleic acid molecule used in gene therapy, wherein the immunostimulatory reactions/effects are induced by activation of IFIT1/2, IRF9, TLR3, TLR7, TLR8 or PKR.

1 . A coupled peptide for use in the reduction of side-effects in form of immunostimulatory reactions/effects which occur with gene therapy, comprising a peptide coupled to a nucleic acid molecule for use in gene therapy. 
     
     
         2 . The coupled peptide according to  claim 1 , wherein the immunostimulatory reactions/effects are induced by activation of IFIT1/2, IRF9, TLR3, TLR7, TLR8 or PKR. 
     
     
         3 . The coupled peptide according to  claim 1 , wherein the immunostimulatory reactions/effects lead to redness, inflammation, flu-like symptoms and/or fever. 
     
     
         4 . The coupled peptide according to  claim 1 , wherein the peptide is specifically cleavable by proteases. 
     
     
         5 . The coupled peptide according to  claim 1 , wherein the peptide is non-specifically cleavable. 
     
     
         6 . The coupled peptide according to  claim 1 , wherein the nucleic acid molecule comprises further chemical modifications. 
     
     
         7 . The coupled peptide according to  claim 1 , wherein the peptide comprises the cleavage sequence for Legumain (cleavage sequence: Ala-Ala-Asn), Caspase-4 (cleavage sequence: Leu-Glu-Val-Asp-Gly; SEQ ID NO:1), uPA (cleavage sequence: Glu-Gly-Arg), Fap (cleavage sequence: Gly-Pro), KLK (cleavage sequence: Val-Gln-Gln-Lys-Ser; SEQ ID NO:2), PLAP (cleavage sequence: Pro-His-Ile-Tyr-Val; SEQ ID NO:3), Cathepsin-E (cleavage sequence: (Gly-Gly-Ala-Phe-Leu-Val-Leu-Pro; SEQ ID NO:4; and cleavage sequence: Ala-Leu-Ala-Phe-Ser-Leu-Ala-Ala; SEQ ID NO:5 respectively), CMV protease (cleavage sequence: Pro-Ser-Val-Ser-Ala; SEQ ID NO:6), or EBV protease (cleavage sequence: Gly-Ser-Ala-Ser-Ala, SEQ ID NO:7). 
     
     
         8 . The coupled peptide according to  claim 6 , wherein the cleavage sequence comprises a sequence having a deletion, substitution, insertion and/or addition of 1 to 10 amino acids in the sequence, which is selected from the group consisting of Ala-Ala-Asn, SEQ ID NO:1, Glu-Gly-Arg, Gly-Pro, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6 and SEQ ID NO:7, and which can be cleaved by the corresponding protease. 
     
     
         9 . The coupled peptide according to  claim 1 , wherein the nucleic acid molecule is selected from the group consisting of mRNA, siRNA, miRNA, saRNA, RNA, PNA, DNA, LNA, mixtures of different nucleotides and aptameres. 
     
     
         10 . The coupled peptide according to  claim 1 , wherein the nucleic acid molecule is an siRNA. 
     
     
         11 . The coupled peptide according to  claim 10 , wherein the siRNA comprises a nucleotide sequence which is selected from the group consisting of: 
       
         
           
                 
                 
               
                     
                   (5-3) UCA UAU UCG ACU UUG GUU GCC 
                 
                     
                   (polo-like kinase (PLK); SEQ ID NO:8); 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 9) 
                 
                     
                   (5-3) UCA AAC UCC AUC AUG AUC U 
                 
                     
                   and/or 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 10) 
                 
                     
                   (5-3) UCC AUC AUG AUC UUC UGG A 
                 
                     
                   (CHMP); 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 11) 
                 
                     
                   (5-3) UUC AUA AAC ACA GUU CUC C 
                 
                     
                   (PDCD); 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 12) 
                 
                     
                   (5-3) UCA AAU UGA GGC ACU GUG C 
                 
                     
                   (RFWD); 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 13) 
                 
                     
                   (5-3) UUU CUU CAG AGC AGG AGC A, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 14) 
                 
                     
                   (5-3) AUA CAC ACC CUU UGC CUC A 
                 
                     
                   and/or 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 15) 
                 
                     
                   (5-3) AUU UCA GGC UCA UAU UCC U 
                 
                     
                   (ATAP); 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 16) 
                 
                     
                   (5-3) CAC AAU UCC CAC UUU GAG C, 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 17) 
                 
                     
                   (5-3) GUU ACC CAC AAU UCC CAC U 
                 
                     
                   and/or 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 18) 
                 
                     
                   (5-3) UUU CUU CUC UUU GUC UGG G 
                 
                     
                   (AGAP); 
                 
                     
                   and 
                 
                     
                     
                 
                     
                   (SEQ ID NO: 19) 
                 
                     
                   (5-3) UAU UCU CCA AAC AAU GUG C 
                 
                     
                   (RCHY). 
                 
             
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
                
               
            
           
         
       
     
     
         12 . The coupled peptide according to  claim 1 , wherein the coupled peptide is covalently or non-covalently linked to at least one molecule selected from the group consisting of cell-penetrating peptides, enzyme substrates, polyethyleneimines, nanocontainers, nanoparticles, lipids, and receptor-ligand complexes, for introduction into cells or for targeted introduction into specific cells. 
     
     
         13 . The coupled peptide according to  claim 5 , wherein the peptide is non-specifically cleavable by modification of pH value thereof. 
     
     
         14 . A gene therapy method, comprising administering to a subject in need thereof a coupled peptide according to  claim 1  thereby to reduce side-effects of the gene therapy in the form of immunostimulatory reactions/effects. 
     
     
         15 . The method according to  claim 14 , wherein the immunostimulatory reactions/effects are induced by activation of IFIT1/2, IRF9, TLR3, TLR7, TLR8 or PKR. 
     
     
         16 . The method according to  claim 14 , wherein the immunostimulatory reactions/effects lead to redness, inflammation, flu-like symptoms and/or fever. 
     
     
         17 . A gene therapy method, comprising administering to a subject in need thereof a coupled peptide according to  claim 4  thereby to reduce side-effects of the gene therapy in the form of immunostimulatory reactions/effects. 
     
     
         18 . A gene therapy method, comprising administering to a subject in need thereof a coupled peptide according to  claim 5  thereby to reduce side-effects of the gene therapy in the form of immunostimulatory reactions/effects. 
     
     
         19 . The method according to  claim 18 , wherein the peptide is non-specifically cleavable by modification of pH thereof. 
     
     
         20 . A gene therapy method, comprising administering to a subject in need thereof a coupled peptide according to  claim 6  thereby to reduce side-effects of the gene therapy in the form of immunostimulatory reactions/effects. 
     
     
         21 . A gene therapy method, comprising administering to a subject in need thereof a coupled peptide according to  claim 7  thereby to reduce side-effects of the gene therapy in the form of immunostimulatory reactions/effects. 
     
     
         22 . A gene therapy method, comprising administering to a subject in need thereof a coupled peptide according to  claim 8  thereby to reduce side-effects of the gene therapy in the form of immunostimulatory reactions/effects. 
     
     
         23 . A gene therapy method, comprising administering to a subject in need thereof a coupled peptide according to  claim 9  thereby to reduce side-effects of the gene therapy in the form of immunostimulatory reactions/effects. 
     
     
         24 . A gene therapy method, comprising administering to a subject in need thereof a coupled peptide according to  claim 10  thereby to reduce side-effects of the gene therapy in the form of immunostimulatory reactions/effects. 
     
     
         25 . A gene therapy method, comprising administering to a subject in need thereof a coupled peptide according to  claim 11  thereby to reduce side-effects of the gene therapy in the form of immunostimulatory reactions/effects. 
     
     
         26 . A gene therapy method, comprising administering to a subject in need thereof a coupled peptide according to  claim 12  thereby to reduce side-effects of the gene therapy in the form of immunostimulatory reactions/effects.