Mannose-6-phosphate bearing peptides fused to lysosomal enzymes

Abstract

A targeted therapeutic including a lysosomal enzyme and a lysosomal targeting moiety that is a peptide containing at least one N-linked glycosylation site. Methods of producing the targeted therapeutic may include nucleotide acids encoding the same and host cells co-expressing GNPT. Pharmaceutical compositions comprising the targeted therapeutic and methods of using the same to treat a lysosomal storage disease.

1 . A targeted therapeutic comprising:
 a lysosomal enzyme; and   a lysosomal targeting moiety, wherein the lysosomal targeting moiety is a peptide containing at least one N-linked glycosylation site.   
     
     
         2 . (canceled) 
     
     
         3 . The targeted therapeutic of  claim 1 , wherein the peptide contains N-linked glycosylation containing a M6P group. 
     
     
         4 . (canceled) 
     
     
         5 . The targeted therapeutic of  claim 1 , wherein the lysosomal enzyme is an N-Acetylglucosaminidase (Naglu) protein. 
     
     
         6 . The targeted therapeutic of  claim 1 , wherein the Naglu protein comprises an amino acid sequence at least 80% identical to SEQ ID NO.: 1. 
     
     
         7 .- 9 . (canceled) 
     
     
         10 . The targeted therapeutic of  claim 1 , wherein the lysosomal targeting moiety is a peptide selected from the group consisting of prosaposin, cellular repressor of E1A-stimulate genes, leukemia inhibitory factor, and fragments thereof. 
     
     
         11 .- 12 . (canceled) 
     
     
         13 . The targeted therapeutic of  claim 10 , wherein the peptide comprises a sequence at least 95% identical to SEQ ID NO. 4, 6, or 9. 
     
     
         14 .- 17 . (canceled) 
     
     
         18 . The targeted therapeutic of  claim 1 , wherein the lysosomal targeting moiety and the lysosomal enzyme is fused via a linker. 
     
     
         19 . The targeted therapeutic of  claim 1 , wherein the linker comprises a sequence of 
       
         
           
                 
               
                   (SEQ ID NO.: 15) 
                 
                   GAPGGGGGAAAAAGGGGGGAPGGGGGAAAAAGGGGGGAPGGGGGAAAAAG 
                 
                     
                 
                   GGGGGAP.  
                 
             
                
                
                
                
               
            
           
         
       
     
     
         20 . (canceled) 
     
     
         21 . The targeted therapeutic of  claim 1 , wherein the fusion protein is at least 95% identical to the amino acid sequence of SEQ ID NO. 18, 19 or 20. 
     
     
         22 .- 24 . (canceled) 
     
     
         25 . The targeted therapeutic of  claim 1 , wherein the fusion protein comprises a sequence at least 95% identical to the amino acid sequence of SEQ ID NO. 23, 24 or 25. 
     
     
         26 . (canceled) 
     
     
         27 . A nucleic acid encoding the targeted therapeutic fusion protein of  claim 1 . 
     
     
         28 . A vector comprising the nucleic acid sequence of  claim 27 . 
     
     
         29 . A host cell comprising the vector of  claim 28 . 
     
     
         30 .- 33 . (canceled) 
     
     
         34 . The host cell of  claim 29 , wherein the host cell co-expresses N-acetylglucosamine-1-phosphoTransferase (GNPT). 
     
     
         35 . The host cell of  claim 34 , wherein the host cell comprises an exogenous nucleic acid encoding GNPT. 
     
     
         36 . The host cell of  claim 34 , wherein the host cell has activated expression of endogenous GNPT. 
     
     
         37 . A method for producing a targeted therapeutic fusion protein, comprising:
 a) culturing cells comprising a nucleic acid encoding a fusion protein of  claim 1 , wherein the cells co-express N-acetylglucosamine-1-phosphoTransferase (GNPT); and   b) recovering the fusion protein produced by the cells.   
     
     
         38 . A pharmaceutical composition comprising the targeted therapeutic of  claim 1 , and a pharmaceutical acceptable carrier. 
     
     
         39 . A method of treating a lysosomal storage disease comprising administering to a subject in need of treatment a pharmaceutical composition of  claim 38 . 
     
     
         40 . The method of  claim 39 , wherein the lysosomal storage disease is Sanfilippo syndrome type B. 
     
     
         41 . (canceled)