Acid alpha-glucosidase and fragments thereof

Abstract

Targeted acid alpha-glucosidase therapeutics that localize to the lysosome are provided. The targeted therapeutics include a therapeutic agent, GAA, and a targeting moiety that binds a receptor on an exterior surface of the cell, permitting proper subcellular localization of the targeted therapeutic upon internalization of the receptor. Nucleic acids, cells, and methods relating to the practice of the invention are also provided.

1 . A polypeptide comprising an amino acid sequence at least 50% identical to amino acid residues 70-790 of human acid alpha-glucosidase (GAA) or a fragment thereof and a targeting domain that binds an extracellular domain of a receptor on the surface of a target cell, wherein the polypeptide is not associated with a sequence at least 50% identical to amino acid residues 880-952 of human GAA.  
     
     
         2 . (canceled)  
     
     
         3 . The polypeptide of  claim 1 , wherein the targeting domain permits localization of the polypeptide in a human lysosome.  
     
     
         4 - 16 . (canceled)  
     
     
         17 . A targeted therapeutic comprising a therapeutic agent comprising at least a portion of human acid alpha-glucosidase (GAA) and a peptide tag, wherein the peptide tag is directly or indirectly linked to an amino acid residue selected from the group consisting of amino acid residues 68, 69, 70, 71, 72, 789, 790, 791, 792, and 793 of human GAA.  
     
     
         18 . The targeted therapeutic of  claim 17 , wherein the amino acid residue is selected from the group consisting of amino acid residues 68-72 of human GAA.  
     
     
         19 . The targeted therapeutic of  claim 17 , wherein the therapeutic agent comprises amino acid residues 70-952 of human GAA.  
     
     
         20 . The targeted therapeutic of  claim 19 , wherein the peptide tag is linked to amino acid residue 70 of human GAA.  
     
     
         21 . The targeted therapeutic of  claim 17 , wherein the peptide tag is a ligand for an extracellular receptor.  
     
     
         22 . The targeted therapeutic of  claim 21 , wherein the peptide tag is a targeting domain that binds an extracellular domain of a receptor on the surface of a target cell and, upon internalization of the receptor, permits localization of the therapeutic agent to a human lysosome.  
     
     
         23 . The targeted therapeutic of  claim 22 , wherein the targeting domain comprises a urokinase-type plasminogen receptor moiety.  
     
     
         24 . The targeted therapeutic of  claim 22 , wherein the peptide tag comprises amino acid residues 48-55 of human IGF-II.  
     
     
         25 . The targeted therapeutic of  claim 22 , wherein the peptide tag comprises amino acid residues 8-28 and 41-61 of human IGF-II, or a sequence variant thereof that binds the human cation-independent mannose-6-phosphate receptor.  
     
     
         26 . The targeted therapeutic of  claim 22 , wherein the peptide tag comprises amino acid residues 8-87 of human IGF-II or a sequence variant thereof, wherein the sequence variant binds the human cation-independent mannose-6-phosphate receptor.  
     
     
         27 . The targeted therapeutic of  claim 26 , wherein the peptide tag comprises an R68A mutation of human IGF-II.  
     
     
         28 . The targeted therapeutic of  claim 24 , wherein the targeting domain comprises a C-terminally truncated form of human IGF-II or a sequence variant thereof, wherein the truncation begins at position 62.  
     
     
         29 . The targeted therapeutic of  claim 17 , further comprising a spacer between the therapeutic agent and the peptide tag.  
     
     
         30 . The targeted therapeutic of  claim 29 , wherein the spacer comprises glycine residues.  
     
     
         31 . The polypeptide of  claim 29 , wherein the spacer comprises an α-helical structure.  
     
     
         32 . The polypeptide of  claim 29 , wherein the spacer comprises an amino acid sequence at least 50% identical to Gly-Gly-Gly-Gly-Thr-Val-Gly-Asp-Asp-Asp-Asp-Lys (SEQ ID NO:1).  
     
     
         33 . A nucleic acid sequence comprising an open reading frame of a polypeptide comprising an amino acid sequence at least 50% identical to amino acid residues 70-790 of human acid alpha-glucosidase (GAA) or a fragment thereof, wherein the open reading frame does not include an amino acid sequence at least 50% identical to amino acid residues 880-952 of human GAA.  
     
     
         34 . (canceled)  
     
     
         35 . (canceled)  
     
     
         36 . The nucleic acid sequence of  claim 33 , wherein the polypeptide further comprises a targeting domain that binds an extracellular domain of a receptor on the surface of a target cell and, upon internalization of the receptor, permits localization of the polypeptide to a human lysosome.  
     
     
         37 - 46 . (canceled)