Methods and compositions for treatment of pompe disease

A chimeric polypeptide comprising:

• (i) a GAA polypeptide comprising a mature acid alpha-glucosidase (GAA) polypeptide and (ii) an internalizing moiety that binds DNA with a KD of less than 100 nM and/or promotes transit across cellular membranes via an equilibrative nucleoside transporter 2 (ENT2) transporter;
• wherein the chimeric polypeptide does not comprise the full length, GAA precursor polypeptide set forth in SEQ ID NO:
• 1.

A nucleic acid construct, comprising a nucleotide sequence that encodes a mature GAA polypeptide, operably linked to a nucleotide sequence that encodes a internalizing moiety, wherein the nucleic acid construct encodes a chimeric polypeptide having the internalizing activity of the internalizing moiety, and wherein the nucleic acid construct does not encode a chimeric polypeptide comprising a GAA precursor polypeptide of approximately 110 kilodaltons.

A nucleic acid construct, comprising a nucleotide sequence that encodes a GAA polypeptide, operably linked to a nucleotide sequence that encodes a internalizing moiety, wherein the nucleic acid construct encodes a chimeric polypeptide comprising:

• (i) a mature acid alpha-glucosidase (GAA) polypeptide and (ii) an internalizing moiety that promotes transit across cellular membranes via an equilibrative nucleoside transporter 2 (ENT2) transporter and/or that binds DNA with a KD of less than 100 nM;
• wherein the chimeric polypeptide does not comprise the full length, GAA polypeptide set forth in SEQ ID NO:
• 1.

A method of treating Pompe disease in a subject in need thereof, comprising administering to the subject an effective amount of a chimeric polypeptide comprising:

• (i) a mature acid alpha-glucosidase (GAA) polypeptide and (ii) a internalizing moiety;
• wherein the chimeric polypeptide has acid alpha-glucosidase activity, and wherein the chimeric polypeptide does not comprise a GAA precursor polypeptide of approximately 110 kilodaltons.

A method of increasing acid alpha-glucosidase enzyme activity in a cell, comprising contacting the cell with a chimeric polypeptide comprising:

• (i) a mature acid alpha-glucosidase (GAA) polypeptide and (ii) a internalizing moiety;
• wherein the chimeric polypeptide does not comprise a GAA precursor polypeptide of approximately 110 kilodaltons.

A method of increasing acid alpha-glucosidase enzyme activity in a cell, comprising contacting the cell with a chimeric polypeptide comprising:

• (i) an acid alpha-glucosidase (GAA) polypeptide comprising a mature GAA polypeptide and (ii) an internalizing moiety that promotes transit across cellular membranes via an equilibrative nucleoside transporter 2 (ENT2) transporter and/or binds to DNA with a KD of less than 100 nM;
• wherein the chimeric polypeptide does not comprise the full length, GAA polypeptide set forth in SEQ ID NO:
• 1.

The method of any of claims 118-127, wherein said subject in need thereof is a subject having pathologic cytoplasmic glycogen accumulation prior to initiation of treatment with said chimeric polypeptide.

A method of decreasing glycogen accumulation in cytoplasm of muscle cells, comprising:

• contacting muscle cells with a chimeric polypeptide, which chimeric polypeptide comprises (i) a mature acid alpha-glucosidase (GAA) polypeptide and (ii) a internalizing moiety;
• wherein the chimeric polypeptide has acid alpha-glucosidase activity, and wherein the chimeric polypeptide does not comprise a GAA precursor polypeptide of approximately 110 kilodaltons.

A method of decreasing glycogen accumulation in cytoplasm and lysosomes of muscle cells, comprising contacting muscle cells with a chimeric polypeptide, which chimeric polypeptide comprises (i) a mature acid alpha-glucosidase (GAA) polypeptide and (ii) a internalizing moiety;

• wherein the chimeric polypeptide has acid alpha-glucosidase activity, and wherein the chimeric polypeptide does not comprise a GAA precursor polypeptide of approximately 110 kilodaltons.

A method of decreasing glycogen accumulation in cytoplasm, lysosomes, and autophagic vacuoles of muscle cells, comprising contacting muscle cells with a chimeric polypeptide, which chimeric polypeptide comprises (i) a mature acid alpha-glucosidase (GAA) polypeptide and (ii) a internalizing moiety;

• wherein the chimeric polypeptide has acid alpha-glucosidase activity, and wherein the chimeric polypeptide does not comprise a GAA precursor polypeptide of approximately 110 kilodaltons.

A method of decreasing glycogen accumulation in cytoplasm and lysosomes of muscle cells, comprising contacting muscle cells with a chimeric polypeptide, which chimeric polypeptide comprising:

• (i) an acid alpha-glucosidase (GAA) polypeptide and (ii) an internalizing moiety that promotes transit across cellular membranes via an equilibrative nucleoside transporter 2 (ENT2) transporter;
• wherein the chimeric polypeptide does not comprise the full length, GAA precursor polypeptide set forth in SEQ ID NO:
• 1.

A method of decreasing glycogen accumulation in cytoplasm, lysosomes, and autophagic vacuoles of muscle cells, comprising contacting muscle cells with a chimeric polypeptide, which chimeric polypeptide comprising:

• (i) an acid alpha-glucosidase (GAA) polypeptide and (ii) an internalizing moiety that promotes transit across cellular membranes via an equilibrative nucleoside transporter 2 (ENT2) transporter;
• wherein the chimeric polypeptide does not comprise the full length, GAA precursor polypeptide set forth in SEQ ID NO:
• 1.

A chimeric polypeptide comprising:

• (i) a mature acid alpha-glucosidase (GAA) polypeptide and (ii) an antibody or antigen binding fragment the promotes delivery of the chimeric polypeptide into cells;
• and wherein the chimeric polypeptide does not comprise a GAA precursor polypeptide of approximately 110 kilodaltons.

A chimeric polypeptide comprising:

• (i) an acid alpha-glucosidase (GAA) polypeptide and (ii) an antibody or antigen binding fragment the promotes delivery of the chimeric polypeptide into cells and can transit a cellular membranes via an equilibrative nucleoside transporter 2 (ENT2) transporter;
• wherein the chimeric polypeptide does not comprise the full length, GAA precursor polypeptide set forth in SEQ ID NO:
• 1.

A composition comprising a chimeric polypeptide comprising:

• (i) an acid alpha-glucosidase (GAA) polypeptide and (ii) an antibody or antigen binding fragment that binds to DNA and that promotes delivery of the chimeric polypeptide into cells;
• wherein the chimeric polypeptide does not comprise the full length, GAA precursor polypeptide set forth in SEQ ID NO:
• 1.

A chimeric polypeptide comprising the amino acid sequence set forth in SEQ ID NO:

• 11 or the amino acid sequence set forth in SEQ ID NO:
• 11, in the absence of one or more epitope tags.

A chimeric polypeptide comprising the amino acid sequence set forth in SEQ ID NO:

• 12 or the amino acid sequence set forth in SEQ ID NO:
• 12, in the absence of one or more epitope tags.