Methods and materials for treatment of pompe's disease

• US 9,249,399 B2
• Filed: 03/14/2013
• Issued: 02/02/2016
• Est. Priority Date: 03/15/2012
• Status: Active Grant

• Current Assignee: Oxyrane UK Limited
• Original Assignee: Oxyrane UK Limited
• Inventors: Vervecken, Wouter, Ludo Stout, Jan Robert, Pynaert, Gwenda Nolla, Telesphore Alida Maria Piens, Kathleen Camilla
• Primary Examiner(s): Mondesi, Robert
• Assistant Examiner(s): EPSTEIN, TODD MATTHEW
• Application Number:
  US13/831,368
• Publication Number:
  US 20130243746A1
• Time in Patent Office: 1,055 Days
• Field of Search: None
• US Class Current: 1/1
• CPC Class Codes: A61K 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• CPC Class Codes: A61K 38/43 Enzymes; Proenzymes; Derivatives thereof
• CPC Class Codes: A61K 38/47 acting on glycosyl compounds (3.2), e.g. cellulases, lactases
• CPC Class Codes: A61K 47/64 Drug-peptide, drug-protein or drug-polyamino acid conjugates, i.e. the modifying agent being a peptide, protein or polyamino acid which is covalently bonded or complexed to a therapeutically active agent peptidic linkers A61K47/65
• CPC Class Codes: A61P 21/00 Drugs for disorders of the muscular or neuromuscular system
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: C07K 14/65 Insulin-like growth factors, i.e. somatomedins, e.g. IGF-1, IGF-2
• CPC Class Codes: C07K 2319/70 containing domain for protein-protein interaction
• CPC Class Codes: C12N 9/2408 acting on alpha -1,4-glucosidic bonds
• CPC Class Codes: C12N 9/2451 acting on alpha-1,6-glucosidic bonds
• CPC Class Codes: C12N 9/62 from Aspergillus
• CPC Class Codes: C12P 21/02 having a known sequence of two or more amino acids, e.g. glutathione
• CPC Class Codes: C12P 21/06 produced by the hydrolysis of a peptide bond, e.g. hydrolysate products preparing foodstuffs by protein hydrolysis A23J3/00
• CPC Class Codes: C12Y 302/01003 Glucan 1,4-alpha-glucosidase (3.2.1.3), i.e. glucoamylase
• CPC Class Codes: C12Y 302/0102 Alpha-glucosidase (3.2.1.20)

Abstract

This document relates to molecular complexes having acid alpha glucosidase activity and at least one modification that results in enhanced ability of the molecular complex to be transported to the interior of a mammalian cell.

Claims

1. A method for making a molecular complex having acid alpha glucosidase (GAA) activity, said method comprising contacting a GAA protein having at least 90% sequence identity to the amino acid sequence set forth in SEQ ID NO:

1 with a protease having at least 85% sequence identity to the amino acid sequence set forth in SEQ ID NO;

8, wherein said protease cleaves said protein at one or more sites between amino acid 60 and amino acid 65 of SEQ ID NO;

1 to produce a molecular complex having GAA activity and comprising at least two polypeptides.

2. The method of claim 1, wherein said contacting is performed in vitro.

3. The method of claim 1, further comprising proteolysis of the protein at one or more sites between amino acid 719 and amino acid 746 of the amino acid sequence set forth in SEQ ID NO:

1.

4. The method of claim 1, further comprising proteolysis of the protein at one or more sites between amino acid 137 and amino acid 151 of the amino acid sequence set forth in SEQ ID NO:

1.

5. The method of claim 1, wherein the contacting occurs in a recombinant fungal cell.

6. The method of claim 5, wherein the fungal cell is a cell of Yarrowia lipolytica, Arxula adeninivorans, or a methylotrophic yeast.

7. The method of claim 6, wherein the methylotrophic yeast is a cell of a genus selected from the group consisting of Candida, Hansenula, Oogataea, Pichia, and Torulopsis.

8. The method of claim 1, further comprising altering the molecular complex with at least one modification that results in enhanced ability of the molecular complex to be transported to the interior of a mammalian cell, wherein:

the at least one modification comprises the recognition domain of human insulin-like growth factor II;

or

at least one of said polypeptides comprises one or more phosphorylated N-glycans and wherein said modification comprises uncapping and demannosylation of at least one phosphorylated N-glycan.

9. The method of claim 8, wherein at least 40% of the N-glycans on said polypeptide are uncapped and demannosylated.

10. The method of claim 8, wherein at least 60% of the N-glycans on said polypeptide are uncapped and demannosylated.

11. The method of claim 8, wherein at least 80% of the N-glycans on said polypeptide are uncapped and demannosylated.

12. The method of claim 8, wherein at least 90% of the N-glycans on said polypeptide are uncapped and demannosylated.