DUALLY DERIVATIZED CHITOSAN NANOPARTICLES AND METHODS OF MAKING AND USING THE SAME FOR GENE TRANSFER IN VIVO

• US 20180344875A1
• Filed: 08/10/2018
• Published: 12/06/2018
• Est. Priority Date: 09/25/2013
• Status: Active Grant

• Current Assignee: enGene, Inc.
• Original Assignee: enGene, Inc.
• Inventors: GAO, Jun, HSU, Eric, CHEUNG, Anthony
• Granted Patent: US 10,456,481 B2
• Time in Patent Office: Days
• CPC Class Codes: A61K 48/005 characterised by an aspect of the 'active' part of the composition delivered, i.e. the nucleic acid delivered
• CPC Class Codes: A61K 9/5161 Polysaccharides, e.g. alginate, chitosan, cellulose derivatives; Cyclodextrin
• CPC Class Codes: C08B 37/003 Chitin, i.e. 2-acetamido-2-deoxy-(beta-1,4)-D-glucan or N-acetyl-beta-1,4-D-glucosamine; Chitosan, i.e. deacetylated product of chitin or (beta-1,4)-D-glucosamine; Derivatives thereof
• CPC Class Codes: C12N 15/88 using microencapsulation, e.g. using amphiphile liposome vesicle

Abstract

Provided herein is chitosan-derivative nanoparticle comprising chitosan functionalized with a cationic amino acid and a hydrophilic polyol; and methods of making and using same, e.g., for gene delivery in vivo.

Claims

1. A chitosan-derivative nanoparticle comprising chitosan functionalized with arginine (Arg) and a hydrophilic polyol (HP) of Formula VII:

2. The chitosan-derivative nanoparticle of claim 1,

comprising a hydrophilic polyol of Formula VII:

3. The chitosan-derivative nanoparticle of claim 2, wherein the hydrophilic polyol has a carboxyl group.

4. The chitosan-derivative nanoparticle of claim 3, wherein the hydrophilic polyol is threonic acid.

5. The chitosan-derivative nanoparticle of claim 2, wherein the hydrophilic polyol is a saccharide selected from the group consisting of glyceraldehyde, threose, erythrose, ribose, arabinose, xylose, lyxose, allose, glucose, altrose, mannose, gulose, idose, galactose, and talose.

6. The chitosan-derivative nanoparticle of claim 2, wherein the hydrophilic polyol is selected from the group consisting of 2,3-dihydroxylpropanoic acid;

2,3,4,5,6,7-hexahydroxylheptanal;

2,3,4,5,6-pentahydroxylhexanal;

2,3,4,5-tetrahydroxylhexanal; and

2,3-dihydroxylpropanal.

7. The chitosan-derivative nanoparticle of any of the preceding claims, comprising chitosan-nucleic acid polyplexes comprising chitosan molecules having an average molecular weight of less than 65 kDa before functionalization, on average less than 400 glucosamine monomer units, an N/P ratio of 2 to 20, an average hydrodynamic diameter of less than 500 nm, an average zeta potential of at least 0 mV at an acidic pH, a PDI of less than 0.5, and a nucleic acid concentration greater than 0.5 mg/ml, wherein said chitosan-nucleic acid polyplexes are substantially free of precipitated polyplex, and said chitosan-nucleic acid polyplexes are size stable in that they increase in average diameter by less than 100% at room temperature for at least 6 hours, at least 12 hours, at least 24 hours, or at least 48 hours.

8. The chitosan-derivative nanoparticle of claim 7, wherein said chitosan-nucleic acid polyplexes increase in average diameter by less than 50% at room temperature for at least 6 hours, at least 12 hours, at least 24 hours, or at least 48 hours.

9. The chitosan-derivative nanoparticle of any of the preceding claims, wherein the nanoparticle has an Arg final functionalization degree:

HP final functionalization degree ratio of between about 1;

1 to about 10;

1.

10. The chitosan-derivative nanoparticle of claim 9, wherein the Arg final functionalization degree:

HP final functionalization degree ratio is between about 3;

1 to about 7;

1.

11. The chitosan-derivative nanoparticle of claim 10, wherein the Arg final functionalization degree:

HP final functionalization degree ratio is about 5;

1.

12. A composition comprising the nanoparticle according to any one of claims 1-11, wherein said chitosan is complexed with a nucleic acid to form a dually derivatized (DD) chitosan nucleic acid polyplex.

13. The composition according to claim 12, wherein said nucleic acid is DNA or RNA.

14. The composition according to claim 12 or claim 13, wherein the amine to phosphate ratio of said DD-chitosan nucleic acid polyplex is between 2 to 100.

15. The composition according to claim 14, wherein the amine to phosphate ratio of said DD-chitosan nucleic acid polyplex is between 2 to 50.

16. The composition according to claim 15, wherein the amine to phosphate ratio of said DD-chitosan nucleic acid polyplex is between 2 to 30.

17. The composition according to claim 16, wherein the amine to phosphate ratio of said DD-chitosan nucleic acid polyplex is between 2 to 15.

18. A method of delivering a nucleic acid molecule to a cell comprising contacting said cell with a composition according to any one of claims 12 to 17.

19. The method of claim 18, wherein said cell is in vivo.