Multiplexed supramolecular assemblies for non-viral delivery of genetic material

Patent metadata

Field	Value
Publication number	US-9572895-B2
Application number	US-201113814688-A
Country	US
Kind code	B2
Filing date	Aug 5, 2011
Priority date	Aug 6, 2010
Publication date	Feb 21, 2017
Grant date	Feb 21, 2017

Abstract

The invention provides a composition comprising two or more of a) genetic material; b) a condensing agent; c) a membrane destabilizing agent; d) a targeting/stabilizing agent, and optionally e) an ionic cross-linking agent. The components of the composition can self-assemble to form particles via electrostatic and/or lipophilic interactions. Methods of making and using the composition are also provided. The compositions can be used, for example, to transfer genetic material to cells in vitro or in vivo.

First claim

What is claimed is:

1. A particulate composite composition, wherein the particulate composite is a particle comprising:
a) genetic material that includes DNA or RNA;
b) an amine-containing condensing agent that binds to the genetic material through electrostatic interactions, wherein the condensing agent comprises a compound of Formula I:

wherein each m is independently 1 or 2; p is 1, 2, 3, 4, or 5; [linker] is an ether, ester, amide, carbonyl, or direct bond; and alkyl is a straight chain or branched C 4 -C 30 (alkyl) group optionally having one, two or three sites of unsaturation;
c) a membrane destabilizing agent that binds to the genetic material through electrostatic interactions; and
d) a targeting/stabilizing agent wherein the targeting/stabilizing agent that binds to the condensing agent, the membrane destabilizing agent, or both, and the targeting/stabilizing agent comprises a lipid component and a ligand linked together by a polyethylene glycol (PEG) chain; wherein particle has a diameter of about 40 nm to about 4 μm.

2. The composition of claim 1 wherein the DNA or RNA in the particle has about 10 to about 5000 base pairs.

3. The composition of claim 1 wherein the condensing agent includes both cationic and lipophilic properties.

4. The composition of claim 1 wherein the condensing agent further comprises chitosan, chitosan having methyl substitutions on nitrogen, chitosan conjugated to one or more straight chain or branched C 4 -C 30 (alkyl) groups optionally having one, two or three sites of unsaturation, or chitosan having methyl substitutions on nitrogen and one or more straight chain or branched C 4 -C 30 (alkyl) groups optionally having one, two or three sites of unsaturation, conjugated to the chitosan.

5. The composition of claim 4 wherein the C 4 -C 30 (alkyl) groups are conjugated to the chitosan through ether, ester, amide, or carbonyl groups, or through direct bonds.

6. The composition of claim 1 wherein the membrane destabilizing agent is a cationic membrane destabilizing agent.

7. The composition of claim 1 wherein the membrane destabilizing agent comprises a polypeptide that includes about 4 to about 200 amino acids.

8. The composition of claim 1 wherein the ligand of the targeting/stabilizing agent comprises a small molecule, a protein, a glycoprotein, a peptide, a sugar, a saccharide, or an aptamer.

9. The composition of claim 8 wherein the ligand of the targeting/stabilizing agent comprises one or more of folate, galactose, mannose, biotin, cysteamine, transferrin, lactoferrin, arginine-glycine-aspartic acid (RGD), chemiluminescence (CL) aptamer, prostate-specific membrane antigen (PSMA) aptamer, or nucleolin aptamer.

10. The composition of claim 1 wherein the lipid component of the targeting/stabilizing agent comprises a carbon chain that has about 8 to about 30 carbon atoms, which optionally includes one, two or three sites of unsaturation.

11. The composition of claim 1 wherein the polyethylene glycol (PEG) chain of the targeting/stabilizing agent comprises a PEG moiety having the formula —O(CH 2 CH 2 O) x — where x is 2 to about 1500.

12. The composition of claim 1 wherein the genetic material, the condensing agent, the membrane destabilizing agent, and the targeting/stabilizing agent self-assemble to form the particle when combined in an aqueous or alcoholic solvent system.

13. The composition of claim 1 wherein the composition comprises particles of about 150 nm in diameter to about 300 nm in diameter.

14. The composition of claim 1 wherein the composition includes an ionic cross-linker electrostatically bonded to the condensing agent.

15. The composition of claim 14 wherein the ionic cross-linker is a tripolyphosphate salt, heparin, hyaluronic acid, poly(γ-glutamic acid), or a combination thereof.

16. A method to transfer genetic material into a cell comprising contacting a cell with an effective amount of a composition as described in claim 1 , wherein the particulate composite of the composition enters the cell and releases the genetic material, thereby transferring the genetic material to the cell.

17. The method of claim 16 wherein the contacting is in vivo.

18. The method of claim 16 wherein the genetic material is RNA and the composition improves RNA interference efficiency in vivo, compared to procedures employing polyethyleneimine or Lipofectamine2000 transfection reagent.

19. The method of claim 16 wherein the composition is delivered to a mammal by oral, intranasal, pulmonary, intraperitoneal, intravenous, or intramuscular administration.

20. A particulate composite composition, wherein the particulate composite is a particle comprising:
a) genetic material that includes DNA or RNA;
b) an amine-containing condensing agent that binds to the genetic material through electrostatic interactions, wherein the condensing agent comprises a compound of Formula I:

wherein each m is independently 1 or 2; p is 1, 2, 3, 4, or 5; [linker] is an ether, ester, amide, carbonyl, or direct bond; and alkyl is a straight chain or branched C 4 -C 30 (alkyl) group optionally having one, two or three sites of unsaturation, or the condensing agent comprises chitosan having methyl substitutions on nitrogen, chitosan conjugated to one or more straight chain or branched C 4 -C 30 (alkyl) groups optionally having one, two or three sites of unsaturation, or chitosan having methyl substitutions on nitrogen and one or more straight chain or branched C 4 -C 30 (alkyl) groups optionally having one, two or three sites of unsaturation, conjugated to the chitosan;
c) a membrane destabilizing agent that binds to the genetic material through electrostatic interactions; and
d) a targeting/stabilizing agent wherein the targeting/stabilizing agent that binds to the condensing agent, the membrane destabilizing agent, or both, and the targeting/stabilizing agent comprises a lipid component and a ligand linked together by a polyethylene glycol (PEG) chain; wherein particle has a diameter of about 40 nm to about 4 μm.

Assignees

• Cheng Jianjun
• Gabrielson Nathan
• Lichen Yin
• Univ Illinois

Inventors

• Cheng Jianjun
• Gabrielson Nathan
• Lichen Yin

Classifications

• A61K48/00

  Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy · CPC title

• A61K9/1075

  Microemulsions or submicron emulsions; Preconcentrates or solids thereof; Micelles, e.g. made of phospholipids or block copolymers (A61K9/0026 takes precedence) · CPC title

• A61K9/5123

  Organic compounds, e.g. fats, sugars · CPC title

• A61K48/0041

  the non-active part being polymeric · CPC title

• A61K48/0025 Primary

  wherein the non-active part clearly interacts with the delivered nucleic acid · CPC title