COMPOSITIONS AND METHODS FOR LESS IMMUNOGENIC PROTEIN-LIPID COMPLEXES

• US 20090053297A1
• Filed: 07/28/2008
• Published: 02/26/2009
• Est. Priority Date: 11/30/2000
• Status: Active Grant

• Current Assignee: The Research Foundation for The State University of New York (State University of New York)
• Original Assignee: The Research Foundation for The State University of New York (State University of New York)
• Inventors: Ramani, Karthik, Straubinger, Robert, Miclea, Razvan D., Balu-Iyer, Sathy V.
• Granted Patent: US 8,110,218 B2
• Time in Patent Office: Days
• US Class Current: 424/450
• CPC Class Codes: A61K 31/74 Synthetic polymeric materials
• CPC Class Codes: A61K 38/37 Factors VIII
• CPC Class Codes: A61K 9/1075 Microemulsions or submicron emulsions; Preconcentrates or solids thereof; Micelles, e.g. made of phospholipids or block copolymers A61K9/0026 takes precedence
• CPC Class Codes: A61K 9/1271 Non-conventional liposomes, e.g. PEGylated liposomes, liposomes coated with polymers liposome as conjugate A61K47/6911
• CPC Class Codes: A61K 9/1272 with substantial amounts of non-phosphatidyl, i.e. non-acylglycerophosphate, surfactants as bilayer-forming substances, e.g. cationic lipids with cholesterol as the only non-phosphatidyl surfactant A61K9/127; lipids as modifying agent A61K47/543
• CPC Class Codes: A61K 9/1274 Non-vesicle bilayer structures, e.g. liquid crystals, tubules, cubic phases, cochleates; Sponge phases

Abstract

The present invention provides compositions and methods for reducing the immunogenicity and increasing the circulating half-life of therapeutic proteins such as Factor VIII. The compositions comprise lipidic structures such as liposomes, micelles and cochleates comprising a negatively charged lipid and polyethylene glycol derivatized phosphatidyl ethanolamine.

Claims

1. A pharmaceutical composition comprising lipidic structures, wherein the lipidic structures comprise:

a. phosphatidyl ethanolamine (PE) derivatized with polyethylene glycol (PEG) (PEG-derivatized PE);

b. an amphipathic lipid selected from the group consisting of phosphatidylcholine (PC), phosphatidylglycerol (PG), and combinations thereof, and, optionally, PE;

c. a negatively charged lipid selected from the group consisting of phosphatidylserine (PS), phosphatidic acid (PA) and combinations thereof; and

d. Factor VIII,

wherein the immunogenicity of Factor VIII is reduced over the immunogenicity of free Factor VIII.

2. The pharmaceutical composition of claim 1, wherein the lipidic structure is selected from the group consisting of liposomes, micelles, cochleates, and combinations thereof.

3. The pharmaceutical composition of claim 1, wherein the ratio of the amphipathic lipid to the negatively charged lipid is between 50:

50 to 90;

10, and the amount of PEG derivatized PE is from 1-15 mole %.

4. The pharmaceutical composition of claim 1, wherein the amphipathic lipid is PC and the negatively charged lipid is PS.

5. The pharmaceutical composition of claim 2, wherein the ratio of PC to PS is 70:

30.

6. The pharmaceutical composition of claim 1, wherein the amphipathic lipid is PC and PE and the negatively charged lipid is PS and the ratio of PC:

PS;

PE is 80;

10;

10 or 70;

10;

20.

7. The pharmaceutical composition of claim 2, wherein the ratio of PC:

PS;

PEG-derivatized PE is 70;

30;

15.

8. The pharmaceutical composition of claim 1, wherein the liposomes further comprise PE that is not derivatized with PEG in the amount of 1-10 mole %.

9. The pharmaceutical composition of claim 1, wherein the two acyl chains of the phospholipids have between 12-22 carbon atoms in each chain, wherein the two acyl chain have the same or different number of carbon atoms.

10. The pharmaceutical composition of claim 8, wherein the acyl chain is selected from the group consisting of myristic acid, palmitic acid, stearic acid.

11. The pharmaceutical composition of claim 1, further comprising 0.5-30 mole % cholesterol.

12. The pharmaceutical composition of claim 1 wherein there is a spacer of 6-12 carbon atoms between the PE and the PEG moieties of the PEG derivatized PE.

13. A method of making the pharmaceutical composition of claim 1, comprising:

a. preparing liposomes comprising PC, PS and activated PE;

b. adding Factor VIII; and

c. adding activated PEG such that the activated PEG binds to the activated PE.

14. The method of claim 18, wherein a spacer arm comprising 6-12 carbon atoms is attached to either PEG or PE.

15. A method of reducing the immunogenicity of an administered Factor VIII protein comprising the steps of:

a. preparing a formulation wherein the protein is associated with or incorporated into lipidic structures selected from the group consisting of liposomes, micelles, cochleates, and combinations thereof, wherein the lipid structures comprise a negatively charged lipid and PEG derivatized PE; and

b. administering the formulation to an individual suffering from a bleeding disorder,

wherein the immunogenicity of the administered protein in less than the immunogenicity of free Factor VIII protein and wherein the circulating half life of the administered protein is more than the circulating half-life of free Factor VIII protein.