Formulation of stabilized cationic transfection agent(s) /nucleic acid particles

1. A composition comprising stabilized particles comprising a complex of at least one cationic transfection agent and a nucleic acid, and at least one nonionic surface-active agent, wherein said agent stabilizes the size of said particles to less than or equal to 160 nanometers;

and wherein the at least one nonionic surface-active agent is chosen from (a) a polyoxyalkylene of the formula:

HO(CH 2 CH 2 O) a (CH(CH 3 )CH 2 O) b (CH 2 CH 2 O) c H

 wherein a, b, and c are, independently, a number from 20 to 100, and

(b) a polyethylene glycol comprising a dendritic benzyl polyether head.

2. The composition according to claim 1 , wherein the at least one cationic transfection agent and the nucleic acid are present in a charge ratio of between 1 and 6.

3. The composition according to claim 2 , wherein the at least one cationic transfection agent and the nucleic acid are present therein in a charge ratio of less than 4.

4. The composition according to claim 1 , wherein the at least one non-ionic surface-active agent is a compound of the formula:

OH(CH 2 CH 2 O) a (CH(CH 3 )CH 2 O) b (CH 2 CH 2 O) c H,

wherein a is 75, b is 30, and c is 75.

5. The composition according to claim 1 , wherein the at least one non-ionic surface-active agent is present at a concentration ranging from 0.01% to 10% weight/volume of said composition.

6. The composition according to claim 5 , wherein the at least one non-ionic surface active agent is present at a concentration ranging from 0.02% to 5% weight/volume of said composition.

7. The composition according to claim 1 , wherein the cationic transfection agent is a lipofectant.

8. The composition according to claim 7 , wherein the lipofectant is an amphiphilic molecule comprising at least one lipophilic region and a hydrophilic region.

9. The composition according to claim 7 , wherein the composition is a lipid mixture comprising cationic liposomes.

10. The composition according to claim 7 , wherein the lipofectant is a cationic lipid.

11. The composition according to claim 7 , wherein the lipofectant comprises at least one polyamine region of the formula:

wherein m is a number greater than or equal to 2 and n is a number greater than or equal to 1, wherein when n is greater than 1, m is independently a number greater than or equal to 2, and wherein said polyamine region is covalently bonded to a lipophilic region of a saturated or unsaturated hydrocarbon chain of cholesterol type, or a natural or synthetic lipid capable of forming lamellar or hexagonal phases.

12. The composition according to claim 11 , wherein the polyamine region is spermine or an analogue thereof that binds nucleic acid.

13. The composition according to claim 7 , wherein the lipofectant is of the formula:

wherein R is a lipophilic region represented by the formula:

wherein

X and X′ are, independently of each other, an oxygen atom, a methylene group —(CH 2 ) q — wherein q is 0, 1, 2 or 3, or an amino group —NH— or —NR′ wherein R′ is a C 1 to C 4 alkyl group;

Y and Y′ are, independently of each other, a methylene group, a carbonyl group or a group C═S;

R 3 , R 4 and R 5 are, independently of each other, a hydrogen atom or a substituted or unsubstituted C 1 to C 4 alkyl radical;

p is a number from 0 to 5;

R 6 is a cholesterol derivative or an alkylamino group —NR 1 R 2 wherein R 1 and R 2 are, independently of each other, a saturated or unsaturated C 12 to C 22 aliphatic radical, wherein said radical is linear or branched.

14. The composition according to claim 7 , wherein the lipofectant is a cationic lipid comprising at least one guanidinium or amidinium group or a mixture thereof.

15. The composition according to claim 1 , wherein the cationic transfection agent is a cationic polymer.

16. The composition according to claim 15 , wherein said cationic polymer is a compound of the formula (I):

wherein R is a hydrogen atom or a group of formula:

wherein n is a number from 2 to 10, and p and q are numbers wherein the sum p+q is such that the average molecular weight of the polymer ranges from 100 to 10 7 Da.

17. The composition according to claim 15 , wherein said cationic polymer is a polyethylene imine having an average molecular weight of 50,000 Da (PEI50K), 22,000 Da (PEI22K), or 800,000 Da (PEI800K).

18. The composition according to claim 1 , wherein said nucleic acid is a deoxyribonucleic acid.

19. The composition according to claim 1 , wherein said nucleic acid is a ribonucleic acid.

20. The composition according to claim 18 , wherein the nucleic acid is chemically modified.

21. The composition according to claim 1 , wherein said nucleic acid is an antisense nucleic acid.

22. The composition according to claim 1 , wherein said nucleic acid comprises a therapeutic gene.

23. The composition according to claim 1 , further comprising an adjuvant selected from the group consisting of dioleoylphosphatidylethanolamine (DOPE), oleoylpalmitoylphosphatidylethanolamine (POPE), di-stearoyl, -palmitoyl, and -myristoyl phosphatidylethanolamines optionally substituted with 1 to 3 N-methyl groups, phosphatidylglycerols, diacylglycerols, glycosyldiacylglycerols, cerebrosides, sphingolipids and asialogangliosides.

24. The composition according to claim 1 , further comprising a targeting element.

25. The composition according to claim 24 , wherein said targeting element is an antibody directed against a cell surface molecule; a membrane receptor ligand selected from the group consisting of insulin, transferrin, folic acid and a growth factor; cytokines; vitamins; lectins; proteins with an RGD unit; peptides containing a tandem array of RGD units wherein said peptides are linear or cyclic; polylysine peptides; natural ligand peptides; and synthetic ligand peptides.

26. A process for making the composition according to claim 1 , comprising forming particles by bringing at least one transfecting agent and a nucleic acid into contact in the presence of a sufficient quantity of at least one nonionic surface-active agent to stabilize the particles formed at a size of less than about 160 nm;

and wherein the at least one nonionic surface-active agent is chosen from

(a) a polyoxyalkylene of the formula:

HO(CH 2 CH 2 O) a (CH(CH 3 )CH 2 O) b (CH 2 CH 2 O) c H

 wherein a, b, and c are, independently, a number from 20 to 100, and

(b) a polyethylene glycol comprising a dendritic benzyl polyether head.

27. The process according to claim 26 , wherein the nucleic acid or the at least one transfecting agent is mixed before said contact with the at least one nonionic surface-active agent.

28. The composition according to claim 7 , wherein the lipofectant is of the formula:

wherein

R 1 , R 2 and R 3 are, independently of each other, a hydrogen atom or a group —(CH 2 ) q —NRR′, wherein each q is, independently, 1, 2, 3, 4, 5 or 6, and each R and R′ is, independently of each other, a hydrogen atom or a group —(CH 2 ) q′ —NH 2 , wherein q′ is independently, 1, 2, 3, 4, 5 or 6;

m, n and p are, independently of each other, a number between 0 and 6, wherein when n is greater than 1, each m is capable of taking different values and each R 3 is capable of having different meanings within their respective definitions;

R 4 represents a group of formula:

 wherein R 6 and R 7 are, independently of each other, a hydrogen atom or a saturated or unsaturated C 10 to C 22 aliphatic radical, with the proviso that R 6 and R 7 are not both hydrogen atoms;

u is a number from 0 to 10, wherein when u is greater than 1, R 5 , X, Y and r are capable of having different meanings within the different units;

X is oxygen, sulphur, or an amine group which is monoalkylated;

Y is a carbonyl group or a methylene group;

R 5 is hydrogen or a natural amino acid side chain which is optionally substituted; and

r is a number from 1 to 10, wherein when r is equal to 1, R 5 is a substituted or unsubstituted natural amino acid side chain, and when r is greater than 1, R 5 is hydrogen.

29. The composition according to claim 23 , wherein the cerebroside is a galactocerebroside.

30. The composition according to claim 23 , wherein the sphingolipid is a sphingomyelin.

31. The composition according to claim 1 , wherein said cationic transfection agent is lipofectamine, dioctadecylamidoglycyl spermine (DOGS), palmitoylphosphatidylethanolamine 5-carboxyspermylamide (DPPES), 2,5-bis(3-aminopropylamino)pentyl(dioctadecylcarbamoylmethoxy)acetate or 1,3-bis(3-aminopropylamino)-2-propyl(dioctadecylcarbamoylmethoxy)acetate,

{H 2 N(CH 2 ) 3 } 2 N(CH 2 ) 4 N{(CH 2 ) 3 NH 2 }(CH 2 ) 3 NHCH 2 COGlyN[(CH 2 ) 17 CH 3 ] 2 ,

H 2 N(CH 2 ) 3 NH(CH 2 ) 4 NH(CH 2 ) 3 NHCH 2 COGlyN[(CH 2 ) 17 CH 3 ] 2 ,

or

H 2 N(CH 2 ) 3 NH(CH 2 ) 4 NH(CH 2 ) 3 NHCH 2 COArgN[(CH 2 ) 17 CH 3 ] 2 .

32. The process according to claim 27 , wherein the at least one transfecting agent is a lipofectant.

33. The process according to claim 26 , wherein the at least one non-ionic surface-active agent is a compound of the formula:

OH(CH 2 CH 2 O) a (CH(CH 3 )CH 2 O) b (CH 2 CH 2 O) c H,

wherein a is 75, b is 30, and c is 75.

34. The process according to claim 26 , wherein the at least one non-ionic surface-active agent is present at a concentration ranging from 0.01% to 10% weight/volume of said composition.

35. The process according to claim 34 , wherein the surface active agent is present at a concentration ranging from 0.02% to 5% weight/volume of said composition.