COMPOSITIONS AND METHODS FOR DELIVERING NUCLEIC ACID TO A CELL
Provided are novel compositions useful for delivering nucleic acids to cells. Also provided are methods for making and using such compositions.
1 . A method for preparing a liposome comprising a nucleic acid and a lipid component, the method comprising:
combining a lipid component and a nucleic acid component in a mixture comprising:
water;
a water-miscible organic solvent; and
a polyamine;
under conditions such that a liposome comprising the nucleic acid and the lipid component is formed.
2 . The method of claim 1 , wherein the polyamine is an oligoethyleneimine, a polyethyleneimine, or a polyaminoC 2 -C 10 alkane.
3 . The method of claim 2 , wherein the polyamine is selected from the group consisting of spermine, spermidine, and putrescine.
4 . The method of claim 3 , wherein the polyamine is spermine.
5 . (canceled)
6 . (canceled)
7 . (canceled)
8 . (canceled)
9 . The method of claim 1 , wherein the lipid component comprises a non-cationic lipid.
10 . The method of claim 1 , wherein the non-cationic lipid is a neutral lipid.
11 . The method of claim 10 wherein the neutral lipid comprises DOPC, DOPE, cholesterol, or PEG-DSG.
12 . The method of claim 9 , wherein the liposome further comprises a cationic lipid.
13 . The method of claim 12 , wherein the cationic lipid comprises DOTAP or DOSPA.
14 . The method of claim 1 , wherein the water-miscible organic solvent comprises methanol, ethanol, 1-propanol, or 2-propanol.
15 . (canceled)
16 . (canceled)
17 . (canceled)
18 . (canceled)
19 . The method of claim 18 , wherein the ratio of polyamine nitrogen to nucleic acid phosphate groups (N/P) is between about 0.8 and about 1.5.
20 . (canceled)
21 . (canceled)
22 . (canceled)
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24 . (canceled)
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26 . (canceled)
27 . A composition comprising a liposome in an aqueous medium, the liposome having an interior and an exterior, wherein the liposome comprises:
a nucleic acid;
a polyamine; and
a lipid component;
wherein the lipid component comprises a neutral phospholipid and essentially no cationic lipid, and wherein the nucleic acid and the lipid component are present at a ratio of from 5 nmol lipid per microgram of the nucleic acid to 20, 30, 40, 50, 60, 70, 80, 90, or 100 (optionally 5-20 or 30, preferably 10) nmol lipid per microgram of the nucleic acid, and wherein the liposome is from 30 to 500 nanometers in diameter.
28 . The composition of claim 27 , wherein the polyamine is oligoethyleneimine, polyethyleneimine, or a polyaminoC 2 -C 10 alkane.
29 . The composition of claim 27 , wherein the polyamine is selected from the group consisting of spermine, spermidine, and putrescine.
30 . The composition of claim 29 , wherein the polyamine is spermine.
31 . (canceled)
32 . (canceled)
33 . (canceled)
34 . (canceled)
35 . The composition of claim 27 , wherein the non-cationic lipid comprises DOPC, DOPE, cholesterol, or PEG-DSG.
36 . (canceled)
37 . The composition of claim 27 , wherein the lipid component comprises a cationic lipid and a non-cationic lipid.
38 . The composition of claim 37 , wherein the cationic lipid comprises DOTAP or DOSPA.
39 . The composition of claim 27 , wherein the liposome is from 70 to 300 nanometers in diameter.
40 . A method of using of a liposome of claim 27 , the method comprising:
attaching an internalizing antibody or a fragment thereof, which antibody or fragment binds to a specific cell surface antigen, to the exterior of the liposome, wherein the liposome with the antibody attached is internalized by a cell expressing at least 100,000 or at least 1,000,000 molecules of the antigen when contacted and incubated with the cell under internalizing conditions.
41 . (canceled)
42 . The composition of claim 27 , wherein the liposome further comprises an internalizing antibody or a fragment thereof attached to the exterior of the liposome, wherein the antibody or fragment binds to a specific cell surface antigen.
43 . A method of delivering a nucleic acid to a cell, the method comprising:
contacting the cell with the composition of claim 42 ;
wherein the liposome with the internalizing antibody or a fragment thereof attached is internalized by a cell expressing at least 100,000 or at least 1,000,000 molecules of the antigen when contacted and incubated with the cell under internalizing conditions.
44 . A method of treating a patient in need thereof with a nucleic acid, the method comprising administering to the patient an effective amount of a composition of claim 42 under conditions such that the patient is treated for a condition responsive to nucleic acid therapy.
45 . A composition prepared by the method of claim 1 , the composition comprising a liposome in an aqueous medium, wherein the liposome comprises:
a nucleic acid;
a polyamine; and
a lipid component.
46 . The composition of claim 27 or claim 45 , wherein the composition comprises no more than about 5 mol % or 1 mol % DOPE.
47 . (canceled)
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