Oligonucleotide-facilitated coalescence
View Patent ↗Coalescence of cells or other membrane-bound entities is facilitated by anchoring an outwardly projecting first oligonucleotide in one member and an outwardly projecting second oligonucleotide, complementary to the first, in a second member and incubating under hybridizing conditions. Liposomes may be coalesced with cells to deliver hydrophilic agents thereto, such as DNA probes or drugs. Kits containing complementary oligonucleotides containing hydrophobic anchoring moieties may be used.
1. A method for fusing a first cell with a member selected from the group consisting of a second cell, lipid bilayer vesicles and liposomes, comprising:
(a) incorporating into the plasma bilayer membrane of said first cell an outwardly projecting first anchored oligonucleotide containing a first hydrophobic anchoring moiety attached to the 5′ or 3′ terminus of the first oligonucleotide,
(b) incorporating into the bilayer membrane of said member an outwardly projecting second anchored nucleotide, complementary to said first anchored nucleotide, containing a second hydrophobic anchoring moiety attached to the 3′ or 5′ terminus of the second oligonucleotide,
(c) mixing said first cell and said member under conditions promoting hybridization of said first and second outwardly projecting anchored oligonucleotides, and
(d) incubating the mixture of said first cell and said member under conditions producing fusion thereof.
2. The method according to claim 1 , wherein the conditions of step (d) include a fusion aid.
3. The method according to claim 1 , wherein the member is a liposome containing an entrapped hydrophilic substance to be inserted into said first cell.
4. The method according to claim 3 , wherein said hydrophilic substance is selected from the group consisting of DNA, drugs, therapeutic agents, chemotherapeutic agents, antisense agents, stains and oligonucleotide probes.
5. The method according to claim 1 , wherein each of the first and second terminal hydrophobic anchoring moieties is selected from the group consisting of cholesterol, a fatty acid, a hydrophobic peptide and a lipid.
6. The method according to claim 1 , wherein at least one of said first and second terminal hydrophobic anchoring moieties is a hemifusion destabilizer.
7. The method according to claim 1 , wherein said incubation is carried out in the presence of a hemifusion destabilizing agent.
8. The method according to claim 2 , wherein said first and second members are cells.
9. The method according to claim 2 wherein the fusion aid is selected from the group consisting of osmotic shock, electric shock, and the addition of calcium ions.
10. The method according to claim 6 wherein said at least one anchoring moiety spans both bilayer membrane layers.
11. The method according to claim 10 wherein both said first and second anchoring moieties span both bilayer membrane layers.
12. The method according to claim 6 , wherein the conditions of step (d) include a fusion aid.
13. The method according to claim 1 , wherein the member is a liposome, and wherein the second hydrophobic anchoring moiety is a hemifusion destabilizer that spans both bilayer membrane layers.
14. The method according to claim 13 wherein the conditions permitting fusion are the conditions promoting hybridization.