Exosome transfer of nucleic acids to cells
Methods for introducing nucleic acids to cells via exosomes for use in gene modulation and therapy, such as for gene silencing and to introduce genetic material into cells to compensate for abnormal genes or to induce or repress a process in the recipient cell.
1. A method of transferring genetic material, comprising in this order:
a) isolating exosomes produced and released by a donor cell;
b) modifying the isolated exosomes to contain selected nucleic acid constructs; and
c) administering the exosomes containing the selected nucleic acid constructs to recipient cells, wherein administering the exosomes results in the transfer of the specific nucleic acid constructs from the exosomes to the cytoplasm of the recipient cells;
wherein the isolated exosomes do not contain any substantial amount of nucleic acids; and further
wherein the selected nucleic acid constructs are added to the exosomes that do not contain any substantial amount of nucleic acids.
2. The method of claim 1 , wherein the selected nucleic acid constructs are selected from the group consisting of mRNA, tRNA, rRNA, siRNA, microRNA, regulating RNA, non-coding and coding RNA, DNA fragments, and DNA plasmids.
3. The method of claim 1 , wherein the recipient cells are selected from the group consisting of cells from recipients having inherited diseases in hematopoietic, non-hematopoietic, stem cells, and organs.
4. The method of claim 1 , wherein the recipient cells comprise CD4T-cells for treatment of HIV infected T-cells.
5. The method of claim 1 , wherein the selected nucleic acid constructs are transferred to treat diseases, induce or repress cell death, change cellular ageing, induce tolerance, re-direct existing immune responses, or change intracellular activity or cellular behavior.
6. The method of claim 1 , wherein the exosomes are isolated by a method selected from the group consisting of isolation from a specific donor cell, isolation from a person with a particular disease or condition; and isolation from a genetically modified donor cell.
7. The method of claim 6 , further comprising genetically modifying the donor cell to erase production of a nucleic acid, or to up-regulate or down-regulate production of a nucleic acid.
8. The method of claim 1 , further comprising introducing the selected nucleic acid constructs into the exosomes using a method selected from the group consisting of transformation, transfection and microinjection of the selected genetic material directly into exosomes.
9. The method of claim 1 , wherein the selected nucleic acid constructs are administered to the recipient cells by a method selected from the group consisting of addition of the exosomes to cell cultures in vitro, intravenous injection of the exosomes, in vivo administration, and administration targeted to particular cells in the body.
10. The method of claim 1 , wherein the exosomes are isolated from a patient with a malignant disease; and are further processed to contain selected nucleic acid constructs, and then are reintroduced into the patient.
11. The method of claim 1 , wherein the exosomes are obtained from tumor tissue.
12. The method of claim 1 , wherein the exosomes are obtained from intestinal epithelial cells, lymphocytes, mast cells or dendritic cells.
13. The method of claim 1 , wherein the exosomes are obtained from in vitro growing cells.
14. The method of claim 13 , wherein the exosomes are isolated from cell culture supernatant.
15. The method of claim 1 , wherein the exosomes are isolated from body fluid.
16. The method of claim 15 , wherein the body fluid comprises blood plasma, urine or bronchoalveolar fluid.