Directed apoptosis in COX-2 overexpressing cancer cells through expression targeted gene delivery
View Patent ↗The present invention provides methods and constructs for selectively expressing an Apoptosis-Inducing Gene (AIG) in a population of cells that overexpress cyclooxygenase-2 (COX-2) to induce apoptosis in the cell. To achieve this goal a chimeric gene construct is used that comprises a cyclooxygenase-2 promoter (COX-2 promoter) that is operably linked to at least one AIG such that the COX-2 promoter is activated in cells that overexpress COX-2, thereby resulting in transcription and translation of the AIG, which in turn activates apoptosis in the cell. Thus, apoptosis is selectively induced in only those cells capable of overexpressing COX-2.
1. A method of causing apoptosis in a diseased cell that overexpresses a protein, the method comprising delivering a chimeric gene construct to the cell, wherein the chimeric gene construct comprises an upstream regulatory element, which is activated by the protein overexpressed in the cell, operably linked to genes that encode Caspase-3 and Caspase-9, and expressing Caspase-3 and Caspase-9 in the cell that overexpresses the protein, wherein the combined production of Caspase-3 and Caspase-9 causes the cell to undergo apoptosis.
2. The method of claim 1 , wherein the step of delivering the chimeric gene construct to the cell comprises delivering the chimeric gene construct by a non-viral delivery vehicle.
3. The method of claim 2 , wherein the non-viral delivery vehicle is selected from the group consisting of poly(ethylenimine), lipofectin, lipofectamine, polylysine, and alginate.
4. The method of claim 2 , wherein the non-viral delivery vehicle is poly(ethylenimine).
5. The method of claim 1 , wherein the step of delivering the chimeric gene construct to the cell comprises delivering the chimeric gene construct by a viral delivery vehicle.
6. The method of claim 5 , wherein the viral delivery vehicle is a viral vector selected from the group consisting of an adenovirus, an adeno-associated virus, a lentivirus and a retrovirus.
7. The method of claim 1 , wherein the upstream regulatory element is selected from the group consisting of an cyclooxygenase promoter, a tumor necrosis factor promoter, an interleukin-2 promoter, an interleukin-21 promoter, and an interleukin-23 promoter.
8. The method of claim 1 , wherein the protein which activates the upstream regulatory element is selected from the group consisting of a tumor necrosis factor, an interleukin-2, an interleukin-21, and an interleukin-23.
9. The method of claim 1 , wherein the diseased cell is selected from the group consisting of a cancer cell, a tumor cell, and an inflammatory cell.
10. The method of claim 1 , wherein the method further comprises expressing at least one additional gene for production of an apoptosis agent selected from the group consisting of Caspase-1, Caspase-2, Caspase-4, Caspase-5, Caspase-6, Caspase-7, Caspase-8, Caspase-10, Granzyme A, Granzyme B, Fas ligand, TRAIL and APO3L.
11. The method of claim 1 , further comprising delivering to the cell an oligomerizing agent selected from the group consisting of a cyclosporin A-type ligand, a tetracycline, a steroid ligand, a Tet-On/Tet-Off system, an ecdysone-dimerizer system, an antiprogestin-dimerizer system, and a courmarin-dimerizer system.
12. A method of causing apoptosis in a diseased cell that overexpresses cyclooxygenase-2, the method comprising delivering a chimeric gene construct to the cell, wherein the chimeric gene construct comprises a cyclooxygenase-2 promoter, which is activated by the cyclooxygenase-2, operably linked to genes that encode for production of Caspase-3 and Caspase-9, and expressing Caspase-3 and Caspase-9 in the diseased cell that overexpresses cyclooxygenase-2, wherein the combined production of Caspase-3 and Caspase-9 causes the cell to undergo apoptosis.
13. The method of claim 12 , wherein the step of delivering the chimeric gene construct to the cell comprises delivering the chimeric gene construct by a non-viral delivery vehicle.
14. The method of claim 13 , wherein the non-viral delivery vehicle is selected from the group consisting of poly(ethylenimine), lipofectin, lipofectamine, polylysine, and alginate.
15. The method of claim 13 , wherein the non-viral delivery vehicle is poly(ethylenimine).
16. The method of claim 12 , wherein the step of delivering the chimeric gene construct to the cell comprises delivering the chimeric gene construct by a viral delivery vehicle.
17. The method of claim 16 , wherein the viral delivery vehicle is a viral vector selected from the group consisting of an adenovirus, an adeno-associated virus, a lentivirus, and a retrovirus.
18. The method of claim 12 , wherein the diseased cell is selected from the group consisting of a cancer cell, a tumor cell and an inflammatory cell.
19. The method of claim 12 , wherein the method further comprises expressing at least one additional gene for production of an apoptosis agent selected from the group consisting of Caspase-1, Caspase-2, Caspase-4, Caspase-5, Caspase-6, Caspase-7, Caspase-8, Caspase-10, Granzyme A, Granzyme B, Fas ligand, TRAIL and APO3L.
20. The method of claim 12 , further comprising delivering to the cell an oligomerizing agent.
21. A method of causing apoptosis in a diseased cell that overexpresses a protein, the method comprising delivering a first and a second chimeric gene construct to the cell, wherein the first chimeric gene construct comprises an upstream regulatory element, which is activated by the protein overexpressed in the cell, operably linked to a Caspase-3 gene and the second chimeric gene construct comprises the upstream regulatory element operably linked to a Caspase-9 gene, and expressing Caspase-3 and Caspase-9 in the cell that overexpresses the protein, wherein the combined production of Caspase-3 and Caspase-9 causes the cell to undergo apoptosis.
22. The method of claim 21 , wherein the step of delivering the chimeric gene constructs to the cell comprises delivering the chimeric gene constructs by a non-viral delivery vehicle.
23. The method of claim 22 , wherein the non-viral delivery vehicle is selected from the group consisting of poly(ethylenimine), lipofectin, lipofectamine, polylysine, and alginate.
24. The method of claim 22 , wherein the non-viral delivery vehicle is poly(ethylenimine).
25. The method of claim 21 , wherein the step of delivering the chimeric gene constructs to the cell comprises delivering the chimeric gene constructs by a viral delivery vehicle.
26. The method of claim 25 , wherein the viral delivery vehicle is a viral vector selected from the group consisting of an adenovirus, an adeno-associated virus, a lentivirus, and a retrovirus.
27. The method of claim 21 , wherein the upstream regulatory element is selected from the group consisting of an cyclooxygenase promoter, a tumor necrosis factor promoter, an interleukin-2 promoter, an interleukin-21 promoter, and an interleukin-23 promoter.
28. The method of claim 21 , wherein the protein which activates the upstream regulatory element is selected from the group consisting of a tumor necrosis factor, an interleukin-2, an interleukin-21, and an interleukin-23.
29. The method of claim 21 , wherein the diseased cell is selected from the group consisting of a cancer cell, a tumor cell, and an inflammatory cell.
30. The method of claim 21 , wherein the method further comprises expressing at least one additional gene for production of an apoptosis agent selected from the group consisting of Caspase-1, Caspase-2, Caspase-4, Caspase-5, Caspase-6, Caspase-7, Caspase-8, Caspase-10, Granzyme A, Granzyme B, Fas ligand, TRAIL and APO3L.
31. The method of claim 21 , further comprising regulating apoptosis using an oligomerizing agent selected from the group consisting of a cyclosporin A-type ligand, a tetracycline, a steroid ligand, a Tet-On/Tet-Off system, an ecdysone-dimerizer system, an antiprogestin-dimerizer system, and a courmarin-dimerizer system.
32. A method of causing apoptosis in a diseased cell that overexpresses cyclooxygenase-2, the method comprising delivering a first and a second chimeric gene construct to the cell, wherein the first chimeric gene construct comprises a cyclooxygenase-2 promoter, which is activated by the protein overexpressed in the cell, operably linked to a Caspase-3 gene and the second chimeric gene construct comprises the cyclooxygenase-2 promoter operably linked to a Caspase-9 gene, and expressing Caspase-3 and Caspase-9-in the diseased cell that overexpresses cyclooxygenase-2, wherein the combined production of Caspase-3 and Caspase-9 causes the cell to undergo apoptosis.
33. The method of claim 32 , wherein the step of delivering the chimeric gene construct to the cell comprises delivering the chimeric gene construct by a non-viral delivery vehicle.
34. The method of claim 33 , wherein the non-viral delivery vehicle is selected from the group consisting of poly(ethylenimine), lipofectin, lipofectamine, polylysine, and alginate.
35. The method of claim 33 , wherein the non-viral delivery vehicle is poly(ethylenimine).
36. The method of claim 32 , wherein the step of delivering the chimeric gene construct to the cell comprises delivering the chimeric gene construct by a viral delivery vehicle.
37. The method of claim 36 , wherein the viral delivery vehicle is a viral vector selected from the group consisting of an adenovirus, an adeno-associated virus, a lentivirus, and a retrovirus.
38. The method of claim 32 , wherein the diseased cell is selected from the group consisting of a cancer cell, a tumor cell and an inflammatory cell.
39. The method of claim 32 , wherein the method further comprises expressing at least one additional gene for production of an apoptosis agent selected from the group consisting of Caspase-1, Caspase-2, Caspase-4, Caspase-5, Caspase-6, Caspase-7, Caspase-8, Caspase-10, Granzyme A, Granzyme B, Fas ligand, TRAIL and APO3L.
40. The method of claim 32 , further comprising delivering to the cell an oligomerizing agent.