Claims
- 1. An adenoviral vector comprising an adenoviral genome comprising (i) at least one deletion in a region of the adenoviral genome selected from the group consisting of E1, E2A and E4, (ii) at least one deletion in the VAI gene of the adenoviral genome, alone or in further combination with at least one deletion in the VAII gene of the adenoviral genome, and, optionally, (iii) a polymerase II (pol II) construct comprising a pol II promoter operably linked to a coding region and/or a polymerase III (pol III) construct comprising a pol III promoter operably linked to a coding region.
- 2. The adenoviral vector of claim 1, which comprises at least one deletion in each of two regions of the adenoviral genome selected from the group consisting of E1, E2A and E4.
- 3. The adenoviral vector of claim 1, which comprises at least one deletion in each of the E1, E4, VAI and VAII regions of the adenoviral genome.
- 4. The adenoviral vector of claim 1, wherein the coding region operably linked to the pol III promoter encodes an antisense RNA molecule, a ribozyme or an RNA interfering molecule.
- 5. The adenoviral vector of claim 1, wherein the pol III promoter is a VA RNA promoter, a transfer RNA promoter or a small nuclear RNA promoter.
- 6. The adenoviral vector of claim 1, wherein the adenoviral vector is packaged as a virion.
- 7. An adenoviral vector comprising an adenoviral genome comprising (i) at least one deletion in a region of the adenoviral genome selected from the group consisting of E1, E2A and E4, (ii) a recombinant VAI gene, alone or in further combination with a recombinant VAII gene, wherein the recombinant gene comprises either of a regulatable promoter in place of the native promoter or a mutated native promoter and 5′ to the mutated native promoter, a pol II promoter, and, optionally, (iii) a pol II construct comprising a pol II promoter operably linked to a coding region and/or a pol III construct comprising a pol III promoter operably linked to a coding region.
- 8. The adenoviral vector of claim 7, which comprises at least one deletion in each of two regions of the adenoviral genome selected from the group consisting of E1, E2A and E4.
- 9. The adenoviral vector of claim 7, wherein the coding region operably linked to the pol III promoter encodes an antisense RNA molecule, a ribozyme, a small interfering RNA (siRNA), or any other RNA interfering molecule.
- 10. The adenoviral vector of claim 7, wherein the pol III promoter is a VA RNA promoter, a transfer RNA promoter or a small nuclear RNA promoter.
- 11. The adenoviral vector of claim 7, wherein the adenoviral vector is packaged as a virion.
- 12. An adenoviral vector comprising an adenoviral genome comprising (i) at least one deletion in a region of the adenoviral genome selected from the group consisting of E1, E2A and E4, (ii) a dominant negative, double-stranded, RNA-dependent protein kinase (PKR), and, optionally, (iii) a pol II construct comprising a pol II promoter operably linked to a coding region and/or a pol III construct comprising a pol III promoter operably linked to a coding region.
- 13. The adenoviral vector of claim 12, which comprises at least one deletion in each of two regions of the adenoviral genome selected from the group consisting of E1, E2A and E4.
- 14. The adenoviral vector of claim 12, wherein the coding region operably linked to the pol III promoter encodes an antisense RNA molecule, a ribozyme, an siRNA, or any other RNA interfering molecule.
- 15. The adenoviral vector of claim 12, wherein the pol III promoter is a VA RNA promoter, a transfer RNA promoter or a small nuclear RNA promoter.
- 16. The adenoviral vector of claims 12, wherein the adenoviral vector is packaged as a virion.
- 17. A system comprising an adenoviral vector of claim 1 and a cell line that complements the adenoviral vector, wherein the cell line comprises the adenoviral vector.
- 18. The system of claim 17, wherein the cell line that complements the adenoviral vector comprises and expresses an adenoviral VAI gene and/or VAII gene.
- 19. The system of claim 18, wherein the promoter of the adenoviral VAI gene is a native promoter.
- 20. The system of claim 18, wherein the promoter of the adenoviral VAI gene is a regulatable promoter.
- 21. The system of claim 20, wherein the promoter of the adenoviral VAI gene comprises an element that binds to a tetracycline repressor.
- 22. The system of claim 21, wherein the element that binds to a tetracycline repressor interferes with the split binding sites of the transcription factor TFIIID.
- 23. A system comprising an adenoviral vector of claim 7 and a cell line that complements the adenoviral vector, wherein the cell line comprises the adenoviral vector.
- 24. The system of claim 23, wherein the cell line that complements the adenoviral vector comprises and expresses an adenoviral VAI gene and/or VAII gene.
- 25. The system of claim 24, wherein the promoter of the adenoviral VAI gene is a native promoter.
- 26. The system of claim 24, wherein the promoter of the adenoviral VAI gene is a regulatable promoter.
- 27. The system of claim 26, wherein the promoter of the adenoviral VAI gene comprises an element that binds to a tetracycline repressor.
- 28. The system of claim 27, wherein the element that binds to a tetracycline repressor interferes with the split binding sites of the transcription factor TFIIID.
- 29. A system comprising the adenoviral vector of claim 12 and a cell line that complements the adenoviral vector, wherein the cell line comprises the adenoviral vector.
- 30. A system comprising an adenoviral vector comprising (i) an adenoviral genome comprising at least one deletion in a region of the adenoviral genome selected from the group consisting of E1, E2A and E4, and, optionally, a pol II construct comprising a pol II promoter operably linked to a coding region and/or a pol III construct comprising a pol III promoter operably linked to a coding region, and (ii) a cell line that complements the adenoviral vector, wherein the cell line expresses a dominant negative, double-stranded PKR and comprises the adenoviral vector.
- 31. The system of claim 30, in which the adenoviral vector comprises at least one deletion in each of two regions of the adenoviral genome selected from the group consisting of E1, E2A and E4.
- 32. The system of claim 30, in which the coding region operably linked to the pol III promoter encodes an antisense RNA molecule, a ribozyme, an siRNA, or any other RNA interfering molecule.
- 33. The system of claim 30, in which the pol III promoter is a VA RNA promoter, a transfer RNA promoter or a small nuclear RNA promoter.
- 34. The system of claim 30, in which the adenoviral vector is packaged as a virion.
- 35. A method of making an adenoviral vector of claim 1, which method comprises (i) introducing at least one deletion in a region of the adenoviral genome selected from the group consisting of E1, E2A and E4, (ii) introducing at least one deletion in the VAI gene of the adenoviral genome, alone or in further combination with at least one deletion in the VAII gene of the adenoviral genome, and, optionally, (iii) introducing a pol II construct comprising a pol II promoter operably linked to a coding region and/or a pol III construct comprising a pol III promoter operably linked to a coding region into the adenoviral vector, with the proviso that (i) and (ii) can be carried out in either order.
- 36. The method of claim 35, which method further comprises (iv) propagating the adenoviral vector in a complementing cell line.
- 37. A method of making an adenoviral vector of claim 3, which method comprises (i) introducing at least one deletion in each of the E1, E4, VAI and VAII regions of the adenoviral genome, and, optionally, (ii) introducing a pol II construct comprising a pol II promoter operably linked to a coding region and/or a pol III construct comprising a pol III promoter operably linked to a coding region into the adenoviral vector, with the proviso that (i) and (ii) can be carried out in either order.
- 38. The method of claim 37, which method further comprises (iii) propagating the adenoviral vector in a complementing cell line.
- 39. A method of making an adenoviral vector of claim 7, which method comprises (i) introducing at least one deletion in a region of the adenoviral genome selected from the group consisting of E1, E2A and E4, (ii) either (a) introducing a mutation into the native promoter of the VAI gene and/or the VAII gene, and 5′ to the mutated native promoter, introducing a pol II promoter, or (b) substituting a regulatable promoter for the native promoter of the VAI gene and/or the VAII gene, and, optionally, (iii) introducing a pol II construct comprising a pol II promoter operably linked to a coding region and/or a pol III construct comprising a pol III promoter operably linked to a coding region into the adenoviral vector, with the proviso that (i) and (ii) can be carried out in either order.
- 40. The method of claim 39, which method further comprises (iv) propagating the adenoviral vector in a complementing cell line, which does not express VAI and VAII, in the presence of an inducer of the regulatable promoter.
- 41. A method of making an adenoviral vector of claim 12, which method comprises (i) introducing at least one deletion in a region of the adenoviral genome selected from the group consisting of E1, E2A and E4, (ii) introducing an oligonucleotide sequence encoding a dominant negative, double-stranded PKR kinase into the adenoviral vector and, optionally, (iii) introducing a pol II construct comprising a pol II promoter operably linked to a coding region and/or a pol III construct comprising a pol III promoter operably linked to a coding region into the adenoviral vector, with the proviso that (i) and (ii) can be carried out in either order.
- 42. The method of claim 41, which method further comprises (iv) propagating the adenoviral vector in a complementing cell line.
- 43. A method of expressing a pol II construct comprising a pol II promoter operably linked to a coding region and/or a pol III construct comprising a pol III promoter operably linked to a coding region in a mammalian cell, which method comprises contacting the mammalian cell with an adenoviral vector of claim 1, which comprises a pol II construct comprising a pol II promoter operably linked to a coding region and/or a pol III construct comprising a pol III promoter operably linked to a coding region, whereupon the mammalian cell internalizes the adenoviral vector and expresses the pol II construct and/or the pol III construct in the mammalian cell.
- 44. The method of claim 43, wherein the mammalian cell is in vivo.
- 45. A method of expressing a pol II construct comprising a pol II promoter operably linked to a coding region and/or a pol III construct comprising a pol III promoter operably linked to a coding region in a mammalian cell, which method comprises contacting the mammalian cell with an adenoviral vector of claim 3, which comprises a pol II construct comprising a pol II promoter operably linked to a coding region and/or a pol III construct comprising a pol III promoter operably linked to a coding region, whereupon the mammalian cell internalizes the adenoviral vector and expresses the pol II construct and/or the pol III construct in the mammalian cell.
- 46. The method of claim 45, wherein the mammalian cell is in vivo.
- 47. A method of expressing a pol II construct comprising a pol II promoter operably linked to a coding region and/or a pol III construct comprising a pol III promoter operably linked to a coding region in a mammalian cell, which method comprises contacting the mammalian cell with an adenoviral vector of claim 7, which comprises a pol II construct comprising a pol II promoter operably linked to a coding region and/or a pol III construct comprising a pol III promoter operably linked to a coding region, whereupon the mammalian cell internalizes the adenoviral vector and expresses the pol II construct and/or the pol III construct in the mammalian cell.
- 48. The method of claim 47, wherein the mammalian cell is in vivo.
- 49. A method of expressing a pol II construct comprising a pol II promoter operably linked to a coding region and/or a pol III construct comprising a pol III promoter operably linked to a coding region in a mammalian cell, which method comprises contacting the mammalian cell with an adenoviral vector of claim 12, which comprises a pol II construct comprising a pol II promoter operably linked to a coding region and/or a pol III construct comprising a pol III promoter operably linked to a coding region, whereupon the mammalian cell internalizes the adenoviral vector and expresses the pol II construct and/or the pol III construct in the mammalian cell.
- 50. The method of claim 49, wherein the mammalian cell is in vivo.
- 51. A method of making a cell line that complements the adenoviral vector of claim 1, which method comprises (i) introducing into the cell those regions of an adenoviral genome selected from the group consisting of E1, E2A and E4 that have been disrupted in the adenoviral vector, and (ii) introducing into the cell an adenoviral VAI coding region operably linked to a promoter and/or an adenoviral VAII coding region operably linked to a promoter desirably wherein there is insufficient overlap between the adenoviral genome of the adenoviral vector and those regions of the adenoviral genome and the adenoviral VAI coding region and/or the adenoviral VAII coding region, as have been introduced into the cell, for generation of replication-competent adenovirus, with the proviso that (i) and (ii) can be carried out in either order.
- 52. The method of claim 51, wherein the promoter operably linked to the adenoviral VAI coding region and/or the adenoviral VAII coding region is a native promoter.
- 53. The method of claim 51, wherein the promoter operably linked to the adenoviral VAI coding region and/or the adenoviral VAII coding region is a regulatable promoter.
- 54. The method of claim 53, wherein the promoter comprises an element that binds to a tetracycline repressor.
- 55. The method of claim 54, wherein the tetracycline repressor, when bound to the element, interferes with the split binding sites of the transcription factor TFIIID.
- 56. The method of claim 54, wherein the promoter is a VA promoter and the element that binds to a tetracycline repressor is incorporated 5′ to Box A of the VA promoter.
- 57. The method of claim 54, wherein the promoter is a VA promoter and the element that binds to a tetracycline repressor is incorporated between Box A and Box B of the VA promoter.
TECHNICAL FIELD OF THE INVENTION
[0001] This patent application is a continuation of International Patent Application No. PCT/US02/29111, filed Sep. 13, 2002, which designates the U.S., which claims the benefit of U.S. Provisional Patent Application No. 60/318,997, filed Sep. 13, 2001.
Provisional Applications (1)
|
Number |
Date |
Country |
|
60318997 |
Sep 2001 |
US |
Continuations (1)
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Number |
Date |
Country |
Parent |
PCT/US02/29111 |
Sep 2002 |
US |
Child |
10778832 |
Feb 2004 |
US |