Patent
Respiratory syncytial virus vaccines expressing protective antigens from promoter-proximal genes
| العنوان: | Respiratory syncytial virus vaccines expressing protective antigens from promoter-proximal genes |
|---|---|
| Patent Number: | 6,923,971 |
| تاريخ النشر: | August 02, 2005 |
| Appl. No: | 09/887469 |
| Application Filed: | June 22, 2001 |
| مستخلص: | Recombinant respiratory syncytial virus (RSV) having the position of genes shifted within the genome or antigenome of the recombinant virus are constructed by insertion, deletion or rearrangement of genes or genome segments within the recombinant genome or antigenome and are useful for eliciting an anti-RSV immune response. Shifting the position of genes in this manner provides for a selected increase or decrease in expression of the gene. In one embodiment, expression of RSV glycoproteins is upregulated by shifting one or more glycoprotein-encoding genes to a more promoter-proximal position. Genes of interest for manipulation to create gene position-shifted RSV include any of the NS1, NS2, N, P, M, SH, M2(ORF1), M2(ORF2), L, F or G genes or a genome segment that may be part of a gene or extragenic. Additional mutations and nucleotide modifications are provided within gene position-shifted RSV to yield desired phenotypic and structural effects. |
| Inventors: | Krempl, Christine D. (Rockville, MD, US); Collins, Peter L. (Rockville, MD, US); Murphy, Brian R. (Bethesda, MD, US); Buchholz, Ursula (Insel Riems, DE); Whitehead, Stephen S. (Gaithersburg, MD, US) |
| Assignees: | The United States of America as represented by the Department of Health & Human Services (Washington, DC, US) |
| Claim: | 1. An isolated infectious recombinant respiratory syncytial virus (RSV) comprising a major nucleocapsid (N) protein, a nucleocapsid phosphoprotein (P), a large polymerase protein (L) a M2(ORF1) RNA polymerase elongation factor, and a partial or complete recombinant RSV genome or antigenome having one or more shifted RSV gene(s) or genome segment(s) within said recombinant genome or antigenome that is/are positionally shifted to a more promoter-proximal or promoter-distal position relative to a position of said RSV gene(s) or genome segment(s) within a wild type RSV genome or antigenome. |
| Claim: | 2. The isolated infectious recombinant RSV of claim 1 , wherein said one or more shifted gene(s) or genome segment(s) is/are shifed to a more promoter-proximal or promoter-distal position by deletion or insertion of one or more displacement polynucleotide(s) within said partial or complete recombinant RSV genome or antigenome. |
| Claim: | 3. The isolated infectious recombinant RSV of claim 2 , wherein said displacement polynucleotide(s) comprise(s) one or more polynucleotide insert(s) of between 150 nucleotides (nts) and 4,000 nucleotides in length which is inserted in a non-coding region (NCR) of the genome or antigenome or as a separate gene unit (GU), said polynucleotide insert lacking a complete open reading frame (ORF) end specifying an attenuated phenotype in said recombinant RSV. |
| Claim: | 4. The isolated infectious recombinant RSV of claim 3 , wherein said polynucleotide insert(s) comprises one or more RSV gene(s) or genome segment(s). |
| Claim: | 5. The isolated infectious recombinant RSV of claim 2 , wherein said displacement polynucleotide(s) comprise(s) one or more RSV gene(s) or genome segment(s) selected from RSV NS1, NS2, N, P, M, SH, M2(ORF1), M2(ORF2), L, F and G genes and genome segments and leader, trailer and intergenis regions of the RSV genome end segments thereof. |
| Claim: | 6. The isolated infectious recombinant RSV of claim 2 , wherein said displacement polynucleotide(s) comprise(s) one or more bovine RSV (BRSV) or human RSV (HRSV) gene(s) or genome segment(s) selected from RSV NS1, NS2, N, P, M, SH, M2(ORF1), M2(ORF2), L, F and G gene(s) or genome segment(s) and leader, trailer and intergenic regions of the RSV genome or segments thereof. |
| Claim: | 7. The isolated infectious recombinant RSV of claim 6 , wherein said displacement polynucleotide(s) is/are deleted to form the recombinant RSV genome or antigenome to cause a positional shift of said one or more shifted RSV gene(s) or genome segment(s) within said recombinant genome or antigenome to a more promoter-proximal position relative to a position of said RSV gene(s) or genome segment(s) within a wild type RSV genome or antigenome. |
| Claim: | 8. The isolated infectious recombinant RSV of claim 7 , wherein said displacement polynucleotide(s) that is/are deleted to form the recombinant RSV genome or antigenome comprise one or more RSV NS1, NS2, SH, M2(ORF2), or G gene(s) or genome segment(s) thereof. |
| Claim: | 9. The isolated infectious recombinant RSV of claim 8 , wherein a displacement polynucleotide comprising a RSV NS1 gene is deleted to form the recombinant RSV genome or antigenome. |
| Claim: | 10. The isolated infectious recombinant RSV of claim 8 , wherein a displacement polynucleotide comprising a RSV NS2 gene is deleted to form the recombinant RSV genome or antigenome. |
| Claim: | 11. The isolated infectious recombinant RSV of claim 8 , wherein a displacement polynucleotide comprising a RSV SH gene is deleted to form the recombinant RSV genome or antigenome. |
| Claim: | 12. The isolated infectious recombinant RSV of claim 8 , wherein a displacement polynucleotide comprising RSV M2(ORF2) is deleted to form the recombinant RSV genome or antigenome. |
| Claim: | 13. The isolated infectious recombinant RSV of claim 8 , wherein a displacement polynucleotide comprising a RSV G gene is deleted to form the recombinant RSV genome or antigenome or antigenome. |
| Claim: | 14. The isolated infectious recombinant RSV of claim 8 , wherein the RSV F and G genes are both deleted to form the recombinant RSV genome or antigenome or antigenome. |
| Claim: | 15. The isolated infectious recombinant RSV of claim 8 , wherein the RSV NS1 and NS2 genes are both deleted to form the recombinant RSV genome or antigenome or antigenome. |
| Claim: | 16. The isolated infectious recombinant RSV of claim 8 , wherein the RSV SH and NS2 genes are both deleted to form the recombinant RSV genome or antigenome or antigenome. |
| Claim: | 17. The isolated infectious recombinant RSV of claim 8 , wherein the RSV SH, NS1 and NS2 genes are all deleted to form the recombinant RSV genome or antigenome or antigenome. |
| Claim: | 18. The isolated infectious recombinant RSV of claim 7 , wherein said displacement polynucleotide(s) comprise(s) one or more deletion(s) within a nontranslated sequence at the beginning or end of an RSV open reading frame or in an intergenic region or 3′ loader or 5′ trailer portion of the RSV genome. |
| Claim: | 19. The isolated infectious recombinant RSV of claim 18 , wherein said displacement polynucleotides comprise a partial gene deletion. |
| Claim: | 20. The isolated infectious recombinant RSV of claim 19 , wherein said partial gene deletion is a partial deletion of the SH gene. |
| Claim: | 21. The isolated infectious recombinant RSV of claim 20 , wherein said partial deletion of the SH gene comprises a deletion within the SH downstream non-translated region. |
| Claim: | 22. The isolated infectious recombinant RSV of claim 21 , which is RSV 6120 having a deletion of 112 nucleotides at positions 4499-4610 in the recombinant RSV antigenome. |
| Claim: | 23. The isolated infectious recombinant RSV of claim 7 , wherein said displacement polynuoleotide(s) is/are selected from one or more region(s) of a downstream untranslated sequence of an RSV gene. |
| Claim: | 24. The isolated infectious recombinant RSV of claim 23 , wherein said downstream untranslated sequence(s) is/are from NS1 (positions 519-563), NS2 (positions 1003-1086), P (positions 3073-3230), M (positions 4033-4197), F(positions 7387-7539), and/or M2 (positions 8433-8490) genes. |
| Claim: | 25. The isolated infectious recombinant RSV of claim 7 , wherein said displacement polynucleotide(s) is/are selected from one or more region(s) of a upstream untranslated sequence of an RSV gene. |
| Claim: | 26. The isolated infectious recombinant RSV of claim 25 , wherein said one or more upstream untranslated sequences is/are from NS1 (positions 55-96), NS2 (positions 606-624) and/or SH (positions 4231-4300). |
| Claim: | 27. The isolated infectious recombinant RSV of claim 7 , wherein said displacement polynucleotide comprises a deletion of nucleotides 4683 to 4685 of the RSV G gene. |
| Claim: | 28. The isolated infectious recombinant RSV of claim 7 , wherein said displacement polynucleotide(s) is/are selected from one or more RSV intergenic sequences. |
| Claim: | 29. The isolated infectious recombinant RSV of claim 7 , wherein said displacement polynucleotide(s) is/are selected from nucleotides within the RSV 5′ trailer region. |
| Claim: | 30. The isolated infectious recombinant RSV of claim 29 , wherein a portion of the 5′ trailer region that immediately follows the L gene is reduced in size by 75 nucleotides, 100 nucleotides, 125 nucleotides, or more, leaving intact the 5′ genomic terminus. |
| Claim: | 31. The isolated infectious recombinant RSV of claim 7 , wherein said displacement polynucleotide(s) is/are selected from nucleotides within the RSV 3′ leader region. |
| Claim: | 32. The isolated infectious recombinant RSV of claim 31 , wherein a portion of the 3′ trailer region that excludes a core portion of the viral promoter located within the first 11 nucleotides of the 3′ leader is deleted. |
| Claim: | 33. The isolated infectious recombinant RSV of claim 7 , wherein a partial or complete deletion from one or any combination of the RSV NS1, NS2, SH, F and/or M2 genes yields an adjustable reduction in genome length of between 1-806 nucleotides. |
| Claim: | 34. The isolated infectious recombinant RSV of claim 7 , wherein a partial or complete deletion from one or any combination of RSV intergenic regions yields an adjustable reduction in genome length of between 1-198 nucleotides. |
| Claim: | 35. The isolated infectious recombinant RSV of claim 7 , wherein a partial or complete deletion from one or any combination of RSV intergenic regions yields an adjustable reduction in genome length of between 1-198 nucleotides. |
| Claim: | 36. The isolated infectious recombinant RSV of claim 6 , wherein said displacement polynucleotide(s) is/are added, substituted, or rearranged within the recombinant RSV genome or antigenome to cause a positional shift of said one or more shifted RSV gene(s) or genome segment(s) within said recombinant genome or antigenome to a more promoter-proximal or promoter-distal position relative to a position of said RSV gene(s) or genome segment(s) within a wild type RSV genome or antigenome. |
| Claim: | 37. The isolated infectious recombinant RSV of claim 36 , wherein said displacement polynucleotide(s) added, substituted, or rearranged within the recombinant RSV genome or antigenome comprise(s) one or more RSV NS1, NS2, SH, M2(ORF2), F, and/or G gene(s) or genome segment(s) thereof. |
| Claim: | 38. The isolated infectious recombinant RSV of claim 36 , wherein said displacement polynucleotide(s) comprise(s) one or more RSV gene(s) or genome segment(s) encoding one or more RSV glycoprotein(s) or immunogenic domain(s) or epitope(s) thereof. |
| Claim: | 39. The isolated infectious recombinant RSV of claim 38 , wherein said displacement polynucleotide(s) is/are selected from gene(s) or genome segment(s) encoding RSV F, G, and/or SH glycoprotein(s) or immunogenic domain(s) or epitope(s) thereof. |
| Claim: | 40. The isolated infectious recombinant RSV of claim 1 , wherein one or more RSV glycoprotein gene(s) or genome segments of RSV F, G and SH is/are added, substituted or rearranged within said recombinant RSV genome or antigenome to a position that is more promoter-proximal compared to a wild type gene order position of said one or more RSV glycoprotein gene(s). |
| Claim: | 41. The isolated infectious recombinant RSV of claim 40 , wherein the RSV glycoprotein gene G is rearranged within said recombinant RSV genome or antigenome to a gene order position that is more promoter-proxirnal compared to the wild type gene order position of G. |
| Claim: | 42. The isolated infectious recombinant RSV of claim 41 , wherein thu RSV glycoprotein gene G is shifted to gene order position 1 withIn said recombinant RSV genome or antigenome. |
| Claim: | 43. The isolated infectious recombinant RSV of claim 40 , wherein the RSV glycoprotein gene F is rearranged within said recombinant RSV genome or antigenome to a gene order position that is more promoter-proximal compared to the wild type gene order position off. |
| Claim: | 44. The isolated infectious recombinant RSV of claim 43 , wherein the RSV glycoprotein gene F is shifted to gene order position 1 within said recombinant RSV genome or antigenome. |
| Claim: | 45. The isolated infectious recombinant RSV of claim 40 , wherein both RSV glycoprotein genes G and F are rearranged within said recombinant RSV genome or antigenome to gene order positions that are more promoter-proximal compared to the wild type gene order positions of G and F. |
| Claim: | 46. The isolated infectious recombinant RSV of claim 45 , wherein the RSV glycoprotein gene G is shifted to gene order position 1 and the RSV glycoprotein gene F is shifted to gene order position 2 within said recombinant RSV genome or antigenome. |
| Claim: | 47. The isolated infectious recombinant RSV of claim 40 , wherein one or more RSV NS1, NS2, SH, M2(ORF2), or G gene(s) or genome segment(s) thereof is/are deleted in the recombinant RSV genome or antigenome. |
| Claim: | 48. The isolated infectious recombinant RSV of claim 40 , wherein a displacement polynucleotide comprising a RSV NS1 gene is deleted to form the recombinany RSV genome or antigenome. |
| Claim: | 49. The isolated infectious recombinant RSV of claim 40 , wherein a displacement polynucleotie comprising a RSV NS2 gene is deleted to form the recombinant RSV genome or antigenome. |
| Claim: | 50. The isolated infectious recombinant RSV of claim 40 wherein a displacement polynucleotide comprising a RSV SH gene is deleted to form the recombinant RSV genome or antigenome. |
| Claim: | 51. The isolated infectious recombinant RSV of claim 50 , wherein the RSV glycoprotein gene G is rearranged within said recombinant RSV genome or antigenome to a gene order position that is more promoter-proximal compared to the wild type gene order position of G. |
| Claim: | 52. The isolated infectious recombinant RSV of claim 51 , wherein the RSV glycoprotein gene G is shifted to gene order position 1 within said recombinant RSV genome or antigenome. |
| Claim: | 53. The isolated infectious recombinant RSV of claim 50 , wherein the RSV glycoprotein gene F is rearranged within said recombinant RSV genome or antigenome to a gene order position that is more promoter-proximal compared to the wild type gene order position of F. |
| Claim: | 54. The isolated infectious recombinant RSV of claim 53 , wherein the RSV glycoprotein gene F is shifted to gene order position 1 within said recombinant RSV genome or antigenome. |
| Claim: | 55. The isolated infectious recombinant RSV of claim 50 , wherein both RSV glycoprotein genes G and F are rearranged within said recombinant RSV genome or antigenome to gene order positions that are more promoter-proximal compared to the wild type gene order positions of G and F. |
| Claim: | 56. The isolated infectious rocombinant RSV of claim 55 , wherein the RSV glycoprotein gene G is shifted to gene order position 1 and the RSV glycoprotcin gene F is shifted to gene order position 2 within said recombinant RSV genome or antigenome. |
| Claim: | 57. The isolated infectious recombinant RSV of claim 40 , wherein the RSV SH end NS2 genes are both deleted to form the recombinant RSV genome or antigenome or antigenome. |
| Claim: | 58. The isolated infectious recombinant RSV of claim 57 , wherein both RSV glycoprotein genes G and F are rearranged within said recombinant RSV genome or antigenome to gene order positions that are more promoter-proximal compared to the wild type gene order positions of G and F. |
| Claim: | 59. The isolated Infectious recombinant RSV of claim 58 , wherein the RSV glycoprotein gene G im shifted to gene order position 1 and the RSV glycoprotein gene F is shifted to gene order position 2 within said recombinant RSV genome or antigenome. |
| Claim: | 60. The isolated infectious recombinant RSV of claim 40 , wherein the RSV SH, NS1 and NS2 genes are all deleted to form the recombinant RSV genome or antigenome or antigenome. |
| Claim: | 61. The isolated infectious recombinant RSV of claim 60 , wherein both RSV glycoprotein genes G and F are rearranged within said recombinant RSV genome or antigenome to gene order positions that are more promoter-proximal compared to the wild type gene order positions of G and F. |
| Claim: | 62. The isolated infectious recombinant RSV of claim 61 , wherein the RSV glycoprotein gene G is shifted to gene order position 1 and the RSV glycoprotein gene F is shifted to gene order position 2 within said recombinant RSV genome or anitgenome. |
| Claim: | 63. The isolated infectious recombinant RSV of claim 1 which is a complete virus. |
| Claim: | 64. The isolated infectious recombinant RSV of claim 1 which is a subviral particle. |
| Claim: | 65. The isolated infectious recombinant RSV of claim 2 , wherein said displaeament polynucleotide is added within or deleted from a noncoding region of the recombinant RSV genome or antigenome. |
| Claim: | 66. The isolated infectious recombinant RSV of claim 1 , wherein the recombinant genome or antigenome incorporates antigenic determinants from one or both subgroup A and subgroup B human RSV. |
| Claim: | 67. A method for stimulating the immune system of an individual to elicit an immune response against RSV which comprises administering to the individual an immunologically sufficient amount of the recombinant RSV of claim 1 combined with a physiologically acceptable carrier. |
| Claim: | 68. The method of claim 67 , wherein the recombinant RSV is administered in a dose of 10 3 to 10 6 PFU. |
| Claim: | 69. The method of claim 67 , wherein the recombinant RSV is administered to the upper respiratory tract. |
| Claim: | 70. The method of claim 67 , wherein the recombinant RSV is administered by spray, droplet or aerosol. |
| Claim: | 71. The method of claim 67 , wherein the recombinant RSV is administered to an individual seronegative for antibodies to RSV or possessing transplacentally acquired maternal antibodies to RSV. |
| Claim: | 72. The method of claim 67 , wherein the recombinant RSV elicits an immune response against either human RSV A or RSV B. |
| Claim: | 73. The method of claim 67 , wherein the recombinant RSV elicits an immune response against both human RSV A and RSV B. |
| Claim: | 74. The method of claim 67 , wherein the recombinant RSV elicits an immune response against either human RSV A or RSV B and is co-administered with an immunologically sufficient amount of a second attenuated RSV capable of eliciting an immune response against human RSV A or RSV B, whereby an immune response is elicited against both human RSV A and RSV B. |
| Claim: | 75. The method of claim 74 , wherein the recombinant RSV and second attenuated RSV are administered simultaneously as a mixture. |
| Claim: | 76. An immunogenic composition to elicit an immune response against RSV comprising an immunologically sufficient amount of the recombinant RSV of claim 1 in a physiologically acceptable carrier. |
| Claim: | 77. The immunogenic composition of claim 76 , formulated in a dose of 10 3 to 10 6 PFU. |
| Claim: | 78. The immunogenic composition of claim 76 , formulated for administration to the upper respiratory tact by spray, droplet or aerosol. |
| Claim: | 79. The immunogenic composition of claim 76 , wherein the recombinant RSV elicits an immune response against either human RSV A or RSV B or both human RSV A and RSV B. |
| Claim: | 80. A method for producing an infectious attenuated recombinant RSV particle from one or wore isolated polynucleotido molceules encoding said RSV, comprising: expressing in a cell or cell-free lysato an expression vector comprising an isolated polynucleotide comprising a recombinant RSV genome or antigenome having one or more shifted RSV gene(s) or genome segment(s) within said recombinant genome or antigenome that is/are positionally shifted to a more pmmoter-proxinial or promoter-distal position relative to a position of said RSV gene(s) or genome segment(s) within a wild type RSV genome or antigenome, and RSV N, P, L and M2ORF1 RNA polymerase elongation factor proteins. |
| Claim: | 81. The method of claim 80 , wherein tie recombinant RSV genome or antigenome and the N, P, L and M2ORF1 RNA polymorase elongation factor proteins are expressed by two or more different expression vectors. |
| Current U.S. Class: | 4242/111 |
| Patent References Cited: | 6264957 July 2001 Collins |
| Other References: | Ball et al, Phenotypic Consequences of Rearranging the P, M and G genes of Vesicular Stomatitis Virus, J. Virology, 1999, 73:4705-4712. cited by examiner Collins et al, Production of infectious human respiratory syncytial virus, PNAS USA, 1995, 92:11563-11567. cited by examiner |
| Assistant Examiner: | Chen, Stacy B. |
| Primary Examiner: | Housel, James |
| Attorney, Agent or Firm: | Graybeal Jackson Haley |
| رقم الانضمام: | edspgr.06923971 |
| قاعدة البيانات: | USPTO Patent Grants |
| الوصف غير متاح. |