WO1996009074B1

WO1996009074B1 - Use of a non-mammalian dna virus to express an exogenous gene in a mammalian cell

Info

Publication number: WO1996009074B1
Application number: PCT/US1995/011456
Authority: WO
Filing date: 1995-09-08
Publication date: 1996-05-09
Anticipated expiration: 1997-03-23

Abstract

Disclosed is a method of expressing an exogenous gene in a mammalian cell, involving infecting the cell with a non-mammalian virus, such as a baculovirus, whose genome carries an exogenous gene, and growing the cell under conditions such that the gene is expressed. Exogenous genes are delivered to mammalian cells by use of a transfer vector such as that described in the figure. Also disclosed is a method of treating a gene deficiency disorder in a mammal by providing to a cell a therapeutically effective amount of a virus whose genome carries an exogenous gene and growing the cell under conditions such that the exogenous gene is expressed in the mammal.

Claims

AMENDED CLAIMS

[received by the International Bureau on 25 March 1996 (25.03.96); original claims 18 and 58 amended; new claims 63-103 added;

remaining claims unchanged (9 pages)]

10. The method cf claim.5, wherein said

baculovirus is in the budded form.

11. The method of claim 1, wherein said genome further comprises a promoter of a long-terminal repeat of a transposable element.

12. The method of claim 1, wherein said genome further comprises a promoter of a long-terminal repeat of a rerrovirus.

13. The method of claim 12, wherein said retrovirus is Rous Sarmoma Virus.

14. The method of claim 1, wherein said genome further comprises an integrative terminal repeat of an adeno-associated virus.

15. The method or claim 14, wherein said genome further comprises an adeno-associated virus rep gene.

16. The method of claim 1, wherein said genome further comprises a cell-immortalizing sequence.

17. The method of claim 1, wherein said genome further comprises an origin of replication. 18. The method of claim 17, wherein said origin of replication comprises an Epstein Barr virus origin of replication.

19. The method of claim 1, wherein said genome further comprises a polyadenylaticn signal and an RNA splicing signal. medicament for treating a gene deficiency disorder in a mammal.

55. The use of claim 54, wherein said virus is an invertebrate virus. 56. The use of claim 55, wherein said

invertebrate virus is an insect virus.

57. The use of claim 56, wherein said insect virus is a baculovirus.

58. The use of claim 54, wherein said gene encodes a gene product selected from the group consisting of fumarylacetoacetate hydrolase, phenylalanine

hydroxylase, alpha-1 antitrypsin, glucose-6-phosphatese, low-density-lipoprotein receptor, porphobilinogen

deaminase, carbamoyl synthetase I, ornithine

transcarbamylase, arginosuccinate synthetase,

arginosuccinate lyase. arginase, factor VIII, factor IX, cystathione β-synthase, branched chain ketoacid

decarboxylase, albumin, isovaleryl-CoA dehydrogenase, propionyl CoA. carboxylase, methyl malonyl CoA mutase, glucaryl CoA dehydrogenase, insulin, β-glucosidase, and pyruvate carboxylase, hepatic phosphorylase,

phosphorylase Kinase, glycine decarboxylase, H-protein, T-protein, Menkes disease protein, the product of

Wilson's disease gene pWD and CFTR. 59. Use of a non-mammalian DNA virus whose ganome comprises a carcinoma-therapeutic gene selected from the group consisting of tumor necrosis factor, thymidine kinase, diphtheria toxin chimeras, and cytosine diaminase in the preparation of a medicament for treating

hepatocellular carcinoma in a mammal.

60. The use of claim 59, wherein said non-mammalian DNA virus is a baculovirus.

61. The use of claim 59, wherein said carcinoma-tharapeutic gene is operably linked to an α-fetoprotein promoter.

62. The method of claim 1, wherein said exogenous gene is selected from the group consisting of lacZ genes, chloramphenicol acetyltransferase genes, alkaline phosphatase genes, luciferase genes, and green fluorescent protein genes.

63. A nucleic acid comprising:

a genome of a non-mammalian DNA virus;

an exogenous mammalian gene; and

an exogenous mammalian-active promoter, wherein said gene is operably linked to said promoter.

64. The nucleic acid of claim 63, wherein said genome is the genome of an insect virus.

65. The nucleic acid of claim 64, wherein said genome is the genome of a baculovirus.

66. The nucleic acid of claim 65, wherein said genome is the genome of an Autographa californica multiple nuclear

polyhedrosis virus.

67. The nucleic acid of claim 63, wherein said mammalian-active promoter is selected from the group consisting of

mammalian promoters, promoters of long-terminal repeats of retroviruses, promoters of long-terminal repeats of transposable elements, the simian Virus 40 early promoter, the sytomegalovirus IE promoter, and the adenovirus major late promoter.

68. The nucleic acid of claim 67, wherein said promoter is a mammalian promoter.

69. The nucleic acid of claim 63, wherein said promoter is selected from the group consisting of cell-type-specific

promoters, stage-specific promoters, inducible promoters and tissue-specific promoters.

70. The nucleic acid of claim 69, wherein said promoter is a liver-specific promoter. 71. The nucleic acid of claim 70, wherein said liver-specific promoter is selected from the group consisting of hepatitis B promoters, hepatitis A promoters, hepatitis C

promoters, albumin promoters, α-1-antitrypsin promoters, pyruvate kinase promoters, phosphenol pyruvate carfaoxykinase promoters, transferrin promoters, transthyretin promoters, α-fetoprotein promoters, α-fibrinogen promoters, and β-fibrinogen promoters.

72. The nucleic acid or claim 70, wherein said liver- specific promoter is selected from the group consisting of low density lipoprotein receptor promoters, α2-macroglobulin

promoters, α1-antichymotrypsin promoters, α2-HS glycoproiein promoters, haptoglobin promoters, ceruloplasmin promoter;, plasminogen promoters, complement protein promoters, 63

complement activator promoters, β-lipoprotein promoters, and α1- acid glycoprotein promoters.

73. The nucleic acid of claim 63, further comprising a mammalian origin of replication.

74. The nucleic acid of claim 63, further comprising an integrative terminal repeat.

75. The nucleic acid of claim 63, wherein said genome lacks a functional polyhedron gene.

76. The nucleic acid of claim 63, wherein said gene is a human gene. 77. The nucleic acid of claim 63, wherein, said gene is a therapeutic gene.

78. The nucleic acid of claim 62, wherein said gene encodes a gene product selected from the group consisting of carbamoyl synthetase I, ornithine transcarbamylase, arginosuccinate

synthetase, arginosuccinate lyase. arginase fumarylacetoacetate hydrolase, phenylalanine hydroxylase, alpha-1 antitrypsin, glucose-6-phosphatase, low-density-lipoprotein receptor,

porphobilinogen deaminase, arginase, factor VIII, factor IX, cystathione β-sγnthase, branched chain ketoacid decarboxylase, albumin, isovaleryl-CoA dehydrogenase, propionyl CoA carboxylase, methyl malonyl CoA mutase, glutaryl coA dehydrogenase, insulin, β-glucosidase, and pyruvate carboxylase, hepatic phosphorylase, phosphorylase kinase, glycine decarboxylase, H-protein, T-protein, Menkes disease protein, the product of Wilson's disease gene pWD, growth factors, interferons, CFTR, tumor suppressors, herpes simplex virus thymidine kinase, and transcription factors.

79. A nucleic acid comprising:

a genome of a non-mammalian DNA virus;

an exogenous antisense RNA gene, the RNA encoded by said gene being complementary to a nucleic acid of a gena that is expressed in a cell at an undesirably high level; and

an exogenous mammalian-active promoter, whereir said gene is operably linked to said promoter.

80. The nucleic acid of claim 79, wherein said genome is the genome of an insect virus.

81. The nucleic acid of claim 79, wherein said genome is the genome of a baculovirus. 82. The nucleic acid of claim 79, wherein said promoter is selected from the group consisting of mammalian promoters, promoters of long-terminal repeats of rerroviruses, and promoters of long-terminal repeats of transposable elements, the s mian Virus 40 early promoter, the cytomegalovirus IE promoter and the adenovirus major late promoter.

83. A cell that contains a nucleic acid, wherein said nucleic acid comprises:

a genome of a non-mammalian DNA virus;

an exogenous mammalian gene; and

84. The cell of claim 83, wherein said genome is tie genome of an insect virus. 85. The cell of claim 85, wherein said genome is tie genome of a baculovirus.

86. The cell of claim 83, wherein said promoter is selected from the group consisting of mammalian promoters, promotsrs of long-terminal repeats or retroviruses, and promoters of long-terminal repeats of transposable elements, the Simian Virus 40 early promoter, the cytomegalovirus IE promoter, and the adenovirus major late promoter.

87. The cell of claim 83, wherein said cell is a primary cell.

88. The call of claim 83, wherein said cell is a human cell. 39. The cell of claim 83, wherein said cell is sel ected from the group consisting of hepatocytes, kidney cells, NIH3T3 cells, HeLa cells, Cos7 cells, C₂C₁₂ myctubes, C₂C₁₂ myoblasts, CHO/dhfr- cells, lung cells, and PC12 cells. 90. The cell of claim 89, wherein said cell is a hapatocyte selected from the group consisting of HepG2 cells, Sk-Hep-1 cells, Hep3B cells, FTO2B cells, and Hepa 1-6 cells.

91. The cell of claim 83, wherein said cell is selected from the group consisting of Ramos cells, Jurkat cells, HL60 cells, and K-562 cells.

92. The cell of claim 83, wherein said promoter is selected from the group consisting of cell-type-specific promoters, tissue-specific promoters, stage-specific promoters, and

inducible promoters.

93. The cell of claim 83, wherein said promoter is a liver-specific promoter.

54. The cell of claim 83 , wherein said gene is a human gene .

95. The call of claim 83, wherein said gene encoders a gene product selected from the group consisting ot carbamoyl

synthetase I, ornithine transcarbamylase, arginosuccinate

synthetase. arginosuccinate lyase, arginase fumarylacetoacetate hydrolase, phenylalanine hydroxylase, alpha-1 antitrypsin, glucose-6-phosphatase, low-density-lipoprotein receptor,

porphobilinogen deaminase, arginase, factor vni, factor IX, cystathione β-synthase, branched chain ketcacid decarboxylase, albumin, isovaleryl-CoA dehydrogenase, propionyl coA carboxylase, methyl malonyl CoA mutase, glutaryl CoA dehydrogenase, insulin, β-glucosidase, and pyruvate carboxylase, hepatic phosphorylase, phosphorylase kinase, glycine decarboxylaee, H-protein, β-protein, Menkes disease protein, the product of Wilson's disease gene pWD, growth factors, interferons, CPTR, tumor suppressors, herpes simplex virus thymidine kinase, and transcription factors.

96. A nucleic acid comprising:

a genome of a non-mammalian DNA virus;

an exogenous cancer therapeutic gene selected from the group consisting of tumor necrosis factor genes, thymidine kinase genes, chimeric diphtheria toxin genes, and cytosine dianinase genes; and

97. The nucleic acid of claim 96, wherein said genome is the genome of an insect virus.

98. The nucleic acid of claim 97, wherein said genome is the genome of a baculovirus.

99. A nucleic acid comprising;

a genome of a non-nammalian DNA virus;

an exogenous gene selected from the group consisting of RNA decoy genes and ribozyme genes; and

an exogenous mammalian-active promoter. 100. The nucleic acid or claim 99, wherein said genome comprises the genome of a baculovirus.

101. A pharmaceutical composition comprising:

(A) a pharmaceutically acceptable excipient and

(B) a nucleic acid, said nucleic acid comprising:

a genome of a non-mammalian DNA virus;

an exogenous mammalian gene; and

102. The pharmaceutical composition of claim 101, therein said cenome comprises the genome of an insect virus.

103. The pharmaceutical composition of claim 102, wherein said genome comprises the genome of a baculovirus.