Baculovirus system for the expression of a gene therapy vector
The invention relates to a recombinant baculoviral genome useful for the production of viral vectors for gene therapy, allowing said production from a single infection.
1. A recombinant baculovirus genome comprising:
(i) one or more expression cassettes comprising adeno-associated virus (AAV) rep and cap genes under the control of a very late promoter suitable for expression of said genes in an insect host cell, and
(ii) a recombinant genome of an AAV vector,
wherein said one or more expression cassettes is(are) inserted into the egt locus;
wherein said recombinant genome of an AAV vector is inserted into the polyhedrin locus; and
wherein the chitinase, cathepsin and p10 genes are deleted.
2. The recombinant baculovirus genome as claimed in claim 1 , said rep and cap genes being contained in a single expression cassette.
3. The recombinant baculovirus genome as claimed in claim 1 , said baculovirus genome being derived from AcMNPV.
4. The recombinant baculovirus genome as claimed in claim 1 , wherein the AAV recombinant genome comprises a heterologous gene encoding a protein, an interfering RNA or an antisense RNA, which is therapeutic.
5. The recombinant baculovirus genome as claimed in claim 1 , said recombinant baculovirus genome being a bacmid.
6. A recombinant baculovirus containing a recombinant baculovirus genome as claimed in claim 1 .
7. A eukaryotic or prokaryotic cell containing the recombinant baculovirus genome as claimed in claim 1 .
8. The cell as claimed in claim 7 , said cell being an insect cell derived from the Spodoptera frugiperda or Trichoplusia ni lines.
9. A method for producing a recombinant virus comprising culturing a cell with one or more recombinant baculovirus as claimed in claim 6 under conditions which allow the infection of the cell with the baculovirus and the production of said recombinant virus.
10. A eukaryotic cell infected with the recombinant baculovirus of claim 6 .
11. The cell as claimed in claim 7 , said cell being selected from the group consisting of Sf21, Sf9, TN 5B1-4 and High Five cells.
12. The cell as claimed in claim 10 , said cell being an insect cell derived from the Spodoptera frugiperda or Trichoplusia ni lines.
13. The cell as claimed in claim 10 , said cell being selected from the group consisting of SF21, SF9, TN 5B1-4 and High Five cells.
14. A recombinant baculovirus genome comprising:
(i) one or more expression cassettes comprising adeno-associated virus (AAV) rep and cap genes under the control of a very late promoter suitable for expression of said genes in an insect host cell, and
(ii) a recombinant genome of an AAV vector,
wherein said one or more expression cassettes is(are) inserted into the egt locus; and
wherein said recombinant genome of an AAV vector is inserted into the polyhedrin locus.