Host cells containing multiple integrating vectors
AssigneeGala Design, Inc.
InventorsRobert D. Bremel, Linda U. Miller, Gregory T. Bleck
Life of this patent
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Abstract
The present invention relates to the production of proteins in host cells, and, more particularly to host cells containing multiple integrated copies of an integrating vector. Suitable integrating vectors for use in the present invention include retrovirus vectors, lentivirus vectors, transposon vectors, and adeno-associated virus vectors. Methods are provided in which the host cells are prepared by using the integrating vectors at a high multiplicity of infection. The host cells are useful for producing pharmaceutical proteins, variants of proteins for use in screening assays, and for direct use in high throughput screening.
Claims32
A clonally selected host cell comprising a genome, said genome comprising from about 20 to about 50 integrated integrating vectors, wherein said integrating vectors comprise at least one exogenous gene operably linked to a promoter, said gene encoding a secreted protein, wherein said host cell does not produce infectious retroviral particles.
A method for transducing host cells comprising:
- 1) providing:
- a) a host cell comprising a genome, and
- b) a plurality of retroviral vectors;
- and
- 2) contacting said host cell with said plurality of retroviral vectors at a multiplicity of infection of grater than 100 under conditions such that at least two retroviral vectors integrate into said genome of said host cell, wherein said retroviral vectors comprise at least one exogenous gene operably linked to a promoter, said gene encoding a secreted protein, wherein said host cell does not produce infectious retroviral particles;
- 3) clonally selecting a transduced host cell.
A method of producing a protein of interest comprising:
- 1) providing a clonally selected host cells comprising a genome, said genome comprising from about 20 to about 50 integrated copies of at least one integrating vector comprising an exogenous gene operably linked to a promoter, wherein said exogenous gene encodes a secretable protein of interest, and
- 2) culturing said host cells under conditions such that said protein of interest is produced at rate of from 10 picograms per cell per day to about 50 picograms per cell per day.
A method comprising:
- 1) providing:
- a) a host cell comprising a genome comprising at least one integrated exogenous gene;
- and
- b) a plurality of retroviral vectors comprising said exogenous gene, said gene encoding a secreted protein, wherein said host cell does not produce infectious retroviral particles;
- and
- 2) contacting said host cell with said plurality of retroviral vectors at a multiplicity of infection of greater than 100 such that at least two of said retroviral vectors integrate into said genome of said host cell.
Dependencies are inferred from the claim text.