Method for producing a cell for protein production by treating a cell overexpressing a taurine transporter with methotrexate
AssigneeChugai Seiyaku Kabushiki Kaisha
InventorsHisahiro Tabuchi, Tomoya Sugiyama
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Abstract
The present invention provides a cell capable of high-yield production of polypeptides and a method for producing the same. The present invention relates to a method for producing a cell capable of high-yield production of a desired polypeptide, wherein a strongly taurine transporter-expressing cell into which DNA encoding the desired polypeptide has been introduced is cultured in the presence of a high concentration of methotrexate and a cell capable of high-yield production of the desired polypeptide is selected from among surviving cells.
Claims8
A method for producing a cell capable of increasing production of a desired antibody by expressing a taurine transporter within the cell, comprising:
- (i) preparing a taurine transporter-expressing cell transfected with
- (a) a DNA encoding a taurine transporter selected from the group consisting of a human, mouse, rat, hamster, bovine and dog taurine transporter,
- (b) a DNA encoding the desired antibody, and
- (c) a DNA encoding dihydrofolate reductase,
- (ii) subculturing the taurine transporter-expressing cell in a culture medium comprising methotrexate (MTX) at a concentration of 200 nM, wherein the medium is replaced every three to five days of the subculturing with a medium comprising MTX at a concentration of 200 nM,
- (iii) subculturing the taurine transporter-expressing cell in a culture medium comprising MTX at a concentration of 20 nM in order to proliferate the cell, wherein the subculturing of (ii) is before the subculturing of (iii);
- and
- (iv) selecting a cell from among surviving cells that exhibits increased production of the desired antibody and that is more MTX-resistant relative to a cell that is not transfected with the DNA encoding the taurine transporter.
A method for enhancing the amount of antibody production by a cell, wherein the cell is transfected with (i) a DNA encoding a taurine transporter selected from the group consisting of a human, mouse, rat, hamster, bovine and dog taurine transporter, (ii) a DNA encoding a desired antibody and (iii) a DNA encoding dihydrofolate reductase, the method comprising:
- (i) subculturing the transfected cell is subcultured in a culture medium comprising MTX at a concentration of 200 nM, wherein the medium is replaced every three to five days of the subculturing with medium comprising MTX at a concentration of 200 nM;
- and
- (ii) subculturing the transfected cell in a culture medium comprising MTX at a concentration of 20 nM, wherein the subculturing of (i) is before the subculturing of (ii),
- thereby enhancing the amount of antibody production by the cell.
A method for producing a cell capable of increasing production of a desired antibody by expressing a taurine transporter within the cell, comprising:
- (i) preparing a taurine transporter-expressing cell transfected with
- (a) a DNA encoding a taurine transporter selected from the group consisting of a human, rat, hamster, bovine and dog taurine transporter,
- (b) a DNA encoding the desired antibody, and
- (c) a DNA encoding dihydrofolate reductase,
- (ii) subculturing the taurine transporter-expressing cell in a culture medium comprising MTX at a concentration of 200 nM, wherein the medium is replaced every three to five days of the subculturing with medium comprising MTX at a concentration of 200 nM,
- (iii) subculturing the taurine transporter-expressing cell in a culture medium comprising MTX at a concentration of 20 nM in order to proliferate the cell, wherein the subculturing of (ii) is before the subculturing of (iii);
- and
- (iv) selecting a cell from among surviving cells that exhibits increased production of the desired antibody and that is more MTX-resistant relative to a cell that is not transfected with the DNA encoding the taurine transporter.
Dependencies are inferred from the claim text.