Production of chimeric antibodies by homologous recombination
First Claim
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1. A method for the production of a cell line expressing a modified antibody molecule comprising:
- (a) transfecting an antibody producing lymphoid cell line with a target vector comprising;
(i) a replacement gene to modify a portion of the genomic sequence of the immunoglobulin genes of the lymphoid cell line, and(ii) a target sequence homologous to a second DNA sequence adjacent to the immunoglobulin sequence to be converted, so that the replacement gene modifies the immunoglobulin sequence via site specific homologous recombination with genomic chromosomal DNA in vivo; and
(b) selecting a transfectant which produces the modified antibody molecule.
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Abstract
A process for producing chimeric antibodies using novel recombinant DNA vectors and homologous recombination in vivo is described. The recombinant DNA constructs of the invention can be used to transfect antibody producing cells so that targeted homologous recombination occurs in the transfected cells leading to gene modification and the production of chimeric antibody molecules by the transfected cells.
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Citations
21 Claims
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1. A method for the production of a cell line expressing a modified antibody molecule comprising:
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(a) transfecting an antibody producing lymphoid cell line with a target vector comprising; (i) a replacement gene to modify a portion of the genomic sequence of the immunoglobulin genes of the lymphoid cell line, and (ii) a target sequence homologous to a second DNA sequence adjacent to the immunoglobulin sequence to be converted, so that the replacement gene modifies the immunoglobulin sequence via site specific homologous recombination with genomic chromosomal DNA in vivo; and (b) selecting a transfectant which produces the modified antibody molecule. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 13, 14, 15, 16, 17, 20, 21)
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11. A method for the production of a modified antibody molecule, comprising:
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(a) culturing a modified-antibody-producing cell line which was prepared by transfecting an antibody-lymphoid-producing cell line with a target vector comprising; (i) a replacement gene to convert a portion of the genomic sequence of the immunoglobulin genes of the lymphoid cell line, and (ii) a target sequence homologous to a second DNA sequence adjacent to the immunoglobulin sequence to be converted so that the replacement gene modifies the immunoglobulin sequence via site specific homologous recombination with genomic chromosomal DNA in vivo and the modified antibody is expressed by the transfected cell line; and (b) isolating the modified antibody molecule from the culture.
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- 18. The method according to claim 22 in which the lymphoid cell line to be transfected produces monoclonal antibody L20.
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