High-level expression of a recombinant antibody from myeloma cells using a glutamine synthetase gene as an amplifiable selectable marker.
We report a method for introducing a glutamine synthetase (GS) selectable marker into myeloma cells in which transfectants are selected by growth in a glutamine-free medium. Vector amplification can subsequently be selected using the specific inhibitor of GS, methionine sulphoximine (MSX). Using this system, DNA sequences encoding a chimeric B72.3 IgG4 antibody were expressed from hCMV-MIE promoters in NSO myeloma cells. A cell line was isolated after a single round of selection for vector amplification which contains approximately 4 copies of the vector, secretes 10-15 pg/cell/day cB72.3 antibody during exponential growth and can accumulate 560 mg/l antibody in a fed-batch air-lift fermentation system. Productivity is stable in the absence of MSX selection.
Bebbington
C R
CR
Celltech Ltd., Slough, U.K.
Renner
G
G
Thomson
S
S
King
D
D
Abrams
D
D
Yarranton
G T
GT
D000818:Animals; D001709:Biotechnology / Q000379:methods; D002460:Cell Line; D002678:Chimera; D006224:Cricetinae; D004797:Enzyme-Linked Immunosorbent Assay; D005819:Genetic Markers; D005822:Genetic Vectors; D005974:Glutamate-Ammonia Ligase / Q000096:biosynthesis / Q000235:genetics*; D007074:Immunoglobulin G / Q000096:biosynthesis / Q000145:classification / Q000235:genetics*; D008717:Methionine Sulfoximine / Q000494:pharmacology; D051379:Mice; D009101:Multiple Myeloma; D012333:RNA, Messenger / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000145:classification; D015183:Restriction Mapping; D014162:Transfection
D005819:Genetic Markers; D007074:Immunoglobulin G; D012333:RNA, Messenger; D011994:Recombinant Proteins; D008717:Methionine Sulfoximine; D005974:Glutamate-Ammonia Ligase