Method for selecting CHO cell for production of glycosylated antibodies
The invention provides an antibody comprising human IgG1 or IgG3 heavy chain constant domains that are glycosylated with a sugar chain at Asn297, said antibody being characterized in that the amount of fucose within said sugar chain is at least 99%, and in addition the amount of NGNA is 1% or less and/or the amount of N-terminal alpha 1,3 galactose is 1% or less, and uses thereof.
1. A method for the selection of a CHO cell that recombinantly produces a monoclonal antibody comprising human IgG1 or IgG3 heavy chain constant domains, wherein the heavy chain constant domains are glycosylated with a sugar chain at Asn297, wherein the sugar chain comprises at least 99% fucose, wherein if the sugar chain comprises N-glycolylneuraminic acid then the amount of N-glycolylneuraminic acid is 1% or less of the sugar chain and wherein if the sugar chain comprises N-terminal alpha-1,3-galactose then the amount of N-terminal alpha-1,3-galactose is 1% or less of the sugar chain;
the method comprising:
(1) cultivating a CHO cell transfected with an expression vector that encodes the heavy and light chains of a monoclonal antibody comprising human IgG1 or IgG3 heavy chain constant domains, wherein two DHFR alleles are deleted from the genome of the CHO cell;
(2) applying DHFR selection pressure;
(3) picking single clones;
(4) expanding the clones;
(5) producing antibodies from the clones;
(6) analyzing glycosylation of the antibodies produced from the clones; and
(7) selecting a clone that produces an antibody wherein the heavy chain constant domains are glycosylated with a sugar chain at Asn297, wherein the sugar chain comprises at least 99% fucose, wherein if the sugar chain comprises N-glycolylneuraminic acid then the amount of N-glycolylneuraminic acid is 1% or less of the sugar chain and wherein if the sugar chain comprises N-terminal alpha-1,3-galactose then the amount of N-terminal alpha-1,3-galactose is 1% or less of the sugar chain.