Influence of variable N-glycosylation on the cytolytic potential of chimeric CD19 antibodies.
To investigate the influence of N-linked oligosaccharides at asparagines-297 on the cytolytic potential of chimeric CD19 antibodies, three distinct variants were generated by production in different expression systems. The same chimeric CD19 antibody was produced in Sf21 insect cells, human 293 T cells, and 293 T cells expressing a co-transfected beta1,4-N-acetylglucosaminyltransferase III (GnTIII). The N-glycan structures and the cytolytic potential of the antibodies produced in these three systems were directly compared. After expression in insect cells, the antibody carried paucimannosidic N-linked oligosaccharides, distinct from the complex biantennary carbohydrate moieties attached to the product from human cells. After co-expression with GnTIII in human cells, the antibody carried an eightfold greater percentage of oligosaccharides with a bisecting N-acetylglucosamine (78.7% versus 9.6%) and a 30-fold increased proportion of bisecting, defucosylated oligosaccharides (15.9% versus 0.5%). The insect cell product triggered stronger antibody-dependent cellular cytotoxicity (ADCC) of a human leukemia-derived cell line than the product from non-re-engineered 293 T cells and was equally effective at 50- to 100-fold lower concentrations. The antibody from glyco-engineered 293 T cells had comparable lytic activity as the insect cell product. Both mediated significant ADCC at lower effector-to-target cell ratios than the antibody from non-re-engineered 293 T cells, and both were highly effective against primary blasts from pediatric leukemia patients. The data demonstrate the influence of the N-glycosylation pattern on the ADCC activity of chimeric CD19 antibodies and point to the importance of suitable expression systems for the production of highly active therapeutic antibodies.
Genetics, University of Erlangen--Nuremberg, D 91058 Erlangen, Germany.
Karin
K
Barbin
Julia
J
Stieglmaier
Domenica
D
Saul
Kristin
K
Stieglmaier
Bernhard
B
Stockmeyer
Matthias
M
Pfeiffer
Peter
P
Lang
Georg H
GH
Fey
D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000276:immunology; D000918:Antibody Specificity; D000920:Antibody-Dependent Cell Cytotoxicity / Q000276:immunology*; D018941:Antigens, CD19 / Q000276:immunology*; D002460:Cell Line; D006031:Glycosylation; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology; D007167:Immunotherapy / Q000379:methods; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000276:immunology; D013329:Structure-Activity Relationship
D000911:Antibodies, Monoclonal; D018941:Antigens, CD19; D007074:Immunoglobulin G; D011993:Recombinant Fusion Proteins