CHARACTERIZATION OF MONOCLONAL-ANTIBODY GLYCOSYLATION - COMPARISON OFEXPRESSION SYSTEMS AND IDENTIFICATION OF TERMINAL ALPHA-LINKED GALACTOSE

CAMPATH-1H is a recombinant humanized murine monoclonal immunoglobulin (IgG(1)) which recognizes the CDw52 antigen of human lymphocytes, and has been the subject of clinical trials for the treatment of non-Hodg kin's lymphoma and rheumatoid arthritis, Peptide mapping by liquid chr omatography-mass spectrometry was used to confirm the predicted amino acid sequences and profile glycosylation for two CAMPATH isotypes expr essed in a murine myeloma cell. line (NSO) and a single isotype expres sed in both Chinese hamster ovary (CHO) and NSO lines, The three major glycoforms identified in CAMPATH are fucosylated biantennary structur es, containing zero, one, or two galactose residues, Glycosylation of the IgG(1) form of CAMPATH expressed in CHO cells is consistent with n ormal human IgG, However, IgG(1) and IgG(4) expressed in NSO cells inc lude two potentially immunogenic glycoforms which contain either one o r two nonreducing terminal alpha-linked galactose residues, Oligosacch aride structures were characterized by a combination of tandem mass sp ectrometry, methylation analysis, and exoglycosidase digestion. The st rategy used here is designed to be widely applicable to recombinant gl ycoproteins. (C) 1997 Academic Press.