Carbohydrates engineered at antibody constant domains can be used for site-specific conjugation of drugs and chelates

• Published in: Journal of immunological methods Vol. 213; no. 2; pp. 131 - 144
• Main Authors: Qu, Zhengxing, Sharkey, Robert M, Hansen, Hans J, Shih, Lisa B, Govindan, Serengulam V, Shen, Jian, Goldenberg, David M, Leung, Shui-on
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 15.04.1998; Elsevier
• Subjects: Amino Acid Sequence; Antibiotics, Antineoplastic - chemistry; Antibodies; Antibodies, Monoclonal - chemistry; Antibodies, Monoclonal - genetics; Antigens, CD - immunology; Antigens, Differentiation, B-Lymphocyte - immunology; Base Sequence; Binding Sites; Biological and medical sciences; Carbohydrate Sequence; Carbohydrates - chemistry; Cell Adhesion Molecules; Chelating Agents - chemistry; Dextrans - chemistry; Doxorubicin - chemistry; General pharmacology; Glycosylation; Human; Humans; Immunoconjugates; Immunoconjugates - chemistry; Immunotherapy; Lectins; Medical sciences; Molecular Sequence Data; Mutagenesis, Site-Directed; Oligosaccharides; Oligosaccharides - chemistry; Peptides - chemistry; Pharmaceutical technology. Pharmaceutical industry; Pharmacology. Drug treatments; Sequence Homology, Amino Acid; Sialic Acid Binding Ig-like Lectin 2; Substrate Specificity; Human; DTPA, diethylenetriamine pentaacetic acid; Immunoconjugates; Immunotherapy; Antibodies; DOX, doxorubicin; Oligosaccharides; CHO, carbohydrate; ABS, antigen binding site; Performance evaluation; Site directed mutagenesis; Glycosylation; Monoclonal antibody; Immunoconjugate; In vitro; Modeling
• ISSN: 0022-1759; 1872-7905
• DOI: 10.1016/S0022-1759(97)00192-0

To improve the efficiency of site-specific conjugation of chelates and drugs to antibodies, and to minimize the incidence of immunoreactivity perturbation to the resultant immunoconjugates, Asn-linked oligosaccharide moieties were designed and engineered into the constant domains of a humanized anti-CD22 monoclonal antibody, hLL2. From 10 potential glycosylation mutants, two CH 1 domain glycosylation sites, HCN1 and HCN5, were identified that were positioned favorably for glycosylation. The carbohydrate (CHO) chains attached at these sites were differentially processed so that HCN5-CHOs were physically larger than HCN1–CHOs. Although both the CH 1-appended CHOs, and the LL2 V κ-appended CHOs conjugated efficiently with small chelates, the HCN5–CHOs, due to the structural and positional superiority, appear to be a better conjugation site for large drug complexes, such as 18 kDa doxorubicin (DOX)–dextran.