Design and development of masked therapeutic antibodies to limit off-target effects: application to anti-EGFR antibodies.
Therapeutic antibodies frequently cause side effects by binding antigen in non-target tissues. Here we demonstrate a novel molecular design of antibodies that addresses this problem by reversibly "masking" antibody complementarity determining regions until they reach diseased tissues containing disease-associated proteases. Specifically, two distinct single-chain Fv (scFv) fragments derived from antibodies against the epidermal growth factor receptor (cetuximab and 425) were fused a protease susceptible linker to their epitopes, which were engineered to encourage intermolecular association. Surface plasmon resonance and flow cytometry were used to confirm that the masked complex poorly interacts with native antigen, whereas protease treatment restores antigen recognition. Minimally, the "masked" scFvs possesses an eight-fold lower association with the epitope compared with the individual scFvs unmasked by proteolytic cleavage. This molecular design may have general utility for targeted release of therapeutic antibodies at disease sites.
Department of Molecular Medicine, Beckman Research Institute at the City of Hope, Duarte, CA, USA.
Joshua M
JM
Donaldson
Csaba
C
Kari
Ruben C
RC
Fragoso
Ulrich
U
Rodeck
John C
JC
Williams
D000911:Antibodies, Monoclonal / Q000008:administration & dosage / Q000737:chemistry* / Q000276:immunology; D061067:Antibodies, Monoclonal, Humanized; D000915:Antibody Affinity; D045744:Cell Line, Tumor; D000068818:Cetuximab; D022801:Complementarity Determining Regions / Q000737:chemistry; D003432:Cross-Linking Reagents / Q000737:chemistry* / Q000378:metabolism; D019281:Dimerization; D016503:Drug Delivery Systems; D015195:Drug Design; D004357:Drug Synergism; D000939:Epitopes / Q000235:genetics / Q000276:immunology; D066246:ErbB Receptors / Q000037:antagonists & inhibitors / Q000235:genetics / Q000276:immunology*; D006801:Humans; D006868:Hydrolysis; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism*; D009363:Neoplasm Proteins / Q000037:antagonists & inhibitors / Q000276:immunology / Q000378:metabolism; D009842:Oligopeptides / Q000737:chemistry* / Q000378:metabolism; D017354:Point Mutation; D011355:Prodrugs / Q000008:administration & dosage / Q000737:chemistry*; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000008:administration & dosage / Q000737:chemistry* / Q000276:immunology; D057127:Single-Chain Antibodies / Q000008:administration & dosage / Q000737:chemistry* / Q000276:immunology; D013379:Substrate Specificity; D020349:Surface Plasmon Resonance
C513566:425(scFv)-ETA'; D000911:Antibodies, Monoclonal; D061067:Antibodies, Monoclonal, Humanized; D022801:Complementarity Determining Regions; D003432:Cross-Linking Reagents; D000939:Epitopes; D009363:Neoplasm Proteins; D009842:Oligopeptides; D011355:Prodrugs; D011993:Recombinant Fusion Proteins; D057127:Single-Chain Antibodies; C512478:EGFR protein, human; D066246:ErbB Receptors; D020780:Matrix Metalloproteinase 9; D000068818:Cetuximab