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Alteration of electrostatic surface potential enhances affinity and tumor killing properties of anti-ganglioside GD2monoclonal antibody hu3F8
Zhao Q.; Ahmed M.; Guo H.-F.; Cheung I.Y.; Cheung N.-K.V.
2015-05-22
Source PublicationJournal of Biological Chemistry
ISSN1083351X 00219258
Volume290Issue:21Pages:13017-13027
Abstract

Ganglioside GD2 is highly expressed on neuroectodermal tumors and an attractive therapeutic target for antibodies that have already shown some clinical efficacy. To further improve the current antibodies, which have modest affinity, we sought to improve affinity by using a combined method of random mutagenesis and in silico assisted design to affinity-mature the anti-GD2 monoclonal antibody hu3F8. Using yeast display, mutants in the Fv with enhanced binding over the parental clone were FACS-sorted and cloned. In silico modeling identified the minimal key interacting residues involved in the important charged interactions with the sialic acid groups of GD2. Two mutations, D32H (L-CDR1) and E1K (L-FR1) altered the electrostatic surface potential of the antigen binding site, allowing for an increase in positive charge to enhance the interaction with the negatively charged GD2-pentasaccharide headgroup. Purified scFv and IgG mutant forms were then tested for antigen specificity by ELISA, for tissue specificity by immunohistochemistry, for affinity by BIACORE, for antibody-dependent cell-mediated cytotoxicity (ADCC) and complement-mediated cytotoxicity in vitro, and for anti-tumor efficacy in xenografted humanized mice. The nearly 7-fold improvement in affinity of hu3F8 with a single D32H (L-CDR1) mutation translated into ã12-fold improvement in NK92MI-transfected CD16-mediated ADCC, a 6-fold improvement in CD32-mediated ADCC, and a 2.5-fold improvement in complement-mediated cytotoxicity while maintaining restricted normal tissue cross-reactivity and achieving substantial improvement in tumor ablation in vivo. Despite increasing GD2 affinity, the double mutation D32H (L-CDR1) and E1K (L-FR1) did not further improve antitumor efficacy.

DOI10.1074/jbc.M115.650903
URLView the original
Indexed BySCIE
Language英語English
WOS Research AreaBiochemistry & Molecular Biology
WOS SubjectBiochemistry & Molecular Biology
WOS IDWOS:000354975700004
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Cited Times [WOS]:22   [WOS Record]     [Related Records in WOS]
Document TypeJournal article
CollectionFaculty of Health Sciences
AffiliationMemorial Sloan-Kettering Cancer Center
First Author AffilicationCancer Centre
Recommended Citation
GB/T 7714
Zhao Q.,Ahmed M.,Guo H.-F.,et al. Alteration of electrostatic surface potential enhances affinity and tumor killing properties of anti-ganglioside GD2monoclonal antibody hu3F8[J]. Journal of Biological Chemistry,2015,290(21):13017-13027.
APA Zhao Q.,Ahmed M.,Guo H.-F.,Cheung I.Y.,&Cheung N.-K.V..(2015).Alteration of electrostatic surface potential enhances affinity and tumor killing properties of anti-ganglioside GD2monoclonal antibody hu3F8.Journal of Biological Chemistry,290(21),13017-13027.
MLA Zhao Q.,et al."Alteration of electrostatic surface potential enhances affinity and tumor killing properties of anti-ganglioside GD2monoclonal antibody hu3F8".Journal of Biological Chemistry 290.21(2015):13017-13027.
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