Asymmetrical Fc engineering greatly enhances antibody-dependent cellular cytotoxicity (ADCC) effector function and stability of the modified antibodies.

Abstract

Antibody-dependent cellular cytotoxicity (ADCC) is mediated through the engagement of the Fc segment of antibodies with Fcγ receptors (FcγRs) on immune cells upon binding of tumor or viral antigen. The co-crystal structure of FcγRIII in complex with Fc revealed that Fc binds to FcγRIII asymmetrically with two Fc chains contacting separate regions of the FcγRIII by utilizing different residues. To fully explore this asymmetrical nature of the Fc-FcγR interaction, we screened more than 9,000 individual clones in Fc heterodimer format in which different mutations were introduced at the same position of two Fc chains using a high throughput competition AlphaLISA® assay. To this end, we have identified a panel of novel Fc variants with significant binding improvement to FcγRIIIA (both Phe-158 and Val-158 allotypes), increased ADCC activity in vitro, and strong tumor growth inhibition in mice xenograft human tumor models. Compared with previously identified Fc variants in conventional IgG format, Fc heterodimers with asymmetrical mutations can achieve similar or superior potency in ADCC-mediated tumor cell killing and demonstrate improved stability in the CH2 domain. Fc heterodimers also allow more selectivity toward activating FcγRIIA than inhibitory FcγRIIB. Afucosylation of Fc variants further increases the affinity of Fc to FcγRIIIA, leading to much higher ADCC activity. The discovery of these Fc variants will potentially open up new opportunities of building the next generation of therapeutic antibodies with enhanced ADCC effector function for the treatment of cancers and infectious diseases.

• Authors: Zhi Liu, Kannan Gunasekaran, Wei Wang, Vladimir Razinkov, Laura Sekirov, Esther Leng, Heather Sweet, Ian Foltz, Monique Howard, Anne-Marie Rousseau, Carl Kozlosky, William Fanslow, Wei Yan
• Journal: The Journal of biological chemistry (J Biol Chem) Vol. 289 Issue 6 Pg. 3571-90 (Feb 07 2014) ISSN: 1083-351X [Electronic] United States
• PMID: 24311787 (Publication Type: Journal Article)

Chemical References

• Antibodies, Neoplasm
• FCGR3A protein, human
• Immunoglobulin Fc Fragments
• Immunoglobulin G
• Receptors, IgG

Topics

• Animals
• Antibodies, Neoplasm (genetics, immunology, pharmacology)
• Antibody-Dependent Cell Cytotoxicity (drug effects, genetics)
• Cell Line, Tumor
• Humans
• Immunoglobulin Fc Fragments (genetics, immunology, pharmacology)
• Immunoglobulin G (genetics, immunology, pharmacology)
• Mice
• Mice, SCID
• Mutation
• Neoplasms
• Protein Engineering
• Receptors, IgG (genetics, immunology)
• Xenograft Model Antitumor Assays