High affinity promotes more effective ADCC by anti-HER2/neu monoclonal antibodies

Abstract

2538 Background: Although monoclonal antibodies have emerged as useful cancer therapeutics, the antibody structural features and biologic properties that maximize therapeutic benefit are not fully understood. Antibody-dependent cellular cytotoxicity (ADCC) is believed to be a major mechanism of some anti-cancer antibodies. Herein, we describe the relationship between antibody affinity and the biologic properties of ADCC. Methods: A series of human IgG1 antibodies was created from the anti-HER2/neu C6.5 scFv and its affinity mutants (affinity ranging from 10−7 to 10−11 M). The IgGs were tested for their ability to elicit ADCC using a standard 51Cr-release assay. The ability of these IgGs to inhibit tumor cell proliferation and to induce apoptosis was evaluated using a colorimetric cell proliferation assay and homogeneous caspase assay/Annexin-V-FLUOS assay, respectively. Three tumor cell lines with differing levels of HER2/neu expression were used as target cells, and unactivated human PBMC from healthy donors were the effector cells. Results: There was a clear influence of both intrinsic and functional affinity on ADCC, with higher levels of peak cytotoxicity as apparent affinity approached 10−11 M. Among the antibodies with apparent affinity of 10−11 M, antibodies with higher intrinsic affinities elicited more ADCC. There also was an affinity-dependent increase in cytotoxicity at lower antibody concentrations and lower effector: target ratios. These antibodies elicited more cytotoxicity against tumor cells expressing high levels of HER2/neu than against cells expressing lower amounts of HER2/neu. None of these IgGs inhibited tumor cell proliferation, or induced apoptosis. Conclusions: Additional studies are needed to determine why, at the same functional avidity, ADCC is improved in antibodies with higher intrinsic affinities. These findings justify the examination of high affinity antibodies for ADCC promotion, although the impaired tumor targeting associated with high affinity may be an important confounding factor in the design of unconjugated anti-tumor antibodies. A careful examination of antibody structure: function relationships is required to develop optimized therapeutic unconjugated antibodies. No significant financial relationships to disclose.