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Engineered antibody Fc variant with selectively enhanced Fc?RIIb binding over both Fc?RIIa(R131) and Fc?RIIa(H131).

ABSTRACT: Engaging inhibitory Fc?RIIb by Fc region has been recently reported to be an attractive approach for improving the efficacy of antibody therapeutics. However, the previously reported S267E/L328F variant with enhanced binding affinity to Fc?RIIb, also enhances binding affinity to Fc?RIIa(R131) allotype to a similar degree because Fc?RIIb and Fc?RIIa(R131) are structurally similar. In this study, we applied comprehensive mutagenesis and structure-guided design based on the crystal structure of the Fc/Fc?RIIb complex to identify a novel Fc variant with selectively enhanced Fc?RIIb binding over both Fc?RIIa(R131) and Fc?RIIa(H131). This novel variant has more than 200-fold stronger binding affinity to Fc?RIIb than wild-type IgG1, while binding affinity to Fc?RIIa(R131) and Fc?RIIa(H131) is comparable with or lower than wild-type IgG1. This selectivity was achieved by conformational change of the C(H)2 domain by mutating Pro to Asp at position 238. Fc variant with increased binding to both Fc?RIIb and Fc?RIIa induced platelet aggregation and activation in an immune complex form in vitro while our novel variant did not. When applied to agonistic anti-CD137 IgG1 antibody, our variant greatly enhanced the agonistic activity. Thus, the selective enhancement of Fc?RIIb binding achieved by our Fc variant provides a novel tool for improving the efficacy of antibody therapeutics.

SUBMITTER: Mimoto F

PROVIDER: S-EPMC3785249 | biostudies-literature | 2013 Oct

REPOSITORIES: biostudies-literature

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Engineered antibody Fc variant with selectively enhanced FcγRIIb binding over both FcγRIIa(R131) and FcγRIIa(H131).

Mimoto F F Katada H H Kadono S S Igawa T T Kuramochi T T Muraoka M M Wada Y Y Haraya K K Miyazaki T T Hattori K K

Protein engineering, design & selection : PEDS 20130605 10

Engaging inhibitory FcγRIIb by Fc region has been recently reported to be an attractive approach for improving the efficacy of antibody therapeutics. However, the previously reported S267E/L328F variant with enhanced binding affinity to FcγRIIb, also enhances binding affinity to FcγRIIa(R131) allotype to a similar degree because FcγRIIb and FcγRIIa(R131) are structurally similar. In this study, we applied comprehensive mutagenesis and structure-guided design based on the crystal structure of the ...[more]

PMID: 23744091

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Project description:Chronic lymphocytic leukemia (CLL) is common in both developed and developing nations where the need for inexpensive and convenient administration of therapy is apparent. Ocaratuzumab is a novel Fc-engineered humanized IgG1 anti-CD20 monoclonal antibody (mAb) designed for effective antibody-dependent cell-mediated cytotoxicity (ADCC) at very low concentrations that may facilitate sub-cutaneous (vs. intravenous) dosing. Here, we report ocaratuzumab's potency against CLL cells. In vitro assessment of ocaratuzumab's direct cytotoxicity (DC), complement-dependent cytotoxicity (CDC), antibody-dependent cellular phagocytosis (ADCP), and ADCC was performed on CLL cells. Ocaratuzumab induced DC, CDC, and ADCP similarly to rituximab or ofatumumab (anti-CD20 mAbs). However, ocaratuzumab showed an advantage in NK cell-mediated ADCC over these antibodies. In allogeneic ADCC, [E:T (effector:target) ratios = 25:1, 12:1, 6:1], ocaratuzumab (10 µg/mL) improved ADCC by ~3-fold compared with rituximab or ofatumumab (P<0.001 all tested E:T ratios). Notably, the superiority of ocaratuzumab-induced ADCC was observed at low concentrations (0.1-10 ug/ml; P<0.03; allogeneic assays). In extended allogeneic ADCC E:T titration, ocaratuzumab (0.1 µg/mL) demonstrated 19.4% more cytotoxicity than rituximab (E:T = 0.38:1; P = 0.0066) and 21.5% more cytotoxicity than ofatumumab (E:T = 1.5:1; P = 0.0015). In autologous ADCC, ocaratuzumab (10 µg/mL) demonstrated ~1.5-fold increase in cytotoxicity compared with rituximab or ofatumumab at all E:T ratios tested (E:Ts = 25:1,12:1,6:1; all P<0.001). Obinutuzumab, a glyco-engineered anti-CD20 mAb, showed no improvement in ADCC activity compared with ocaratuzumab. The enhanced ADCC of ocaratuzumab suggests that it may be effective at low concentrations. If supported by clinical investigation, this feature could potentially allow for subcutaneous dosing at low doses that could expand the potential of administering chemoimmunotherapy in developing countries.

Dataset Information

Engineered antibody Fc variant with selectively enhanced Fc?RIIb binding over both Fc?RIIa(R131) and Fc?RIIa(H131).

Publications

Engineered antibody Fc variant with selectively enhanced FcγRIIb binding over both FcγRIIa(R131) and FcγRIIa(H131).

Similar Datasets

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