Inhibitory FcγRIIb-Mediated Soluble Antigen Clearance from Plasma by a pH-Dependent Antigen-Binding Antibody and Its Enhancement by Fc Engineering

Abstract

Fc engineering can modulate the Fc–FcγR interaction and thus enhance the potency of Abs that target membrane-bound Ags, but it has not been applied to Abs that target soluble Ags. In this study, we revealed a previously unknown function of inhibitory FcγRII in vivo and, using an Ab that binds to Ag pH dependently, demonstrated that the function can be exploited to target soluble Ag. Because pH-dependent Ab dissociates Ag in acidic endosome, its Ag clearance from circulation reflects the cellular uptake rate of Ag/Ab complexes. In vivo studies showed that FcγR but not neonatal FcR contributes to Ag clearance by the pH-dependent Ab, and when Fc binding to mouse FcγRII and III was increased, Ag clearance was markedly accelerated in wild-type mice and FcR γ-chain knockout mice, but the effect was diminished in FcγRII knockout mice. This demonstrates that mouse FcγRII efficiently promotes Ab uptake into the cell and its subsequent recycling back to the cell surface. Furthermore, when a human IgG1 Fc variant with selectively increased binding to human FcγRIIb was tested in human FcγRIIb transgenic mice, Ag clearance was accelerated without compromising the Ab half-life. Taken together, inhibitory FcγRIIb was found to play a prominent role in the cellular uptake of monomeric Ag/Ab immune complexes in vivo, and when the Fc of a pH-dependent Ab was engineered to selectively enhance human FcγRIIb binding, the Ab could accelerate soluble Ag clearance from circulation. We assume such a function would enhance the therapeutic potency of Abs that target soluble Ags.

Entire CD3ε, δ, and γ humanized mouse to evaluate human CD3–mediated therapeutics

“…Recent advances in antibody-engineering technology make it possible to develop not only target-specific therapeutics but also antibodies with various functions, such as bispecific target-binding, agonistic activity, conditional target-binding properties, and effector cell redirection826272829. Of these antibody-based therapeutics, TRABs are attracting attention in the field of cancer immunotherapy8.…”

Sweeping antibody as a novel therapeutic antibody modality capable of eliminating soluble antigens from circulation

“…When the Fc variant was engineered further to introduce the E233D/G237D/P238D/H268D/P271G/A330R mutation (mentioned earlier in Section ‘Sweeping antibody with Fc engineering to increase FcγR binding’, as variant v12), it achieved more than 200‐fold increased binding affinity to FcγRIIb, while binding affinity to human FcγRIIa histidine 131 allotype was 0.068‐fold lower, and binding affinity to human FcγRIIa arginine 131 allotype was only twofold higher than that of wildtype human IgG1. A pH‐dependent antibody with this v12 variant accelerated the antigen clearance from circulation compared to wildtype IgG1 in human FcγRIIb transgenic mice . It was reported that platelet activation mediated by FcγRIIa may cause thrombosis ; hence, selective human FcγRIIb binding can be beneficial, both from the point of sweeping efficiency and from that of safety in an antibody with a pH‐dependent antigen‐binding property.…”

“…Recently, we have demonstrated that mouse FcγR but not FcRn contributes to the uptake into the cell of an antibody–antigen complex with wildtype human and mouse IgG1 . When the FcγR binding of a pH‐dependent anti‐IL‐6 receptor antibody was silenced, hsIL‐6R accumulation in plasma was more than that of the antibody with wildtype IgG1.…”

“…Based on this finding, we have developed an FcγRIIb sweeping antibody, in which the Fc region of a pH‐dependent antibody was engineered to have selectively increased human FcγRIIb binding, to enhance the uptake rate of antibody–antigen complexes ( Fig. D ).…”

Pharmacokinetic and Pharmacodynamic Considerations in the Design of Therapeutic Antibodies

“…This increased the number of cycles of antigen binding and lysosomal degradation, which, coupled with Fc engineering to increase binding to FcRn, significantly improved both PK and PD in cynomolgus monkeys. Additionally, they showed that the Fc gamma receptor FcγRIIb is a major contributor in the cellular uptake of monomeric immune complexes . Subsequently, the authors expanded the sweeping technology to accelerate the clearance of pH‐dependent anti‐IL‐6R by engineering the Fc to selectively enhance human FcγRIIb (instead of FcRn) binding.…”