Anti-CTLA-4 therapy requires an Fc domain for efficacy

Ipilimumab, a monoclonal antibody that recognizes cytotoxic T lymphocyte antigen ( CTLA)- 4, was the first approved “checkpoint”-blocking anticancer therapy. In mouse tumor models, the response to antibodies against CTLA-4depends entirely on expression of the Fcγ receptor (FcγR), which may facilitate antibody-dependent cellular phagocytosis, but the contribution of simple CTLA-4 blockaderemains unknown. To understand the role of CTLA-4 blockadein the complete absence of Fc-dependent functions, we developed H11, a high-affinity alpaca heavy chain-only antibody fragment (VHH) against CTLA-4. The VHH H11 lacks an Fc portion, binds monovalently to CTLA-4, and inhibits interactions between CTLA-4and its ligand by occluding the ligand-binding motif on CTLA-4as shown crystallographically. We used H11 to visualize CTLA-4expression in vivo using whole-animal immuno-PET, finding that surface-accessible CTLA-4is largely confined to the tumor microenvironment. Despite this, H11-mediated CTLA-4 blockadehas minimal effects on antitumor responses. Installation of the murine IgG2a constant region on H11 dramatically enhances its antitumor response. Coadministration of the monovalent H11 VHH blocks the efficacy of a full-sized therapeutic antibody. We were thus able to demonstrate that CTLA-4–binding antibodies require an Fc domain for antitumor effect.