Lack of blocking activity in anti-CTLA-4 antibodies reduces toxicity, but not anti-tumor efficacy

Anti-CTLA-4 antibodies such as ipilimumab were among the first immune-oncology agents to show significantly improved outcomes for patients. However, existing anti-CTLA-4 therapies fail to induce a response in a majority of patients and can induce severe, immune-related adverse events. It has been assumed that checkpoint inhibition, i.e., blocking the interaction between CTLA-4 and its ligands, is the primary mechanism of action for ipilimumab. In this study we present evidence that checkpoint inhibition is not a primary mechanism of action for efficacy of anti-CTLA-4 antibodies. Instead, the primary mechanism for efficacy is FcR-mediated Treg depletion in the tumor microenvironment. First, we identified a monoclonal antibody (mAb) that binds to CTLA-4 at an epitope that differs from ipilimumab’s by only a few amino acids, yet has limited checkpoint inhibitor activity. Surprisingly, the weak checkpoint inhibitor has superior anti-tumor activity compared to ipilimumab in a murine model. The weak checkpoint inhibitor also induces less Treg proliferation and has increased ability to induce in vitro FcR signaling and in vivo depletion of intratumoral Tregs. Further experiments showed that the enhanced FcR activity of the weak checkpoint inhibitor likely contributes to its enhanced anti-tumor activity. Importantly, we also showed that weak checkpoint inhibition was associated with lower toxicity in murine models. Our work suggests that new anti-CTLA-4 drugs should be optimized for Treg depletion rather than checkpoint inhibition. ### Competing Interest Statement E.L.S., K.P.C., E.K.W., M.A.A., E.B., Y.Y.C., G.L.C., C.E., B.K.G., A.G., J.L., R.L., V.A.M., R.A.M., A.R.N., J.S., S.K.S., J.F.S., K.S., B.T., N.P.W., Y.W.L., A.S.A., and D.S.J. are/were employees of GigaGen, Inc. and received cash salary and equity shares for their work. L.T. was a consultant for PHASTAR, Inc., and received cash payments from GigaGen for statistical consulting.