Anti-tumor effect of CTLA-4 antibody is independent of checkpoint blockade

Abstract Antibodies targeting CTLA-4 are emerging as an important class of cancer therapeutics. It is assumed that these antibodies cause tumor rejection by blocking negative signaling from the CTLA-4-B7 interactions to enhance the priming of naïve T cells in lymphoid organs. However, recent findings have shown that the effectiveness of CTLA-4 antibody critically depends on the Fc domain and the host Fc receptors. It remains unclear if the blocking function of CTLA-4 antibody is required for its anti-tumor activity. To address this, here we have selected a non-blocking anti-CTLA-4 antibody (D138) and assessed its binding property and antitumor activity in comparison with the therapeutic CTLA-4 antibody ipilimumab. Crystal structures of CTLA-4 complexed with these antibodies show that D138 binds to a distinctly different site to that of ipilimumab on the CTLA-4 surface. D138 binding did not block the association of cells expressing CTLA-4 and B7 whereas ipilimumab did. Subsequent antitumor assay revealed that D138 was similarly effective as ipilimumab in inhibiting tumor growth in mice. This antitumor activity required Fc function for efficacy and was correlated with selective reduction of intratumor regulatory T (Treg) cells, resulting in a significant increase in the ratio of CD8+ over Treg cells. Overall these data clearly demonstrate that blocking CTLA-4-B7 interaction is not required for CTLA-4 antibody mediated antitumor activity, opening prospects of developing non-blocking CTLA-4 antibodies or simple binders towards other Treg surface markers for Treg-targeted immunotherapy.