Anti–CTLA-4 antibody induces selective depletion of T reg cells within tumor lesions in a manner that is dependent on the presence of Fc gamma receptor-expressing macrophages within the tumor microenvironment.

Tumors are composed of multiple cell types besides the tumor cells themselves, including innate immune cells such as macrophages. Tumor-associated macrophages (TAMs) are a heterogeneous population of myeloid cells present in the tumor microenvironment (TME). Here, they contribute to immunosuppression, enabling the establishment and persistence of solid tumors as well as metastatic dissemination. We have found that the pattern recognition scavenger receptor MARCO defines a subtype of suppressive TAMs and is linked to clinical outcome. An anti-MARCO monoclonal antibody was developed, which induces anti-tumor activity in breast and colon carcinoma, as well as in melanoma models through reprogramming TAM populations to a pro-inflammatory phenotype and increasing tumor immunogenicity. This anti-tumor activity is dependent on the inhibitory Fc-receptor, FcγRIIB, and also enhances the efficacy of checkpoint therapy. These results demonstrate that immunotherapies using antibodies designed to modify myeloid cells of the TME represent a promising mode of cancer treatment.

The in vivo biological activities of IgG antibodies result from their bifunctional nature, in which antigen recognition by the Fab is coupled to the effector and immunomodulatory diversity found in the Fc domain. This diversity, resulting from both amino acid and glycan heterogeneity, is translated into cellular responses through Fcγ receptors (FcγRs), a structurally and functionally diverse family of cell surface receptors found throughout the immune system. Although many of the overall features of this system are maintained throughout mammalian evolution, species diversity has precluded direct analysis of human antibodies in animal species, and, thus, detailed investigations into the unique features of the human IgG antibodies and their FcγRs have been limited. We now report the development of a mouse model in which all murine FcγRs have been deleted and human FcγRs, encoded as transgenes, have been inserted into the mouse genome resulting in recapitulation of the unique profile of human FcγR expression. These human FcγRs are shown to function to mediate the immunomodulatory, inflammatory, and cytotoxic activities of human IgG antibodies and Fc engineered variants and provide a platform for the detailed mechanistic analysis of therapeutic and pathogenic IgG antibodies.

CD40, a member of the TNF receptor (TNFR) superfamily, is expressed on antigen presenting cells (APCs) and is essential for immune activation. While agonistic CD40 antibodies have been developed for immunotherapy, their clinical efficacy has been limited. We have found that co-engagement of the Fc domain of agonistic CD40 monoclonal antibodies (mAbs) with the inhibitory Fcγ receptor FcγRIIB is required for immune activation. Direct comparison of anti-CD40 mAbs enhanced for activating FcγR binding hence capable of cytotoxicity, or FcγRIIB binding revealed that enhancing FcγRIIB binding conferred immunostimulatory activity and considerably greater anti-tumor responses. This unexpected requirement for FcγRIIB in enhancing CD40 mediated immune activation has direct implications for the design of agonistic, anti-TNFR antibodies as therapeutics.

Summary While engagement of the inhibitory Fcγ-Receptor (FcγR) IIB is an absolute requirement for in vivo antitumor activity of agonistic mouse anti-CD40 monoclonal antibodies (mAbs), a similar requirement for human mAbs has been disputed. By using a mouse model humanized for its FcγRs and CD40, we revealed that FcγRIIB-engagement is essential for the activity of the human CD40 mAbs, while engagement of the activating FcγRIIA inhibits this activity. By engineering Fc variants with selective enhanced binding to FcγRIIB, but not to FcγRIIA, significantly improved antitumor immunity was observed. These findings highlight the necessity of optimizing the Fc domain for this class of therapeutic antibodies by using appropriate pre-clinical models that accurately reflect the unique affinities and cellular expression of human FcγR.