Dissecting mechanisms of antibody-Fc interactions that shape innate immune antiviral responses. - PROJECT SUMMARY Antibodies are critical mediators of immune protection against pathogens, simultaneously acting to neutralize entry and activate innate immune cells through interaction between the antibody Fc domain and Fc-receptors (FcRs) on innate immune cells. Leveraging Fc-mediated activation in monoclonal antibody-based therapies is increasingly being investigated for the treatment of infectious diseases but the precise mechanisms and breadth of functions that can be induced by range of FcR-bearing immune cells is still unclear. However, these efforts are limited by the fact that we do not fully understand the spectrum of effector functions and effector programs that are induced via different FcRs, which could greatly impact efficacy and safety of mAb-based therapeutics. Moreover, as some innate immune cells such as monocytes express multiple FcRs, understanding how these pathways interact will help in the development of rational antibody design strategies to effectively harness innate immune cells against a variety of pathogens. There are four activating FcRs that bind IgG and are expressed on innate immune cells (FcγR1, FcγR2A, FcγR3A, and FcγR3B) and one inhibitory receptor (FcγR2B). The interplay between these receptors can impact Fc-effector functions, especially in monocytes that express multiple FcRs. Moreover, antibody-mediated activation of non-classical monocytes that express 3 activating FcγRs must balance protection and pathology, as antibody-dependent enhancement of infection/disease has been observed in some contexts. Thus, defining the transcriptional profiles of FcR-mediated activation of will be critical to better predict in vivo activity of immunotherapies and antibodies induced by vaccination or infection. To that end, we propose to begin to generate an ‘FcR Effector Atlas’ that defines the anti-pathogen pathways that are induced in distinct innate immune cell types mediated by different FcRs. The Atlas can be used to map the pathways induced across all innate immune cells to help guide development of optimally efficacious and safe antibody therapeutics against infectious diseases. By combining Fc-engineered antibodies that facilitate individual or combinatorial activation of specific FcRs and nascent transcriptomics, the goal of this proposal is pioneer an approach to generate the Atlas in nonclassical monocytes. We will use SARS-CoV-2-specfic Fc-engineered monoclonal antibodies as a model system to specifically engage different FcRs on monocytes to define the FcR-mediated transcriptional network using capped small RNA-sequencing (csRNA-seq) that enables transcriptional profiling of nascent RNA transcriptional start sites. Further, we will functionally link induction of specific pathways to restriction of SARS- CoV-2 infection. Together, the approaches and results generated from this proposal will contribute to our fundamental understanding of how antibodies leverage innate immunity and can be used to guide development of highly effective and safe immunotherapeutics for infectious diseases and beyond.
