Functional characterization of SARS-CoV-2 myeloid cell receptors as an immunopathogenic mechanisms of COVID-19

Project Summary The rapid spread of SARS-CoV-2 and the resulting Coronavirus Disease 2019 (COVID-19) pose a global pandemic. SARS-CoV-2 and its family members share some puzzling, unique pathological features, most notably acute respiratory distress syndrome (ARDS), cytokine release syndrome (CRS), and lymphopenia, despite excessive myeloid cell-dominant inflammation, which has been correlated with COVID-19 severity. How the virus engages and dysregulates the immune system is currently unknown; although angiotensin-converting enzyme 2 (ACE2) is the canonical SARS-CoV-2 receptor, immune cells, particularly myeloid cells, express little, if any, levels of ACE2 despite evidence for direct viral engagement. Based on our extensive experience in the discovery and characterization of immune-modulatory receptor-ligand interactions, we sought to address how SARS-CoV-2 interacts with myeloid cells, which play essential roles in both virus innate sensing and the modulation of host immunity, by developing a myeloid cell receptor-focused ectopic expression screen. In ongoing experiments, we have identified several novel glycan-dependent host interaction partners for SARS-CoV-2 Spike (S) protein, including several C-type lectins and Tweety Family Member 2 (TTYH2). Pulmonary single-cell RNA sequencing (scRNA-seq) analysis in COVID-19 patients indicates a myeloid cell-dominant expression of these receptors as opposed to ACE2. In the preliminary studies, we have shown that these molecules interact mostly through regions outside of the ACE2 receptor-binding domain (RBD), suggesting that they may provide a novel function outside of virus entry. Although these receptors do not support the active replication of authentic SARS-CoV-2, the direct virus-myeloid cell engagement induces a robust pro-inflammatory response, which is blocked by receptor-decoy proteins and a picomolar-affinity anti- spike bispecific nanobody that also blocks virus infection through ACE2. Together, our findings provide the first evidence for direct immune modulation by SARS-CoV-2, potentially targeted for therapeutic benefit. Given these findings, we hypothesize that novel SARS-CoV-2 virus-receptor interactions in myeloid cells constitute a pathogenic pathway for COVID-19, serving as signaling receptors that directly drive myeloid cell dysregulation. These new direct virus-immune interactions may also have heretofore unexplored functions to affect other cells indirectly. These hypotheses will be addressed within the following Specific Aims: (1) to determine the functional contribution of each myeloid cell receptor in the induction of pro-inflammatory responses upon SARS-CoV-2 engagement, and (2) to determine the role of these interactions in the dysregulation of adaptive immunity and SARS-CoV-2 trans-infection through ACE2 receptor. Our study on novel virus myeloid cell receptors that govern aberrant immune responses would greatly contribute to our understanding of COVID- 19 pathogenesis, revealing new therapeutic targets against COVID-19 for the benefit of humanity at large.