Emergence of Disease Specific Cardiac Macrophages in Immune Checkpoint Inhibitor Myocarditis
Immune checkpoint inhibitors (ICIs), antibodies targeting PD-1/PD-L1 or CTLA4 have revolutionized cancer management but are associated with devastating immune-related adverse events (irAEs) including myocarditis. The main risk factor for ICI myocarditis is the use of combination PD-1 and CTLA4 inhibition. ICI-myocarditis is often fulminant and is pathophysiologically characterized by myocardial infiltration of T lymphocytes and macrophages. While much has been learned regarding the role of T-cells in ICI-myocarditis, little is understood regarding the identity, transcriptional diversity, and functions of infiltrating macrophages. We employed an established murine ICI myocarditis model (Ctla4+/-Pdcd1-/- mice) to explore the cardiac immune landscape using single-cell RNA-sequencing, immunostaining, and molecular imaging. We observed marked increases in CCR2+ monocyte-derived macrophages and CD8+ T-cells in this model. The macrophage compartment was heterogeneous and displayed marked enrichment in an inflammatory CCR2+ subpopulation expressing Cxcl9, Cxcl10, Gbp2b, and Fcgr4 that originated from CCR2+ monocytes.  Importantly, a similar macrophage population expressing CXCL9, CXCL10, and CD16a (human homologue of mouse FcgR4) were found selectively in patients with ICI myocarditis compared to other forms of heart failure and myocarditis. In silico prediction of cell-cell communication suggested interactions between T-cells and Cxcl9+Cxcl10+ macrophages via IFN-? and CXCR3 signaling pathways. Depleting CD8+T-cells and blockade of IFN-? signaling blunted the emergence of Cxcl9+Cxcl10+ macrophages in the heart and attenuated myocarditis suggesting that this interaction was necessary for disease pathogenesis. Collectively, these data demonstrate that ICI-myocarditis is associated with the emergence of a specific population of inflammatory macrophages and suggest the possibility that IFN-? blockade may serve as an effective treatment for this devastating condition. Overall design: To reveal the cardiac  immune landscape of immune chechpoint inhibitors (ICI) myocarditis, we performed single-cell RNA-sequencing (scRNA-seq) of heart tissue from control (Ctla4+/+Pdcd1-/-, n=4) and ICI myocarditis (Ctla4+/-Pdcd1-/-, n=10) mice. We constructed five libraries (control: L1, L2; disease: L3, L4, L5) from DRAQ5+DAPI- cells purified from pooled enzymatically digested hearts by FACS.