Single-cell profiling of human heart and blood in immune checkpoint inhibitor-associated myocarditis

2507 Background: Myocarditis due to immune checkpoint inhibitors (ICIs) is uncommon; however, myocarditis due to ICIs leads to severe morbidity and even death in 20-40% of cases. The molecular underpinnings of ICI-associated myocarditis are poorly understood, and there is an unmet clinical need to identify therapeutic targets and biomarkers that can aid in disease management. Methods: Heart tissue was obtained through endomyocardial biopsy or autopsy of patients receiving ICIs and was profiled with paired single-cell RNA sequencing (scRNA-seq) and T cell receptor sequencing (TCR) using the 10x Genomics Chromium system. A control dataset was constructed using scRNAseq data of heart tissue from patients receiving ICIs but without myocarditis and a published dataset from healthy patients not receiving ICIs. Peripheral blood mononuclear cells (PBMCs) were collected at the time of myocarditis diagnosis in a larger cohort of patients and analyzed with ICI-treated controls. The CITE-Seq protocol was used to measure paired scRNA-seq, TCR, and surface proteomics in PBMCs, using serial timepoints where available. Results: Heart tissue from 13 patients with myocarditis, including three fatal cases, and seven controls yielded 77,712 single cells. Blood profiling from 27 patients with ICI myocarditis and ICI-treated controls across 54 samples yielded over 230,000 cells. ICI myocarditis tissue demonstrated an increased T cell infiltrate (OR 8.94, FDR = 0.0021). Expression of multiple inflammatory pathways, most notably interferon responses, was up-regulated across multiple immune and non-immune cell types in the setting of myocarditis, providing important pathophysiological insights. T cell clones were also found to be shared between blood and heart, enabling the identification of putative pathogenic T cell subsets. Conclusions: Increased intramyocardial T cells and the activation of interferon response gene networks were seen in the setting of ICI myocarditis. These preliminary findings highlight potential pathological pathways in ICI myocarditis that could serve as biomarkers or therapeutic targets.