CXCR6⁺ T Cells Drive Immune Checkpoint Inhibitor Myocarditis.

Abstract

Myocarditis is a severe complication of immune checkpoint inhibitors (ICIs). The major risk factor for ICI myocarditis is the use of combination ICI treatment, especially when relatlimab, a novel anti-LAG-3 (lymphocyte-activation gene 3) antibody, is combined with anti-PD-1 (programmed cell death protein 1) therapy. Although pathogenic T cells are necessary for ICI myocarditis, the specific signaling and T-cell populations that drive cardiac infiltration have not been fully elucidated, especially in setting of anti-LAG-3/PD-1 treatment. We used VigiBase, an international pharmacovigilance database, to assess the risk of myocarditis with anti-LAG-3 compared with other ICI treatment regimens. We identified a mouse model of LAG-3/PD-1-associated ICI myocarditis through genetic deletion of immune checkpoints LAG-3 and PD-1 (<i>Lag3</i>^<i>-/</i>-, <i>Pdcd1</i>^<i>-/</i>- mice) and performed rigorous cardiac phenotyping using histology, flow cytometry, electrocardiography, single-cell RNA sequencing, and antibody-induced cellular depletion. We found an increased risk of myocarditis with anti-LAG-3 combination treatment clinically, confirming early clinical trial data. <i>Lag3</i>^<i>-/</i>-, <i>Pdcd1</i>^<i>-/</i>- mice were found to develop severe cardiac inflammation by histology with increased cardiac macrophages and clonal T cells, which was associated with the development of spontaneous arrhythmias leading to premature death by 6 to 8 weeks. We identified CXCR6 (C-X-C motif chemokine receptor 6) as a key marker of activated cardiac T cells in this model, along with analogous signals in other preclinical models and patient data. CXCR6 marked a heterogenous group of cardiac T cells, including distinct clusters of <i>Gzmk</i>, <i>Gzmb</i>, <i>Cd4</i>, and actively dividing T cells. CXCL16 (C-X-C motif chemokine ligand 16), the sole known ligand for CXCR6, was similarly upregulated in the cardiac macrophage population. Treatment with anti-CXCR6 antibody prevented premature lethality, attenuated arrhythmias, and reduced the histological severity of myocarditis, demonstrating that CXCR6⁺ T cells are necessary for disease pathogenesis. Our findings suggest that ICI myocarditis is driven by an expansion of CXCR6⁺ T cells and identifies CXCR6 as a putative therapeutic target for this highly morbid condition.