Combined in vivo and in cellulo approach to study the role of endothelial cells and the NLRP3 inflammasome in cardiac dysfunction associated with immune-mediated myocarditis

Immunotherapy has led to considerable clinical improvement in cancer patients, but is associated with serious side effects such as myocarditis, which is associated with a high mortality risk. Immune checkpoint inhibitor-induced myocarditis (ICI-M) is characterised by T cells and macrophages infiltration within the myocardium, which lead to necrosis of cardiac tissue. The NLRP3 inflammasome pathway, associated with interferon-γ (IFN-γ), is overexpressed within the myocardium of ICI-M patients.

To study the pathophysiological mechanisms of ICI-M using a preclinical model to investigate the electrophysiological disorders associated with myocardium immune infiltration, and to unravel the molecular mechanisms in a cellular model using IPSC from ICI-M patient.

The preclinical ICI-M model was established by subcutaneous injection of murine melanocytic cells into BALB/c mice, treated with anti-PD1/anti-CTLA-4 combination therapy. IPSC-derived cardiomyocytes (CM) and endothelial cells (EC), generated from ICI-M patient and healthy donor, were exposed to IFN-γ.

A decrease in tumour volume was observed in mice exhibiting ECG disturbances, with a reduction in QRS and T wave amplitude, as well as a reduction in repolarisation time. Impaired cardiac function was associated with increased CD3 transcript and overexpression of PD-L1 in the myocardium. In hiPSC-derived CM and EC, genes involved in immune response (PD-L1, MHC-II) and in NLRP3 inflammasome (GBP5, GBP6, NLRC5) were up-regulated by IFN-γ. A large number of dysregulated genes were found in EC compared to CM after stimulation. The nature of the inflammasome activated by IFN-γ was different between the two cell types, NLRP3 in hiPSC-CM and AIM2 in hiPSC-EC. The hiPSC-EC from ICI-M patient showed specific regulations, in the immune response (PD-L2, TLR2), and in secreted pro-inflammatory cytokines (CCL2, CCL5, IL-1b), associated with NLRP3 inflammasome.

Mice treated with ICI show ECG disturbances correlated with myocardial inflammation. Our cellular model highlighted the major role played by hiPSC-EC in the response to IFN-γ and the specific regulation of the NLRP3 inflammasome. We would like to further investigate the role of endocardium and microvascularisation in the mechanisms of ICI-M.