Beyond the periphery: the thymus as the central modulator of Anti-PD-1 induced cardiac dysfunction
M Kocsis, A Kulin, L Szabo, T Gergely, N V Sayour, V Toth, D Nagy, T Kovacs, Z S Hegedus, L Varga, B Agg, P Ferdinandy, Z V VargaAbstract
Introduction
Immune checkpoint inhibitors (ICIs) have revolutionized oncology, yet their use is compromised by immune-related adverse events (irAEs), among which cardiotoxicity remains the most lethal. Current dogma attributes these events to the peripheral reactivation of exhausted autoreactive T-cells. However, recent clinical observations linking thymic epithelial tumours and hyperplasia with severe myocarditis suggest a critical, upstream role for the thymus (1, 2, 3). We hypothesize that anti-PD-1 therapy actively disrupts central tolerance mechanisms within the thymus and serves as a priming mechanism to the already established disruption of peripheral tolerance and irAE mechanisms.
Purpose
The aim of this study was to determine whether active thymic function is a prerequisite for ICI-induced cardiotoxicity and to test whether thymic involution either physiological (aging) or pharmacological (5-Azacytidin, AZA) protects against cardiac dysfunction.
Methods
We compared young (12-week) versus aged (16-month) C57BL/6J mice treated with anti-PD-1 (200 µg i.p., 6 dose in 2 weeks). Pharmacological thymic atrophy was induced in young mice with AZA (2.5 mg/kg daily). Our endpoints were high-resolution echocardiography, myocardial qRT-PCR, bulk thymic RNAseq with GO enrichment, spatial RNAscope (Ets1, Ccl21a, Cxcl9), morphological assessment, histology/PCNA/CD25 immunhistochemistry (IHC), flow cytometry of the thymus and spleen, and organ toxicity screens.
Results
Anti-PD-1 in young mice induced a reproducible decline in systolic function but not in aged. Young thymuses showed robust transcriptional reprogramming (212 differentially expressed genes including leukocyte activation, T-cell differentiation, ribosomal biogenesis), medullary enrichment of Cxcl9 and spatial disruption of Cd3e/Ccl21a. Flow cytometry and IHC revealed expansion of DN2/DN3 and DP thymocytes, increased CD25 and PCNA, and reduced intrathymic Tregs. Cardiac tissue of affected mice had elevated Cxcl9/Cxcl10, Ifn-γ and Il23 without overt myocarditis. Physiological and pharmacological thymic involution with 5-azacitidine attenuated anti-PD-1-induced systolic dysfunctions and decreased cardiac chemokine/cytokine levels.
Conclusion
Expanding the current peripheral-centric paradigm, we identify the thymus as a key driver complementing peripheral mechanisms of ICI-cardiotoxicity. Anti-PD-1 disrupts central tolerance by lowering TCR thresholds and impairing selection, driving a cardiac-specific inflammatory loop dependent on active thymic output. Physiological (aging) or pharmacological (AZA) involution prevents this phenotype, suggesting that thymic activity modulates susceptibility. Thus, thymic activity-based stratification and targeted modulation could serve as a potential target in reducing immune mediated side effect and identify high risk patients.