2903 - Radiation Combined with Anti-PD-1 Inhibitor and Myocardial Injury via NLRP3 Inflammasome in Mice

J. Wang¹, Y. Wu¹, Y. Zhou², N. Zhang³, and S. Wang¹; ¹Department of Radiation Oncology, the Fourth Hospital of Hebei Medical University, Shijiazhuang, China, ²Hebei General Hospital, Shijiazhuang, China, ³Shijiazhuang People’s Hospital, Shijiazhuang, China

Purpose/Objective(s): The combination of immune checkpoint inhibitors and thoracic radiotherapy has demonstrated clinical activity, yet cardiac adverse events, although rare, are potentially fatal. This study evaluates the acute cardiac damage effects induced by radiation at different doses and radiation concurrent with an anti-PD-1 inhibitor (aPD-1). Materials/

Methods: Cardiac irradiation was delivered in a mouse model concurrently with aPD-1 (anti-mouse PD-1) to evaluate cardiac toxicity by mortality, transthoracic echocardiography, cardiac troponin I (cTNI) level, NT-proBNP, and histological analysis with HE & Masson staining. Radiation with 6-MV X-ray was delivered at a single fraction of 20 Gy (RT-20), 30 Gy (RT-30), and 45 Gy in three weekly fractions (RT-45). Anti-PD-1 inhibitor was administered intraperitoneal injections every second day at 10µg/g. Cardiac damage was assessed on the 21st day after the first irradiation. NLRP3 inflammasome activation in cardiomyocytes was assessed by western blot (WB) and immunofluorescence (IF). IL-1ß and IL-18 levels in serum were tested by ELISA.
Results: Acute mortality was 80% and 40% within 21 days in the RT-30 group and RT-30+aPD-1 group, respectively, while 0 in the other groups. On the 21st day, a significant decrease in left ventricular ejection fraction (LVEF) was observed in the RT-45 group (58.66%±2.69 vs. 38.40%±3.54, P<0.001) but was improved in the RT-45+aPD-1 group (47.28%±1.01 vs. 38.40%±3.54, P<0.01), with no significant change with 20 Gy. Cardiac troponin I, a marker of myocardial injury, significantly increased after radiation with 45 Gy and 30 Gy and was higher than each concurrent with aPD-1 (1652.00 pg/ml±215.80 for RT-45 vs. 903.9 pg/ml±96.76 for RT-45+aPD-1; 2649.00 pg/ml±69.69 for RT-30 vs. 1027.00 pg/ml±56.33 for RT-30+aPD-1, P<0.0001 for both). No statistical difference was observed between RT-20 and RT-20+aPD-1. Histological damage and deposition of collagen following radiation were mainly observed in the endocardium. Less myocardial necrosis was observed in the aPD-1 group. However, histological damage was alleviated in the combined group. NLRP3 inflammasome activation with increased levels of NLRP3 and Caspase-1 was identified by WB. Consistently, levels of IL-1ß and IL-18 in mice serum were elevated, especially in the radiation group. Co-localization of NLRP3 and WGA, a protein that specifically binds to the myocardial cell membrane, by IF images indicated that NLRP3 increased in cardiomyocytes.
Conclusion: Acute cardiac toxicity should be concerned, especially with cardiac biological effective dose >100 Gy. Cardiac damage induced by radiation concurrent with aPD-1 is complicated. Compared to radiotherapy or immune checkpoint inhibitor alone, the severity and mechanism are currently undefined and require further verification in future studies. NLRP3 inflammasome activation in cardiomyocytes might be one of the mechanisms of radiation concurrent with aPD-1-induced myocardial injury.