318 - Radiation Activates the AIM2 Inflammasome to Enhance Immunotherapy by Triggering IL-1ß Signaling and DC Activation in Colorectal Cancer

Purpose/Objective(s):

Microsatellite stable (MSS) colorectal cancer (CRC) responds poorly to PD-1 blockade monotherapy but can benefit from its combination with radiotherapy. Radiation induces antitumor immunity through multiple cytosolic nucleic acid sensing pathways. However, the role of the DNA-sensing absent in the melanoma 2 (AIM2) inflammasome and its downstream IL-1ß remains unclear. This study aims to investigate the function of the AIM2 inflammasome in combined radiotherapy and immunotherapy and to explore the underlying mechanisms.

Materials/Methods:

Two CT26 cell lines with distinct sensitivities to combined therapy were generated, by in vivo passaging for four cycles in BALB/c mice treated with 8Gy×3Fx radiation and PD-1 antibody. One cell line was sensitive, and the other resistant. Bulk RNA-seq, WB, and IHC were performed on two cell lines to investigate the correlation between the AIM2-IL-1ß pathway and the response to combined therapy. Flow cytometry and lentivirus-mediated shRNA targeting AIM2 were used to assess the impact of AIM2 on DC function. AIM2 expression in tumor samples was examined using scRNA-seq data from a clinical trial in which patients with locally advanced MSS rectal cancer underwent neoadjuvant chemoradiotherapy and PD-1 blockade.

Results:

RNA-seq revealed higher expression of AIM2 and a preferential enrichment of cytosolic DNA-sensing pathway in sensitive tumor cells. CIBERSORT analysis further showed that the expression level of AIM2 was positively correlated with the abundance of tumor-infiltrating immune cells including CD8+ T cells (total, Tcm, and Tem), non-regulatory and memory CD4+T cells, and activated DCs. Similar correlations were observed in colorectal samples from the TCGA database. Higher levels of AIM2 localized primarily to the nucleus of sensitive tumor cells were confirmed using WB and IHC. Importantly, 24h after 8Gy radiation, AIM2 was recruited to and densely concentrated within cytosolic micronuclei clusters resulting from nuclear damage in sensitive tumor cells. The AIM2 inflammasome was then activated evidenced by increased levels of cleaved caspase-1 and IL-1ß. Co-culture with BMDCs in vitro showed that irradiated sensitive tumor cells were more potent in inducing DC activation partially dependent on AIM2, as AIM2 knockdown reduced this effect. Consistent results were observed in DCs isolated from tumors and draining lymph nodes of mice treated with 8Gy×3Fx radiation and PD-1 antibody. Lastly, we checked samples from rectal cancer patients and found higher expression of AIM2 in responders to neoadjuvant therapy compared to non-responders.

Conclusion:

Overall, we demonstrated that radiation-induced activation of the AIM2 inflammasome enhanced antitumor responses by triggering IL-1ß signaling and subsequent DC activation. Targeting the AIM2-IL-1ß axis may represent a therapeutic strategy to improve combined radiotherapy and immunotherapy in MSS CRC.