Abstract 2812: NL-201, a de novo CD25-independent IL-2Rβγ agonist, synergizes with radiation to generate potent antitumor immunity
Authors: Huntoon Kristin , Lee DaeYong , Li Xuefeng , Wang Yifan , Jiang Wen , Kim Betty 
Published: April 2023

Abstract
There is a growing interest in combining radiotherapy (RT) with immunotherapeutic agents to improve efficacy. One such approach is RT in combination with high-dose recombinant interleukin 2 (IL-2), which elicits anti-tumor immune responses by stimulating T-cell and NK cell populations. Although effective for some patients, the overall clinical benefit of high-dose recombinant IL-2 is limited by toxicity and the expansion of regulatory T cells (Tregs) through signaling events mediated by IL2Rα (CD25). Here we examined the cytosolic DNA sensor cyclic GMP-AMP synthase-stimulator of interferon genes (cGAS-STING) pathway dependency of type I interferon (IFN) response and anti-tumor effect of RT in combination with NL-201, a highly potent and stable CD25-independent IL-2 and IL-15 receptor agonist with enhanced affinity for IL2Rβ and IL2Rγc. In mice implanted with E0771 syngeneic tumors, NL-201 in combination with RT enhanced activation of the STING pathway. This was associated with increased type I IFN production as well as increased activation and number of tumor associated dendritic cells (DCs). In STING (-/-) mice implanted with E0771 tumors, Type I IFN production, tumor-growth inhibition (TGI), and survival advantage was significantly decreased and there were significantly fewer activated CD4+ and CD8+ T-cells. Similarly, CD11c+ knock-out mice implanted with E0771 tumors and treated with the combination exhibited similar TGI, survival, and IFN production compared to vehicle treated mice. Based on these results, we tested the ability of the combination of RT and NL-201 to enhance systemic immunotherapy with an anti-PD-1 checkpoint inhibitor in mice implanted with 4T1 breast tumor cells. Tumor-bearing mice treated with the triple combination exhibited superior TGI and survival, along with increased effector memory T-cell populations as compared to all other treatment groups. These results support our previous findings that the CD25-independent IL-2 and IL-15 receptor agonist, NL-201, synergizes with RT, likely via the direct activation of STING. Genetic knockout of STING in tumor-bearing mice abrogated the activity of the NL-201 and RT combination, thus confirming the crucial role of innate immune sensors in the combination treatment’s immune activation and resulting antitumor effect. These results also describe the important role of STING-mediated activation and tumor penetration of DCs, which were crucial for anti-tumor response. Additionally, when combined with anti-PD-1 antibodies, NL-201+RT combination treatment significantly improved TGI of metastatic breast cancers that were previously resistant to checkpoint blockade. This work provides rationale for the continued exploration of RT in combination with CD25-independent IL-2Rβγ agonism as a means to enhance the systemic immune response to treat immunologically challenging tumors.
Citation Format: Kristin Huntoon, DaeYong Lee, Xuefeng Li, Yifan Wang, Wen Jiang, Betty Kim. NL-201, a de novo CD25-independent IL-2Rβγ agonist, synergizes with radiation to generate potent antitumor immunity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2812.