IL-9 stimulates an anti-tumor immune response and facilitates immune checkpoint blockade in the CMT167 mouse model

Abstract

<h3>Objectives</h3><p>There is mounting evidence that interleukin-9 (IL-9) is associated with various cancers although its function in lung cancer remains elusive. This study aimed to elucidate the role(s) of IL-9 in lung cancer and the mechanisms involved.</p><h3>Materials and methods</h3><p>Expression of IL-9 receptor (IL-9R) in two murine lung cancer cell lines: CMT167 and Lewis lung carcinoma (LLC) were assessed and syngeneic murine lung cancer models were established. Tumor growth, intratumoral immune responses and downstream signaling pathways in tumor-bearing mice were analyzed upon IL-9 treatment. Human lung cancer cell lines A549 and H1975 were included for <em>in vitro</em> validation. Synergistic effects and immune responses of IL-9 in combination with anti-PD-1 were studied.</p><h3>Results</h3><p>IL-9R expression was only detected in CMT167 but not LLC cells. IL-9 suppressed CMT167 tumor growth and enhanced anti-tumor T cell responses, both of which were absent in IL-9R-deficient LLC model and lost upon IL-9R knockdown in CMT167 model. In CMT167 tumors, while IL-9 increased CD4⁺ and CD8⁺ T cells and dendritic cells, the cytotoxic T subset was the key driver of IL-9-induced tumor suppression. Consistently, in CMT167 and A549 cells, IL-9/IL-9R signaling promoted MHC class I upregulation. Inhibition of ERK signaling abolished IL-9-mediated MHC class I upregulation in CMT167 cells. IL-9 induced expression of PD-1 and PD-L1 on CD8⁺ T lymphocytes and CMT167 cells respectively. Combined IL-9 treatment with PD-1 blockade further upregulated tumor-infiltrating CD8⁺ T cell frequencies and synergistically suppressed tumor growth in CMT167 model.</p><h3>Conclusion</h3><p>IL-9 suppresses tumor growth by promoting tumor-derived MHC class I presentation and enhancing cytotoxic T cell immunity. Expression of IL-9R might be used as a biomarker for identification of potential target population susceptible to IL-9 treatment. Our study proposes IL-9 as a promising therapeutic immunomodulatory agent that can be used in combination with PD-1 blockade in lung cancer.</p>