Combining PYROXD2 Overexpression with Anti-PD-1 Immunotherapy Suppresses NSCLC by Promoting NK Cell Infiltration

Background Non-small cell lung cancer (NSCLC) remains a leading cause of cancer-related mortality. The limited efficacy of current immunotherapies underscores an urgent need for novel therapeutic targets. PYROXD2 has been implicated as a potential tumor suppressor, yet its functional role in NSCLC and the mechanisms by which it modulates the immune microenvironment remain poorly understood. Methods Bioinformatics analysis was employed to determine the expression profile and prognostic value of PYROXD2. Its expression in NSCLC tissues and cell lines was validated by qRT-PCR, Western blot, and immunohistochemistry. In vitro and in vivo experiments were conducted; in the in vivo setting, an anti-programmed cell death protein 1 (anti-PD-1) combination treatment was included. Flow cytometry, Transwell co-culture assays, and transcriptome sequencing were utilized to explore its regulatory effects on the tumor microenvironment (TME) and associated signaling pathways. Results PYROXD2 was significantly downregulated in NSCLC tissues and cell lines, and its low expression predicted poor prognosis. Both in vitro and in vivo experiments confirmed that PYROXD2 overexpression markedly inhibited the proliferation, migration, and invasion of NSCLC cells and reduced tumor growth in vivo. Mechanistically, PYROXD2 overexpression inhibited the phosphoinositide 3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) signaling pathway, leading to increased interferon-γ (IFN-γ) and decreased tumor necrosis factor-α (TNF-α) levels in tumor tissues, which in turn promoted natural killer (NK) cell infiltration and reduced the accumulation of regulatory T cells (Tregs). Furthermore, anti-PD-1 treatment further enhanced the anti-tumor effect of PYROXD2 overexpression in vivo, as shown by greater tumor growth inhibition in the combination group. Notably, IFN-γ neutralization attenuated NK cell recruitment and partially reversed the anti-tumor effects of PYROXD2, indicating that the PI3K/AKT–IFN-γ–NK cell axis is critical for PYROXD2-mediated tumor suppression. Conclusion This study identifies a novel tumor-suppressive function of PYROXD2 in NSCLC and reveals a key immunomodulatory mechanism, providing a potential combination partner for anti-PD-1 immunotherapy in NSCLC