<p>Leukemia inhibitory factor (LIF) is an established oncogenic driver in multiple cancers. However, its functional contribution and mechanistic underpinnings in lung squamous cell carcinoma (LSCC) remain incompletely defined. To delineate the role of LIF in LSCC progression, and elucidate the underlying molecular mechanisms. LIF protein expression in clinical LSCC tissues was assessed by immunohistochemistry (IHC) and Western blotting. LIF gain-of-function (using recombinant LIF protein) and loss-of-function (via LIF knockdown) assays were performed in vitro to assess its impact on the malignant phenotypes of LSCC cells. The changes in signaling pathways were analyzed by Western blotting and immunofluorescence. The functional role of LIF was further validated in patient-derived organoid models and a nude mouse xenograft model. LIF protein was significantly upregulated in LSCC tissues, and its high expression was correlated with unfavorable clinical outcomes. Recombinant LIF (rLIF) significantly enhanced the proliferation, migration, and invasion of LSCC cells in vitro in a dose-dependent manner, and the dosage of 200 ng/mL exerted the most pronounced effects. Conversely, LIF knockdown suppressed these malignant phenotypes. LIF activated the PI3K/AKT pathway, leading to inhibitory phosphorylation of GSK-3β at Ser 9, which consequently stabilized β-catenin and facilitated its nuclear accumulation. Furthermore, rLIF also promoted the growth of patient-derived organoids, and upregulated the expression of Ki-67 and α-SMA. In contrast, LIF knockdown significantly attenuated the growth of LSCC xenografts in vivo, and this effect was partially reversed by exogeneous rLIF. The oncogenic effects of LIF were associated with increased expression levels of p-AKT, p-GSK-3β, and β-catenin. LIF mediates LSCC progression by activating the PI3K/AKT/GSK-3β/β-catenin pathway. Therefore, LIF and its effectors warrant further study as potential diagnostic biomarkers and therapeutic targets in LSCC.</p>