Pharmacological inhibition of JNK-MAPK disrupts cigarette smoke-induced RUNX2/Galectin-3 -driven EMT and cancer stemness in lung adenocarcinoma cells

Biochem Pharmacol. 2025 Dec;242(Pt 3):117399. doi: 10.1016/j.bcp.2025.117399. Epub 2025 Oct 3.

Abstract

Cigarette smoke (CS), a major driver of lung cancer (LC), promotes epithelial-mesenchymal transition (EMT) and stemness resulting in metastasis, therapy resistance, and recurrence, but the precise mechanism is elusive. Building on our earlier identification of Runt related transcription factor-2 (RUNX2) and Galectin-3 (Gal-3) as mediators of CS-induced EMT, in this study, we aimed to identify a potential molecular mechanism and delineate the upstream regulators of RUNX2 using A549 lung adenocarcinoma cells and human small airway epithelial cells (SAECs) cultured at the air-liquid interface (ALI). CSE exposure markedly elevated intracellular reactive oxygen species (ROS), assessed via Dichloro-dihydro-fluorescein diacetate (DCFH-DA) assay, and promoted invasive behavior (Boyden chamber assay), spheroid formation, and colony formation, the hallmarks of cancer stemness. Expression analysis via RT-qPCR, immunoblotting, and immunocytochemistry revealed that CSE upregulated EMT and stemness-associated markers, notably via upregulating RUNX2 and Galectin-3, at both transcriptional and translational levels through the involvement of c-Jun N-terminal kinase- Mitogen-Activated Protein Kinase (JNK-MAPK) pathways. A specific pharmacological inhibitor of JNK (SP600125) significantly attenuated CSE-induced RUNX2 and Galectin-3 (Gal-3) expression, and also reversed CSE-driven EMT marker alterations, suppressed transcriptional EMT perturbations, and reduced proinflammatory cytokines, including monocyte chemoattractant protein-1 (MCP-1), interleukin-8 (IL-8), and tumor necrosis factor-alpha (TNF-α). In conclusion, this study identifies that ROS/JNK/RUNX2/Gal-3 axis drives CS-induced oncogenic plasticity, suggesting that targeted inhibition of this pathway could be an effective strategy for mitigating CS-related LC progression.

Keywords: Cigarette smoke; Epithelial-mesenchymal transition (EMT); Galectin-3; JNK; Lung cancer; RUNX2; Reactive oxygen species (ROS); Stemness.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • A549 Cells
  • Adenocarcinoma of Lung* / metabolism
  • Adenocarcinoma of Lung* / pathology
  • Blood Proteins
  • Cigarette Smoking* / adverse effects
  • Core Binding Factor Alpha 1 Subunit* / genetics
  • Core Binding Factor Alpha 1 Subunit* / metabolism
  • Epithelial-Mesenchymal Transition* / drug effects
  • Epithelial-Mesenchymal Transition* / physiology
  • Galectin 3* / genetics
  • Galectin 3* / metabolism
  • Galectins
  • Humans
  • JNK Mitogen-Activated Protein Kinases* / antagonists & inhibitors
  • JNK Mitogen-Activated Protein Kinases* / metabolism
  • Lung Neoplasms* / drug therapy
  • Lung Neoplasms* / metabolism
  • Lung Neoplasms* / pathology
  • MAP Kinase Signaling System / drug effects
  • MAP Kinase Signaling System / physiology
  • Neoplastic Stem Cells* / drug effects
  • Neoplastic Stem Cells* / metabolism
  • Neoplastic Stem Cells* / pathology
  • Protein Kinase Inhibitors* / pharmacology
  • Reactive Oxygen Species / metabolism
  • Smoke* / adverse effects

Substances

  • RUNX2 protein, human
  • Core Binding Factor Alpha 1 Subunit
  • Galectin 3
  • LGALS3 protein, human
  • Smoke
  • Reactive Oxygen Species
  • Protein Kinase Inhibitors
  • JNK Mitogen-Activated Protein Kinases
  • Blood Proteins
  • Galectins