EML4–ALK Fusion Rewires Transcriptomic, miRNA, and CAF-Associated Programs in Non-Small Cell Lung Cancer

This study establishes an integrative framework that combines paired mRNA/miRNA profiling with immune microenvironmental features to clarify how EML4–ALK fusions shape transcriptomic and post-transcriptional networks in Non-small cell lung cancer (NSCLC). Using paired mRNA-seq and miRNA-seq data generated from the same patients, we compared fusion-positive and fusion-negative NSCLC across three interconnected layers: (i) transcriptome architecture, including differential expression, pathway, and network analyses; (ii) the miRNA–mRNA regulatory axis, encompassing dysregulated miRNAs, target repression and sponging, and fusion-specific regulatory pairs; and (iii) the tumor microenvironment, with emphasis on immune and stromal infiltration, particularly cancer-associated fibroblast (CAF)-linked extracellular matrix (ECM) and adhesion programs. Our analyses revealed a distinct reprogramming pattern in fusion-positive NSCLC, marked by activation of metabolic and proteostasis pathways, including N-glycan metabolism coupled to ER export, together with attenuation of immune–stromal communication, adhesion, ECM, calcium signaling, and PI3K/VEGF-axis transcription relative to fusion-negative NSCLC. We also identified fusion-associated microRNA perturbations, including an exclusively upregulated miR-3065-centered regulatory hub predicted to repress ECM- and adhesion-related genes (PDGFRB, CTSK, COL4A2, SPARC, FBN1, and LUM) in fusion-positive tumors, in contrast to broader miRNA network rewiring in fusion-negative tumors targeting ciliary and mitotic hubs. Tumor microenvironment analysis further distinguished the subtypes, with fusion-positive tumors showing reduced CAF infiltration relative to fusion-negative tumors and concordant gene–CAF associations. By linking mechanistic insight with candidate biomarkers and targetable pathway nodes, this work provides a basis for precision strategies in both fusion-positive and fusion-negative cohorts and broadens the therapeutic perspective beyond kinase inhibition alone.