ANGPTL4 contributes to osimertinib resistance through NDRG1-associated extracellular matrix remodeling in EGFR-mutant NSCLC organoid models.

backgroundAs a first-line treatment for EGFR-mutated non-small cell lung cancer (NSCLC), acquired resistance to osimertinib has become a major clinical challenge. Traditional two-dimensional cell culture models have significant limitations in simulating the tumor microenvironment, making it difficult to fully elucidate complex resistance mechanisms. In recent years, three-dimensional organoid models have emerged as a crucial platform for studying resistance mechanisms due to their superior ability to preserve tumor tissue architecture and heterogeneity. Angiopoietin-like protein 4 (ANGPTL4), a secreted glycoprotein, has been implicated in regulating extracellular matrix remodeling and is associated with chemotherapy resistance in multiple cancers. However, its specific role in osimertinib resistance remains unclear. Therefore, this study investigates the mechanism of ANGPTL4 in lung cancer resistance using organoid models.

methodsThis study established a three-dimensional osimertinib-resistant organoid model that highly mimics the tumor microenvironment. Transcriptome sequencing and bioinformatics analysis were used to screen key resistance mediators. Functional gain-of-function/loss-of-function experiments validated candidate gene functions. Immunoprecipitation, molecular docking, and single-cell RNA sequencing data were employed to explore the underlying mechanisms in depth. Finally, virtual screening was conducted to identify potential inhibitors targeting this resistance pathway.

resultsThis study revealed that ANGPTL4 was upregulated in osimertinib-resistant models and functionally contributed to reduced osimertinib sensitivity. Mechanistically, ANGPTL4 interacted with NDRG1 and was associated with activation of extracellular matrix-related programs. Knockdown of ANGPTL4 restored osimertinib sensitivity, whereas ANGPTL4 overexpression promoted a resistant phenotype. Additional validation in H1975-derived models further supported the reproducibility of ANGPTL4-associated resistance. Knockdown of ANGPTL4 in resistant models restored drug sensitivity, whereas overexpression induced resistance. NDRG1 is an essential molecule for ANGPTL4-regulated ECM pathway activation, exhibiting significant colocalization with ANGPTL4 in clinical samples. Knocking down NDRG1 in ANGPTL4-overexpressing drug-resistant models effectively restored drug sensitivity. Furthermore, virtual screening of 2.08 million small molecules yielded five lead compounds with high binding affinity for ANGPTL4.

conclusionsThis study supports ANGPTL4 as an important contributor to osimertinib resistance in EGFR-mutant NSCLC and suggests that ANGPTL4/NDRG1-associated ECM remodeling may represent a relevant resistance-associated pathway. Computational virtual screening further nominated candidate ANGPTL4-binding molecules, which require future biochemical and functional validation. It not only provides novel insights into resistance mechanisms but also identifies potential therapeutic candidates for overcoming resistance, highlighting the significant value of 3D organoid models in investigating resistance mechanism.