Establishment of lung cancer organoid lines from malignant pleural effusion: medium optimization and specimen quality-control analysis

Lung cancer organoids (LCO) derived from malignant pleural effusion (MPE) offer unique advantages for modeling advanced disease, but existing methods suffer from variable success rates and limited capacity for long-term culture, hindering the establishment of stable organoid lines. Here, we systematically optimized the generation of MPE-derived LCO lines through a dual-strategy approach. Medium supplementation with 10%–25% pleural effusion supernatant (PES) significantly promoted organoid growth. Analysis of 35 clinicopathological parameters identified hemorrhagic effusion, positive liquid-based cytology and positive cell block as factors associated with successful organoid establishment. In a multivariable model, hemorrhagic effusion (adjusted OR 3.47, 95% CI 1.18–10.2) and positive liquid-based cytology (adjusted OR 6.92, 95% CI 1.43–33.4) remained associated with success after adjustment, though these exploratory findings require external validation. Using this optimized system, we successfully generated 30 lung cancer organoid lines representing major histological subtypes, demonstrating the broad applicability of our method. Collectively, this study establishes an efficient workflow for generating LCO lines from malignant pleural effusion, providing a useful technical basis for downstream applications.