An endothelial-centered regulatory framework reveals context-dependent roles of MYLK in lung adenocarcinoma

Background: Lung adenocarcinoma (LUAD) is shaped by the tumor microenvironment, yet endothelial cell (EC) regulatory programs and their biological roles remain insufficiently defined. Methods: We analyzed scRNA-seq data to map EC-associated programs and applied hdWGCNA to identify EC modules and communication patterns. Network pharmacology integrated EC-module genes with LUAD-related targets to prioritize MYLK. MYLK expression and function were evaluated by RT-qPCR, immunohistochemistry, and gain-/loss-of-function assays in endothelial and LUAD cell models. We then performed network-based in silico knockout in LUAD tumors (GSE164789) and exploratory immune-cell eQTL analysis. Results: EC modules were enriched for junction organization, angiogenesis, and immune-related pathways, with extensive epithelial-stromal-endothelial interactions. Network pharmacology nominated MYLK as an EC-linked LUAD candidate. MYLK expression was reduced in LUAD and associated with unfavorable clinical outcomes. In endothelial cells, MYLK perturbation altered junction integrity and trans-endothelial tumor cell migration; in LUAD cells, MYLK gain/loss affected migration, invasion, and proliferation. In silico knockout of MYLK produced regulatory shifts enriched for tight junction organization, endothelial apoptosis, angiogenesis, vascular permeability, and vascular/cancer-related pathways. Immune-cell eQTL analysis identified an association between increased MYLK expression in dendritic cells and elevated LUAD risk. Conclusions: These findings define an endothelial-centered regulatory framework in LUAD and highlight the context-dependent, cell-type-specific roles of MYLK at the tumor-endothelial-immune interface.