Phosphoproteomics landscape of lung DTP

Osimertinib is the first-line therapy for EGFR-mutated non-small cell lung cancer, but acquired resistance commonly develops, linked to a drug-tolerant persister (DTP) cell state that promotes survival and eventual resistance. Using data-independent acquisition mass spectrometry (DIA-MS), we mapped dynamic proteomic and phosphoproteomic changes in DTPs. While osimertinib initially blocks EGFR signaling, DTPs show activation in ribosome synthesis and protein translation pathways, with resistance emerging via EGFR reactivation and anti-apoptotic mechanisms, including YAP1 and mTOR-BAD hyperphosphorylation. Elevated phosphorylation of CDK1 substrates in DTPs was observed, and inhibiting CDK1-mediated SAMHD1 activation impaired DTP growth. This study highlights key DTP mechanisms, suggesting biomarkers and therapeutic targets to delay resistance.