GSTA1 conferred tolerance to osimertinib and provided strategies to overcome drug-tolerant persister in EGFR-mutant lung adenocarcinoma.

The generation of drug-tolerant persister (DTP) cancer cells remains a major challenge for lung adenocarcinoma (LUAD) patients treated with EGFR-tyrosine kinase inhibitors (TKIs), as these cells eventually lead to drug resistance and disease progression. However, the mechanisms underlying DTP formation are poorly understood, limiting treatment options when DTP state or resistance emerges following TKI therapy. In this study, utilizing samples from LUAD patients receiving front-line osimertinib therapy, including baseline, DTP, and stable resistance states, we dissected the cellular and transcriptomic features of TKI-induced DTP cells. These cells exhibited an active drug-metabolizing phenotype, characterized by significantly increased expression of Glutathione S-Transferase Alpha 1 (GSTA1), which was regulated by Radial Spoke Head Component 1 (RSPH1). Mechanistically, we demonstrated that elevated GSTA1 expression in cancer cells promoted osimertinib degradation. Furthermore, RSPH1+ DTP cells interacted with macrophages via Protein S (PROS1)-AXL signaling to establish an immunosuppressive tumor microenvironment (TME), thereby contributing to persister formation. We investigated the RSPH1-CALML4-GSTA1 regulatory axis and found that PROS1 expression was also governed by this axis, suggesting that GSTA1 acted as an upstream regulator of the PROS1-AXL signaling pathway. We evaluated the feasibility of combination therapy using osimertinib and the GSTA1 inhibitor curzerene in both osimertinib-induced DTP and acquired resistance mouse models. Notably, this strategy demonstrated superior efficacy and safety compared to chemotherapy or AXL inhibitor combination in both settings. Collectively, our study elucidated novel mechanisms contributing to the TKI-induced DTP state and provided a promising combination strategy to overcome drug tolerance and resistance in osimertinib-treated LUAD patients.