SRC
Activation Drives Acquired Pemetrexed Resistance in Non‐Small Cell Lung Cancer and Is Targetable by Dasatinib
Jeongmin Seo, Kui‐Jin Kim, Se Hyun Kim, Ji Hea Sung, Sung‐Hyun Hwang, Woochan Park, Minsu Kang, Eun Hee Jung, Sang‐A Kim, Koung Jin Suh, Ji Yun Lee, Ji‐Won Kim, Jeong‐Ok Lee, Jin Won Kim, Yu Jung Kim, Keun‐Wook Lee, Jee Hyun Kim, Soo‐Mi Bang, Ji‐Hyun Kwon, Jong Seok Lee ABSTRACT
Purpose
Pemetrexed, a standard antifolate chemotherapy for non‐squamous non‐small cell lung cancer (NSCLC), is often limited by acquired resistance. We investigated whether SRC kinase mediates pemetrexed resistance and whether SRC inhibition can suppress survival signaling in pemetrexed‐resistant NSCLC cells.
Methods
Pemetrexed‐resistant cell lines (PC9 R, NCI‐H460 R, and NCI‐H3122 R) were generated through chronic pemetrexed exposure. Protein phosphorylation profiling and Western blotting identified key signaling alterations. SRC was abrogated using transient and doxycycline‐inducible CRISPR/Cas9 systems, and reciprocal siRNA knockdown of SRC and thymidylate synthase (TS) was performed. Three SRC inhibitors—dasatinib, bosutinib, and saracatinib—were assessed using cell viability, colony formation, apoptosis, and cell cycle assays.
Results
All pemetrexed‐resistant cell lines demonstrated elevated phosphorylated SRC (p‐SRC Y416) and enhanced downstream signaling. SRC knockout reduced p‐SRC Y416, p‐STAT3, and p‐FAK, while increasing markers of DNA damage (γ‐H2AX) and apoptosis (cleaved caspase‐3), impairing cell viability and clonogenicity. SRC knockdown reduced TS expression in two resistant models, whereas TS knockdown did not suppress p‐SRC Y416. Among SRC inhibitors, dasatinib showed the most potent, dose‐dependent suppression of SRC signaling. Dasatinib also induced G0/G1 arrest, DNA damage, and apoptosis, thereby reducing colony formation.
Conclusion
SRC activation is a critical mechanism of acquired pemetrexed resistance in NSCLC and may contribute to resistance through TS‐associated survival signaling. SRC inhibition, particularly with dasatinib, disrupts pro‐survival pathways and promotes DNA damage‐mediated cell death in pemetrexed‐resistant cells. These findings support further investigation of SRC inhibitors, alone or with pemetrexed, as a biomarker‐guided therapeutic strategy for SRC‐activated pemetrexed‐resistant NSCLC.