A Fluorescence-Based High-Throughput Screening Method for EGFR Inhibitors Using an EGFR-Targeted Environment-Sensitive Fluorescent Probe

Fluorescence technology offers a highly sensitive, rapid, and efficient approach for drug screening at the cellular level, enabling the direct quantification of drug-target binding interactions through precise measurement of fluorescence intensity changes. In our previous research, a small-molecule fluorescent probe targeting epidermal growth factor receptor (EGFR) was successfully developed, featuring strong targeting specificity, low background signal, and an "off-on" response feature. Based on the EGFR-overexpressing A431 cell line, this study established a cell-based screening system for EGFR inhibitors and successfully identified flavokawain A (FKA), a novel potential EGFR inhibitor. FKA notably reduces the expression levels of p-EGFR and its downstream molecule p-Erk in the EGFR signaling pathway. It demonstrates a potent inhibitory effect on the proliferation of A431 cells, effectively suppressing clone formation and cellular migration, and exhibits a strong apoptosis-inducing effect against A431 cells. This fluorescence-based high-throughput screening method has demonstrated its effectiveness in identifying novel EGFR inhibitors from a wide range of natural materials. It holds significant application potential and plays a crucial role in expediting the discovery of EGFR inhibitors from medicinal sources.