Discovery ofNovel 2,4,5-Trisubstituted PyrimidineDerivatives as Potent and Selective FGFR Inhibitors against GatekeeperMutants for the Treatment of NSCLC

Fibroblast growth factor receptors (FGFRs) play a critical role in the regulation of cancer cell proliferation, differentiation, and migration. However, the development of acquired resistance to FGFR inhibitors remains a major challenge in treating non-small cell lung cancer (NSCLC), particularly due to mutations at the gatekeeper residue. In this study, we report the discovery of a series of irreversible FGFR inhibitors targeting gatekeeper mutations in FGFR1–3, utilizing a 2,4,5-trisubstituted pyrimidine scaffold. Through rational design, structure–activity relationship optimization, and pharmacokinetic evaluation, compound ng <b>12l</b> emerged as a promising candidate. It demonstrated potent inhibition of FGFR1–3 gatekeeper mutations <i>in vitro</i> along with favorable pharmacokinetic properties. The efficacy of <b>12l</b> in targeting FGFR1 gatekeeper mutations was confirmed in assays using L6-FGFR1^V561M/F cells. Furthermore, in xenograft models using both H1581 and L6-FGFR1^V561M cells, <b>12l</b> exhibited robust anti-tumor activity with minimal toxicity. These findings position <b>12l</b> as a promising therapeutic agent for overcoming gatekeeper-mediated resistance in NSCLC.