In this study, a series of 4-aniline quinazoline derivatives bearing hydrogen sulfide (HS) donors were designed, synthesized and evaluated for biological activities. The synthesized compounds were screened for the enzymatic activities against EGFR and EGFR mutants by kinase target-based cell screening method. The results demonstrate that most compounds exhibit selectively inhibitory activities against TEL-EGFR-L858R-BaF3, especially compound 9h with GI = 0.008 μM (TEL-EGFR-L858R-BaF3), 0.0069 μM (TEL-EGFR-C797S-BaF3), >10 μM (BaF3), >10 μM (TEL-EGFR-BaF3) and 6.03 μM (TEL-EGFR-T790M-L858R-BaF3). The results from anti-proliferative assays in two NSCLC cell lines indicate that synthetic derivatives (9g, 9h, 15e and 15f) with HS donor ACS81 display greater anti-proliferative potency against NSCLC cell line H3255 bearing EGFR mutant (L858R) with GI values ranging from 0.3486 to 1.348 μM. In addition, compound 9h exhibits weak anti-proliferative effects on other tumor cells (HepG2, MCF-7, HT-29 and A431) and has lower toxic effect on HUVEC cells than AZD9291 (positive control). Meanwhile, compound 9h inhibits the phosphorylation of EGFR in H3255 cells in a dose-dependent manner. Cell cycle analysis reveals that compound 9h suppresses the proliferation of cells by inducing cell cycle arrest in G0-G1 phase. The result of HS release evaluation suggests that the HS release of compound 9h is significantly more and faster than other compounds.Copyright © 2020. Published by Elsevier Masson SAS.