Targeting L858R/T790M/C797S mutant EGFR is a major challenge in the new-generation EGFR tyrosine kinase inhibitors development for conquering drug resistant NSCLC. In this study, a series of novel 9-heterocyclyl substituted 9H-purine derivatives were designed as EGFR tyrosine kinase inhibitors. Among these compounds, D4, D9, D11 and D12 showed significantly potent anti-proliferation and EGFR inhibition activity. In particular, the most potent compound D9 showed anti-proliferation against HCC827 and H1975 cell lines with the IC values of 0.00088 and 0.20 μM, respectively. And D9 inhibited the EGFR with an IC value of 18 nM. Furtherly, D9 could significantly suppress the EGFR phosphorylation, induce the apoptosis, arrest cell cycle at G0/G1, and inhibit colony formation in HCC827 cell line by a concentration-dependent manner. Molecular docking indicated that the introduction of a cyclopropylsulfonamide group in D9 led to the formation of additional two hydrogen bonds with mutant Ser797 which played key roles in generating efficient EGFR inhibitory activity. These findings strongly indicated that 9-heterocyclyl substituted 9H-purine derivatives were promising L858R/T790M/C797S mutant EGFR-TKIs. The introduction of extra hydrogen bond interaction with mutant Ser797 is efficient method for the design of the fourth-generation EGFR-TKIs.