To clarify whether arsenic could exert inhibitory effects on tumor cells in pleural effusions of patients with non-small cell lung cancer (NSCLC), 36 NSCLC pleural effusion samples were collected from Changzheng Hospital and Ruijin Hospital, from 2019 to 2022. The genotype of epidermal growth factor receptor (EGFR) was identified. Tumor cells were isolated and treated with arsenic trioxide (ATO) or/and gefitinib. Additionally, six patients were intrapleurally administrated with ATO. Results showed that 25 samples boreEGFRwild type (WT) and 11 harboredEGFRmutations, including 6 with L858R, 3 with ΔE746-A750, and 2 with T790M. ATO diminished the number of tumor cells from patients with WT and mutantEGFR, down-regulated the expression or phosphorylation of EGFR, pmTOR, PI3K, PTEN, and p4E-BP1, and up-regulated the expression of LC3. Immunofluorescent experiments showed that ATO enhanced LC3 and P62. By contrast, gefitinib was only effective in those harboringEGFRsensitizing mutations. Notably, in patients with intrapleural ATO injection, the pleural effusion underwent a bloody to pale yellow color change, the volume of the pleural effusion was reduced, and the number of the tumor cells was significantly reduced. In conclusion, arsenic is effective against NSCLC with variousEGFRgenotypes in vitro and in vivo, and potentially circumvents gefitinib resistance.