Epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR‑TKIs) have shown promising effects against the growth of non-small cell lung cancer (NSCLC) cells harboring EGFR mutations (EGFR‑mts). However, many patients with NSCLC that are accepted for EGFR‑TKI treatment followed by chemotherapy possess an unknown EGFR status including wild-type EGFR (EGFR‑wt). Little is known about the potential effects of EGFR‑TKI treatment prior to chemotherapy. We investigated the effects and underlying molecular events of 4 weeks of continuous exposure to EGFR‑TKIs in the EGFR‑wt NSCLC line H1299. This treatment dramatically increased the IC50 of several relevant chemotherapeutic agents: cisplatin (DDP) (29.25±6.1 µM for gefitinib, 43.25±14.87 µM for erlotinib, and 6.92±1.15 µM for parental), paclitaxel (11.16±3.36 µM for gefitinib, 9.16±1.41 µM for erlotinib, and 2.09±0.44 µM for parental), gemcitabine (47.18±6.2 µM for gefitinib, 40.36±11.1 µM for erlotinib, and 16.00±3.38 µM for parental) and pemetrexed (11.78±4.07 µM for gefitinib, 15.97±7.23 µM for erlotinib, and 4.72±1.9 µM for parental). This chemoresistance was critically dependent on the activation of the mediator signal transducer and activator of transcription 3 (STAT3). In cells exposed to EGFR‑TKIs for 4 weeks, activation of STAT3 was found to be unrelated to EGFR and to be independent of IL‑6 and ‑22. Treatment with the STAT3 inhibitor NSC 74859 was able to reverse the TKI exposure-induced chemoresistance in EGFR‑wt NSCLC cells. Similar phenomena were observed in H1975 cells harboring EGFR L858R and T790M mutations. Based on the observed molecular events following long exposure of an EGFR‑wt NSCLC cell line to an EGFR‑TKI, this study indicates that such drugs should be not recommended for EGFR‑wt patients who can undergo chemotherapy. This study also suggests that STAT3 inhibitors may aid in the treatment NSCLC patients who exhibit EGFR‑TKI resistance due to an acquired T790M mutation.