Objective:To identify the effect of oestrogen receptor(ER) antagonist,Tamoxifen,combination with EGFR tyrosine kinase inhibitor,Gefitinib,on the proliferation and apoptosis in non-small cell lung cancer(NSCLC) cell lines and the related molecular mechanism,providing theoretical evidence for the molecular targeted treatment combination with endocrine treatment in NSCLC. Methods:The cell lines were treated with gefitinib and tamoxifen alone or in combination. The cell proliferation were detected with MTT,the cell apoptosis with flow cytometry and the expression of EGFR and ERβ with real-time PCR and Western blot. In addition,the effect of oestrogen and Tamoxifen on the expression of EGFR,the effect of EGF and Gefitinib on the expression of ERβ were detected,respectively. The effect of oestrogen on the phospho-EGFR were also detected. Results:The combination of Tamoxifen and Gefitinib increased the inhibition and apoptosis in EGFR and ERβ positive cell lines A549 and H1650,while showed no significant changes in EGFR and ER negative cell line H520. It was found that oestrogen could promote the phospho-EGFR protein expression,meanwhile,EGFR protein expression was down-regulated in response to estrogen and up-regulated in response to Tamoxifen in vitro. Conversely,ERβ expression was decreased in response to EGF and increased in response to Gefitinib. Conclusion:The combination of Tamoxifen and Gefitinib can promote its antiproliferative effects on EGFR and ERβ positive lung cell lines. These results suggested that this might be involved in cross-signaling between the EGFR/ER pathways in NSCLC. In conclusion,it provided the rationale to combine Gefitinib with hormone therapy for lung cancer treatment.