Although the relationships between CXCR4 and EGFR expression and survival in non‑small cell lung cancer (NSCLC) have been studied independently, dual CXCR4/EGFR tumor status and its relationship with survival has not been previously investigated. In the present study, we examined the relationship between CXCR4 expression, EGFR expression and dual CXCR4/EGFR expression and survival in patients with NSCLC (n=125) using immunohistochemical techniques. Overall survival was estimated using Kaplan-Meier and Cox proportional hazards models adjusting for patient age, tumor stage and type of treatments. Patients with CXCR4-positive tumors were significantly associated with distant metastasis and tended to have poorer prognosis compared to patients with CXCR4-negative tumors (HR=2.172, 95% CI=1.229‑3.839). No significant association between EGFR expression and survival was found; however co-expression of CXCR4/EGFR was a significant prognostic factor of worse overall survival (HR=2.741, 95% CI=1.330‑5.741). Furthermore, we showed that EGF enhanced the expression of CXCR4 in NSCLC cells through the PI-3K pathway, and treatment of NSCLC cells with EGFR phosphorylation inhibitor, AG1478, resulted in downregulation of the expression of CXCR4. These results suggest an important interaction between CXCR4 and EGFR intra-cellular pathways that may activate signals of tumor progression and may provide a valid explanation for the poor overall survival rate of patients whose co-expression of CXCR4 and EGFR is detected in tissue sections. Based on EGFR and CXCR4 expression, new molecular subtypes of NSCLC established in the present study can be used for customization of NSCLC treatment. Our results also showed that EGFR and CXCR4 are potential therapeutic targets for NSCLC and that simultaneous inhibition of EGFR and CXCR4 in NSCLC patients with concomitant expression of both CXCR4 and EGFR may be an effective treatment strategy.