Objective: To observe the effects of recombinant adenovirus TRAIL (AdS-TRAIL & Ad5F35-TRAIL) on apoptosis of non-small cell lung (NSCLC) cells, so as to assess the value of Ad-TRAIL in gene therapy of NSCLC. Meth-ods: CAR and CD46 expression levels in lung cancer cell lines (A549, Z793, QG56 and NCI-H520) and the primary lung cancer cells from samples of 10 NSCLC patients were assayed by flow cytometry analysis. The lung cancer cell lines and primary lung cancer cells were infected with Ad5-TRAIL & Ad5F35-TRAIL adenoviral vectors at MOI 10 or 50, re-spectively; the percentage of apoptosis cells labeled by Annexin V-FITC in different cells were measured by flow cytometry 48 h after transfection. Results: The expression of CD46 were higher than that of CAR in all the lung cancer lines (A549, Z793, QG56 and NCI-H520) and the primary lung cancer cells. Significant apoptosis was observed in Z793 and QG56 cells transfected with Ad5-TRAIL or Ad5F35-TRAIL at MOI 10, with the apoptosis rate being (1.76±2.10)% (Ad5-TRAIL), (15.96±2.89) % (Ad5F35-THAIL) and (6.05±1.58) % (Ad5-TRAIL), (10.11±1.26) % (Ad5F35-TRAIL), respectively, compared to no adenovirus-transfected cells ([2.33±0.37] % and [5.95±1.89]%, respectively, P < 0.05). Less than 10% of apoptosis cells were detected in NCI-H520 cells transfected with Ad5- or Ad5F35-TRAIL at MOI 50 ([12.89±3.2] % for AdS-TRAIL and [9.08±1.35]% for Ad5F35-TRAIL, respectively) compared to no adenovirus-transfected cells ([7.04±2.17] %, P > 0.05). Moreover, apoptosis induced by Ad5- or Ad5F35-TRAIL transfection in A549 cells was not detected both at MOI 10 and 50. About half of the primary lung cancer cells from 10 patients induced apoptosis after transfected with Ad5-TRAIL or Ad5F35-TRAIL vector. A higher percentage of apoptotic cells were found in Ad5F35-TRAIL group than those in Ad5-TRAIL and control groups. Conclusion: Ad5-TRAIL can induce apoptosis of NSCLC cells in vitro, and Ad5F35-TRAIL is more potent than Ad5-TRAIL, so Ad5F35-TRAIL is more suitable for gene therapy of NSCLC.