[Antitumor Activity of TRAIL-Mu3 Protein in vitro and in vivo and the Mechanisms]

Abstract

Objective: TRAIL-Mu3 was obtained by mutating the N-terminus of human tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) gene to an eight continuous arginine sequence. The present study was designed to explore the antitumor effect of this soluble mutant protein and the underlying mechanisms.

Methods: The inhibitory effect of TRAIL-Mu3 on the proliferation of lung cancer cell lines NCI-H460, A549, NCI-H1299 and calu-1 was tested by CCK8 assay. The apoptotic rates of A549 and NCI-H460 treated by TRAIL-Mu3 were detected by flow cytometer (FCM). The expressions of apoptosis related proteins death receptor (DR) 4, DR5, Caspase-3, Caspase-8 and X-linked inhibitor of apoptosis protein (XIAP) were detected by Western blot .Moreover, a subcutaneous xenograft tumor mouse model of NCI-H460 was established and treated with TRAIL-Mu3 daily or every other day or three times a week. The expressions of DR4, DR5, Caspase-3, Caspase-8 and XIAP were detected by immunohistochemical staining.

Results: The in vitro study demonstrated that as compared to the TRAIL, the TRAIL-Mu3 was more toxic and pro-apoptotic by up-regulation of the expression and activity of DR4, Caspase-3 and Caspase-8. Also, the animal study showed a similar antitumor effect between treatment with TRAIL-Mu3 every other day and three time a week, which was better than daily use. All treatments significantly suppressed the growth of xenograft tumor, increased the expression or activity of DR4 and Caspase-3, and down-regulated the expression of XIAP ( P<0.05).

Conclusion: TRAIL-Mu3 could improve antitumor activity in vivo and in vitro through elevating DR4 expression, activating Caspase-3/-8, and inhibiting XIAP activation.

Keywords: Apoptosis; Arginine; Lung cancer; Mutant; TRAIL.