The use of survivinT34A mutant targeted disruption of survivin, the strongest inhibitor of apoptosis protein overexpressed in tumors, has proved a promising strategy for advanced cancers. However, hyperthermia, as a cytotoxic enhancer, regularly activates the expression of survivin to counteract the heat-induced antitumor activity. Here, we investigated the combinational antitumor effect by using liposome-encapsulated mouse survivinT34A and hyperthermia in mouse models. We observed that the combination treatment of surivinT34A and hyperthermia significantly increased the growth inhibition and apoptosis of tumor cells in vitro compared with single treatment or other controls, which was similar to the effect of survivin silencing in combination with hyperthermia. Moreover, the inhibition of tumor growth in vivo was also remarkably enhanced by combination of surivinT34A and hyperthermia when compared with other treatments. Naturally, the tumor tissues in combination treatment presented the larger necrosis-like areas, more apoptotic cells and less microvessel density. Our findings suggest that the antitumor efficacy of survivin disruption can be enhanced by hyperthermia, which might be a new feasible approach for cancer therapy.