The development of a drug delivery strategy that can not only cross the blood-brain barrier (BBB) rapidly, but also target the glioma and reach the core of glioma is essential and important for glioma treatment. To achieve this goal, we established a dual-targeting liposomal system modified with TAT (AYGRKKRRQRRR) and T7 (HAIYPRH), in which the specific ligand T7 could target BBB and brain glioma tumor and the nonspecific ligand TAT could enhance the effect of passing through BBB, and elevate the penetration into the tumor. The dual-targeting effects were evaluated by both in vitro and in vivo experiments. To identify the targeting effect, in vitro cellular uptake and BBB model were performed. Tumor spheroid penetration was performed to evaluate the penetration characteristics of the dual-targeting liposomes. In vivo pharmacokinetic studies were utilized to evaluate the influence of T7 and TAT peptides on the behavior of nanoparticle drug delivery system, and tissue distribution was further utilized to evaluate the glioma-targeting efficiency of the dual-targeting liposomes.