In regard to gene vectors for cancer gene therapy, their percolation into the tumor tissue should be essential for successful outcome. Here, we studied the tumor penetrability of nonviral vectors (polyplexes) after incubation with the multicellular tumor spheroid (MCTS) models and intratumoral (i.t.) injection into subcutaneous tumors. As a result, polyethylene glycolated (PEGylated), core-shell type polyplexes (polyplex micelles) showed facilitated percolation and improved transfection inside the tumor tissue, whereas conventional polyplexes from cationic polymers exhibited limited percolation and localized transfection. Furthermore, the transfection of hypoxia-responsive plasmid demonstrated that polyplex micelles allowed the transfection to the hypoxic region of the tumor tissue in both in vitro and in vivo experiments. To the best of our knowledge, our results demonstrated for the first time that polyplex micelles might show improved tumor penetrability over cationic polyplexes, thereby achieving transfection into the inside of the tumor tissue.