A variety of three-dimensional DNA assemblies have been proposed as drug carriers owing to their good biocompatibility and easy fabrication. In this study, inspired by the structure of cockleburs, a novel aptamer-tethered DNA assembly was developed for effective targeted drug delivery. The Apt-nanocockleburs were fabricated via a facile process of DNA base pairing: four complementary DNA single strands, including one aptamer-ended strand and three sticky-end strands, were applied to pair with each other. The main body of the nanocockleburs can load doxorubicin (Dox) whilst the covered aptamer spines bind to the target MCF-7 cells. The self-assembled Apt-nanocockleburs exhibit higher cell uptake as well as increased cytotoxicity to MCF-7 cells than DNA nanocockleburs without aptamers. This study provided a DNA constructing platform to produce new drug carriers with high selectivity for cancer targeted drug delivery.