EpCAM-Antibody-Labeled Noncytotoxic Polymer Vesicles for Cancer Stem Cells-Targeted Delivery of Anticancer Drug and siRNA Jing Chen, †,‡,§ Qiuming Liu, ‡,§ Jiangang Xiao, ‡ and Jianzhong Du* ,†,‡ † Shanghai Tenth People’s Hospital, Tongji University School of Medicine, 301 Middle Yanchang Road, Shanghai 200072, China ‡ Key Laboratory of Advanced Civil Engineering Materials of Ministry of Education, School of Materials Science and Engineering, Tongji University, 4800 Caoan Road, Shanghai 201804, China * S Supporting Information ABSTRACT: Cancer stem cells (CSCs) have the capability to initiate tumor, to sustain tumor growth, to maintain the heterogeneity of tumor, and are closely linked to the failure of chemotherapy due to their self-renewal and multilineage differentiation capability with an innate resistance to cytotoxic agents. Herein, we designed and synthesized a novel anti-EpCAM (epithelial cell adhesion molecule)-monoclonal-antibody-labeled CSCs-targeting, non- cytotoxic and pH-sensitive block copolymer vesicle as a nanocarrier of anticancer drug and siRNA (to overcome CSCs drug resistance by silencing the expression of oncogenes). This vesicle shows high delivery efficacy of both anticancer drug doxorubicin hydrochloride (DOX·HCl) and siRNA to the CSCs because it is labeled by the monoclonal antibodies to the CSCs-surface- specific marker. Compared to non-CSCs-targeting vesicles, the DOX·HCl or siRNA loaded CSCs-targeting vesicles exhibited much better CSCs killing and tumor growth inhibition capabilities with lower toxicity to normal cells (IC 50,DOX decreased by 80%), demonstrating promising potential applications in nanomedicine. ■ INTRODUCTION Tumor recurrence is one of the most important challenges in cancer treatment. In the past two decades, the growing understanding of cancer has made it clear that many cancers, including hematopoietic and solid tumors, may be driven by a small subpopulation of cancer stem cells (CSCs). The CSCs were first discovered in acute myeloid leukemia, 1 and were subsequently identified in various solid tumors and confirmed to play a critical role in the tumor occurrence, deterioration, metastasis, and recurrence. 2-12 The surface antigen epithelial cell adhesion molecule (EpCAM) is increasingly recognized as one of the most important surface specific markers of CSCs for a variety of tumors. 2,8,12-15 EpCAM positive (EpCAM+) cells can be isolated from cancer cell lines, which have the ability to self- renew, to initiate tumor formation, and are intrinsically resistant to therapy. These specific characteristics of the EpCAM+ cells are involved in multiple cell signal transduction pathways, which increased the difficulties of screening sensitive drugs to kill CSCs. 16-19 Fortunately, small interfering RNA (siRNA) has been explored to overcome the drug resistance of cancer cells by silencing the expression of genes in signal transduction pathways. 20-23 Nevertheless, the successful siRNA therapy needs efficient, stable, easy making, and low cytotoxic siRNA delivery carriers. 24-27 Nanoscale carriers with targeting units are promising for drugs/siRNAs delivery in modern pharmaceutics. The nano- carriers, 28-30 such as polymer vesicles, 31-41 will help the drug/ siRNA to prevent nonspecific distributing, inactivated, cleared shortly after administration before reaching action sites, and creating toxic effects in healthy organs. 22,42-44 However, the current cancer treatment strategies may affect the bulk of the tumor cells but leave CSCs behind. 45-48 Fortunately, CSCs- targeting drug/siRNA delivery systems such as polymer vesicles may present a promising strategy for solving the above problem in cancer treatment. Herein, we report a novel noncytotoxic and pH-sensitive polymer vesicle for specifically targeting EpCAM positive CSCs and for efficient intracellular drug/siRNA delivery. The vesicle is self-assembled from a pH-responsive triblock copolymer, poly(ethylene oxide)-block-poly[2-(diisopropylamino)ethyl methacrylate]-block-poly(acrylic acid) (PEO 43 -b-PDPA 76 -b- PAA 17 ). The anti-EpCAM monoclonal antibodies (EpCAM- Abs) are labeled on the vesicle coronas, leading to a specific recognition between vesicles and CSCs. As shown in Scheme 1, the biocompatible PEO is designed as the mixed coronas with hydrophilic PAA chains, while the pH-sensitive PDPA chains form the membrane of the vesicle. The PAA chains in the outer coronas can be decorated with Streptavidin (SA), and then be bound with biotin-labeled EpCAM antibodies. This kind of EpCAM-antibodies-labeled pH-sensitive polymer vesicles with positive charges could lead to a significant improvement in the loading and delivery of Received: January 10, 2015 Published: May 19, 2015 Article pubs.acs.org/Biomac © 2015 American Chemical Society 1695 DOI: 10.1021/acs.biomac.5b00551 Biomacromolecules 2015, 16, 1695-1705