A-B-C Triblock Copolymer Micelles for Intracellular Delivery of Cancer-Targeted siRNA

摘要

Successful clinical use of synthetic siRNAs for gene therapy via a pathway called RNA interference (RNAi) is still limited by a variety of factors in the delivery process. These factors include recognition and uptake by the targeted cell type, efficient escape from the intracellular endosomal cavities, and release of the siRNA payload into the cytosol where the RNAi mechanism can be initiated. siRNA on its own is incapable of enduring this journey intact and thus various protective vehicles have been designed for the safe and efficient delivery of siRNA for gene silencing. Among the most promising of the non-viral vectors studied thus far in the literature are synthetic polymers, designed to protect and deliver the siRNA cargo intracellularly with minimal toxicity to the host. In this work, we will explore the A-B-C triblock copolymer PEG-PnBA-PDMAEMA, which forms micelle-like aggregates in aqueous buffer, providing an unconventional architectural platform for studying siRNA delivery properties. The in vitro and in vivo performance (toxicity, gene silencing, biodistribution, tumor accumulation, etc.) of the PEG-PnBA-PDMAEMA micelle/siRNA complexes (micelleplexes) are compared relative to more traditional polycation-based systems (e.g., PDMAEMA, PEG-PDMAEMA) to determine the role of nano-carrier architecture on delivery behavior. These three systems are very similar chemically but are expected to have distinct delivery behaviors due to their architectural dissimilarities and differing degrees of PEGylation. We observed an overall improvement in the gene silencing and tumor accumulation efficiencies with the micelleplex system with no additional toxicity than the PDMAEMA and PEG-PDMAEMA complexes under the same conditions. This proves that the micelleplex concept affords net-positive benefits to the nano-carrier based on its architecture which the PDMAEMA and PEG-PDMAEMA systems are not able to provide. However, in spite of its obvious edge over basic polyctations, the absolute gene silencing efficiency and levels of tumor accumulation for the micelleplexes are lower than desirable for a potent delivery vehicle. Lack of endosome-escaping capacity and intracellular degradability are believed to be the two most prominent factors limiting the successful delivery of the PEG-PnBA-PDMAEMA micelleplexes. Hence, for the micelleplex concept to be ultimately suitable for the clinic, we have proposed a next-generation polymer, Folate-PEG-PLGA-SS-PEI, which we believe will possess the characteristics necessary to overcome the remaining shortcomings of the PEG-PnBA-PDMAEMA micelle system. With the remaining challenges sufficiently addressed, we are confident that the micelleplex design will make a promising addition to the viable clinical options for synthetic polymer-mediated gene therapy.