Synthesis Of Polymeric Nanocarriers And Their Applications In Protein Delivery

As an emerging therapeutic strategy, protein therapy shows huge potential applications in many fields, such as treatment of diseases including cancer, diabetes and gene defects, vaccine immunity and regenerative medicine. Compared with small molecule drugs and gene therapy, protein drugs own several advantages including strong specificity, low side effects, high safety and broad clinical application range. However, the practical efficiency is always low and its effects get limited by many factors, such as fragile activity, short half-life, low intracellular delivery efficiency and hard to control release. To improve the efficiency of protein delivery, numerous protein delivery systems were explored, for example, liposomes, polymeric nanocarriers and inorganic nanocarriers. In this thesis, we systemically investigated the synthesis of polymeric nanocarriers and their applications in protein delivery, including glucose-responsive polymeric complex micelles for repeated on-off release and protection of insulin and single-protein nanocapsules for prolonging circulation of proteins in vivo. This thesis contains three parts:First, we synthesized two types of glucose-responsive diblock copolymers poly(ethylene glycol)-b-poly(aspartic acid-co-aspartamidophenylboronic acid)(PEGn4-6-P(Asp0.3-co-AspPBA0.7)168) and poly(N-isopropylacrylamide)-b-poly(aspartic acid-co-aspartamidophenylboronic acid)(PNIPAM130-b-P(Aspo.3-co-AspPBAo.7)175). Through ATRP and ROP, We synthesized two types of narrow-dispersed diblock copolymers containing the same PBLA, which are PEG114-b-PBLA168and PNIPAM130-b-PBLA175. After hydrolysis of PBLA and modidication of PAsp on the side chains with PBA in the same PBA/PAsp ratio, we got PEG114-b-P(Aspo.3-co-AspPBAo.7)168and PNIPAM130-b-P(Aspo.3-co-AspPBAo.7)175. The study from light scattering showed both the two polymers hold obvious glucose responsiveness, and the latter also shows thermer-responsiveness. Considering the responsiveness and biodegradability, these two polymers would facilitate the self-assembly for further fabrication of multifunctional complex micelles. Second, we developed a glucose-responsive complex polymeric micelle (CPM) through self-assembly of the two diblock copolymers mentioned above and achieved repeated on-off release and protection of insulin. By controlling the weight ratio between PNIPAM and PEG as6/4(WpNIPAM/WPRG), hydrophilic PNIPAM would become hydrophobic and collapse on the P(Asp-co-AspPBA) core to form a continuous hydrophobic layer when the temperature increased above the LCST of PNIPAM, which also leads to the formation of PEG channels. When the micellar core became completely hydrophilic, Our study showed the CPM with this special core-shell-corona structure would not disaggregate but just only swell. As a result, the CPM exhibits a reversible swelling in response to changes in the glucose concentration, enabling the repeated on-off release of insulin regulated by glucose level. Furthermore, the CPM could effectively protect the encapsulated insulin against protease degradation. Therefore, this glucose-responsive CPM provides a simple and powerful strategy to construct a self-regulated insulin delivery system for diabetes treatment. Besides, the oral delivery of insulin-loaded CPM in rabbits showed it could sharply decrease the blood glucose level, which indicates the CPM hold potential applications for oral insulin delivery.Last, we explored single-BSA nanocapusles (zwitterionic nBSA) based on zwitterioinc2-methacryloyloxyethyl phosphorylcholine (MPC) and achieved remarkably prolonged circulation of protein in vivo. We modified the BSA with polymerizable acryl groups and then obtained nBSA through in situ radical polymerization on protein surface. In vitro study exhibited that zwitterionic nBSA is very stable and has no interaction with FBS and HeLa cells. Furthermore, it could prolong the half-life of BSA in mice to3.5day. As the broad feasibility of the single-protein nanocapsule technique, it could be utilized to encapsulate many proteins. Therefore, MPC-based nanocapsules would widely and significantly improve the Pharmaceutical effect of proteins and further open a new avenue for targeted and intracellular protein delivery, which holds huge potential applications for treatment of threatening diseases such as cancer and gout.