Design,Synthesis,Self-Assembly And In Vitro Antitumor Evaluations Of Novel Polymeric Carriers Based On Ring Topology

The advanced topology of polymers has a significant impact on their physical and chemical properties,self-assembly behavior,and performance of self-assemblies.Therefore,it is of great significance to design and precise synthesis of macromolecules with complicated structure,not only for the basic research of polymer science,but also for the application research of polymer materials.Among various polymers possessing advanced topological structures,cyclic and its derived topologies have received considerable attentions due to their distinctive chemical and physical properties resulting from the absence of polymer chain ends,such as smaller volume of hydrodynamics,etc..However,there are still some challenges in the precise and controllable synthesis of ring-containing polymers,which limits the application of its and the research of its structure-activity relationship.In order to solve the above problems,a series of novel macromolecules containing cycle were designed and synthesized in this dissertation,and the effects of topological structure on self-assembly behavior of polymers in aqueous solution was systematically studied,besides,the therapeutic properties of the self-assembled micelles as drug carriers in vitro were also evaluated.The specific research contents and main results of each chapter of the dissertation are as follows:1.In chapter one,we review the synthesis methods of cyclic polymers,the topological effects of cyclic structures on the self-assembly behavior of polymers,the research status of cyclic derived topological polymers,and the potential applications of cyclic polymers.2.In chapter two,an amphiphilic cyclic binary block copolymer poly?ethylene glycol?-b-poly??-caprolactone??c?PEG-b-PCL??was prepared by combining ring-opening polymerization?ROP?with homodifunctional bimolecular coupling,and the ratio of molecular weight was 2:1 for hydrophilic and hydrophobic segments of the copolymer.Studies on self-assembly behavior show that both cyclic c?PEG-b-PCL?and linear PEG-b-PCL self-assemble to form spherical micelles in aqueous solution,but the micelles formed by c?PEG-b-PCL?have larger particle size and smaller critical micelle concentration?CMC?.A comparison study on the in vitro drug loading and drug release profiles,intracellular endocytosis,in vitro cytotoxicity and degradation properties of the resulting c?PEG-b-PCL?with those of the linear counterpart?PEG-b-PCL?revealed that the c?PEG-b-PCL?micelles have high drug loading,slow drug release rate,high absorption rate in HeLa cells,greater in vitro cytotoxicity to HeLa cells,and slow in vitro degradation.The above properties based on the cyclic polymer carrier are caused by the cyclic topological effect.Therefore,the work not only provides a method for synthesizing cyclic diblock copolymers,but also reveals the influence of cyclic topological structure on the self-assembly behavior of copolymers and performance of self-assembled micelles served as drug carriers.3.Considering that the cyclic diblock structure prepared in the previous chapter restricts the chain motion of the hydrophilic segment of the copolymer,which may be affected the stability of the self-assembled micelles.Therefore,to further improve the stability and drug loading of the polymer micelles,an amphiphilic tadpole-shaped polymer with a hydrophobic macrocyclic cPCL as the tadpole head and a linear hydrophilic chain PEG as the tail,mPEG-b-cPCL,was synthesized by intra-chain Cu?I?-catalyzed azide-alkyne cycloaddition?CuAAc?click cyclization in the third chapter.A comparison study on the self-assembly behaviors,in vitro drug loading and drug release,and degradation properties of the resulting mPEG-b-cPCL with those of the linear mPEG-b-PCL unclosed that mPEG-b-cPCL micelles are a better formulation than the micelles formed by the linear analogues in terms of micelle stability,drug loading capacity,and degradation property.This study consequently uncovered the topological effect of a hydrophobic moiety on the properties of the self-assembled micelles,and developed a complementary alternative to enhance the micelle stability by introducing a cyclic hydrophobic segment.4.Based on the research in chapter 3,the polymer topological structure was further adjusted from tadpole-to dumbbell shape,hoping to improve the performances of self-assembled micelles as drug carriers by increasing the number of hydrophobic rings in the polymer chain.The target dumbbell-shaped copolymer with two hydrophobic macrocyclic building blocks as the bells,cPCL-b-PEG-b-cPCL,was successfully prepared via the inter-chain click coupling of cPCL and Alkynyl-PEG-Alkynyl.Interestingly,a comparison study further revealed that the micelle based on dumbbell-shaped copolymer c PCL-b-PEG-b-cPCL is a better formulation than the micelles of linear analogue?PCL-b-PEG-b-PCL?and tadpole-shaped copolymer(PEG₄₅-b-cPCL)with only one hydrophobic macrocycle in terms of the micelle stability,drug loading capacity and in vitro drug release behaviors.Most importantly,the drug-loaded micelles of dumbbell-shaped copolymers showed greater in vitro cytotoxicity against HeLa cells than those of the linear counterparts and tadpole-shaped copolymers.This paper therefore developed an efficient strategy for the synthesis of dumbbell-shaped amphiphilic copolymers with cyclic moieties as the two bells,and disclosed the enhanced performance of dumbbell-shaped copolymers for controlled drug release because of the simultaneous introduction of two macrocycles.5.According to the previous study of our group,the introduction of both macrocyclic hydrophobic PCL or hydrophilic POEGMA was beneficial to enhance the stability and drug loading capacity of polymeric micelles.Therefore,a well-defined hetero-8-shaped amphiphilic copolymer,cPOEGMA-b-cPCL,was synthesized in this chapter by inter-chain click coupling between c POEGMA and cPCL to figure out whether micellar properties could be further enhanced by cyclization of hydrophilic and hydrophobic segments simultaneously.The potential of the self-assembled micelles of cPOEGMA-b-cPCL for controlled drug release was evaluated by in vitro drug loading and drug release,cellular uptake,cytotoxicity and degradation studies.The results showed that compared with linear POEGMA-b-PCL and tadpole-like cPOEGMA-b-PCL,the micelles formed by 8-shaped cPOEGMA-b-cPCL have higher endocytosis efficiency and in vitro cytotoxicity in HeLa cells.Most importantly,the micelles based on cPOEGMA-b-cPCL showed ultra-high stability mainly because the stronger steric hindrance of cyclic hydrophilic POEGMA segment prevented the access of the hydrogen ions in the aqueous medium to the cyclic hydrophobic PCL moiety for degradation,which greatly delays the two-stage degradation behavior of the cyclic PCL chain found in our previous work.Therefore this study not only developed an efficient strategy for the precise synthesis of well-defined hetero-8-shaped amphiphilic copolymers with biodegradability,but also reports the first modulation of polymer degradation property by topological control of the non-degradable moiety in the polymer construct through advanced macromolecular engineering.6.Based on the work in the previous chapters,in this chapter,a copolymer cPCL-b-PEG-b-cPCL/c?PCL-b-PEG-b-c PCL?,containing dumbbell and?-shaped hybrid structures,was synthesized by intrachain click cyclization of amphiphilic linear precursor,so as to further study the correlation between the polymer topology and its properties.TEM observation and DLS measurement showed that the mixed structure copolymer self-assembled into larger aggregates with a particle size 245.7nm.In addition,the copolymer has a larger CMC value determined by fluorescence spectrum.The stability of micelles was also studied,and the results showed that the micelles were extremely unstable in both water and PBS solution,which may be because the existence of the?-shaped topology greatly limits the chain motion of hydrophilic moiety PEG.In vitro drug loading and drug release studies have shown that the drug loading capacity of the micelles based on the mixed topological copolymer was slightly high,but the drug release rate of the drug-loaded micelles was very slow due to micelle aggregation.In vitro degradation of the copolymer showed that the degradation rate was similar to that of linear polymer.Therefore,the work indicates that the increase of topological restriction degree is not conducive to the improvement of the stability of polymer micelles and their controlled release performance as drug carriers.