Design,Preparation,and Self-assembly Behaviors Of Novel Cyclic Brush Polymers And Their In Vitro Anti-cancer Drug Delivery Behaviors

In recent years,polymers with advanced topological architectures,including dendrimers,star-shaped polymers,segmented hyperbranched polymers and cyclic brush polymers are promising materials for wide applications due to their structure-generated unique properties different from that of the linear analogues.Among these architectures,cyclic brush polymer with nanoscale size,hyperbranched polymer structure,the simultaneous functionalities of the cyclic core and the polymer brushes,facile preparation,ability of drug encapsulation within the interior cyclic core domain stabilized by the outer shell of polymer brushes is probably considered as the most unique topology,which has attracted increasing attention.However,the adaptation of cyclic brush polymer for drug delivery applications remains largely unexplored.For this purpose,in this thesis,cyclic brush polymers with uniformly radiating polymer brushes grafting from a cyclic core template and functional brushes were fabricated via click chemistry and atom transfer radical polymerization（ATRP）.The structure of polymers was characterizaed by nuclear magntic resonance（¹H NMR）spectrometer,Fourier transform infrared（FT-IR）spectrometer and size exclusion chromatography and multi-angle laser light scattering（SEC-MALLS）analyses.The particle size and size distribution were determined by dynamic light scattering（DLS）.The ability to form micelles from cyclic brush polymers was studied by critical aggregation concentration（CAC）determination.The morphology of the self-assembled nanoparticles was observed by the transmission electron microscopy（TEM）.The cytotoxicity of polymers was evaluated in vitro usingthe3-（4,5-dimethylthiazol-2-yl）-5-（3-carboxymethoxyphenyl）-2-（4-sulfophenyl）-2H-tetrazolium（MTS,Promega）assay.The cell uptake behavior was observed by laser scanning fluorescence confocal microscope（LSCM）.The detailed contents and the results of this thesis are as follows,1.Thermo-sensitive cyclic brush polymers with poly（N-isopropylacrylamide）（PNIPAAm）brushes were synthesized by controlled living radical polymerization using cyclic multimacroinitiator.The thermo-induced phase transition behaviors of the resultant cyclic brush polymers with different compositions were investigated in detail by temperature-dependent optical transmittance measurements,and compared with the properties of bottlebrush and linear counterparts.Interestingly,the lower critical solution temperature（LCST）of cyclic brush PNIPAAm could be regulated by the chain length of PNIPAAm brush.Although the bottlebrush polymers with the same composition exhibited similarly structurally dependent LCSTs behaviors to the cyclic brush polymers,the cyclic brush PNIPAAm did show higher critical aggregation concentration（CAC）and enhanced stability against dilution than the bottlebrush counterpart.The readily tailorable Tcps together with the ability to form highly stable nanoparticles makes thermo-sensitive cyclic brush PNIPAAm a promising candidate for controlled drug delivery.2.Based on the previous work,cyclic brush copolymer of poly（2-hydroxyethyl methacrylate-g-poly（N-isopropylacrylamide-st-N-hydroxyethylacrylamide））（cb-P（HEMA-g-P（NIPAAm-st-HEAAm）））comprising a cyclic core of PHEMA and thermo-sensitive brushes of statistical copolymer of P（NIPAAm-st-HEAAm）was designed and synthesized successfully via a graft-from approach using ATRP from a cyclic multimacroinitiator.The composition of the brush was optimized to endow the resulting cyclic brush copolymer with a lower critical solution temperature（LCST）slightly above the physiological temperature,but lower than the localized temperature of tumor tissue,which is suitable for the hyperthermia-triggered anticancer drug delivery.Critical aggregation concentration（CAC）determination revealed better stability for the unimolecular nanoparticle formed by the cyclic brush copolymer than that formed by the bottlebrush analogue.The dramatically increased size with elevated temperatures from below to above the LCST confirmed hyperthermia-induced aggregation for both formulations.Such structural destabilization promoted significantly the drug release at 40^oC.Most importantly,the drug-loaded cyclic brush copolymer showed enhanced in vitro cytotoxicity against HeLa cells than the bottlebrush counterpart.The better stability and higher therapeutic efficacy demonstrated that the thermo-sensitive cyclic brush copolymer is a better formulation than bottle brush copolymer for controlled drug release applications.3.Amphiphiliccyclicbrushcopolymers,poly（2-hydroxyethylmethacrylate-g-poly（ε-caprolactone）-poly（oligoethyleneglycol methacrylate））（P（HEMA-g-PCL-POEGMA））with the hydrophobic PCL middle layer and the hydrophilic POEGMA outer corona were designed and synthesized successfully in this study via a“grafting from”approach using sequential ring-opening polymerization（ROP）and ATRP from a cyclic multimacroinitiator PHEMA.The unimolecular core-shell-corona（CSC）micelles self-assembled from amphiphilic cyclic brush copolymers of P(HEMA-g-PCL₁₈-POEGMA)₅₀ with different chain lengths showed sufficient stability in the physiological condition for prolonged circulation,as well as in the intracellular reducing environment.The drug loading content of micelles self-assembled from the amphiphilic cyclic brush copolymer P(HEMA-g-PCL₁₈-POEGMA₃₀)₅₀ reached 4.60%,which is higher than the thermo-sensitive cyclic brush polymers cb-P（HEMA-g-P（NIPAAm-st-HEAAm））developed in our previous study,demonstrating the ability of cyclic brush P(HEMA-g-PCL₁₈-POEGMA₃₀)₅₀0 to improve the loading capacity of anticancer drugs by introducing hydrophobic PCL middle layer.4.To the best of our knowledge,the preparation of reduction-sensitive cyclic brush polymers for drug delivery applications remains unexplored.For this purpose,a reduction-sensitive amphiphilic cyclic brush copolymer,poly（2-hydroxyethyl methacrylate-g-poly（ε-caprolactone）-disulfide link-poly（oligoethyleneglycol methacrylate））（P（HEMA-g-PCL-SS-POEGMA））with reducible block junctions bridging the hydrophobic PCL middle layer and the hydrophilic POEGMA outer corona was designed and synthesized successfully in this study via a“grafting from”approach using sequential ROP and ATRP from a cyclic multimacroinitiator PHEMA.The unimolecular core-shell-corona（CSC）micelles self-assembled from reducible cyclic brush copolymers of P(HEMA-g-PCL₁₈-SS-POEGMA₃₀)₅₀ showed sufficient stability in the physiological condition for prolonged circulation,while efficient destabilization in the intracellular reducing environment for promoted drug release.A further evaluation of the delivery efficacy by an in vitro drug release study and an in vitro cytotoxicity study confirmed greater cumulative drug release from the doxorubicin（DOX）-loaded reduction-sensitive micelles of cyclic brush copolymers P(HEMA-g-PCL₁₈-SS-POEGMA₃₀)₅₀0 in the presence of 10 mM dithiothreitol（DTT）and higher therapeutic efficacy of this formulation relative to the reduction-insensitive counterparts.The overall results demonstrate the reducible cyclic brush copolymers developed herein provides an elegant solution to the tradeoff between extracellular stability and intracellular high therapeutic efficacy toward efficient anti-cancer drug delivery.