Study On Topological Polymers With Acid-Cleavable Features Prepared By Ring-Opening Polymerization And “Click” Chemistry For P H-Triggered Drug Delivery

pH-responsive polymers can potentially be exploited in a broad range of drug delivery applications as pH values in different tissues and cellular compartments vary tremendously. For example, the tumor extracellular environment is more acidic（pH 6.5） than blood and normal tissues（pH 7.4）, and the pH values of endosome and lysosome are even lower at 5.0-5.5. The encapsulation or conjugation of anticancer drugs into pH-sensitive polymers can significantly improve their solubility. The hydrolysis of acid-labile groups in the acidic microenvironment of cancer cells would accelerate the dissociation of the acid-cleavable polymers and further result in a rapid release of the encapsulated or conjugated drugs. These anticancer drug delivery systems remain stable during circulation but are sensitive to the signal caused by disease and release an appropriate amount of drugs in response. The present work is mainly focused on the design and preparation of the topological polymers, such as the linear, brush-type and star-block copolymer, with acid-cleavable features via a combination of ring-opening polymerization and “Click” chemistry. These copolymers are mainly constructed by the biocompatible and biodegradable poly（ε-caprolactone）（PCL）, poly（ethylene glycol）（PEG） and polyphosphoester（PPE） blocks and further used to fabricate controlled drug delivery systems. The present work can be summarized as follows:（1） Precise modular synthesis and characterization of a novel acid-cleavable amphiphilic star-block copolymer（mPEG-acetal-PCL-acetal-）3. A series of well-defined three-armed star-block copolymers containing poly（ethylene glycol） monomethyl ether（mPEG） and poly（ε-caprolactone）（PCL） blocks linked with acid-cleavable acetal groups, designated as（mPEG-acetal-PCL-acetal-）3 or（mPEG-a-PCL-a-）3, have been prepared via a “coupling-onto” method based on ring-opening polymerization（ROP） and Cu（I）-catalyzed azide-alkyne cycloaddition（CuAAC） “Click” chemistry. Two polymer precursors were first synthesized, including an acetal- and azide-functionalized PEG（mPEG-a-N₃）, as well as an acetal- and propargyl-containing PCL（PA-PCL-a-Cl）. Conjugation of these preformed polymer building blocks has been achieved via CuAAC “Click” reaction to form a diblock copolymer with two acetal linkages（mPEG-a-PCL-a-Cl）. After transferring the chlorine group to azide group, the subsequent CuAAC “Click” reaction between mPEG-a-PCL-a-N₃ and a tri-functional core molecule triprop-2-ynyl benzene-1,3,5-tricarboxylate（TPBTC） afforded the well-defined three-arm star-block copolymers. The chemical compositions and structures, as well as the molecular weights and molecular weight distributions（PDIs） of these copolymers have been fully characterized by 1H NMR, FT-IR, and GPC measurements. Differential scanning calorimetry（DSC） and wide-angle X-ray diffraction（WAXD） analyses demonstrated that the thermal behaviors of the star-block copolymers strongly depended on the relative lengths of PEG and PCL blocks in the arms.（2） A structure-property study of acid-cleavable star-block copolymer（mPEG-acetal-PCL-acetal-）3 and used for pH-triggered drug delivery. The star-block copolymer can self-assemble into micelles and encapsulate the hydrophobic anticancer drug doxorubicin（DOX） simultaneously. The acetal linkages can be cleaved in endosomal and lysosomal acidic media once the DOX-loaded micelles are internalized by cancer cells. The shedding of hydrophilic PEG shells would result in the dissociation of micelles and the subsequent fast release of DOX. The self-assembly behaviors of these amphiphilic star-block copolymers were investigated by a fluorescence probe method, dynamic light scattering（DLS） and transmission electron microscopy（TEM） analyses. Moreover, the acid-cleavable properties of these star-block copolymers were systematically studied by 1H NMR, GPC, and DLS measurements, and the results indicated that they were relatively stable in neutral pH media, but could be degraded under acidic conditions. The in vitro DOX release studies, MTT assays, live cell imaging system and flow cytometry analysis revealed that these polymeric micelles could efficiently deliver and release DOX into HeLa cells.（3） Injectable hydrogels by inclusion complexation between three-armed star copolymer（m PEG-acetal-PCL-acetal-）3 and a-cyclodextrin for pH-triggered drug delivery. We focus on developing the acid-cleavable star-block copolymer to an injectable hydrogel that is based on inclusion complexes between（mPEG-a-PCL-a-）3 and α-cyclodextrin（α-CD）. The gelation times for the hydrogels were tested by a vial-tilting method, and the results indicated that these gels have a fast gelation process. WAXD and DSC analyses demonstrated that the formation of a channel-type structure of a crystalline necklace-like complex induced the gelation process. The morphologies of various lyophilized hydrogels were observed by scanning electron microscope（SEM） measurement. The rheological measurements were employed to investigate the dynamic rheological properties, “gel-sol” transition process and viscosities of the supramolecular hydrogels. In addition, Doxorubicin hydrochloride（DOX·HCl）, as a model drug, was encapsulated into the hydrogels, and then was released from drug-loaded hydrogels in a pH-dependent behavior.（4） Core cross-linked polyphosphoester micelles with folate-targeted and acid-cleavable features for pH-triggered drug delivery. In this part, we have developed a novel folate-conjugated core cross-linked polyphosphoester micelles with acid-cleavable acetal groups（ACCL-FA） via a combination of ring-opening polymerization（ROP） and CuAAC “Click” chemistry and used for pH-triggered drug delivery. The synthesis routes can be divided into four steps: i) synthesis of an azide and acetal-functionalizced TEG（N₃-a-TEG-a-N₃）; ii) preparation of a polyphosphoester-based diblock copolymer（PBYP-b-PEEP） with the alkynyl groups in the pendants via the sequential ROP reaction of 2-（but-3-yn-1-yloxy）-2-oxo-1,3,2-dioxaphospholane（BYP） and 2-ethoxy-2-oxo-1,3,2-dioxaphospholane（EOP） monomers using IPA as the initiator and DBU as the organic catalyst; iii) introduction of an targeted folate group at the chain end of PBYP-b-PEEP; and iv) formation of the ACCL-FA micelles by CuAAC “Click” reaction between N₃-a-TEG-a-N₃ and PBYP-b-PEEP-FA. The chemical compositions and structures, as well as the molecular weights and PDIs of these copolymers have been fully characterized by 1H NMR, 31 P NMR, FT-IR and GPC measurements. The self-assembly behaviors of these amphiphilic star-block copolymers were investigated by a fluorescence probe method, dynamic light scattering（DLS） and transmission electron microscopy（TEM） analyses. In addition, the in vitro DOX release studies, MTT assays, live cell imaging system and flow cytometry analysis revealed that these polymeric micelles could efficiently deliver and release DOX into KB cells.（5） Synthesis and characterization of a novel acid-cleavable polymeric prodrug polyphosphoester–bortezomib. A new bortezomib-conjugated polyphosphoester prodrug with acid-cleavable groups（P（BYPBTZ-co-EEP）） was developed via a combination of ring-opening polymerization（ROP） and CuAAC “Click” chemistry. The synthesis routes of the polymeric prodrug can be divided into four steps: i) preparation of a polyphosphoester-based random copolymer P（BYP-co-EEP） with the alkynyl groups in the pendants via the ROP reaction of BYP and EOP monomer mixture using IPA as the initiator; ii) the clickable azidoacetylcatechol（AAC） was prepared via a nucleophilic substitution reaction between chloroacetylcatechol（CAC） and sodium azide（NaN₃）; iii) formation of the AAC functionalized polyphosphoester-based random copolymer P（BYPAAC-co-EEP） by CuAAC “Click” reaction between P（BYP-co-EEP） and AAC; iv) introduction of bortezomib at the side chain of P（BYPAAC-co-EEP） to prepare P（BYPBTZ-co-EEP）. The chemical compositions and structures of these copolymers have been characterized by 1H NMR, FT-IR, and GPC measurements.