Preparation And Performance Study Of Dual-responsive Chitosan-Based Drug Carriers Containing Disulfide Bonds

According to the World Health Organization,cancer is the second leading cause of death worldwide today.Since tumor cells spread widely in the late stage of cancer,chemotherapy,which mainly relies on anti-cancer drugs to induce apoptosis of tumor cells,has become one of the main methods of cancer treatment.However,there are still some important challenges in drug therapy,such as poor stability of drug carriers in blood circulation,poor permeability around tumor cells,early release of drugs and lack of targeting.Targeting and local delivery chemotherapeutic agents not only help to reduce poison side effects,but also improve the therapeutic efficacy of the drugs.Therefore,p H and glutathione(GSH)dual responsive drug-loaded carboxymethyl chitosan(DOX@CMC-SS-CMC)gels and GSH and hyaluronidase(Halys)dual responsive drug-loaded hyaluronic acid/chitosan(DOX@m HA/CS-SS-CS)microspheres were synthesized in this paper.The purpose of the former is to solve the problem of early drug release and poor permeability around tumor cells.The latter uses the hyaluronic acid shell layer to further solve the problems of poor carrier blood stability and inability to target tumor cells.The main studies and results of this thesis are as follows:(1)DOX@CMC-SS-CMC gels with p H and GSH dual responsiveness were designed and prepared.Carboxymethyl chitosan(CMC),which could be soluble in water,was first synthesized,and then reacted with 2,2’-dithiobenzoyl chloride to generate disulfide-bonded carboxymethyl chitosan(CMC-SS-CMC)gels.The equilibrium dissolution curves of CMC-SS-CMC gels were characterized at different p H and GSH concentrations,demonstrating their dual responsiveness to p H and GSH.The drug release profiles of the drug-loaded gels within 84 h were characterized by UV spectrophotometer.The results showed 13.3% of the drug was released in PBS buffer at p H 7.4,while its drug release rate reached 36.5% at p H 5.5 and 91.5% in PBS buffer with 10 m M GSH.These results demonstrated that DOX@CMC-SS-CMC gels possessed p H and GSH dual responsiveness,allowing drug release at low p H and high GSH concentrations.(2)Core-shell hyaluronic acid/chitosan(m HA/CS-SS-CS)microspheres with targeted and charge reversal stepwise Halys and GSH dual responsive properties were prepared.CS-SS-CS microspheres were first synthesized by the reverse microemulsion method and then the CS-SS-CS microspheres were self-assembled with methacrylated hyaluronic acid to form m HA/CS-SS-CS microspheres by electrostatic interaction.The size distribution and surface Zeta potential magnitude of the microspheres were characterized by DLS.This shift from positive to negative charge demonstrated that the core-shell structured microspheres possessed charge reversal properties.The stability of m HA/CS-SS-CS microspheres was measured at different Na Cl,p H,GSH and Halys concentrations,and the results showed that the m HA/CS-SS-CS microspheres exhibited Halys and GSH dual-responsive properties.(3)The in vitro cytotoxicity and drug release of DOX@m HA/CS-SS-CS microspheres were investigated.MTT assay characterized the in vitro cytotoxicity of different concentrations of m HA/CS-SS-CS microspheres and DOX@m HA/CS-SSCS microspheres on He La cells at 24 h and 48 h.Blank microspheres exhibited lower cytotoxicity,while drug-coated microspheres showed greater cytotoxicity.Next,the drug release profiles of DOX@CS-SS-CS microspheres were tested at different GSH concentrations,and the results showed that the DOX release rate was 91.2% after 84 h in PBS buffer of 10 m M GSH.The drug release profiles of DOX@m HA/CS-SS-CS microspheres were tested at different GSH+Halys concentrations,and the DOX release rate was 88.0% in PBS buffer of 10 m M GSH+150 unit/m L Halys.The main mechanism of drug release is that when DOX@m HA/CS-SS-CS microspheres are targeted to the tumor site,the high content of Halys causes the m HA to be degraded,exposing the positively charged DOX@CS-SS-CS microspheres inside.This makes the internal charge DOX@CS-SS-CS microspheres exposed,which binds to the negatively charged cell membrane and endocytosis into the tumor cell.The high content of GSH inside the tumor cells caused the disulfide bonds of the microspheres to break and releasing the DOX inside.This indicated the successful prepared dual-responsive drug carriers had potentials in tumor therapy.