Construction Of Intrinsic Stimuli-responsive Nanocarriers And Inhibition Of Liver Cancer By Drug Release

Stimuli-responsive nano-carriers have attracted more and more attention due to their unique functional characteristics,such as improving the bioavailability of poorly soluble drugs,achieving the “on-demand” delivery to tumors and reducing the side effects of drugs.However,many challenges still exist in the application of combination therapy via these nano-carriers.In order to construct a more intelligent carrier system with improved drug loading capacity and controlled release ability,series of self-assembled nano drug delivery systems in response to the intrinsic stimuli were designed in this work.Firstly,a pH-responsive polymer self-assembly drug delivery system was constructed by means of drug bonding and gene electrostatic assembly.Doxorubicin(DOX)was connected with chitosan-polyethyleneimine polymer(CS-PEI)via a pH-sensitive linker 4-Hydrazinobenzoic acid(HBA).Bcl-2 si RNA was loaded to CS-PEI-HBA-DOX by electrostatic adsorption.Glycyrrhizic acid(GA)was modified to the carriers as targeting unit to form the pH responsive co-delivery system glycyrrhizic acid-chitosan-polyethyleneimine-4-Hydrazinobenzoic acid-doxorubicin(GA-CS-PEI-HBA-DOX@si RNA).The results demonstrated the particle size and surface charge of micelles were closely related to the polymer/nucleic acid(N/P)ratio.The particle size and the zeta potential values of the GA-CS-PEI-HBA-DOX-@si RNA micelle increased with the increase of N/P value.The charge reversion characteristic of the micelle offered an effective way for long circulation in bloodstream(pH 7.4)and easy cell uptake in tumor tissues(pH 6.8),as the positively charged micelles were easily caught by the reticuloendothelial system(RES).In the process of somatic circulation,GA-CS-PEI-HBA-DOX@si RNA micelles stably combined to DOX and si RNA,thus preventing drug leakage.In the slightly acidic conditions of tumor cells,the controlled targeting release of drugs was achieved by means of hydrazone bond cleavage and micelle structure destruction.The DOX and Bcl-2 si RNA played a synergistic effect to improve the therapeutic effect of tumors.The target ing ability to liver cancer cells was enhanced by GA receptor mediated endocytosis.Secondly,a redox-responsive polymer self-assembly drug delivery system was constructed by hydrophobic inclusion method.Polyhexolactone(PCL)with low molecular weight,as a hydrophobic core,was linked with carboxymethyl chitosan through a disulfide bonds linker(cystamine)to synthesize the amphiphilic polymer PCL-SS-CS.The DOX and the pheophorbide a(PHA)were embedded in the hydrophobic core of the PCL-SS-CS-GA micelle through the hydrophobic interaction,and the GA was modified to the micelle surface as a target ligand.The redox-responsive chemical drugs and photodynamic therapy drugs co-delivery system PCL-SS-CS-GA@DOX/PHA was obtained.The micelle had obvious GSH response sensitivity to drug release.In 0.01 m M GSH solution,the binding state with DOX and PHA can be maintained well,and the side effects on normal tissues and organs can be reduced.In 10 m M GSH solution,the loaded DOX and PHA were released as the results of breakage of disulfide bond and destruction of micelle structure.In order to solve the problem of poor drug loading efficiency of polymer micelle delivery system,a small molecular predrug self-assembly nano-drug delivery system was constructed.Capetabine(CAP),curcumin(CCM)and lysine-modified curcumin(LCM)were used to react with Borotezomi(BTZ)and the phenylboric acid connector-modified DOX(DOX-PBA)and PHA(PHA-PBA)to synthesize prodrug compounds with boric acid ester bond.The prodrug micelles were self-assembled in aqueous solution by spiral membrane evaporation and solvent volatilization method.The critical micelle concentration of the small molecular prodrug compound was significantly related to the hydrophilicity/hydrophobicity of the original drug compound.The prepared LCM-PBA-PHA micelles were sensitive to low pH,high ROS and ATP concentration.In addition,the degree of micelle disintegration and drug release were more complete under dual or triple stimuli conditions.The inhibitory effects on tumor cells,pharmacokinetic properties,targeted delivery properties and combined tumor treatment effects of stimuli-responsive nanodrug vectors were studied by human hepatoma cell model HepG2 in vitro and a HepG2 xenograft model in vivo.The stimuli-responsive nano drug carrier prepared in this article exhibited high potency of drug co-delivery in vitro and in vivo.After intravenous administration,it can significantly prolong the average residence time and half-life of the drug,enhance the degree of drug enrichment at the tumor sit e and increase the bioavailability.GA-CS-PEI-HBA-DOX@si RNA micelles can fully exert the combined effects of DOX and Bcl-2 si RNA,and the combined inhibition of HepG2 tumors in vitro and in vivo is significantly higher than that of a single drug therapy.Due to the synergistic effect of chemotherapy and photodyn amic therapy,DOX and PHA delivered by PCL-SS-CS-GA significantly improved the therapeutic effect on HepG2 tumors.The prepared LCM-PBA-PHA@si RNA micelles can fully exert the combination therapy effect of si RNA gene therapy and PHA photodynamic therapy,and the inhibition rate of the HepG2 xenograft model tumor reached up to 86.8%.