Study Of Multifunctional SiRNA Nano-delivery Systems Toward Enhancement Of Tumor Photothermal Therapy And Reversal Of Tumor Drug Resistance

Cancer is one of the major diseases that seriously threaten human life and health presently.The traditional treatment methods for cancer mainly include chemotherapy,radiotherapy and surgical resection.Although these methods have certain therapeutic effects,they all have significant defects.In recent years,the rise and development of nanomedicine technology has provided a new approach to the treatment of cancer,which taking advantage of nano-drug delivery system to deliver therapeutic drugs to the tumor site to destroy tumor cells,exhibiting obvious advantages over traditional methods.Nano-drug delivery system could prolong the blood circulation time of drugs,increase the enrichment of drugs at the tumor site,control the drug release,and reduce systemic toxic and side effects.However,the ideal therapeutic effect could not be achieved by relying on monotherapy due to the molecular complexity of tumor cells and the complicacies of tumor microenvironment.Therefore,it is urgent to design multi-functional nano-drug delivery systems and combine the advantages of different therapeutic methods to achieve efficient synergetic therapy for malignant tumors.Gene therapy and phototherapy have been widely researched and applied with the development of nanomedicine technology.In this paper,we were committed to multifunctional siRNA nano-delivery system toward enhancement of tumor photothermal therapy and reversal of tumor drug resistance.In chapter 1,we present a brief literature overview on the status of tumor therapy,drug-resistant tumor therapy,gene therapy,gene delivery barriers,gene delivery carriers,photothermal therapy(PTT)and photodynamic therapy(PDT).In chapter 2,we designed a spherical polypeptide with topological structure as siRNA delivery vector for the enhancement of tumor photothermal tharpy.The dendritic macromolecular PAMAM was used as initiator to initiate ring opening polymerization(ROP)of POB-L-Glu-NCA and introduce guanidine groups into the side chain via efficient "click chemistry" reaction,which ultimately yield spherical and helical polypeptides(DPP).Compared to linear polypeptides(LPP),DPP were demonstrated to have stronger transmembrane activity,higher siRNA uptake efficiency,and lower cytotoxicity,achieving better gene silencing effect.Then DPP were used to co-load siRNA and pohothermal reagent for synergistic therapy toward breast cancer.Photo thermal reagent ICG was loaded into the hydrophobic cavity of PAM AM,and siPKM2 inhibiting tumor glycolysis was encapsulated by electrostatic adsorption.In order to neutralize excessive positive charge,reduce the non-specific phagoc ytosis of reticuloendothelial system(RES)and prolong the blood circulation time,human albumin(HSA)was coated on DPP surface to form an uniform and stable D-I/P@HSA nanocomplexes(NCs).D-I/P@HSA NCs could accumulate at the tumor site through passive targeting and DPP could achieve the effective silencing of PKM2,and further destroy the glycolysis pathway and decrease the generation of ATP.The decrease of ATP could not only stimulate the apoptosis of tumor cells,but also reduce the production of heat shock protein(HSPs)by tumor cells under photothermal stimulation(0.8 W/cm2),so as to enhance the sensitivity of tumor cells to PTT.It was proved that D-I/P@HSA NCs achieved good synergistic therapeutic effect on MCF-7 tumor through gene therapy and enhanced PTT by various in vitro and in vivo experiments.In chapter 3,we designed a multistep ROS-responsive NCs,which could release si-gp and chemotherapy drug successively according to the difference of ROS concentration,so as to realize the programmed therapy toward multi-drug resistant tumors.Firstly,we synthesized cRGD modified,PEGylated tellurium crosslinked PEI(RPPT).Gel electrophoresis and other in vitro experiments demonstrated that RPPT could degrade in H2O2 with low concentration(100 ?M).Then we synthesized a prodrug(TK-Dox2)of doxorubicin dimer containing thioketal bonds.In vitro experiment showed that TK-dox2 was not toxic and required a higher concentration of ROS(50 mM)to be activated.Subsequently we prepared the multistep ROS-responsive NCs to treat MCF-7/ADR using RPPT and TK-Dox2.Firstly,TK-Dox2 and photosensitizer Ce6 were co-loaded into the negatively charged mesoporous silicon nanoparticle(MSN),and then a layer of RPPT was coated on the surface of the MSN by electrostatic adsorption.Finally,siP-gp was condensed to form multifunctional TDC@M/RPPTP NCs.TDC@M/RPPTP NCs could target tumor cells overexpressing?v?3 integrins and achieve endocytosis.Relatively small amount of ROS at the tumor site could oxidize the tellurium bonds to degrade RPPT,thereby releasing siP-gp and achieving effective silencing of P-gp.Subsequently,we carried out PDT to produce large amount of ROS.On the one hand,ROS could directly kill tumor cells.On the other hand,ROS could cut off the thioketal bond to activate the prodrug.At this moment,the drug-resistance of the tumor has been reversed and the drug could not be pumped out of the cell,so as to improve the utilization of drugs and reduce side effects,which ultimately achieved programmed therapy toward multidrug resistant tumor.In chapter 4,we designed a temperature-sensitive CO release system co-loading the redox-responsive drug TPZ,which could achieve the specific release of CO in tumor location and the activation of TPZ,so as to synergistically kill tumor cells.First,the mesoporous Prussian blue and carbonyl iron were coordinated,and then TPZ was encapsulated in Prussian blue.Cationic polymers(PAH),anionic polymers(PAA),and polyethylene glycol(PEG)were then sequentially encapsulated on the surface of Prussian blue by a layered self-assembly method to increase it's water solubility and stability.When the NCs were enriched at the tumor site,CO could be released under the non-lethal photothermal effect(42?),accelerating the mitochondrial oxygen consumption,which led to hypoxia in tumor cells and activation of TPZ.Meanwhile,the release of CO could promote tumor cells to produce ROS and induce apoptosis,so as to achieve the synergistic anti-tumor effect with TPZ was designed(in cooperation with Li Yongjuan).In chapter 5,we summarized the work of this paper and looked forward to the future research.