Multifactor Synergistically Promoted Photothermal Nanosystem With Drug Targeted Delivery And Controlled Release

Currently,cancer is considered as the second leading cause of human death around the world.According to the latest data from the national cancer center in 2019,the number of cancer deaths in China reached 2.338 million in 2015,and 3.929million people were diagnosed with cancer.The death rate and incidence of malignant tumors have maintained rapid growth.In the past two decades,nanomaterials have been widely studied and applied in biomedicine,especially as drug carriers and medical imaging agents.Photothermal Therapy（PTT）is a non-invasive therapy that used nanomaterials with photothermal conversion property to generate local temperature increase under the stimulation of laser and thus kill tumor cells.Recently,the emergence of nanotechnology for integrated diagnosis and treatment provides a safe and effective method for cancer treatment.It integrates the treatment and imaging technology to achieve accurate and personalized treatment of cancer.In this thesis,photothermal materials were used to prepare multi-factor nano-drug system with targeted functions,imaging functions,drug controlled release functions and combined treatment functions on the basis of integrated diagnosis and treatment platform.The main contents were summarized as the followings:（1）Folic acid-modified Prussian blue/polydopamine nanoparticles as an MRI agent for use in targeted chemo/photothermal therapy.PTT also has its own defects due to uneven laser heat distribution,limited light penetration and cell-protective effects induced by high expression of heat shock protein.In this chapter,Prussian blue nanoparticles（PB NPs）with photothermal effect and imaging function were used as the core,and PB NPs were decorated with a layer of PDA through an in situ dopamine polymerization under alkaline conditions（PD NPs）.Then,PDA was bound with bi-amino polyethylene glycol（NH2-PEG-NH2）via a Michael addition and/or Schiff base reactions（PDG NPs）to endow the PD NPs with good physiological stability.To further enhance nanoparticles accumulation at the tumor site and to reduce the side-effects,Folic acid（FA）was attached at the end of PDG NPs,as FA can bind to the highly expressed folate receptor on the surface of most tumor cells.Finally,PDG-FA-DOX NPs was obtained by usingπ-πstacking and hydrogen bonding interactions to load DOX.The nanoagent was estimated to have an average size of 40 nm with a DOX-loading capacity of 36%,photothermal conversion efficiency of 45.7%and a transverse relaxation rate of 0.366 m M^-1s^-1.In vitro release investigations showed a dual-responsive release by a mild acid and near-infrared（NIR）laser irradiation.PDG-FA NPs illustrated negligible cytotoxicity against the HL-7702cell line and 38.2%cell viability under NIR against the He La cell line.PDG-FA-DOX NPs exhibited 45.2%cell viability.In contrast,the cell viability of PDG-FA-DOX NPs was dramatically decreased to 18.4%under NIR.Exclusive of folic acid,PDG-FA-DOX NPs demonstrated 40.5%cell viability.These results demonstrated the potential of the nanoagent for integrated photothermal therapy（PTT）and chemotherapy,also embracing the FA targeting effect.In vivo MRI confirmed the effective nanoparticle accumulation,while infrared thermal images revealed the dramatically increased temperature under NIR at a tumor site.In vivo combination treatment-induced tumors were nearly completely destroyed without significant body weight loss after 14 days.H&E and Ki67 staining indicated remarkable necrosis and weak cell proliferation in the tumor area.Histologic examination revealed a lower toxicity in the vital organs.Therefore,PDG-FA-DOX NPs are an effective multifunctional diagnostic and therapeutic reagent for MRI imaging and cancer treatment.（2）Chlorin e6 conjugated functionalized Prussian blue nanoparticles for MRI guided combination photodynamic/photothermal therapy.In the previous chapter,the multi-functional diagnostic and treatment reagent prepared by us is loaded with a chemotherapy drug DOX,but it was inevitable that there will be drug leakage in the drug delivery process,causing side effects to normal tissues.Therefore,in this chapter,PDT combined with PTT were adopted as another non-invasive treatment method to completely eliminate the tumor.At first,PD NPs which is composed of polydopamine（PDA）coated Prussian blue nanoparticles（PB NPs）for photothermal preparation,polyethyleneimine（PEI）was attached to the surface of PD to produce the PEI modified PD NPs（PDE NPs）,then the photosensitizer dihydrogen porphin e6（Ce6）was covalently conjugated on PDE NPs to achieve PDE-Ce6.Finally,HA was adsorbed on PDE-Ce6 NPs to improve the cancer-targeting properties（PDE-Ce6-HA NPs）.PDE-Ce6-HA NPs not only exhibited high colloid stability,good biocompatibility and suitable transverse relaxation rate(0.54 m M^-1s^-1),but also high photothermal conversion efficiency（40.4%）and excellent ROS generation efficiency under NIR light irradiation.The confocal microscopy images demonstrated a selective uptake of PDE-Ce6-HA by CD44overexpressed He La cells via HA-mediated endocytosis.Meanwhile,in vitro anti-cancer evaluation verified the significant photodynamic and photothermal combined effects of PDE-Ce6-HA on cancer cells.Moreover,PDE-Ce6-HA led to an increase of T1-MRI contrast in tumor site.Furthermore,in vivo anti-tumor evaluation proved that the PDE-Ce6-HA under both 808 and 670 nm laser demonstrated significantly high tumor growth inhibition effects compared with individual PTT or PDT.Hence,PDE-Ce6-HA is applicable in tumor targeted and MRI-guided photodynamic/photothermal combined treatment.（3）Magnetic theranostic nanosystems enabling photothermal/chemo combination therapy of triple-stimuli responsive drug release with MRIChemotherapy is still the most commonly used method to treat malignant tumors.How to avoid leakage of chemotherapy drugs in the delivery process is an urgent problem to be solved.Multifactor synergistically nanosystem can realize controllable delivery of drugs according to environmental changes.In this chapter,the release of chemotherapeutics were controlled by means of"endogenous stimulation"and"exogenous stimulation"combined with regulation.First of all,a magnetic resonance imaging and therapy（heat）function of field of Fe3O4 NPs were first coated with a layer of PDA to build core-shell Fe3O4@PDA nanoparticles（FP NPs）as MRI agent and enhanced photothermal performance.The cystamine dihydrochloride（CYS）containing reduction sensitive S-S bond was connected to FP NPs surface layer（named as FPC NPs）.Then hyaluronic acid（HA）was conjugated to FPC NPs to produce FPCH NPs.At last,DOX was coated onto FPCH NPs viaπ-πstacking and hydrophobic interactions to obtain the final product of FPCH-DOX NPs.HA shell would improve the cellular uptake of FPCH-DOX NPs through the binding role to the overexpressed CD44 receptor of tumor cells,but also serve as gatekeepers for controlling drug release.The nanocomposite possessed the average diameter of 120nm,saturation magnetization of 28.5 emu g^-1,DOX loading capacity of 7.13%and transverse relaxation rate of 171.76 m M^-1s^-1.The drug release could be triggered by p H,glutathione（GSH）and light irradiation.Prussian blue staining and confocal microscopy demonstrated the nanoplatforms can improve the biocompatibility and cellular uptake in CD44-positive He La cell lines rather than in CD44-negative NIH3T3 normal cell lines.In vitro evaluations demonstrated that the combination therapy of FPCH-DOX NPs brought about as low as 16.2%cell viability,sharply lower than single chemotherapy（55.3%）or PTT（52.1%）.In vivo MRI indicated the tumor accumulation of FPCH-DOX NPs by providing enhanced contrast and in vivo thermal imaging verified their localized photothermal conversion effect in tumor tissue.Importantly,FPCH-DOX NPs presented remarkable anti-tumor efficacy by photothermal-chemo combination therapy.H&E and Ki67 staining showed obvious necrosis and weak cell proliferation at the region of tumor.Thus,FPCH-DOX NPs are promising“all-in-one”nanoplatforms for highly effective cancer theranostics.