The Preparation Of Multifunctional Biomaterials Based On Polydopamine And Their Application In Cancer Treatment

Background and Objective:The development of new combinated treatment methods based on biocompatible materials is the frontier research direction in tumor precision treatment.Polydopamine(PDA)has shown unique advantages in the combined treatment of cancer because of its adhesion,biodegradability,and chemical modification.This thesis intends to construct two different combined treatment modes by using the PDA surface modification method and evaluate the therapeutic effects of the two strategies through animal experiments,to provide new ideas and theoretical basis for the diversified development of tumor combined treatments.They are as follows:In the first part,the integrated nano-drug Ce6@PDA-DCL-PFP targeting PSMA for prostate cancer diagnosis and treatment was constructed by multiple modifications on the surface of PDA nanoparticles,the application potential of this nano-drug for prostate cancer diagnosis and treatment was systematically evaluated through various experiments in vitro and in vivo.In the second part,PLGA microspheres were coated with PDA,and then the surface of PDA coating was chemically modified to introduce tetrazine labeled with iodine-131 to prepare radioactive microspheres—PLGA@PDATz-131I,using tetrazine click release reaction to realize drug release in tumor tissue.Using the orthotopic liver cancer model,the application potential of the microsphere in the treatment of liver cancer was systematically evaluated.Chapter Ⅰ PDA multifunctional nanoparticles targeting PSMA for prostate cancer theranosticsMaterials and Methods:1.PDA nanoparticles were prepared by solution oxidation with dopamine hydrochloride as raw material.Under the weak alkaline condition,benzoquinone group on PDA surface and the terminal amino group of the compound 5(PEG derivative of N-[N-[(S)-1,3-dicarboxypropyl]carbamoyl](S)-1-lysine(DCL))were reacted via Michael addition/Schiff base reaction.PDA-DCL nanoparticles with PSMA targeting function were prepared.2.In the same way,PDA-DCL reacted with terminal sulfhydryl of 1H,1H,2H,2H-perfluorododecanethiol to introduce fluorine chain on the surface of PDA-DCL nanoparticles and then used fluorine-fluorine interaction to adsorb 1H-perfluoropentane with ultrasonic properties to prepare PDA-DCLPFP ultrasonic nanoparticles.The F element of the nanoparticles was quantitatively analyzed by XPS.3.Finally,by loading photosensitizer chlorin e6(Ce6)by π-π stacking,the integrated medicine for diagnosis and treatment-Ce6@PDA-DCL-PFP nanoparticles were prepared,and the maximum loading rate of Ce6 was detected by UV-Vis.PDT and PTT efficiency of particles irradiated by NIR laser were evaluated.4.Evaluate the targeting performance of the nano-drug by cell imaging experiment,and investigate the killing effect of PDT/PTT combined therapy on cells by flow cytometry.5.Evaluate the diagnostic effect of Ce6@PDA-DCL-PFP on prostate cancer by in vivo ultrasound imaging and evaluate the distribution of nanoparticles in vivo by fluorescence imaging of small animals.6.Evaluate the curative effect of PTT/PDT combined therapy with Ce6@PDA-DCL-PFP on prostate cancer by using the tumor inhibition rate of living tumor.Experimental Results:1.The particle size of PDA nanoparticles was 235 nm;the DCL content in PDA-DCL nanoparticles was 10.11%.2.The F content of PDA-DCL after introducing fluorine chain was 1.45%and the F content of PDA-DCL-PFP prepared after further adsorption of fluoride reached 9.97%.3.Ce6@PDA-DCL-PFP nano-drug prepared by multiple modifications had a particle size of 250 nm;The maximum load rate of Ce6 is up to 52.80%.Nanoparticles irradiated by NIR laser shown good PDT performance and its photothermal conversion efficiency was 22.56%.4.Cell imaging results showed that the degree of cell internalization of Ce6@PDA-DCL-PFP was 6.5 times higher than that of non-targeted nanoparticles.The results of flow cytometry showed that the lethal rate of PDT/PTT combined therapy to cells was 83.7%.5.Ce6@PDA-DCL-PFP formed uniform,dense and high-level echoes in tumor tissues when used in tumor imaging,which can realized the ultrasonic diagnosis of prostate cancer.In vivo imaging of small animals showed that Ce6@PDA-DCL-PFP nanoparticles were circulated in vivo for 24 h and the uptake of Ce6@PDA-DCL-PFP nanoparticles reached the maximum.6.Using Ce6@PDA-DCL-PFP to treat tumor with PTT/PDT,the tumor inhibition ratio reached 80.0%.Conclusion:In this thesis,the nano-drug Ce6@PDA-DCL-PFP targeting prostate cancer was successfully prepared by multiple modifications on the surface of PDA nanoparticles.Among them,DCL actively targeted PSMA and improved the degree of intracellular internalization of nanoparticles and the enrichment at tumor sites;The loaded PFP realized the ultrasonic diagnosis of prostate cancer;Using PTT properties of PD A nanoparticles and PDT properties of Ce6,PTT/PDT combined therapy for prostate cancer was successfully realized.In a word,this multifunctional nano drug targeting PSMA provided effective measures for the diagnosis and treatment of prostate cancer,with potential clinical application prospects.Chapter Ⅱ The fabrication of PLGA@PDA-Tz-131I microspheres:radioembolization therapy combined with chemotherapy regulated by click-release reaction for liver cancer therapyMaterials and Methods:1.TCO-PNB ester and Mitomycin C are used as raw materials to synthesize TCO-Mitomycin C prodrug through nucleophilic substitution reaction;Tetrazine compounds were synthesized with nickel triflate as catalyst;iodine-131 labeling by electrophilic substitution.2.PLGA microspheres were prepared by emulsion crosslinking method with PLGA polymer and polyvinyl alcohol as raw materials.PDA coating was modified on the surface of PLGA microspheres,and the thickness of PDA coating was measured by XPS.3.PLGA@PDA-Tz-131I microspheres were prepared by Michael addition/Schiff base reaction between benzoquinone group of PDA coating on the surface of microspheres and the amino group of the compound 18 labeled with iodine-131(Tz-131I)under alkaline condition.The stability of microspheres in serum and PBS was evaluated.4.The release efficiency of TCO-Mitomycin C from microspheres was detected,and the release efficiency of Mitomycin C at different time points(0 min,30 min,2 h,4 h)was monitored by HPLC.5.PLGA@PDA-Tz-131I microspheres were injected into liver through hepatic artery embolization and SPECT/CT fusion imaging was used to monitor the distribution and stability of microspheres in vivo.6.The orthotopic liver cancer model of rats was established by DEN,and the liver cancer model was identified by MR imaging.Hepatic carcinoma model mice were treated with PLGA@PDA-Tz-131I microspheres via hepatic artery embolization combined with chemotherapy regulated by tetrazine click release reaction.The effect of combined treatment was evaluated by MRI imaging,survival time statistics and pathological examination of liver tumors.Experimental Results:1.The compound 14 were successfully synthesized by 4 steps of chemical synthesis,and the total yield was 5.8%.The yield of TCO-Mitomycin C prodrugs was 64.2%.The labeling rate of tetrazine small molecule labeled with iodine-131 was 92.9%.2.The particle size of PLGA microspheres was about 33 μm,which met the particle size requirements of radioactive embolic microspheres.PDA coating thickness on PLGA@PDA microspheres was about 20 nm.3.The iodine-131 labeling ratio of PLGA@PDA-Tz-131I microsphere was 50%.After incubation in PBS and serum for 2 weeks,the proportion of microspheres still stably binding iodine-131 was 90.0%and 84.0%,respectively.4.When the tetrazine compound on the surface of microspheres reacts with TCO-Mitomycin C for 30 min,the release rate of Mitomycin C was 47.5%and the release ratio reached 53.1%after 4 h.5.SPECT/CT image showed that PLGA@PDA-Tz-131I microspheres were distributed in liver tissue after arterial embolization,and iodine-131 signal was detected in liver during continuous scanning for 60 days.6.The success rate of drug induction for hepatic carcinoma model was 81.0%.Transhepatic arterial embolization with PLGA@PDA-Tz-131I microspheres combined with chemotherapy regulated by tetrazine click release reaction significantly reduced tumor volume and inhibited liver cancer metastasis.The survival time of the combined treatment group was prolonged by 45 days compared with that of the control group.Pathological examination results showed that the liver tumor cells in the combined treatment group were necrotic and the normal liver cells were normal in morphology.Conclusion:In this work,PLGA@PDA-Tz-131I microspheres,whose particle size of was about 35 μm,were successfully prepared by PLGA microspheres coated with PDA layer and introducing Tz-131I on the PDA surface.PLGA@PDATz-131I microspheres were used for hepatic artery embolization.On the one hand,internal irradiation treatment for liver cancer was realized,and on the other hand,chemotherapy drugs were released in situ by tetrazine click-release reaction,so that radiotherapy and chemotherapy were simultaneously realized for liver in situ.The experimental results showed that the combined treatment strategy had a significant curative effect on liver tumors,and it is a combined treatment strategy of radioactive embolization and chemotherapy with clinical application prospect.