| Objective: This study aimed to construct and characterize a novel nanomachine(CPDNs)using the cell membrane of castration-resistant prostate cancer(CRPC)as a biomimetic carrier to encapsulate polylactic acid-glycolic acid copolymer(PEG-PLGA)containing chemotherapy drug docetaxel(DTX),and to demonstrate its efficacy in vivo and in vitro for the treatment of CRPC.Methods: This nanostructure(CPDNs)was constructed by encapsulating DTX into PEG-PLGA nanoparticles,which were then modified with CRPC cell membranes.we first presented a methodology for the synthesis of CPDNs.The CPDNs were subsequently characterized by transmission electron microscopy,Fourier transform infrared spectroscopy,ultraviolet-visible spectroscopy and Western blotting.In vitro,the uptake of CPDNs was quantitatively and qualitatively observed by fluorescence labeling.Then,the effects of the CPDNs in the treatment of CRPC in vivo and in vitro were studied by dual-model imaging,fluorescence and radionuclide imaging.In addition,we evaluated the therapeutic efficacy in mice model bearing CRPC tumors.Results: In this study,PLGA was used as the base material to prepare NPs loaded with DTX by modified emulsification and evaporation method,and a novel nanomedical drug CPDNs was synthesized by co-extrusion method for the first time.In this study,DTX nanoparticles(PDNs)of PLGA were used as the core,and purified Du145 cell membrane(CMs)was used as the bionic shell of tumor cells.Finally,CMs was used to modify the surface of PDNs.The appearance of NPs was observed by transmission electron microscopy(TEM).The morphology of CPDNs is spherical core-shell structure.The electron microscope size of CPDNs is 125.4±0.6nm.Due to the presence of CMs in CPDNs,the size of CPDNs is larger than that of PDNs(102.2±0.5nm).Next,the diameter and surface charge of the nanoparticles were measured using the dynamic light scattering(DLS)method.The particle diameter was similar to that measured by transmission electron microscopy,and the surface ZETA potential of CPDNs was close to that of membrane vesicle level potential.The key membrane protein molecules in the CPDNs were identified by Western blotting.The study showed that the key proteins on the Du145 cell membrane were retained in the CPDNs,suggesting that the CPDNs continued the biological function of the Du145 cell membrane.Western blotting detected CD44,E-cadherin,CD147 and CD47.These membrane proteins are activated receptors on the membrane of prostate cancer cells and play a key role in the isotype interaction of tumor cells and the adhesion function of metastasis.Scanning fluorescence microscopy was used to observe the co-localization of CMs(red)and PDNs(green)in Du145 cells.Yellow indicates successful co-localization of CMs and PDNs signals,indicating that CPDNs maintains a particularly stable structure.Next,the release function of DTX in PBS and 1640 cell culture medium was studied to detect the stability of DTX in CPDNs.When DTX concentration was 0.1 mg/ml,the encapsulation rate of DTX was 52.5%,and CPDNs hardly encapsulated more DTX as DTX concentration increased.The release rate of DTX was low in the CPDNs group,indicating that the outer envelope acted as a diffusion barrier to reduce drug diffusion release,indicating that the CPDNs structure was relatively stable.In vitro,to verify the stability of PDNs and CPDNs,the diameter of CPDNs changed little in1640 cell culture medium and PBS cell culture medium,while the size of PDNs increased somewhat in 1640 cell culture medium.Considering that PDNs in 1640 cell culture medium is related to protein binding adsorption,It is further suggested that cell membrane can enhance the structural stability of PDNs in addition to homologous targeting ability.Under fluorescence microscope,the uptake of CPDNs by Du145 cells was significantly higher than that of PDN and erythrocyte membrane coated PDNs(RPDNs).Meanwhile,as compared with Pr EC of normal prostate epithelial cells,there was no significant difference in uptake among the three cells,indicating that CPDNs had specific targeting ability on the same type of Du145 cells.The study verified the stability,biocompatibility and in vitro targeting of CPDNs.The next step was to evaluate the targeting of CPDNs in Du145 tumor mouse model.Studies have shown significant differences in the accumulation of CPDNs in tumors at 0.5-,1-,2-,4-,12-and 24-h in vivo tumor imaging.24 h after injection,the tumor of CPDNs group still showed obvious fluorescence,while the fluorescence of PDNs group was negligible.In addition to tumor,PDNs and CPDNs are also distributed in kidney and liver.Finally,SPECT-CT system was used to verify the advantages of CPDNs in the treatment of CRPC.Corresponding to the in vivo biological imaging data,SPECT-CT more clearly and intuitively demonstrated that CPDNs can penetrate deeply into prostate cancer tissue,which further verified the efficacy of CPDNs homotype targeting prostate cancer and its long-term existence in the circulatory system.In order to further verify the homotype targeting function of CPDNs in the treatment of CRPC in vivo,the chemotherapy effect of CPDNs was comprehensively evaluated by using Du145 tumor model in nude mice.After one week of treatment,there was no significant difference in tumor growth among the groups.However,tumor growth was significantly inhibited in CPDN group compared with DTX group and PDN group.To comprehensively evaluate the chemotherapy effect of CPDNs on CRPC,the study found that the median survival time in the CPDN group was significantly longer than that in the other three groups,and there was no difference between the DTX group and the PDN group.Weight change is an important index to evaluate the systemic toxicity of chemotherapy drugs.The study compared the toxicity of different groups.The study showed that the CPDN group had significantly less weight loss than the other groups,suggesting that CPDNs had fewer toxic side effects and better chemotherapy tolerance.Finally,the changes of CD34 and Ki67 expression levels in prostate cancer tissues after treatment were detected by immunohistochemistry.The results showed that the expression levels of Ki-67 and CD34 proteins in the CPDN group were decreased,suggesting that the CPDN group significantly inhibited the tumorigenicity and angiogenesis of prostate cancer,and had a stronger anti-cancer effect.Through Du145 subcutaneous tumor model in nude mice,the study verified that CPDNs can effectively inhibit prostate cancer in vivo,prolong the median survival time of mice,have less toxicity,improve the level of chemotherapy tolerance,and have a stronger inhibitory effect on tumor.Conclusion: Membrane encapsulation technology is a hot field in nanobiology research.By wrapping nanoparticles in cell membranes,the combination of natural cell membrane properties and nanoparticle properties can be achieved,which not only increases the biocompatibility of nanoparticles,but also achieves better targeting and cycling properties in vivo.In this study,we constructed a novel homologous targeted bionic nanoparticle CPDNs that uses CRPC cell membrane(Du145)to wrap the NPs(PLGA)of docetaxel DTX(PDN),preserving the biological function of prostate cancer cell membrane.The results showed that CPDNs was spherical and had core-shell structure.CPDNs had homologous targeting mechanism with cancer cell membrane and immune escape ability.CPDNS had good targeting ability against CRPC,long blood circulation time,significantly inhibited tumor growth and prolonged survival time in vivo and in vitro.The specific homotype targeting provided by the CRPC membrane coating was validated using in vivo fluorescence and radionuclide dual model imaging,and CPDNs treatment was significantly improved in a mouse model with CRPC tumors.CPDNs has realized targeted chemotherapy for CRPC.As a novel cell-wrapped nanoparticle technology,CPDNS provides a new idea for specific targeted chemotherapy for tumors. |
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