NIR Responsive Tumor Vaccine In Situ Amplifies Photothermal Ablation Triggered Antitumor Immune Responses

Objective:Photothermal ablation not only causes tumor cell death or apoptosis,but also triggers immunogenic death to release tumor-associated antigens.However,the tumor immunosuppressive microenvironment severely weakens the dendritic cells’ ability to recognize and present antigens,and therefore cannot trigger strong anti-tumor immunity.Therefore,a new type of near-infrared light-responsive in situ vaccine(HA-PDA@IQ/DOX HG)was prepared in this paper to mediate a powerful anti-tumor immune response triggered by photothermal ablation.After characterizing the physical and chemical properties,we explored the photothermal conversion,cell phagocytosis,and cell compatibility of HA-PDA@IQ/DOX HG;At the same time,using mouse breast cancer as a tumor model,the therapeutic effect of photothermal ablation in animals and the potential value of enhancing anti-tumor immune response were discussed.Methods:1.First,stabilizer hyaluronic acid(HA)and immune adjuvant imiquimod(IQ)were used as precursors,IQ-loaded hyaluronic acid dopamine nanoparticles(HA-PDA@IQ NPs)were synthesized by one-step oxidative polymerization of dopamine initiated by KMn O4.2.Then,the chemotherapeutic drug doxorubicin(DOX)was adsorbed on the surface of HA-PDA@IQ NPs through π-π stacking to form HA-PDA@IQ/DOX NPs.Finally,HA-PDA@IQ/DOX NPs were blended with temperature-sensitive hydrogel F127 to obtain a near-infrared light-responsive in situ vaccine(HA-PDA@IQ/DOX HG).3.Transmission electron microscope(TEM)and X-ray diffraction(XRD)were used to characterize the physical properties of HA-PDA@IQ/DOX NPs;Elemental mapping and X-ray photoelectron spectroscopy(XPS)were used to characterize the chemical properties of HA-PDA@IQ/DOX NPs.4.Under 808 nm near infrared light(NIR)irradiation,photothermal conversion ability and photothermal stability of HA-PDA@IQ/DOX NPs were explored,At the same time,CCK-8 experiment and living dead cell staining experiment were used to explore its photothermal tumor killing ability.The laser scanning confocal microscope was used to characterize its location after entering the cell,and the CCK-8 experiment and hemolysis experiment were used to detect its biocompatibility..5.The 4T1 subcutaneous breast cancer model was constructed.After in situ injection of HA-PDA@IQ/DOX HG in vivo,flow cytometry was used to analyze memory T cells in mouse lymph nodes and cytotoxic T lymphocytes in spleen.6.Luc-4T1 subcutaneous breast cancer model was constructed,after in situ injection of HA-PDA@IQ/DOX HG in vivo,the small animal live imager was used to monitor the tumor growth of the mice in each treatment group,and the mouse body weight,tumor volume and growth curve were analyzed.Finally,the H-E slicing technique was used to analyze the histocompatibility of the important organs of mice.Result:1.The IQ/DOX-loaded hyaluronic acid polydopamine nanoparticles HA-PDA@IQ/DOX NPs in brown-black powder form were prepared successfully.2.The prepared HA-PDA@IQ/DOX NPs had uniform particle size,about 150 nm,and conformed to gaussian normal distribution.At the same time,HA-PDA@IQ/DOX NPs had an amorphous structure;Its constituent elements include carbon,nitrogen,oxygen,and manganese,and the surface was rich in hydrophilic groups such as-COOH and-OH,which made it have good water dispersibility.3.Under 808 nm near-infrared light(NIR)irradiation,HA-PDA@IQ/DOX NPs could quickly increase the temperature of the aqueous solution,and calculate its photothermal conversion efficiency as high as 41.2%;After repeated irradiation,its heating effect remained stable,And would not weaken with the increase of the number of exposures,showing good repeatability and stability.The results of CCK-8experiment and living dead cell staining experiment showed that HA-PDA@IQ/DOX NPs have good tumor photothermal killing ability.4.Laser scanning confocal results showed that HA-PDA@IQ/DOX NPs will quickly enter the cell and locate in the cytoplasm,but not enter the nucleus;After 2days of incubation with the cells,the cell viability remained above 80% when the concentration was as high as 1200 μg/m L;the hemolysis experiment showed that there was no hemolysis even at the highest concentration.Both experimental results prove that HA-PDA@IQ/DOX NPs have good biocompatibility.5.The results of flow cytometry showed that HA-PDA@IQ/DOX HG could promote the maturation of DC cells,the differentiation of memory T cells in lymph nodes and cytotoxic T lymphocytes in spleen.6.The results of the in vivo imager showed that HA-PDA@IQ/DOX HG could significantly inhibit the growth of tumors.In addition,the weight of the mice did not change significantly.The growth curve results showed that the survival period of mice in the HA-PDA@IQ/DOX HG treatment group could be extended to 60 days.The H-E staining chart showed that HA-PDA@IQ/DOX HG did not cause obvious pathological changes in major organs,indicating that it had good histocompatibility.Conclusion:1.The IQ/DOX-loaded hyaluronic acid polydopamine nanoparticles HA-PDA@IQ/DOX NPs with superior photothermal conversion efficiency,immune activation,good biocompatibility and nuclear magnetic imaging function was prepared successfully.2.HA-PDA@IQ/DOX HG obtained by compounding with temperature-sensitive hydrogel could be injected locally to achieve tumor local administration.This method not only could prolong the treatment window which was conducive to the photothermal ablation of the tumor,but also helped induce the body’s anti-tumor immune response.