| Immune checkpoint blockade(ICB)therapy can relieve the immunosuppression of T lymphocytes by immune checkpoint blocking,so that the host immune system can recognize and eliminate tumor cells.However,the antitumor efficacy of ICB therapy depends to a large extent on the infiltration of cytotoxic T lymphocytes,which means that“cold tumor”with limited T lymphocytes infiltration cannot benefit from this treatment.Therefore,it is urgent to improve the response rate of“cold tumor”to ICB therapy,so as to improve the antitumor effects of ICB therapy.It is reported that doxorubicin(DOX)can not only kill tumor cells directly,but also induce immunogenic cell death(ICD)of tumor cells,enhance the immunogenicity of tumor cells,and increase the infiltration of T lymphocytes in tumor.So the combination of DOX and immune checkpoint inhibitors can effectively activate the antitumor immune response,improve the response rate of tumor to ICB therapy,and enhance the antitumor effects.However,chemotherapeutic agents usually function inside the tumor cell,while immune checkpoint inhibitors are efficacious out of the tumor cell.Therefore,co-delivering an immune checkpoint inhibitor and a chemotherapeutic drug to different sites of a tumor is a key issue to improve the antitumor effects.Given these problems,a pH-responsive Pickering nanoemulsion was constructed in order to improve the antitumor effects,which was used for encapsulation and specified spatial delivery of small molecule immune checkpoint inhibitor HY19991(HY)and chemotherapy drug DOX.The main contents and results of this study are listed as following:(1)A pH-responsive Pickering nanoemulsion was successfully prepared by using the nanogel SNG with pH-triggered hydrophilicity-hydrophobicity switch,and redox-responding properties as stabilizer,isopropyl myristate as the oil phase,which underwent high-speed shearing and ultrasonic treatments.Experimental results showed that PNE was of spherical morphology,and gradually disintegrated at pH 6.5 and 37°C.(2)DOX was loaded into SNG by the solvent evaporation method,and the hydrophobic drug HY was dissolved in the oil phase to construct a Pickering nanoemulsion that co-loaded the chemotherapy agent DOX and programmed cell death protein 1(PD-1)/programmed cell death protein-ligand 1(PD-L1)interaction inhibitor HY(named D/HY@PNE).D/HY@PNE underwent the phase separation of oil and water at pH 6.5 and 37°C.The results of drug release showed that the in vitro release of HY from D/HY@PNE at acidic pH was significantly faster than that at pH 7.4.DOX can release continuously from D/HY@PNE in response to low pH and high glutathione conditions.(3)In vitro cell experiments verified PNE had good biocompatibility.The results of cell uptake showed that D@PNE was internalized by 4T1 cells at pH 6.5 significantly higher than that at pH 7.4.Cytotoxicity experiments showed that D@PNE inhibited the proliferation of 4T1,A549 and B16 cells in a concentration dependent manner.ICD detection experiments showed the extracellular release of adenosine triphosphate、high mobility group protein 1 and the exposure of calreticulin increased after 4T1 cells treated with D@PNE.Dendritic cells maturation was efficiently induced after incubation with D@PNE-pretreated tumor cells,indicating that D@PNE can induce the ICD of tumor cells effectively.(4)In 4T1 tumor-bearing mice model,D/HY@PNE can not only significantly kill tumor cells,but also induce ICD of tumor cells,promote the maturation of dendritic cells,increase the infiltration of CD8~+T cells in the tumor site,and trigger antitumor immune response.In conclusion,the pH-responsive Pickering nanoemulsion can be used for specified spatial co-delivery of different therapeutic or diagnostic reagents,and has a promising application potential in the diagnosis and treatment of tumor. |
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