Core-shell Structures Of Carbon Nanohorns And Liposome For The Co-delivery Of Doxorubicin And Tumor Associated Macrophages Polarization Factor IL-21

Objectives:This study aimed to prepare the core-shell structures of pH-sensitive liposome(PSL)-coated single-walled carbon nanohorns(SWCNHs)for the co-delivery of doxorubicin(DOX)and tumor associated macrophages(TAM)polarization factor IL-21,and to evaluate the physicochemical properties,drug release,cytotoxicity,cellular uptake,intracellular localization,tumor associated macrophages polarization in vitro and the anti-tumor effects in vivo.Methods:Raw SWCNHs were oxidized by H2O2 to form the oxidized SWCNHs(O-SWCNHs).DOX was nocovalently conjugated with the O-SWCNHs via ?-? stacking interaction to form DOX-O-SWCNHs,which was further coated with the PSL loading IL-21 to form IL-21-PSL-DOX-O-SWCNHs.Nanocarriers were characterized by laser paticle size analyzer,transmission electron microscopy,and fourier transform infrared resonance spectroscopy.The drug loading and encapsulation efficacy were determined by ultraviolet-visible spectrophotometry.The cytotoxicity of DOX-O-SWCNHs was evaluated in A549 cells using a WST-1 assay.The cellular uptake and intracellular localization of functionalized SWCNHs were performed by using flow cytometry and confocal laser scanning microscope.The polarization of TAMs was evaluated by RT-PCR.In addition,the tumor targeting,tissue distribution,and anti-tumor effects of IL-21-PSL-DOX-O-SWCNHs were also evaluated in A549 tumor-bearing nude mice.Results:According to a series of characterization,it was revealed that IL-21-PSL-DOX-O-SWCNHs were successfully synthesized.The drug loading and encapsulation efficiency of O-SWCNHs were 75%and 92%,respectively.The in vitro release at pH 5.5 was about 30%,which was faster than that at pH 7.4,presenting a pH sensitive drug release.The blank nanocarriers showed relatively low cytoxicity to A549 cells and 293T cells,showing more than 80%of cell viability.The IC50 value of PSL-DOX-O-SWCNHs for A549 cells was 125 ?g/mL.The cellular uptake and intracellular localization of nanocarriers showed that PSL-DOX-O-SWCNHs could be effectively uptake by A549 cells.PCR results showed that IL-21 could down-regulate the expression of M2 macrophage-related factors and up-regulate the expression of M1 macrophage-related factors.Compared to IL-21-PSL-DOX-O-SWCNHs showed remarkably higher tumor-targetting and distribution effects,and significantly reduced the tumor volume of A549 lung cancer cells-bearing nude mice,showing synergistic effects of DOX and IL-21.Conclusion:Novel functionalized SWCNHs loading DOX and IL-21(IL-21-PSL-DOX-O-SWCNHs)were successfully constructed.The functionalized SWCNHs showed effective anti-tumor effects both in vitro and in vivo,and might provide a theoretical basis for the co-delivery of anti-cancer drug and immune factor.