Redox Sensitive Of Small Molecule Prodrug Combined With Lapachol For Tumor Treatment

It is one of the main methods of tumor treatment that chemotherapeutic drugs act on different stages of cell proliferation,thereby inhibiting or killing tumor cells.However,due to the lack of specific accumulation at the tumor site and poor permeability,chemotherapy drugs have serious systemic toxicity.In this study,PTX-AZT/LAP/AS1411/DSPE-PEG(PZ/LA)nanoparticles were prepared by synthesizing redox-sensitive small molecule prodrug Paclitaxel-Zidovudine PTX-AZT(PZ)connect to AS1411 aptamer and encapsulating LAP.The experiments and results of this study mainly includes five parts:1.Synthesis and characterization of small molecule prodrugsWith cystamine as the linker,carboxylated paclitaxel PTX-SA(PS)and carboxylated zidovudine AZT-SA(ZS)were connected to the two ends through esterification and amide reactions,and the yields of PS and ZS were 92%and 90%respectively.The successful synthesis of the above substances was verified by using 1H-NMR,Fourier transform infrared spectroscopy(FT-IR),mass spectrometry(MS)and high performance liquid chromatography(HPLC)2.Preparation and physical and chemical properties of PTX-AZT/DSPE-PEG(PZ/DP),LAP/DSPE-PEG(LAP/DP),PTX-AZT/LAP/DSPE-PEG(PZ/LAP),PTX-AZT/LAP/AS1411/DSPE-PEG(PZ/LA)nanoparticlesDifferent nanoparticles were prepared by the nanoprecipitation method,and the formula screening of the molar ratio of PTX and LAP was carried out.The drug loading efficiency of LAP in LAP/DP nanoparticle was only 10.6%.However,when coencapulation of PTX and LAP,due to the π-π interaction LAP has a higher drug loading efficiency of 30.78%under the molar ratio of PTX and LAP is 1:1.The particle size is 107.9±0.92 nm,the PDI is 0.184±0.003,the distribution is uniform and the shape is spherical3.Connect to AS1411 adapterAptamers are receptor that act like antibodies.AS 1411 aptamer can specifically recognize the nucleolar protein which is highly expressed in some tumor cells,so as to achieve the purpose of targeted therapy.Based on the principle of base complementary pairing,AS 1411 is connected to a nucleoside drug AZT through hydrogen bonds in A-T base mode.The particle size of the PZ/LA nanoparticles is 153.4±4.18 nm,the PDI is 0.156±0.05,and the CAC value is 0.0104 mg/mL.TEM results show that there is a thin fin out of the nanoparticle.In addition,the in vitro release behavior of nanoparticles was investigated.Under the condition of 10 mM DTT,both PTX and LAP are released faster,showing redox sensitivity.4.Evaluation of the anti-tumor effect of PZ/LA nanoparticles in vitroHuman breast cancer cells(MCF-7 cells)and melanoma cells(B16 cells)were used as models.The cytotoxicity of free drugs and different nanoparticles was investigated by MTT assay,the CI values of PZ and LAP in MCF-7 cells and B16 cells are 0.53 and 0.28.Furthermore,the cytotoxicity of different volume ratio of PZ and LAP show that PZ and LAP has synergistic therapeutic effect.Compare to free C6 and PZ/LAP nanoparticle,because AS1411 can increase the accumulation of nanoparticles in tumor sites PZ/LA has stronger cell uptake ability.When add different concentration of NQO1 coenzyme inhibitor dicumarol,the ROS produce capacity was weaken in varying degrees.In addition,the ability of PTX,PZ and PZ/LA to induce cell apoptosis were investigated by AnnexinV-FITC/PI kit.Finally,the cell cycle arrest test results show that PTX,PZ,PZ/LA can block cells in the G2/M phase to varying degrees.5.Evaluation of the anti-tumor effect of PZ/LA nanoparticles in vivoIn tumor-bearing mouse model established by B16 cells,a certain amount of normal saline(NS),Taxol,PZ/LAP and PZ/LA were injected into the tail vein every two days.After 14 days of administration,major organs and tumor tissues were dissected and used for histopathological section staining analysis.Finally,using the dialkyl carbocyanine dye DiR as the fluorescent substance,the distribution of free DiR and PZ/LA nanoparticles in the mouse body was observed in real time with in vivo imaging.The results of qualitative and quantitative analysis showed that compared with free DiR,PZ/LA nanoparticles accumulated more at the tumor site.The above results indicate that PZ/LA nanoparticles have a good anti-tumor effect in vivo.