Co-delivery Of Doxorubicin And MDR1-siRNA By Mesoporous Silica Nanoparticles Treats Resistant Oral Squamous Carcinoma

Objective: To investigate the effect of mesoporous silica nanoparticles (MSNP)co-delivery system on reversing multi-drug resistance and killing tumor cells,and toconfirm that combined application of DOX and MDR1-siRNA can exert a synergisticantitumor effect.Methods: Firstly, mesoporous silica nanoparticles were prepared by sol-gelmethod in room temperature and coated by cationic polymerpolyethylenimine(PEI)on the surface to stay positively charged, which could facilitate its combination withnegatively charged MDR1-siRNA in the next step.With the obsorption of DOX, theantitumor drug, at the same time, the drug and gene co-delivery system then formed.In vitro, drug loading was evaluated by UV-Vis spectrophotometer; antitumor drugswere co-cultured respectively with oral squamous cells carcinoma cells lines KB andresistant cell lines KB/VCR. Then MTT assay was conducted to measure the halfmaximal inhibitory concentration (IC50); Fluorescence microscope and flowcytometry instrument were used to measue transfection efficiency of the co-deliverysysterm; Real-Time PCR technique was applied to detect the expression level ofMDR1gene; AnnexinV-FITC/7-AAD double staining by flow cytometry instrumentwas conduceted to detect cell apoptosis. Western Blot was conducted to assess theexpression of apoptosis-and autophagy-related protein. In vivo, we fabricated atumor-bearing nude mice model to further evaluate the curative effect of MSNPco-delivery system multi-drug resistant oral squamous carcinoma.Results: The mesoporous silica nanoparticles appeared to have a highdispersibility and homogeneous size by particle size analyzer and transmissionelectron microscopy (TEM). The formed nanoparticles had a diameter distribution of100-200nm, and the surface mesoporous diameter was3-5nm. We measured IC50(KB-DOX) was182.9ng/ml, and IC50(KBV-DOX) was9233.5ng/ml by ultravioletspectrophotometer method. The resistance index of KBV was50.483871. The transfection efficiency was6.73%measured by fluorescence microscope and flowcytometry. The Real-Time PCR test indicated that MDR-1expression in theco-delivery systerm decreased dramatically, compared with drug-loaded nanoparticlesand gene-loaded nanoparticles. The results of AnnexinV-FITC/7-AAD double stainingshowed that the cell apoptosis rates were12.74%and10.08%for drug-loaded andgene-loaded nanoparticles, respectively, while the cell apoptosis rate increased to25.68%for co-delivery nanoparticles. The result of in vivo experiment shows thattumor-inhibitory rates of the drug-loaded nanoparticles and co-delivery systerm were58.67%and81.64%. Western Blot demonstrated an conspicuous up-regulatedexpression of apoptosis-related protein Beclin1and LC3and a decreased expressionof P62in co-delivery group, indicating the occurrence of apoptosis.Conclusions: The results demonstrated that comparing with the pure medicine orgenome, the mesoporous silica nanoparticles co-delivery system can effectivelyreverse multi-drug resistance of tumor and inhibited the growth dramatically. At thesame time, DOX and the MDR1-siRNA can exert synergistic antitumor effect whenapplied in combination.And in the process of the treating resistant oral squamouscarcinoma via co-delivery system, autophagy may play an important role.