Preparation Of N-succinyl-siRNA- Chitosan Nanoparticles And Research On The Activity Against Anti-tumor

Chitosan is a natural polymer and it derives from deacetylated chitin. Chitosan was applied in the study of drug carrier in recent years because of its low toxicity and good biocompatibility. However the applications of chitosans has been limited in fields of medicine and others because of the spatial structure of no rules and the huge molecular mass resulting in its poor solubility. N-succinyl-chitosan is a N-acylating derivative of chitosan and synthesis derived from chitosan and succinic anhydride. Compared to chitosan N-succinyl-chitosan has greatly improved solubility besides the vivo compatibility, non-toxic and strong adhesion as same to chitosan thus it broadens the range of applications of chitosan. So it is an excellent body medium which has open prospects for drug-loaded.N-succinyl-chitosan with DS=58% was synthesized in the HAc/EtOH system. Then we evaluated the antitumor activity of the NSC. NSC was incubated with three kinds of tumor cells:MCF-7, A549, H460. We set three kinds of transfection conditions:time, drug concentration and cells. Through MTT, we get the tumor cell growth inhibition rate, the highest inhibition rate is NSC against H460 for 72h, then we labeled NSC with FITC, incubated with tumor cells, BCA protein kit was used to quantify each cell’s protein, Fluorophotometer was used to learn the uptake amount of FITC-NSC by tumor cells. The uptake rate of NSC was present by uptake amount divided protein amount, we found that the highest uptake rate is also the NSC incubated with H460 for 72h.N-succinyl-chitosan nano was prepared by self-assembly. siRNA was loading in NSC nanoparticles by the electrostatic effection. Experimentally obtained N-succinyl-chitosan nanoparticles was measured by the particle diameter reaches 150-200nm, the surface potential measured by the potential analyzer+8～20mV, nanoparticles system is more stable, TEM analysis of the nanoparticle surface morphology can be seen from the lens photo cashiermorphological integrity of a grain of rice. Agarose gel electrophoresis and UV spectrophotometer was used to test the encapsulation efficiency,1:1 group shows a good EE about 95.28%.N-succinyl-chitosan nanoparticles loading siRNA in vitro antitumor activity was assessed and compared with liposomes loading siRNA. MTT shows that NSC-NPs-siRNA has a higher inhibition rate than NSC and Lipo-siRNA. ELISA proteins kit was used to evaluate the VEGF concentration of H460. The expression level of VEGF was appraised by Real-time PCR. The result shows that NSC combined with siRNA showed a great potential for tumor treatment, it down-regulated the expression level of VEGF and showed a nearly same transfection capability with liposomes.