乳糖化赖氨酸与反义x基因质粒复合物体外对HBV表达的抑制

目的研制一种能把基因药物定向导入肝脏组织的小分子肝靶向药物载体。方法用还原氨化法合成乳糖化赖氨酸(Lac-Lys)共价结合物作为肝靶向药物载体。将含HBV反义x基因的质粒在一定条件下与Lac-Lys制成复合物后转染HepG2.2.15。用酶联免疫吸附试验(ELISA)检测细胞培养上清中的HBsAg和HBeAg。结果经红外光谱定性,成功制备出了Lac-Lys的共价结合物;此共价结合物能与质粒DNA形成复合物,经细胞培养观察可将质粒DNA载入细胞;抗原检测结果显示,含有反义xDNA的质粒在细胞内能有效抑制病毒的复制。xDNA与Lac-Lys的最佳比例是1∶5,最大抑制率是73.2%。结论合成的Lac-Lys可将反义x基因质粒定向载入细胞内,并有效抑制HBsAg和HBeAg的表达。

The inhibition to expression of HBV by complex of Lactosaminated L-Lysine and antisense x gene plasmid in vitro

PurposeTo develop a small molecular carrier to transfer gene drug into liver cell.MethodsThe Lactoseaminated L-Lysine(Lac-Lys) as drug delivery vector of targeting liver was synthesized by reductive amination and was conjugated with plasmid containning antisense x gene,then HepG2.2.15 were transfected. The inhibition to HBV was tested by determining HbsAg and HbeAg produced by HepG2.2.15 cells with ELISA technique.ResultsThe results of analysis for the structure of the compound prepared by re- ductive amination show that the Lac-Lys has been synthesized. This compound can form complex with plasmid. The complex, consisting of xDNA and Lac-Lys, was able to deliver xDNA into cells. The results of antigen test show that the plasmid containing xDNA can inhibit efficiently the copying of HBV. The best proportion of xDNA to Lac-Lys of this complex is 1∶5 and the most inhibition rate is 73.2%.ConclusionThe Lac-Lys can effectively deliver xDNA into HepG2.2.15 cells in vitro and strongly inhibit HBV gene expression.