抗乙型肝炎反义核酸药物的筛选及细胞毒性研究

Study on Screening Anti-HBV Agents from Antisense Oligodeoxynucleotide and Their Toxicity

为了筛选抗乙型肝炎（HBV）反义寡核苷酸药物，根据HBV基因组序列择S基因起始区、SPⅡ帽子区、前核心基因起始区、核心起始区、包装信号起始区等5个基因靶位点，设计并合成硫代反义寡核苷酸（S－asODN），以HePG22215细胞为研究对象，利用ELIAS法、MTT比色法、电子显微镜观察等手段，观察上述s－asODN对HePG22215细胞HBsAg和HBeAg表达以及对细胞的杀伤作用和细胞形态、超微结构的影响，结果显示s－asODN能显著抑制HBsAg和HBeAg的表达，s－asODN抗HBV具有序列特异性、剂量相关性、抗核酸酶性、联合用药协同性。

To screen a new anti-HBV agent, according to seqence of HBV genome, 5 complementary sites were selected and the antisense phosphorothioate oligodeoxynucleotides (s-asODN) were designed and synthesized, they are S gene initiator, SP Ⅱ cap site, Pre-C gene initiator, C gene initiator and gene ε initiator. The complementarity to different HBV genome were assayed for their anti-HBV activity in HepG22215 cell line with ELISA method, and the toxicity of s-asODN also was studied with MTT colorimetric assay and electron microscope. The result shows that s-asODN can inhibit the expressionof HBsAg and HBeAg, and the inhibition shows sequence- specific, dose- related,antinuclease and combination - cooperative, and harmless to host characteristic in the viability, the shape and ultrastucture of cells. The study had laid the foundation of exploring new anti-HBV agents.