半乳糖结合位点封闭式免疫毒素的制备及其特性

Preparation and characterization of immunotoxin with blocked galactose-binding sites of ricin

本文报道B链半乳糖结合位点封闭的蓖麻毒素—胃癌单克隆抗体(McAb)结合物的制备及其细胞毒特征.首先将引入碘乙酰基团的毒素与引入巯基的抗体反应,得到的结合物通过琼脂糖亲和层析柱除去仍保留半乳糖结合能力的部分.ELISA结果表明,结合物稀释至2 nmol/L水平,仍保留抗体结合活性.结合物对靶细胞在体外有较强的选择性杀伤作用,在0.1 pmol/L水平,对人胃癌细胞KATOⅢ的杀伤率为46％,优于McAb—蓖麻毒素A链结合物.结果表明,免疫毒素中B链的存在有助于增强A链的细胞杀伤作用。

In the present study,a specific cytotoxic agent was prepared by linking intact ricin to antibody,in a manner that the galactose-binding sites on the B chain of the ricin were blocked.The ricin was iodoacetyla-ted by N-hydroxy-succinimidyl ester of iodoacetic acid and then treated with thiolated McAb,a murine monoclonal antibody against gastric cancer.The fraction of the conjugate with reduced galactose-binding capacity were separated by affinity chromatography on Sepharose 4B.The binding activity of the antibody in the conjugate retained well at theconcentration of 2 nmol/L.The conjugate showed highly selective cytotoxicity on a gastric cancer cell line KATOⅢ The cytotoxic activity of McAb-ricin was much higher than that of McAb-ricin A chain,suggesting that this conjugate has both high potency,which is often lacked in antibody-A chain conjugate,and high specificity,which is lacked in intact ricin.