目的： 观察抗肝癌重组单链免疫毒素hdsFv-RC-RNase对荷人肝癌裸鼠移植瘤的导向治疗作用,并探讨其临床应用价值。方法： 将原核表达载体TIG-hdsFv-RC-RNase转化E.coli BL21（DE3）plys中，利用IPTG大量诱导表达抗肝癌hdsFv-RC-RNase重组单链免疫毒素。表达产物在非变性条件下经Ni-NTA Agarose亲合层析法纯化并体外复性，利用细胞ELISA法检测其特异性结合体外培养的人肝癌细胞的免疫活性。建立荷人肝癌裸鼠移植瘤动物模型，初步评估其对荷人肝癌裸鼠移植瘤的导向治疗作用。结果： 工程菌经IPTG诱导后，出现一条相对分子质量约为41 000的新生蛋白带，且主要以可溶性形式表达。纯化后表达产物的纯度达到基本均一。细胞ELISA检测结果显示，纯化产物复性后能够与体外培养的人肝癌细胞特异性结合，而对正常肝细胞的结合能力非常弱，两者具有非常显著的差异﹙P＜0.01）。导向治疗结果表明，其对荷人肝癌裸鼠移植瘤治疗有效率为100%，与生理盐水组相比具有非常显著的差异（P＜0.01），抑瘤率达到79.38%。结论： 成功获得了特异性强的有活性的抗肝癌重组单链免疫毒素hdsFv-RC-RNase，其对荷人肝癌裸鼠移植瘤具有一定的导向治疗作用，可能成为抗肝癌导向治疗药物。

Objective: To observe the targeting therapy of the hdsFv-RC-RNase recombinant single chain immunotoxin on the xenograft of the human hepatocellular carcinoma in nude mice and to explore its clinical potentiality. Methods: The prokaryotic expression vector TIG-hdsFv-RC-RNase was transformed into E.coli BL21（DE3）plys to largely express recombinant single chain immunotoxin hdsFv-RC-RNase against hepatocellular carcinoma induced by IPTG. The expressed product was purified via Ni-NTA affinity chromatography under native conditions and mildly refolded. The ELISA was used to analyze its immunological activity of antigen-binding capability. The xenogrft model of the human hepatocellular carcinoma in nude mice was established and the targeting therapy of hdsFv-RC-RNase was evaluated. Results:After induced by IPTG, a new protein band with Mr 41 000 was found in the supernatant of the bacteria and expressed in a soluble form. The expressed product was purified to homogeneity via Ni-NTA affinity chromatography under native conditions. The results of ELISA showed the refolded hdsFv-RC-RNase had the specific antigen-binding capability to the human hepatocellular carcinoma cell, but not to the normal hepatocyte (P＜0.01). The targeting therapy on the xenograft in nude mice indicated that the efficiency of the hdsFv-RC-RNase was 100%(P＜0.01). The tumor inhibition rate reached 79.38%. Conclusion: The recombinant immunotoxin hdsFv-RC-RNase has been obtained sussefully, which has high activity and targeting therapy on hepatocellular carcinoma xinograft in nude mice. It may be used as targeting drug in therapy of hepatocellular carcinoma.

国家自然科学基金资助项目(No.30370810)