单链抗体与力达霉素基因工程强化融合蛋白Fv-LDP-AE的抗肿瘤作用

高瑞娟 李亮 赵春燕 王玉凤 李电东 甄永苏

中国药学杂志 ›› 2010, Vol. 45 ›› Issue (11) : 808-813.

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中国药学杂志 ›› 2010, Vol. 45 ›› Issue (11) : 808-813.
论著

单链抗体与力达霉素基因工程强化融合蛋白Fv-LDP-AE的抗肿瘤作用

  • 高瑞娟,李亮,赵春燕,王玉凤,李电东,甄永苏*
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Antitumor Effect of the Engineered and Energized Fusion Protein Fv-LDP-AE Composed of Lidamycin and Anti-type Ⅳ Collagenase Single-chain Fv Antibody

  • GAO Rui-juan, LI Liang, ZHAO Chun-yan, WANG Yu-feng, LI Dian-dong, ZHEN Yong-su*
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摘要

目的 研究抗Ⅳ型胶原酶单链抗体(Fv)和力达霉素辅基蛋白(LDP)在大肠杆菌表达的融合蛋白(Fv-LDP)与力达霉素(LDM)的活性发色团AE形成的基因工程强化融合蛋白Fv-LDP-AE的生物学活性及体外抗肿瘤作用。方法 采用酶联免疫吸附剂测定、免疫荧光细胞化学染色和明胶酶谱分析Fv-LDP-AE生物学活性;通过MTT法、Hoechst 33342和碘化丙啶共染、DNA ladder和侵袭实验探讨其抗肿瘤作用。结果 Fv-LDP-AE对人肝癌BEL-7402和人结肠癌HT-29细胞的免疫反应呈阳性,可与BEL-7402细胞特异结合,并呈剂量依赖性抑制人高转移性巨细胞肺癌PG细胞内Ⅳ型胶原酶分泌。Fv-LDP-AE对BEL-7402和PG细胞的增殖抑制作用比力达霉素强,可诱导BEL-7402细胞裂亡、凋亡,抑制BEL-7402细胞侵袭。结论 Fv-LDP-AE是一种以Ⅳ型胶原酶为靶点的、具有高效抗肿瘤活性的抗体药物。

Abstract

OBJECTIVE To study the biological activity and in vitro antitumor effect of the engineered and energized fusion protein Fv-LDP-AE composed of the fusion protein Fv-LDP and the active enediyne (AE) of lidamycin (LDM). METHODS Enzyme-linked immunosorbent assay, immunofluorescent cytochemical staining and gelatin-zymography were used to analyze the biological activity of Fv-LDP-AE. The antitumor effects of Fv-LDP-AE were evaluated by MTT method, co-staining of Hoechst 33342 and propidium, DNA ladder and Boyden Chamber assay. RESULTS Fv-LDP-AE showed positive immunoreactivity against, human hepatoma BEL-7402 cells and human colon cancer HT-29 cells, respectively. It specifically bound to BEL-7402 cells and inhibited the secretion of type Ⅳ collagenase in human highly metastatic lung carcinoma PG cells in a dose-dependent manner. Fv-LDP-AE also showed higher growth inhibition toward BEL-7402 and PG cells than LDM. Furthermore,Fv-LDP-AE induced mitotic cell death, apoptosis and blocked the invasion of BEL-7402 cells. CONCLUSION Fv-LDP-AE is a highly potent antitumor antibody-based drug directed against type Ⅳcollagenase.

关键词

Ⅳ型胶原酶 / 单链抗体 / 力达霉素 / 强化融合蛋白 / 生物学活性 / 抗肿瘤作用

Key words

type Ⅳ collagenase / single-chain Fv antibody / lidamycin / energized fusion protein / biological activity / antitumor effect

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高瑞娟 李亮 赵春燕 王玉凤 李电东 甄永苏. 单链抗体与力达霉素基因工程强化融合蛋白Fv-LDP-AE的抗肿瘤作用[J]. 中国药学杂志, 2010, 45(11): 808-813
GO Rui-jun;LI Ling;ZHO Chun-yn;WNG Yu-feng;LI Din-dong;ZHEN Yong-su. Antitumor Effect of the Engineered and Energized Fusion Protein Fv-LDP-AE Composed of Lidamycin and Anti-type Ⅳ Collagenase Single-chain Fv Antibody[J]. Chinese Pharmaceutical Journal, 2010, 45(11): 808-813

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