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。
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.
出处
《中国药学杂志》
CAS
CSCD
北大核心
2010年第11期808-813,共6页
Chinese Pharmaceutical Journal
基金
<重大新药创制>国家科技重大专项课题基金(2009ZX09103-136)
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
type Ⅳ collagenase
single-chain Fv antibody
lidamycin
energized fusion protein
biological activity
antitumor effect
