Development of tumor-targeted therapy using measles virus-displayed antibody library

利用麻疹病毒展示抗体库开发肿瘤靶向治疗

基本信息

批准号：
22659227
负责人：
NAKAMURA Takafumi
金额：
$ 2.06万
依托单位：
The University of Tokyo
依托单位国家：
日本
项目类别：
Grant-in-Aid for Challenging Exploratory Research
财政年份：
2010
资助国家：
日本
起止时间：
2010 至 2011
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-22659227/
关键词：
実験外科学癌ウイルス抗体バイオテクノロジー生体機能利用

项目摘要

We have successfully developed a pseudoreceptor system which allows rescue and propagation of retargeted viruses displaying single chain antibody fragments(scFvs). Thus, receptor choice is not a significant limitation for targeting measles viruses and the use of single chain antibodies for targeting potentially allows us to redirect the virus against any chosen cellular receptor.In this study, we try to develop measles virus-displayed scFv library(virobody library) using the technology for identification of tumor-specific scFv. Human A549 lung, or BxPC-3 pancreatic carcinoma cells were infected with virobody library at an MOI of 0. 01 to 1, one hour later the cells were washed with Opti-MEM medium(Invitrogen) four times. 2 to 5 days after infection, propagated viruses were harvested by repeated freeze-thaw cycles from the GFP-positive infected cells. The biopanning was repeated several times, and finally the tumor-targeted measles virus was cloned from the GFP-positive infected cells. The cDNA encoding scFv from the viral RNA genome was amplified by RT-PCR for sequence analysis. However, the identified scFvs were not functional as a tumor-specific antibody. We have assumed that the diversity of measles virus-displayed scFv library is not sufficient enough to identify tumor-specific scFvs. To address this question, not traditional ligation method but Gateway[○! R] Technology-based cloning method(Invitrogen) was used for generation of plasmid library. Various kind of scFvs were incorporated into the C terminus of a receptor-blind H gene in a full-length cDNA clone of Edmonston measles virus. Furthermore, infectious scFv-displaying measles viruses were rescued from the full-length cDNA clones using helper vaccinia virus expressing T7 RNA polymerase. These modifications dramatically increased the diversity of measles virus-displayed scFv library.

我们已经成功地开发了一个伪受体系统，该系统允许拯救并宣传自动式病毒单链片段（SCFV）。通过该技术，我们尝试开发病毒引起的SCFV文库，以鉴定肿瘤特异性SCFV。。 -PCR用于序列分析，确定的SCFV是肿瘤特异性抗体的功能这个问题不是传统的连接方法对接（○！麻疹病毒此外，使用表达T7 Na聚合酶的辅助病毒病毒从全长的克隆中发出感染性SCFV脱落的麻疹。

项目成果

期刊论文数量（0）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

マイクロRNAによって制御されるウイルスの開発とその応用

microRNA控制病毒的研制及其应用

DOI：
发表时间：
2010
期刊：
影响因子：
0
作者：
Sawada Y;Sugita K;Kabashima R;Hino R;Nakamura M;Koga C;Tokura Y;高橋昭久;中村貴史
通讯作者：
中村貴史

Highly Attenuated Vaccinia Virus as a Potential Oncolytic Agent for Cancer Virotherapy

高度减毒痘苗病毒作为癌症病毒治疗的潜在溶瘤剂

DOI：
发表时间：
2010
期刊：
影响因子：
0
作者：
Fusasaki T;Narita R;Hiura M;Abe S;Tabaru A;Hino R;Matsuyama A;Shimajiri S;Tokura Y;Sasaguri Y;Harada M;Nakamura T
通讯作者：
Nakamura T

MicroRNA Targeting of Oncolytic Viruses for cancer therapy

用于癌症治疗的溶瘤病毒的 MicroRNA 靶向