出版年:

2001

研究生:

黃彥淞

研究生(英文姓名):

Yen Sung Huang

論文名稱:

利用腫瘤或組織特異性啟動子構築E1b缺失腺病毒進行癌症基因治療

英文論文名稱:

Construction of E1b-deleted adenoviruses driven by tumor- or tissue-specific promoters for cancer gene therapy

指導教授:

蕭璦莉

指導教授(英文姓名):

Ai Li Shiau

學位類別:

碩士

校院名稱:

國立成功大學　

系所名稱:

微生物暨免疫學研究所

學號:

S46881068

學年度:

89

語文別:

中文

論文頁數:

62

關鍵詞:

E1b缺失腺病毒 ; 腫瘤裂解 ; 啟動子 ; 基因治療

英文關鍵詞:

E1b- deleted adenovirus ; oncolytic ;
promoter ; gene therapy

被引用次數:

0

[ 摘要 ]

有複製能力的E1b缺失腺病毒可以選擇性地殺死缺乏有功能p53的腫瘤細胞，這種病毒被認為可用來做癌症的基因治療，然而，有些報導指出此種病毒並不絕對在p53缺失的細胞造成細胞病變，因此在本研究中我們試圖利用不同組織或腫瘤專一啟動子去調控病毒E1a基因，試圖改善病毒複製的專一性。E1a是病毒的立即早期基因對病毒複製是很重要的，不同組織或腫瘤專一啟動子，例如:人類transthyretin啟動子，老鼠POU轉錄因子啟動子和HER-2/neu啟動子，構築不同啟動子調控E1a的E1b缺失腺病毒。我們利用AdEasy-1系統在E. Coli中產生重組腺病毒，並利用293細胞進行病毒的放大及定量，我們初步結果顯示人類啟動子調控的E1b缺失腺病毒可以專一地在p53突變的肝細胞中複製並造成細胞病變，但較少在p53正常的肝細胞中造成細胞病變;另外HER-2/neu啟動子調控的E1b缺失腺病毒可以在J82和MBT-2具有p53突變的膀胱細胞中複製，並造成細胞病變;在動物實驗中也見到此病毒有效地阻礙MBT-2腫瘤細胞的生長。另外mOCT4啟動子調控的E1b缺失腺病毒則可專一地在具有p53突變且OCT4過度表現的MBT-2細胞中複製並造成細胞病變。透過以上種種實驗讓我們了解利用啟動子控制E1b缺失腺病毒可以有效提升其複製之專一性。

[ 英文摘要 ]

Replication-competent, E1b-deleted adenoviruses, which can selectively kill tumor cells lacking functional p53, have shown promise for cancer gene therapy. However, some reports indicate that the cytopathic effect of the virus is not absolutely related to mutated p53. In this study, we aimed to improve the specificity of viral replication in tumor cells by using tumor- or tissue-specific promoters to drive E1a gene, the immediate early gene required for adenoviral replication. Various tumor- and tissue-specific promoters, including human transthyretin promoter (hTTRp), mouse POU transcription factor promoter (mOCT4p) and HER-2/neu promoter (hHER2p) were exploited to construct E1b-deleted adenoviruses with the E1a gene under the control of these promoters. The AdEasy-1 system was used to construct these recombinant adenoviruses in E. coli. Recombinant adenoviruses were amplified and titrated in 293 cells. Our results showed that the hTTRp-driven E1b-deleted adenoviruses could replicate in liver cells with p53 mutation, resulting in cytopathic effects, whereas the virus was attenuated in liver cells with wild-type p53. The hHER2p-driven E1b-deleted adenoviruses could replicate in J82 and MBT-2 cells with p53 mutation and HER-2/neu over-expression. In vivo studies demonstrated that hHER2p-driven E1b-deleted adenoviruses could retard the growth of MBT-2 cells. Furthermore mOCT4p-driven E1b-deleted adenovirus could replicate in MBT-2 cells with p53 mutation and OCT4 over-expression. From these observations, application of promoter driven E1b-deleted adenovirus can improve the specificity of viral replication.