Dept. of Medical Biotechnology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran.
Biotechnol Prog. 2010 Mar-Apr;26(2):505-11. doi: 10.1002/btpr.353.
Developing cancer gene therapy constructs based on transcriptional targeting of genes to cancer cells is a new and promising modality for treatment of cancer. Introducing truncated Bid (tBid), a recently known member of the Bcl-2 family, eradicates cancer cells efficiently. For transcriptional targeting of tBid, two dual-specificity promoters, combining cancer specific core promoters and response modules, were designed. These two core promoter modules contained cancer specific promoters of MUC1 and Survivin genes accompanied by hypoxia-responsive elements and estrogen responsive elements (microenvironment condition of breast cancer cells) which were employed to achieve a higher and more specific level of tBid expression in breast cancer cells. Correlation of the level of tBid expression in normal and cancer cell lines with promoter activity was measured by RT-PCR after treatment with hypoxia and estrogen. The level of tBid expression under control of new hybrid promoters was compared with its expression under control of cytomegalovirus (CMV) promoter as a control. Our data revealed that the level of tBid expression in breast cancer cells were nearly 11 times more than normal cells because of the cancer specific promoters, although tBid expression under control of CMV promoter was almost the same in normal and cancer cell lines. Increased apoptosis was detected in the transfected breast cancer cell lines by the Caspase-3 activity assay. The application of these promoters may prove to have the advantage of tumor selective gene therapy in breast cancer cells and low-potential toxicity for normal tissues.
基于基因转录靶向癌症细胞开发癌症基因治疗构建体是治疗癌症的一种新的有前途的方法。引入截断的 Bid（tBid），一种最近发现的 Bcl-2 家族成员，能够有效地消灭癌细胞。为了实现 tBid 的转录靶向，设计了两个双特异性启动子，结合了癌症特异性核心启动子和反应模块。这两个核心启动子模块包含了 MUC1 和 Survivin 基因的癌症特异性启动子，同时伴有缺氧反应元件和雌激素反应元件（乳腺癌细胞的微环境条件），以实现乳腺癌细胞中更高和更特异性的 tBid 表达水平。通过在缺氧和雌激素处理后进行 RT-PCR 测量正常和癌细胞系中 tBid 表达水平与启动子活性的相关性。将新的杂交启动子控制下的 tBid 表达水平与作为对照的巨细胞病毒（CMV）启动子控制下的 tBid 表达水平进行比较。我们的数据表明，由于癌症特异性启动子，乳腺癌细胞中的 tBid 表达水平几乎是正常细胞的 11 倍，尽管 CMV 启动子控制下的 tBid 表达水平在正常和癌细胞系中几乎相同。通过 Caspase-3 活性测定检测到转染的乳腺癌细胞系中凋亡增加。这些启动子的应用可能证明在乳腺癌细胞中具有肿瘤选择性基因治疗的优势，并且对正常组织的潜在毒性较低。
