Pützer B M, Stiewe T, Crespo F, Esche H
Institute of Molecular Biology (Cancer Research), University of Essen, Medical School, Germany.
Gene Ther. 2000 Aug;7(15):1317-25. doi: 10.1038/sj.gt.3301235.
The transfer of pro-apoptotic genes to tumors is one of the most promising strategies for anticancer gene therapy. However, the use of potentially toxic genes, such as tumor suppressor genes or apoptotic genes, needs controllable transgene activation. To achieve regulation of the transgene at a desired time, we developed an adenovirus (Ad) vector, in which the apoptotic activity of the target gene has been made 4-OHT-dependent by fusion to the ligand binding-domain of the estrogen receptor (ER). For evaluation of the system in human tumor cells, we used the E2F1 gene which encodes a transcription factor that triggers massive apoptosis in several human cancers. AdER-E2F1 expressed high levels of transgene over at least 1 week. Upon activation of E2F1 by the ligand 4-hydroxy-tamoxifen (4-OHT) the ER-E2F1 fusion protein correctly translocated from the cytosol to the nucleus, transactivated E2F-dependent promoters, and rapidly induced substantial E2F1-related toxicity. Finally, experiments in nude mice showed tightly regulated tumor growth suppression in vivo. Taken together, our system represents a powerful approach for tight regulation and rapid induction of cytotoxicity as the major criteria for safe gene delivery.
将促凋亡基因导入肿瘤是抗癌基因治疗中最具前景的策略之一。然而，使用潜在有毒基因，如肿瘤抑制基因或凋亡基因，需要可控的转基因激活。为了在期望的时间实现对转基因的调控，我们开发了一种腺病毒（Ad）载体，其中通过与雌激素受体（ER）的配体结合域融合，使靶基因的凋亡活性依赖于4-羟基他莫昔芬（4-OHT）。为了在人肿瘤细胞中评估该系统，我们使用了E2F1基因，它编码一种转录因子，可在多种人类癌症中引发大量细胞凋亡。AdER-E2F1在至少1周内高水平表达转基因。在用配体4-羟基他莫昔芬（4-OHT）激活E2F1后，ER-E2F1融合蛋白正确地从细胞质转运到细胞核，反式激活E2F依赖的启动子，并迅速诱导大量与E2F1相关的毒性。最后，裸鼠实验表明体内肿瘤生长抑制受到严格调控。综上所述，我们的系统代表了一种强大的方法，可实现严格调控和快速诱导细胞毒性，这是安全基因递送的主要标准。
