Brade A M, Ngo D, Szmitko P, Li P X, Liu F F, Klamut H J
Department of Medical Biophysics, University of Toronto, Ontario, Canada.
Cancer Gene Ther. 2000 Dec;7(12):1566-74. doi: 10.1038/sj.cgt.7700267.
Targeting therapeutic gene expression to tumor cells represents a major challenge for cancer gene therapy. The strong transcriptional response exhibited by heat shock genes, along with the beneficial therapeutic effects of hyperthermia have led us to develop a heat-directed gene-targeting strategy for cancer treatment. Heat shock gene expression is mediated in large part by the interaction of heat shock factor 1 with specific binding sites (heat shock elements; HSE) found in the promoters of heat-inducible genes. Here we present a quantitative analysis of heat-inducible gene expression mediated by the wild-type hsp70b gene promoter, as well as a modified hsp70b promoter containing additional HSE sequences. Beta-galactosidase (beta-gal) expression was induced between 50- and 800-fold in a panel of human breast cancer cell lines infected with an adenoviral vector containing the wild-type hsp70b promoter (Ad.70b.betag) following treatment at 43 degrees C for 30 minutes. Infection with an adenoviral vector containing the modified hsp70b promoter (Ad.HSE.70b.betag) resulted in a 200- to 950-fold increase in beta-gal expression under the same conditions, and also provided a 1-2 degrees C decrease in the threshold of activation. Significant increases in the heat responsiveness of the Ad.HSE.70b.betag construct were observed in five of six tumor cell lines tested, as well as under thermotolerant conditions. Finally, we demonstrate that localized heating of a HeLa cell xenograft can effectively target beta-gal gene expression following intratumoral injection of Ad.70b.betag. Adenoviral vectors incorporating heat-inducible therapeutic genes may provide useful adjuncts for clinical hyperthermia.
将治疗性基因表达靶向肿瘤细胞是癌症基因治疗面临的一项重大挑战。热休克基因表现出强烈的转录反应，以及热疗的有益治疗效果，促使我们开发一种用于癌症治疗的热导向基因靶向策略。热休克基因的表达在很大程度上是由热休克因子1与热诱导基因启动子中发现的特定结合位点（热休克元件；HSE）相互作用介导的。在这里，我们对野生型hsp70b基因启动子以及包含额外HSE序列的修饰hsp70b启动子介导的热诱导基因表达进行了定量分析。在一组感染了含有野生型hsp70b启动子的腺病毒载体（Ad.70b.betag）的人乳腺癌细胞系中，在43℃处理30分钟后，β-半乳糖苷酶（β-gal）表达诱导了50至800倍。用含有修饰hsp70b启动子的腺病毒载体（Ad.HSE.70b.betag）感染，在相同条件下β-gal表达增加了200至950倍，并且还使激活阈值降低了1至2℃。在测试的六个肿瘤细胞系中的五个以及在热耐受条件下，均观察到Ad.HSE.70b.betag构建体的热反应性显著增加。最后，我们证明在HeLa细胞异种移植瘤局部加热后，瘤内注射Ad.70b.betag可有效靶向β-gal基因表达。掺入热诱导治疗基因的腺病毒载体可能为临床热疗提供有用的辅助手段。
