Vincent A J, Esandi M C, Avezaat C J, Vecht C J, Sillevis Smitt P, van Bekkum D W, Valerio D, Hoogerbrugge P M, Bout A
Department of Neurosurgery, University Hospital Rotterdam, The Netherlands.
Neurosurgery. 1997 Aug;41(2):442-51; discussion 451-2. doi: 10.1097/00006123-199708000-00023.
Adenoviral gene transfer and killing efficiency using the thymidine kinase (TK)/ganciclovir (GCV) mechanism was evaluated in human cancer cells occurring as central nervous system tumors. The effectiveness of this approach was tested in vitro and in experimental models for brain tumor and leptomeningeal metastases in rats in vivo. Recombinant adenoviruses with different promoters were compared.
Adenoviral vectors harboring a marker (lacZ) or a TK gene were constructed. Transcription of genes was under the control of either the adenovirus Type 2 major late promoter (MLP) or the human cytomegalovirus (CMV) immediate early gene promoter. lacZ expression and GCV killing efficiency after TK gene transfer in several human tumor cells was evaluated in vitro. The 9L rat brain tumor and leptomeningeal metastases models were used to determine the effectiveness of adeno-TK and subsequent GCV treatment in vivo. MLP and CMV containing adenoviral vectors were compared.
Gene expression and the killing of tumor cells were very efficient in all human tumor cell lines tested. The adenovirus containing the CMV promoter showed cytopathic effects in cultured tumor cells at high multiplicity of infections but also greater cell killing efficiency after TK/GCV treatment, as compared to the MLP promoter. Although both the MLP and CMV vectors showed a significant dose-dependent therapeutic effect, animals treated with recombinant adenovirus containing the CMV promoter showed significantly longer survival time (brain tumors) or symptom-free periods (leptomeningeal metastases).
This study demonstrates the therapeutic efficiency and feasibility of the TK/GCV approach in experimental brain tumors and leptomeningeal metastases. It also demonstrates that the promoter driving the transgene in an adenoviral vector influences toxicity and efficiency of treatment.
利用胸苷激酶（TK）/更昔洛韦（GCV）机制评估腺病毒基因转移和杀伤效率，该评估在发生于中枢神经系统肿瘤的人类癌细胞中进行。在体外以及大鼠脑肿瘤和软脑膜转移瘤的体内实验模型中测试了这种方法的有效性。对具有不同启动子的重组腺病毒进行了比较。
构建携带标记基因（lacZ）或TK基因的腺病毒载体。基因转录受腺病毒2型主要晚期启动子（MLP）或人巨细胞病毒（CMV）立即早期基因启动子的控制。在体外评估了几种人类肿瘤细胞中TK基因转移后的lacZ表达和GCV杀伤效率。使用9L大鼠脑肿瘤和软脑膜转移瘤模型来确定腺病毒-TK及随后的GCV治疗在体内的有效性。比较了含有MLP和CMV的腺病毒载体。
在所有测试的人类肿瘤细胞系中，基因表达和肿瘤细胞杀伤都非常有效。与MLP启动子相比，含有CMV启动子的腺病毒在高感染复数时在培养的肿瘤细胞中显示出细胞病变效应，并且在TK/GCV处理后细胞杀伤效率更高。尽管MLP和CMV载体均显示出显著的剂量依赖性治疗效果，但用含有CMV启动子的重组腺病毒治疗的动物存活时间显著延长（脑肿瘤）或无症状期延长（软脑膜转移）。
本研究证明了TK/GCV方法在实验性脑肿瘤和软脑膜转移瘤中的治疗效率和可行性。还证明了腺病毒载体中驱动转基因的启动子会影响治疗的毒性和效率。
