Highly efficient and tumor-restricted gene transfer to malignant gliomas by replication-competent retroviral vectors
Highly efficient and tumor-restricted gene transfer to malignant gliomas by replication-competent retroviral vectors
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DOI：
10.1089/104303403321070810
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发表时间：
2003-01-01
影响因子：
4.2
通讯作者：
Chen, TC
中科院分区：
文献类型：
--
作者：
Wang, WJ;Tai, CK;Chen, TC
基因治疗的第一个大型随机III期试验表明，注射了包装细胞的患者的存活率没有改善，这些包装细胞产生了携带单纯疱疹病毒胸苷激酶基因的传统复制缺陷逆转录病毒载体，这一令人失望的结果被归因于极低的转导效率水平。为了避免这个问题，我们开发了一种改良的复制能力逆转录病毒(RCR)，它能够在体外多个感染周期内转导人胶质瘤细胞系A-172、U-87、T-98G、U-373和U-138以及大鼠胶质瘤细胞系C6和9L，从而在相同剂量下比传统的复制缺陷逆转录病毒载体的转导效率大大提高。而常规逆转录病毒载体以1.0×10~(-5)转导单位(TU)注射到预先建立的U-87皮下肿瘤中，6周时转导效率仅为0.2%，而相同剂量的RCR载体转导效率高达97.2%。将1.0×10~(4)TU的RCR载体注射到预先建立的U-87肿瘤中，2周和3周的转导效率分别为74%和98.1%。然而，值得注意的是，颅内注射RCR载体并没有导致正常脑细胞的可检测到感染。此外，使用灵敏的聚合酶链式反应分析，在任何脑外组织中都没有检测到RCR信号，包括肺、肝、肾、上胃肠道(食道和胃)、下胃肠道(结肠和小肠)、皮肤、脾和骨髓。用携带酵母胞嘧啶脱氨酶自杀基因的RCR载体治疗U-87脑胶质瘤后，再用5-氟胞嘧啶前药给药，60d内存活率为100%，而对照组仅为0%。我们的结果表明，RCR载体可以在恶性人脑胶质瘤中实现显著的治疗转导水平，并且RCR载体在瘤内注射后的传播仅限于肿瘤本身。
The first large randomized phase III trial in gene therapy demonstrated no improvement in the survival of patients injected with packaging cells that produced conventional replication-defective retroviral vectors carrying the herpes simplex virus thymidine kinase gene, a disappointing result that was attributed to extremely poor levels of transduction efficiency. To circumvent this problem, we have developed a modified replication-competent retrovirus (RCR) that is capable of transducing human glioma cell lines A-172, U-87, T-98G, U-373, and U-138 and rat glioma cell lines C6 and 9L, over multiple infection cycles in vitro, resulting in a tremendous enhancement in transduction efficiency over conventional replication-defective retroviral vectors at the same dose. Whereas the transduction efficiency of conventional retroviral vectors injected into preestablished subcutaneous U-87 tumors at a dose of 1.0x10(5) transducing units (TU) was only 0.2% at 6 weeks postinjection, the same dose of RCR vector resulted in up to 97.2% transduction. When RCR vectors at a dose of 1.0x10(4) TU were injected into preestablished intracranial U-87 tumors, transduction efficiency at 2 and 3 weeks was 74 and 98.1%, respectively. Notably, however, intracranial injection of RCR vectors did not result in detectable infection of normal brain cells. Furthermore, using a sensitive polymerase chain reaction assay, no detectable RCR signal could be observed in any extracerebral tissues, including lung, liver, kidney, upper gastrointestinal tract (esophagus and stomach), lower gastrointestinal tract (colon and small intestine), skin, spleen, and bone marrow. Treatment of U-87 intracranial gliomas with RCR vectors carrying the yeast cytosine deaminase suicide gene followed by 5-fluorocytosine prodrug administration resulted in 100% survival over a 60-day follow-up period, compared with 0% survival of control groups receiving vector alone or prodrug alone. Our results demonstrate that RCR vectors can achieve therapeutically significant levels of transduction in malignant human gliomas, and that RCR vector spread after intratumoral injection is restricted to the tumor itself.