Shah Khalid, Tang Yi, Breakefield Xandra, Weissleder Ralph
Center for Molecular Imaging Research, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA.
Oncogene. 2003 Oct 9;22(44):6865-72. doi: 10.1038/sj.onc.1206748.
Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has been shown to induce apoptosis in neoplastic cells. While many previous studies have been performed in cell culture, the delivery and efficiency of TRAIL variants in vivo is less well established. Using dual substrate/reporter bioluminescence imaging (Fluc: firefly luciferase-luciferin and Rluc: Renilla luciferase-coelenterazine), we tested the efficacy of TRAIL using replication-deficient herpes simplex virus (HSV) type 1 amplicon vectors in gliomas. The cDNA for complete TRAIL and the extracellular domain of TRAIL (aa 114-281) were cloned into HSV amplicons and packaged into helper virus-free vectors. Both forms of TRAIL induced similar degrees of apoptosis in human glioma cells (Gli36) in culture within 24 h of infection with TRAIL amplicon vectors. Growth of tumors stably transfected with Fluc (Gli36fluc+) was readily monitored in vivo by bioluminescence imaging following luciferin administration. HSV amplicon vectors bearing the genes for TRAIL and Rluc injected directly into Gli36fluc(+)-expressing subcutaneous gliomas revealed peak Rluc activity 36 h after intratumoral injection as determined by coelenterazine injection followed by imaging. TRAIL-treated gliomas regressed in size over a period of 4 weeks as compared to the mock-injected gliomas. These results show the efficacy of vector delivered TRAIL in treating tumors in vivo and offer a unique way to monitor both gene delivery and efficacy of TRAIL-induced apoptosis in tumors in vivo in real time by dual enzyme substrate (Rluc/Fluc) imaging.
肿瘤坏死因子相关凋亡诱导配体（TRAIL）已被证明可诱导肿瘤细胞凋亡。虽然之前许多研究是在细胞培养中进行的，但TRAIL变体在体内的递送和效率尚未得到充分证实。我们使用双底物/报告基因生物发光成像（Fluc：萤火虫荧光素酶 - 荧光素和Rluc：海肾荧光素酶 - 腔肠素），测试了使用复制缺陷型单纯疱疹病毒1型（HSV）扩增载体递送TRAIL在胶质瘤中的疗效。将完整TRAIL的cDNA和TRAIL的胞外结构域（氨基酸114 - 281）克隆到HSV扩增子中，并包装成无辅助病毒的载体。在感染TRAIL扩增子载体24小时内，两种形式的TRAIL在培养的人胶质瘤细胞（Gli36）中诱导了相似程度的凋亡。在用荧光素给药后，通过生物发光成像可在体内轻松监测稳定转染Fluc的肿瘤（Gli36fluc +）的生长。将携带TRAIL和Rluc基因的HSV扩增子载体直接注射到表达Gli36fluc（+）的皮下胶质瘤中，通过注射腔肠素后成像确定，瘤内注射后36小时Rluc活性达到峰值。与模拟注射的胶质瘤相比，经TRAIL治疗的胶质瘤在4周内体积缩小。这些结果表明载体递送的TRAIL在体内治疗肿瘤的疗效，并提供了一种独特的方法，通过双酶底物（Rluc/Fluc）成像实时监测体内肿瘤中基因递送和TRAIL诱导凋亡的疗效。
