肿瘤特异性自杀基因治疗肝癌由转录有针对性的逆转录病毒

StephenW

Original Article

Gene Therapy (2015) 22, 155–162; doi:10.1038/gt.2014.98; published online 30 October 2014
Tumor-specific suicide gene therapy for hepatocellular carcinoma by transcriptionally targeted retroviral replicating vectors

Y-H Lai1, C-C Lin1, S-H Chen1 and C-K Tai1

1Department of Life Science and Institutes of Molecular Biology and Biomedical Science, National Chung Cheng University, Min-Hsiung, Chia-Yi, Taiwan

Correspondence: Dr C-K Tai, Department of Life Science, National Chung Cheng University, 168 University Road, Min-Hsiung, Chia-Yi 621, Taiwan. E-mail: [email protected]

Received 6 April 2014; Revised 12 September 2014; Accepted 17 September 2014
Advance online publication 30 October 2014

Abstract

Replicating virus vectors are attractive tools for anticancer gene therapy, but the potential for adverse events due to uncontrolled spread of the vectors has been a major concern. To design a tumor-specific retroviral replicating vector (RRV), we replaced the U3 region of the RRV ACE-GFP with a regulatory sequence consisting of the hepatitis B virus enhancer II (EII) and human α-fetoprotein (AFP) core promoter to produce ACE-GFP-EIIAFP, a hepatocellular carcinoma (HCC)-targeting RRV. Similar to ACE-GFP, ACE-GFP-EIIAFP exhibited robust green fluorescent protein (GFP) expression in HCC cells and, most importantly, it exhibited HCC-specific replication and did not replicate in non-HCC tumor cells or normal liver cells. We sequenced the promoter region of ACE-GFP-EIIAFP collected from serial infection cycles to examine the genomic stability of the vector during its replicative spread, and found that the vector could retain the hybrid promoter in the genome for at least six infection cycles. In vitro studies revealed that ACE-CD-EIIAFP and ACE-PNP-EIIAFP, which express the yeast cytosine deaminase and Escherichia coli purine nucleoside phosphorylase, respectively, exert a highly potent cytotoxic effect on HCC cells in the presence of their respective prodrugs. In vivo, ACE-CD-EIIAFP-mediated suicide gene therapy efficiently suppressed HCC tumor growth and no detectable RRV signal was observed in extratumoral tissues. These results suggest that the tumor-specific, suicide-gene-encoding RRV may fulfill the promise of retroviral gene therapy for cancer.

StephenW

原来的文章

基因治疗（2015）22，155-162; DOI：10.1038/ gt.2014.98;网上公布的30月到2014年
肿瘤特异性自杀基因治疗肝癌由转录有针对性的逆转录病毒的复制载体

Ÿ-H Lai1，C-C林1，S-H臣1和C-K Tai1

生命科学与分子生物学研究所和生物医学科学，国立中正大学，民雄，嘉义，台湾教研室

函授：博士CK大，生命科学，国立中正大学，大学路168号，民雄，嘉义621，台湾系。电子邮件：[email protected]

收到2014年4月6日;修订后的2014年9月12日;接受2014年9月17日
推进网上公布二〇一四年十月三十〇日

摘要

复制型病毒载体是抗癌基因治疗有吸引力的工具，但由于载体的无节制扩散不良事件的可能性一直是一个大问题。设计一种肿瘤特异性的逆转录病毒复制的载体（RRV），我们取代RRV的ACE-GFP的U3区域与调节序列选自乙肝病毒增强子II（EII）和人α甲胎蛋白（AFP）的核心启动子，以产生的ACE-GFP-EIIAFP，肝细胞癌（HCC）-targeting RRV。类似于ACE-GFP，ACE-GFP-EIIAFP展出鲁棒绿色荧光蛋白（GFP）的表达在肝癌细胞中，而最重要的是，它显示出肝癌特异性复制和非肝癌肿瘤细胞或正常肝细胞不复制。我们测序的ACE-GFP-EIIAFP从串行感染周期收集在其复制传播来检查载体的基因组稳定性的启动子区域，并发现该载体能保持杂合启动子在基因组中至少6个感染循环。体外研究揭示的ACE-CD-EIIAFP和ACE-PNP-EIIAFP，其表达的酵母胞嘧啶脱氨酶和大肠杆菌嘌呤核苷磷酸化酶，分别施加在各自的前体药物的存在下对肝癌细胞的高度有效的细胞毒作用。在体内，ACE-CD-EIIAFP介导的自杀基因疗法有效地抑制肝癌肿瘤生长和没有可检测的RRV信号中观察到extratumoral组织。这些结果表明，肿瘤特异性，自杀基因编码的RRV可以实现逆转录病毒基因治疗癌症的承诺。