Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 100052, China;
Beijing FivePlus Molecular Medicine Institute, Beijing 100176, China;
Chin J Cancer Res. 2013 Dec;25(6):646-55. doi: 10.3978/j.issn.1000-9604.2013.11.07.
Intratumoral administration of adenoviral vector encoding herpes simplex virus (HSV) thymidine kinase (TK) gene (Ad-TK) followed by systemic ganciclovir (GCV) is an effective approach in treating experimental hepatocellular carcinoma (HCC). However, hepatotoxicity due to unwanted vector spread and suicide gene expression limited the application of this therapy. miR-122 is an abundant, liver-specific microRNA whose expression is decreased in human primary HCC and HCC-derived cell lines. These different expression profiles provide an opportunity to induce tumor-specific gene expression by miR-122 regulation.
By inserting miR-122 target sequences (miR-122T) in the 3' untranslated region (UTR) of TK gene, we constructed adenovirus (Ad) vectors expressing miR-122-regulated TK (Ad-TK-122T) and report genes. After intratumoral administration of Ad vectors into an orthotopic miR-122-deficient HCC mouse model, we observed the miR-122-regulated transgene expression and assessed the antitumor activity and safety of Ad-TK-122T.
Insertion of miR-122T specifically down-regulated transgene expression in vitro and selectively protected the miR-122-positive cells from killing by TK/GCV treatment. Insertion of miR-122T led to significant reduction of tansgene expression in the liver without inhibition of its expression in tumors in vivo, resulting in an 11-fold improvement of tumor-specific transgene expression. Intratumoral injection of Ad vectors mediated TK/GCV system led to a vector dosage-dependent regression of tumor. The insertion of miR-122T does not influence the antitumor effects of suicide gene therapy. Whereas mice administrated with Ad-TK showed severe lethal hepatotoxicity at the effective therapeutic dose, no liver damage was found in Ad-TK-122T group.
miR-122-regulated TK expression achieved effective anti-tumor effects and increased the safety of intratumoral delivery of adenovirus-mediated TK/GCV gene therapy for miR-122-deficient HCC.
瘤内给予携带单纯疱疹病毒胸苷激酶（HSV-TK）基因的腺病毒载体（Ad-TK），然后全身给予更昔洛韦（GCV），这是治疗实验性肝细胞癌（HCC）的有效方法。然而，由于载体的意外传播和自杀基因表达引起的肝毒性限制了该疗法的应用。miR-122 是一种丰富的、肝脏特异性的 microRNA，其在人原发性 HCC 和 HCC 衍生细胞系中的表达降低。这些不同的表达谱为通过 miR-122 调节诱导肿瘤特异性基因表达提供了机会。
通过在 TK 基因的 3'非翻译区（UTR）插入 miR-122 靶序列（miR-122T），我们构建了表达 miR-122 调控的 TK（Ad-TK-122T）和报告基因的腺病毒（Ad）载体。在 miR-122 缺陷的原位 HCC 小鼠模型中瘤内给予 Ad 载体后，我们观察了 miR-122 调控的转基因表达，并评估了 Ad-TK-122T 的抗肿瘤活性和安全性。
miR-122T 的插入特异性地下调了体外转基因的表达，并选择性地保护了 miR-122 阳性细胞免受 TK/GCV 治疗的杀伤。miR-122T 的插入导致体内肝脏中转基因表达的显著减少，而不抑制其在肿瘤中的表达，导致肿瘤特异性转基因表达提高了 11 倍。瘤内注射 Ad 载体介导的 TK/GCV 系统导致肿瘤的剂量依赖性消退。miR-122T 的插入不影响自杀基因治疗的抗肿瘤作用。虽然给予 Ad-TK 的小鼠在有效治疗剂量下表现出严重的致命性肝毒性，但在 Ad-TK-122T 组中未发现肝损伤。
miR-122 调控的 TK 表达实现了有效的抗肿瘤效果，并增加了 miR-122 缺陷 HCC 瘤内给予腺病毒介导的 TK/GCV 基因治疗的安全性。
