Combined gene therapy with oncolytic adenovirus and NK-4 against tumor growth and invasion in hepatocellular carcinoma

过去用于治疗肝细胞癌（HCC）患者的许多方法都不太令人满意。越来越多的证据表明，肝癌细胞的侵袭和转移受到肝细胞生长因子（HGF）拮抗剂NK4的抑制，使用复制能力强的腺病毒进行基因治疗是一种有效的癌症治疗方法。约70%的HCC可产生u-胎蛋白（AFP）， HCC被认为是开发使用不同基因和不同载体的新治疗方法的候选疾病。为了建立有效的抗癌治疗的合理基础，研究了afp启动子驱动的缺乏E1B-55k基因的腺病毒（AdAFP/Rep）和表达NK4的腺病毒载体（AdCMV）治疗的抑制效果。NK4（由大阪大学的T. Nakamura教授和日本东北大学的T. Nukiwa教授提供）靶向产生afp的HCC细胞对肿瘤生长和侵袭的影响在体内和体外进行了研究。AdAFP/Rep能够溶解所有产生afp的HCC细胞系（HuH7, HepG2），而不能溶解原代培养的正常人肝细胞。AdCMV。NK4对产生afp的HCC细胞的增殖有抑制作用，而对正常肝细胞没有抑制作用。AdCMV.NK4可抑制产afp的HCC细胞通过基质基膜组分侵入胶原凝胶。AdAFP/Rep或AdCMV治疗。NK4在移植产生afp的HCC细胞的裸鼠中对肿瘤生长有抑制作用。AdCMV。NK4处理降低了肝癌细胞和植入性皮下肿瘤细胞中血管内皮生长因子（VEGF）、CD31以及基质金属蛋白酶(MMP)-2和MMP-9 mRNA的表达。尽管AdCMV。NK4单独对细胞凋亡的刺激作用并不比AdAFP/Rep和AdCMV联合治疗AdAFP/Rep更有效。NK4诱导细胞和肿瘤的协同促凋亡作用，对隐性肝癌生长和转移的抑制作用明显高于其他治疗。这些发现清楚地表明AdAFP/Rep与AdCMV的联合作用。在实验动物中，NK4可以抑制肝癌细胞的生长、侵袭和转移。因此，它可能为HCC提供更有效的基因治疗。少

The many past approaches used to treat hepatocellular carcinoma (HCC) patients have been less than satisfactory. Accumulating evidence has shown that carcinoma cell invasion and metastasis are inhibited by NK4, an antagonist for hepatocyte growth factor (HGF), and that gene therapy using replication-competent adenovirus is an effective treatment for cancer. HCC, approximately 70% of which can produce u-fetoprotein (AFP), is considered as a candidate disease for the development of a novel treatment using different genes and different vectors. To develop a rational basis for effective anticancer therapy, inhibitory effects of treatment with an AFP-promoter-driven adenovirus deficient of the E1B-55k gene (AdAFP/Rep) and an adenoviral vector expressing NK4 (AdCMV.NK4, provided by Prof. T. Nakamura, Osaka University, and Prof. T. Nukiwa, Tohoku University) to target AFP-producing HCC cells on the tumor growth and invasion were examined in vivo and in vitro. AdAFP/Rep was able to lyse all th … More e AFP-producing HCC cell line (HuH7, HepG2) and not to lyse normal human hepatocytes in primary culture. AdCMV.NK4 played an inhibitory role in the proliferation of the AFP-producing HCC cells, but not normal hepatocytes. Invasion of AFP-producing HCC cells through Matrigel basement membrane components and into collagen gels was inhibited by treatment with AdCMV.NK4. Treatment with AdAFP/Rep or AdCMV.NK4 in nude mice implanted AFP-producing HCC cells resulted in an inhibition in the tumor growth. AdCMV.NK4 treatment decreased mRNA expressions of vascular endothelial growth factor (VEGF) and CD31 as well as matrix metalloproteinase(MMP)-2 and MMP-9 in the HCC cells and implanted subcutaneous tumors. Although AdCMV.NK4 alone did not have a more potent stimulatory effect on apoptosis than that of AdAFP/Rep, the combined treatment with AdAFP/Rep and AdCMV.NK4 induced synergic pro-apoptotic action in the cells and tumors, and inhibited growth and metastasis of invisible HCC much more than each treatment. These findings clearly showed that the combination of AdAFP/Rep and AdCMV.NK4 can inhibit tumor growth, invasion and metastasis of HCC cells in laboratory animals. It may therefore offer a more effective gene therapy for HCC. Less