Potent effect of adenoviral vector expressing short hairpin RNA targeting ribonucleotide reductase large subunit M1 on cell viability and chemotherapeutic sensitivity to gemcitabine in non-small cell lung cancer cells
Potent effect of adenoviral vector expressing short hairpin RNA targeting ribonucleotide reductase large subunit M1 on cell viability and chemotherapeutic sensitivity to gemcitabine in non-small cell lung cancer cells
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DOI：
10.1016/j.ejca.2015.05.013
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发表时间：
2015-11-01
影响因子：
8.4
通讯作者：
Yokomise, Hiroyasu
中科院分区：
文献类型：
--
作者：
Tokunaga, Yoshimasa;Liu, Dage;Yokomise, Hiroyasu
背景资料：核糖核苷酸还原酶大亚基（RRM 1）是负责合成DNA合成过程中使用的脱氧核糖核苷酸的主要酶。它也是吉西他滨（GEM）的细胞靶点。据报道，RRM 1的过表达与对GEM的抗性和许多类型的恶性肿瘤的不良预后相关。方法：构建靶向RRM 1基因的短发夹状siRNA腺病毒载体（Ad-shRRM 1）。使用两种RRM 1过表达的非小细胞肺癌（NSCLC）细胞系，MAC 10和RERF-LC-MA。结果：Ad-shRRM 1能有效下调两种NSCLC细胞中RRM 1 mRNA和蛋白的表达，并显著降低存活细胞的百分率（P < 0.005）。半胱天冬酶3/7分析显示，用Ad-RRM 1转染增加了含有任何类型的RRM 1过表达细胞的培养物中的凋亡细胞的百分比（p < 0.001）。用Ad-shRRM 1治疗对RRM 1过表达的RERF-LC-MA异种移植物产生了有效的抗肿瘤作用（p < 0.05）。此外，Ad-shRRM 1介导的RRM 1抑制特异性增加了每种RRM 1过表达肿瘤细胞对吉西他滨的敏感性。Ad-shRRM 1和GEM联合治疗对细胞增殖的抑制作用明显大于Ad-shRRM 1或GEM单独治疗。结论：RRM 1是一个很有前途的基因治疗靶点，Ad-shRRM 1对过表达RRM 1的NSCLC细胞具有较强的抗肿瘤作用，尤其是抗增殖和促凋亡作用。Ad-shRRM 1与GEM联合治疗可能成为NSCLC患者的一种新的治疗选择。(C)2015爱思唯尔有限公司版权所有。
Background: Ribonucleotide reductase large subunit (RRM1) is the main enzyme responsible for synthesis of the deoxyribonucleotides used during DNA synthesis. It is also a cellular target for gemcitabine (GEM). Overexpression of RRM1 is reportedly associated with resistance to GEM and the poor prognosis for many types of malignant tumours. Aim of the present study is to establish gene therapy against RRM1-overexpressing tumours.Method: An adenoviral vector that encoded a short hairpin siRNA targeting the RRM1 gene (Ad-shRRM1) was constructed. Two RRM1-overexpressing non-small cell lung cancer (NSCLC) lines, MAC10 and RERF-LC-MA, were used. Finally, a human tumour xenograft model in nude mice was prepared by subcutaneously implanting tumours derived from RERF-LC-MA cells.Results: Ad-shRRM1 effectively downregulated RRM1 mRNA and protein in both types of NSCLC cells and significantly reduced the percentage of viable cells as detected by 3-(4,5-dime thylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay (p < 0.005). Caspase 3/7 analysis revealed that transfection with Ad-RRM1 increased the percentage of apoptotic cells in culture containing either type of RRM1-overexpressing cell (p < 0.001). Treatment with Ad-shRRM1 exerted a potent antitumour effect against the RRM1-overexpressing RERF-LC-MA xenografts (p < 0.05). Furthermore, Ad-shRRM1-mediated inhibition of RRM1 specifically increased sensitivity to gemcitabine of each type of RRM1-overexpressing tumour cell. Combination treatment with Ad-shRRM1 and GEM exerted significantly greater inhibition on cell proliferation than Ad-shRRM1 or GEM treatment alone.Conclusion: RRM1 appeared to be a promising target for gene therapy, and Ad-shRRM1 had strong antitumour effects, specifically anti-proliferative and pro-apoptotic effects, against NSCLC cells that overexpressed RRM1. Combination therapy with Ad-shRRM1 and GEM may become a new treatment option for patients with NSCLC. (C) 2015 Elsevier Ltd. All rights reserved.