Constant and steady transgene expression of interferon-γ by optimization of plasmid construct for safe and effective interferon-γ gene therapy
Constant and steady transgene expression of interferon-γ by optimization of plasmid construct for safe and effective interferon-γ gene therapy
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DOI：
10.1002/jgm.2616
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发表时间：
2012-04-01
影响因子：
3.5
通讯作者：
Takakura, Yoshinobu
中科院分区：
文献类型：
--
作者：
Ando, Mitsuru;Takahashi, Yuki;Takakura, Yoshinobu
pmCMV（enh）-hEF-1（prom）-muIFN-γ（一种表达鼠干扰素（IFN）-γ的质粒DNA（pDNA），具有鼠巨细胞病毒（mCMV）增强子和人延伸因子（EF）-1启动子）的流体动力学注射已被证明有效治疗小鼠中的癌症和特应性皮炎。然而，与随后时期的稳定水平相比，注射后不久IFN-γ的初始峰值相当高，这可能导致不必要的不良反应。因此，在本研究中，旨在优化IFN-γ基因转移的疗效/副作用比，我们已经开发了质粒载体编码小鼠IFN-γ的控制下的不同组合的启动子和增强子sequences.Methods的启动子和增强子序列的pmCMV（enh）-hEF-1（prom）-huIFN-γ，一个原型质粒表达人IFN-γ，被替换或删除，以获得各种pDNAs。为了评估转基因表达谱，通过流体动力学注射将每种pDNA递送至小鼠，并定期测量血清IFN-γ浓度。结果原型pmCMV（enh）-hEF-1（prom）-huIFN-γ显示出高但下降的IFN-γ浓度。那些含有hROSA 26启动子表达的转基因在一个更恒定的方式，没有初始高浓度，几乎没有降低bodyweight.Conclusions这些结果表明，hROSA 26启动子，无论是否存在和类型的增强子，是适合实现恒定和稳定的转基因表达水平，有效地避免高浓度的IFN-γ引起的体重减轻。版权所有（C）2012约翰威利父子有限公司
Background Hydrodynamic injection of pmCMV(enh)-hEF-1(prom)-muIFN-gamma, a plasmid DNA (pDNA) expressing murine interferon (IFN)-gamma with a murine cytomegalovirus (mCMV) enhancer and a human elongation factor (EF)-1 promoter, has been proven effective for the treatment of cancer and atopic dermatitis in mice. However, the initial peak of IFN-gamma soon after injection was quite high compared to the steady level for subsequent periods, which could cause unwanted adverse effects. Therefore, in the present study, aiming to optimize the efficacy/side-effect ratio of IFN-gamma gene transfer, we have developed plasmid vectors encoding murine IFN-gamma under the control of different combinations of promoter and enhancer sequences.Methods The promoter and enhancer sequence of pmCMV(enh)-hEF-1(prom)-huIFN-gamma, a prototype plasmid expressing human IFN-gamma, was replaced or deleted to obtain various pDNAs. To assess the transgene expression profile, each pDNA was delivered to mice by hydrodynamic injection and the serum IFN-gamma concentration was measured periodically. On the basis of the results obtained, murine IFN-gamma expressing pDNAs were constructed and the body weight change was monitored as an indicator of adverse effects.Results The prototype pmCMV(enh)-hEF-1(prom)-huIFN-gamma showed a high but declining concentration of IFN-gamma. Those containing hROSA26 promoter expressed the transgene in a more constant manner with no initial high concentrations and scarcely reduced the body weight.Conclusions These results indicate that hROSA26 promoter, irrespective of the presence and type of enhancers, is suitable for achieving constant and steady level of transgene expression and effective in avoiding the body weight loss caused by high concentrations of IFN-gamma. Copyright (C) 2012 John Wiley & Sons, Ltd.