Kawano Hiroki, Nishikawa Makiya, Mitsui Masaru, Takahashi Yuki, Kako Keiko, Yamaoka Kiyoshi, Watanabe Yoshihiko, Takakura Yoshinobu
Department of Biopharmaceutics and Drug Metabolism, Graduate School of Pharmaceutical Sciences, Kyoto University, Sakyo-ku, Kyoto, Japan.
Int J Cancer. 2007 Jul 15;121(2):401-6. doi: 10.1002/ijc.22636.
Plasmid DNA (pDNA) expressing mouse interferon (IFN)-beta or IFN-gamma (pCMV-Mu beta and pCMV-Mu gamma, respectively) has been shown to be effective in inhibiting the growth of colon carcinoma CT-26 cells in the liver (Kobayashi et al., Molecular Therapy 2002;6:737-44). The therapeutic effect of such IFN gene transfer could be significantly increased by the sustained expression of IFNs. In the present study, CpG-reduced pDNA encoding IFN-beta or IFN-gamma (pGZB-Mu beta and pGZB-Mu gamma, respectively) was constructed. pCMV-Mu beta and pCMV-Mu gamma were used as conventional CpG-replete pDNAs. Each pDNA was injected into the tail vein of mice by the hydrodynamics-based procedure. An injection of pGZB-Mu beta resulted in very high IFN-beta activities in the serum for at least 24 hr after injection, whereas the IFN-beta activity after pCMV-Mu beta injection declined quickly. About a 14-fold greater amount of IFN-beta was produced from pGZB-Mu beta than from pCMV-Mu beta. pGZB-Mu beta markedly inhibited the pulmonary metastasis of CT-26 cells. Similar, but more marked results were obtained with pGZB-Mu gamma: it increased the area under the concentration-time curve by more than a 60-fold and the mean residence time of IFN-gamma 4-fold compared with pCMV-Mu gamma. The survival time of the pGZB-Mu gamma-treated mice was significantly (p<0.05) longer than that of the saline- or pCMV-Mu gamma-treated mice. These results indicate that long-term expression of IFN can be achieved by CpG-reduced pDNA and sustained IFN gene expression results in enhanced therapeutic effects of IFN gene transfer against tumor metastasis.
表达小鼠干扰素（IFN）-β或IFN-γ的质粒DNA（分别为pCMV-Muβ和pCMV-Muγ）已被证明可有效抑制结肠癌CT-26细胞在肝脏中的生长（Kobayashi等人，《分子治疗》2002年；6：737-44）。这种IFN基因转移的治疗效果可通过IFN的持续表达而显著提高。在本研究中，构建了编码IFN-β或IFN-γ的低CpG质粒DNA（分别为pGZB-Muβ和pGZB-Muγ）。pCMV-Muβ和pCMV-Muγ用作传统的富含CpG的质粒DNA。通过基于流体动力学的方法将每种质粒DNA注入小鼠尾静脉。注射pGZB-Muβ后，血清中IFN-β活性在注射后至少24小时内保持非常高的水平，而注射pCMV-Muβ后IFN-β活性迅速下降。pGZB-Muβ产生的IFN-β量比pCMV-Muβ多约14倍。pGZB-Muβ显著抑制CT-26细胞的肺转移。pGZB-Muγ也得到了类似但更显著的结果：与pCMV-Muγ相比，它使浓度-时间曲线下面积增加了60多倍，IFN-γ的平均驻留时间增加了4倍。pGZB-Muγ治疗组小鼠的存活时间显著长于生理盐水或pCMV-Muγ治疗组小鼠（p<0.05）。这些结果表明，低CpG质粒DNA可实现IFN的长期表达，持续的IFN基因表达可增强IFN基因转移对肿瘤转移的治疗效果。
