IMB-Z抗EV71的作用机制研究

In recent years, hand,foot and mouth disease (HFMD) caused by EV71 outbreak and result in higher mortality, and now there is no effective drugs and vaccines for the prevention and treatment. IMB-Z found in pre drug screening in vitro exhibits antiviral activity against EV71. Preliminary study found that IMB-Z raised the expression of some interferon induced proteins (ISGs) in cells. It had been reported that ISGs (such as OASL) could be used as anti-EV71 targets, so we infer that IMB-Z may suppress EV71 replication through regulating ISGs expression via signaling pathways. In this study, we will confirm the antiviral efficacy and mechanism of IMB-Z in vivo and in vitro using real-time fluorescent quantitative PCR, western blot, etc. The experiments in vitro: IMB-Z inhibits EV71 replication; IMB-Z may suppress EV71 replication through regulating ISGs expression via signaling pathways, and whether the antiviral effect of IMB-Z can be inhibited by protein inhibitors or siRNA. The experiments in vivo: to evaluate the toxicity，efficacy，antiviral mechanism of IMB-Z in mice. This study will illustrate the anti-EV71 mechanism of IMB-Z, and provide theoretical basis for antiviral drug development.

近年来，EV71 引起手足口病的爆发流行并造成较高死亡率，目前缺乏有效的药物和疫苗用于预防和治疗。前期药物筛选中发现IMB-Z在体外有很好的抗EV71活性。通过前期研究，我们发现IMB-Z能够上调细胞中部分干扰素诱导蛋白（ISGs）的表达，有文献报道ISGs（如OASL）可作为抗EV71靶点，据此推测IMB-Z可能通过信号通路调节ISGs表达，发挥抗EV71 作用。本研究将采用实时荧光定量PCR，westernblot等方法研究IMB-Z在体内外抗EV71的药效及机制。体外实验研究：IMB-Z对EV71复制影响；IMB-Z是否通过信号通路调节ISGs的表达发挥抗病毒作用，抗病毒作用是否被相关蛋白抑制剂或siRNA所抑制。体内实验研究：IMB-Z在小鼠体内的毒性，药效和作用机制。本研究将阐明IMB-Z 抗EV71的作用机制，为抗病毒药物的研发提供一定的基础和理论依据。

肠病毒71型(EV71)是一种单正链RNA病毒，是引起手足口病的主要病原体，目前尚没有有效的抗病毒治疗方法。本课题研究结果表明N-苯并酰胺衍生物IMB-Z通过激活JAK/STAT通路上调干扰素刺激基因（Interferon-stimulated genes, ISGs）的表达来发挥抗EV71的作用。IMB-Z上调的ISGs之一载脂蛋白B信使RNA编辑酶催化多肽样3G (A3G)是一种胞苷脱氨酶，可抑制多种病毒的复制，如人类免疫缺陷病毒I型、乙肝病毒和丙肝病毒。在本研究中，我们发现A3G过表达抑制EV71复制，而下调内源性A3G表达促进EV71复制。IMB-Z对EV71复制的抑制与细胞内A3G水平升高有关，通过RNA干扰降低A3G水平降低了IMB-Z的抗病毒活性。从机制上，我们得到的证据表明，胞苷脱氨酶活性不是A3G抑制EV71复制所必需的。相反，我们证明了A3G可以与病毒3D蛋白和病毒RNA相互作用，并被包装进入子代病毒粒子从而降低子代病毒的复制。综上所述，我们的结果表明A3G是EV71的细胞限制因子，是IMB-Z抗EV71的关键调控蛋白。药物诱导或稳定A3G是一种潜在的治疗EV71的方法，而IMB-Z具有成为治疗EV71感染的药物前景。