Mechanistic Studies of Anti-HIV Activity of a Novel Ring-Expanded Nucleoside

描述（由申请人提供）：我们最近发现了一种新的环扩展核苷（REN），它能在体外有效抑制人类免疫缺陷病毒（HIV）和丙型肝炎病毒（HCV），这两种病毒的IC50值分别在微摩尔和纳摩尔范围内，对宿主细胞几乎没有毒性。由于HCV是HIV感染者的主要合并感染，因此发现一种能够双重抑制HIV和HCV病毒的化合物具有重要的生物医学意义，值得进一步探索。HIV和HCV是目前引起国内外卫生关注的两种可怕病毒。本建议重点研究该化合物抗hiv活性的机制。由于在这方面什么都不知道，我们打算使用这个R21应用程序来快速获取一些关于受REN影响的病毒生命周期阶段的重要初步数据。这些初步数据将作为后续R01申请的基础，用于深入研究特定于病毒生命周期阶段的抗hiv活性机制，进而为广泛的SAR研究奠定基础。这里需要解决的重要问题包括：(a) REN是否对病毒产生或病毒抑制产生影响，(b)它是否抑制病毒生命周期的早期事件，包括病毒进入、逆转录、核定位和整合，或后期事件，包括整合原病毒的转录、核输出、组装和颗粒产生，(c)考虑到许多（如果不是全部）REN的活性形式，REN的体内磷酸化是否对其活性是必要的，已知核苷类似物是其相应的单、二或三磷酸衍生物，并且(d) REN的活性机制是否涉及通过其5'-三磷酸衍生物并入核酸并随后终止链，考虑到许多抗病毒核苷类似物已知通过该机制起作用。此外，我们将选择性合成几种先导化合物的类似物，以观察其抗病毒活性是否能进一步增强。这包括(a)合成铅REN的核糖类似物，以及(b)在-6位延长几个碳原子的烷基链。最后，我们将研究主要的REN及其类似物是否对耐药HIV突变体有效。

DESCRIPTION (provided by applicant): We have recently discovered a novel ring-expanded nucleoside (REN) that potently inhibits both the human immunodeficiency virus (HIV) as well as the hepatitis C virus (HCV) in vitro with IC50 values in the micromolar and nanomolar ranges, respectively, for the two viruses, with little toxicity, if any, to the host cells. Since HCV is a major co-infection in the HIV-infected individuals, the discovery of a compound capable of dual viral inhibition of HIV and HCV, the two dreaded viruses eliciting current national and international health concern, is biomedically significant and merits further exploration. This proposal specifically focuses on investigation of the mechanism of anti-HIV activity of the compound. Since nothing is known on this front, we intend to use this R21 application to quickly acquire some important preliminary data concerning the stage of the viral life cycle that is being affected by the REN. These preliminary data would serve as the basis for a subsequent R01 application for an in-depth investigation of the mechanism of anti-HIV activity specific to the stage of the viral life cycle, and in turn, for the extensive SAR studies. The important questions to be addressed in here include: (a) whether the REN exerts its effect on the virus production or virus inhibition, (b) whether it inhibits the early events in the viral life cycle, including the viral entry, reverse transcription, nuclear localization and integration, or later events, including transcription of the integrated provirus, nuclear export, assembly and particle production, (c) whether in vivo phosphorylation of the REN is necessary for it to be active, considering that the active forms of many, if not all, nucleoside analogues are known to be their corresponding mono-, di-, or triphosphate derivatives, and (d) whether the mechanism of activity of REN involves incorporation into nucleic acids via their 5'-triphosphate derivatives and subsequent chain termination, considering that many antiviral nucleoside analogues are known to operate by this mechanism. In addition, we will synthesize a few selected analogues of the lead compound to see if the antiviral activity could be further enhanced. This includes (a) synthesis of the ribose analogue of the lead REN, and (b) extending the alkyl chain at position-6 by a few more carbon atoms. Finally, we will investigate if the leading REN as well as its analogues are effective against the resistant HIV mutants.