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型扩环核苷（REN），其在体外有效抑制人类免疫缺陷病毒（HIV）和丙型肝炎病毒（HCV），两种病毒的IC 50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（如果有的话）。由于HCV是HIV感染个体中的主要合并感染，因此发现能够双重抑制HIV和HCV（引起当前国家和国际健康关注的两种可怕病毒）的化合物具有生物医学意义并且值得进一步探索。该提案特别侧重于研究该化合物的抗HIV活性机制。由于在这方面还一无所知，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R 01申请的基础，用于深入研究病毒生命周期阶段特定的抗HIV活性机制，进而进行广泛的SAR研究。这里要解决的重要问题包括：（B）它是否抑制病毒生命周期中的早期事件，包括病毒进入、逆转录、核定位和整合，或后期事件，包括整合的前病毒的转录、核输出、组装和颗粒产生，（c）考虑到已知许多（如果不是全部）核苷类似物的活性形式是其相应的单-、二-或三磷酸衍生物，REN的体内磷酸化是否是其活性所必需的，和（d）REN的活性机制是否涉及通过它们的5 ′-三磷酸衍生物和随后的链终止，考虑到已知许多抗病毒核苷类似物通过该机制起作用。此外，我们将合成一些先导化合物的选择性类似物，看看是否可以进一步增强抗病毒活性。这包括（a）合成前导REN的核糖类似物，和（B）将6位的烷基链延长几个碳原子。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.