Therapeutic Antisense RNA and The HIV Rev-RRE Pathway

描述（由申请人提供）：针对HIV感染的基因治疗有望提供传统药物治疗的替代方案。最终，这种疗法可能能够在没有慢性化疗的情况下通过持续干扰病毒复制来预防进行性HIV感染。本提案中描述的实验旨在提供对有效抑制HIV复制的HIV反义结构的作用机制的详细了解。在初步研究中，我们已经获得证据表明，有效的反义抑制可以通过反义RNA通过Rev/RRE途径的运输来解释（并依赖于）。值得注意的是，这些研究还提出了一种新的细胞质反义抑制机制。该应用程序有三个特定目的：1)进一步分析存在于表达rre驱动的反义RNA的细胞中的靶核糖核蛋白（RNP）复合物；2)研究rre驱动的反义抑制与细胞小调控RNA通路的关系；3)确定ADAR编辑机制是否参与反义抑制。这些研究对于进一步开发递送反义RNA的治疗载体具有重要意义，因为它们可能导致这样的结论：确保用于HIV治疗的反义RNA沿着Rev/RRE途径运输具有显着优势。此外，这些研究无疑将使我们进一步了解Rev与细胞机制之间的相互作用。如果我们能够在Rev/RRE， ADAR编辑机制和miRNA的产生之间建立联系，这将具有更重要的意义。这些发现将为进一步研究HIV利用Rev和RRE通过ADAR编辑增加病毒多样性的可能性提供基础。公共卫生相关性：本研究将重点关注一种新的HIV感染治疗方法，其中涉及使用反义RNA。我们将研究当这种RNA在感染细胞中表达时有效抑制HIV的机制，以及这些机制与HIV Rev-RRE介导的核-胞质转运途径的关系。这项工作对开发新的HIV治疗药物以及利用反义策略治疗其他疾病具有重要意义。

DESCRIPTION (provided by applicant): Gene therapy against HIV infection holds the promise of providing alternatives to traditional drug based therapies. Ultimately, such therapy may be able to prevent progressive HIV infection by sustained interference with viral replication in the absence of chronic chemotherapy. The experiments described in this proposal are aimed at providing a detailed understanding of the mechanism of action of an HIV antisense construct that efficiently inhibits HIV replication. In preliminary studies, we have obtained evidence to suggest that efficient antisense inhibition is explained by (and dependent on) trafficking of the antisense RNA through the Rev/RRE pathway. Significantly, these studies also suggest a novel cytoplasmic mechanism for antisense inhibition. The application has three specific aims which are to: 1) Further analyze the target Ribonuclear Protein (RNP) complexes that are present in cells expressing RRE-driven antisense RNA; 2) Investigate how RRE-driven antisense inhibition relates to cellular small regulatory RNA pathways; 3) Determine if the ADAR editing machinery is involved in antisense inhibition. These studies are of importance for further development of therapeutic vectors delivering antisense RNA, since they may lead to the conclusion that there is a significant advantage in making sure that antisense RNA for HIV therapy traffics along the Rev/RRE pathway. In addition, these studies will undoubtedly give us further insight into the interactions between Rev and the cellular machinery. If we can establish a connection between Rev/RRE, the ADAR editing machinery and the generation of miRNA, this would have even more significant implications. Such findings would provide the basis for further studies looking into the possibility that HIV uses Rev and the RRE to increase viral diversity through ADAR editing. PUBLIC HEALTH RELEVANCE: This research will focus on a novel therapeutic approach for HIV infection, which involves the use of antisense RNA. We will investigate the mechanisms underlying the efficient inhibition of HIV that is seen when this RNA is expressed in infected cells and the relationship of these mechanisms to the HIV Rev-RRE mediated nucleo-cytoplasmic transport pathway. The work has significance for the development of novel HIV therapeutic and also for the general development of the use of antisense strategies for treatment other diseases.