Conditionally replicating vectors and the parasitism of HIV-1

项目总结 艾滋病毒-1通过整合并持续存在于受感染细胞的基因组中，继续对全球健康构成威胁。目前的治疗策略有效地减少了艾滋病毒-1的复制并提高了生活质量，然而，病毒变异的存在允许致病变异的持续和最终恢复。 这些变异是HIV-1稳定整合的必然结果，这允许抗药性毒株的自然选择和进化。有趣的是，整合的艾滋病毒可以为有条件的复制载体的传播和传播提供理想的环境。我们已经发现，当有条件复制的载体包含表达靶向于 在LTR转录活性区域，一个额外的长期表观遗传介导的选择压力施加在病毒上。观察到的选择性压力与载体的持久性和病毒的显着转录抑制有关，从而降低了病毒的适合性。此外，我们将利用这些有效的非编码小RNA和最近开发的小RNA靶向基因切除 从人类细胞中切除HIV-1或CCR5片段的复合体，以永久改变病毒和OCR共同受体的表达。在这个项目中，我们将开发有条件的复制载体，它可以针对HIV-1或CCR5进行抑制和/或切除，同时劫持病毒机制以 将抗HIV-1载体传播给其他感染HIV-1的细胞。本质上，我们希望将囚徒困境置于艾滋病毒-1上，并调节病毒的适应性(Morris 2004；Morris and Looney 2005)。我们假设观察到的对HIV-1的长期抑制可以直接受到这种选择压力的调节，并建议通过以下方式检验这一假设：(1)利用可选择的有条件复制的HIV-2载体，(2)确定最佳抑制载体，以及针对HIV或CCR5和/或靶点的小RNA的数量是否重要，以及(3)表征最佳候选载体在人类T细胞和巨噬细胞中各自的长期抑制和/或切除特性。

PROJECT SUMMARY HIV-1 remains a constant threat to global health through its ability to integrate and persist in the genome of infected cells. Current therapeutic strategies effectively reduce HIV-1 replication and improve quality of life, however, existence of viral variants allows for the persistence and eventual recovery of pathogenic variants. These variants are the inevitable outcome of the stable integration of HIV-1, which allows for the natural selection and evolution of drug resistant strains. Interestingly, integrated HIV can provide an ideal environment for the propagation and dissemination of conditionally replicating vectors. We have found that when conditionally replicating vectors contain antiviral genes expressing small non-coding RNAs targeted to transcriptionally active regions of the LTR, an added long-term epigenetic mediated selective pressure is placed on the virus. The observed selective pressure correlates with both the persistence of the vector and significant transcriptional suppression of the virus resulting in reduced viral fitness. Moreover, we will utilize these validated small non-coding RNAs and a recently developed small RNA targeted gene excision complex to excise fragments of HIV-1 or CCR5 from human cells in an effort to permanently alter viral and OCRS co-receptor expression. In this project we will develop conditionally replicating vectors that can both target HIV-1 or CCR5 for suppression and/or excision while simultaneously hijacking the viral machinery to spread the anti-HIV-1 vector to other cells being infected with HIV-1. In essence we wish to place a prisoners dilemma on HIV-1 and modulate viral fitness (Morris 2004; Morris and Looney 2005). We hypothesize that the observed long-term suppression of HIV-1 can be directly regulated by such selective pressures and propose to test this hypothesis by (1) utilizing selectable conditionally replicating HIV-2 vectors, (2) determining the best suppressive vector and whether the number of small RNAs targeted to HIV or CCR5 and/or the targeted loci is important and (3) characterizing the respective long-term suppressive and/or excision properties of the best candidate vectors in human T cells and macrophages.