TARGETED CPG OF CCR5 GENE--NOVEL HIV THERAPY

CCR 5基因编码HIV-1嗜巨噬细胞株的共受体;在缺乏CCR 5的情况下，HIV-1感染被阻断。我们建议通过靶向灭活巨噬细胞祖细胞中的CCR 5基因来使个体对HIV-1具有抗性。我们描述了一种开发靶向甲基转移酶的方法，该方法将对CCR 5基因表达产生强烈而持久的抑制作用。DNA甲基化是导致转座因子和受X失活或基因组印记影响的基因的转录抑制的原因。甲基化模式在S期期间沿着DNA复制。序列特异性甲基化剂的瞬时存在将使靶基因持久失活。 我们将通过将CpG特异性DNA甲基转移酶与锌指蛋白融合来开发针对CCR 5启动子的DNA甲基转移酶，所述锌指蛋白被设计为与CCR 5基因的近端启动子结合。该程序涉及合理设计和选择新的锌指蛋白。该项目的最终目标是开发一种重组腺相关病毒（AAV），其可以递送靶向甲基转移酶以选择性地将CCR 5基因在靶细胞群体中进行转录。这里开发的策略将在其他疾病治疗中得到广泛应用。建议的商业应用：CCR 5基因的靶向甲基化将使个体对HIV感染具有抵抗力。这将对艾滋病毒治疗和预防感染产生直接影响。这项研究更广泛的潜在应用是选择性地抑制与人类疾病有关的任何人类或病毒基因，从而创造出一类新的治疗药物。

The CCR5 gene encodes a co-receptor for macrophage-tropic strains of HIV-1; in the absence of CCR5, HIV-1 infection is blocked. We propose to render individuals resistant to HIV-1 by targeted inactivation of the CCR5 gene in the progenitor cells of macrophages. We describe a method for the development of a targeted methyltransferase that will give strong and enduring inhibition of CCR5 gene expression. DNA methylation is responsible for the transcriptional repression of transposable elements and genes subject to X inactivation or genomic imprinting. Methylation patterns are replicated along with the DNA during S phase. Transient presence of a sequence specific methylating agent will give enduring inactivation of the target gene. We will develop DNA methyltransferases specific to the CCR5 promoter by fusion of CpG-specific DNA methyltransferases to zinc-finger proteins designed to bind to the proximal promoter of the CCR5 gene. The procedure involves rational design and selection of novel zinc finger proteins. The ultimate goal of the project is the development of a recombinant adeno- associated virus (AAV) that can deliver a targeted methyltransferase to selectively inactivate the CCR5 gene in the target cell populations. The strategy developed here will find broad application in other disease therapies. PROPOSED COMMERCIAL APPLICATION: Targeted methylation of the CCR5 gene will render individuals resistant to HIV infection. This will have an immediate impact on HIV therapy and the prevention of infection. The broader potential application of this research is to selectively repress any human or viral gene implicated in human disease, thus creating a new class of therapeutic agents.