Combination gene editing for local and systemic HIV resistance

摘要 基因编辑有望提高抗HIV基因治疗的精确度和安全性 设计造血干细胞和祖细胞(HSPC)，以重建柏林患者的成分 解药。靶向核酸酶技术已经被用于利用NHEJ介导的DNA断裂修复和 从而破坏CCR5基因，而CCR5基因模仿了他治疗的一个方面。然而，将这种治疗扩大到非 接受工程改造的自体HSPC的癌症患者可能需要包括其他方法。要做到这点， 我们正在开发基于同源定向修复(HDR)的DNA断裂修复的替代途径。这可以 被用来将功能增益突变引入细胞限制因子，或指导受控分泌 可溶的抗病毒因子，包括广泛进入的抑制剂eCD4-Ig。此外，针对特定地点的插入 CCR5基因上的eCD4-Ig将艾滋病毒抵抗力与系统性保护结合起来，从而提供一种 联合抗艾滋病毒的方法。在目前的建议中，我们的目标是继续提高安全性和有效性 在HSPC中进行基因编辑，并应用这项技术来提供局部和系统的艾滋病毒抵抗。我们会 将我们已有的抗艾滋病毒方法与设计生产广谱中和的新方向结合起来 抗体，从而提供一种新的合成免疫能力。个人策略和组合 这些方法将在适当的人源化小鼠模型中进行评估，以评估它们对艾滋病毒感染的影响。 和潜伏的储集层。

ABSTRACT Gene editing holds the promise of enhancing the precision and safety of anti-HIV gene therapies based on engineering hematopoietic stem and progenitor cells (HSPC), in order to recreate elements of the Berlin Patient cure. Targeted nuclease technology is already being used to exploit NHEJ-mediated repair of DNA breaks and thereby disrupt the CCR5 gene, which mimics one aspect of his cure. However, extending this treatment to non- cancer patients, receiving engineered autologous HSPC, will likely need to include other approaches. To do this, we are exploiting the alternate pathway of DNA break repair based on homology-directed repair (HDR). This can be used to introduce gain-of-function mutations into cellular restriction factors, or to direct the controlled secretion of soluble anti-viral factors, including the broad entry inhibitor eCD4-Ig. Moreover, the site-specific insertion at eCD4-Ig at the CCR5 locus would combine HIV-resistance with systemic protection and thereby provide a combinatorial anti-HIV approach. In the current proposal we aim to continue to improve the safety and efficacy of gene editing in HSPC, and to apply the technology to provide both local and systemic HIV resistance. We will combine our established anti-HIV approaches with a new direction to engineer production of broadly neutralizing antibodies, and thereby provide a novel synthetic immune capability. Individual strategies and combination approaches will be evaluated in appropriate humanized mouse models, to evaluate their impact on HIV infection and the latent reservoir.