Minimizing transgene clearance

最近的一项研究证明了rAAV介导的scFv免疫粘附素的肌内递送可以促进免疫粘附的概念。 恒河猴可以产生持续的高血清水平的这些抑制剂，并保护研究动物 高剂量SIV攻击的结果然而，该研究中的一些治疗动物产生了抗免疫粘附素， 抗体应答，导致清除抗病毒抑制剂。这种免疫清除 sort在rAAV基因治疗文献中有很好的记载：来自 大型动物模型中的rAAV通常可以引起强烈的转基因定向免疫，导致宿主免疫应答。 表达的转基因的清除。事实上，免疫清除仍然是进一步研究的关键挑战。 rAAV递送疗法的翻译开发。因此，有必要更好地定义免疫 过程有助于抗抑制剂抗体反应，并开发新的策略，以防止 表达的转基因的清除。这是项目2的主要目标。 为实现这些目标，我们建议进行以下研究： 1）。限制rAAV-免疫粘附素在非预期靶细胞中的转导和抗原呈递， 通过评估一组肌肉特异性启动子的启动子强度、组织特异性和 用于包装大小受限的rAAV基因组的可行性。 2）。通过利用内源性miRNAs将rAAV转导从抗原呈递细胞去靶向， 树突细胞特异性转录后转基因沉默。 3）。为了通过亲肝rAAV8介导的免疫粘附素表达诱导持续的全身耐受， 和肝特异性转导。 这些研究将产生优化以限制表达的转基因的清除的AAV载体基因组， 在许多情况下，这对AAV载体的治疗用途至关重要。

A recent study proved the concept that rAAV-mediated intramuscular delivery of scFv immunoadhesins to rhesus macaques can generate sustained high serum levels of these inhibitors, and protect study animals from a high-dose SIV challenge. However, some of the treated animals in that study developed anti-immunoadhesin antibody responses, resulting in clearance of antiviral inhibitors. Immune clearance of this sort has been well documented in the rAAV gene-therapy literature: expression of foreign proteins from rAAVs in large animal models can in general elicit strong trangene-directed immunity, leading to host clearance of the expressed transgene. Indeed immune clearence remains a key challenge to further translational development of rAAV-delivered therapies. It is therefore necessary to better define immune processes contributing to anti-inhibitor antibody responses, and to develop novel strategies to prevent clearance of expressed transgenes. These are the main objectives of Project 2. We proposed the following studies to accomplish these objectives: 1). To limit transduction and antigen presentation of rAAV-immunoadhesin in unintended target cells and tissues by evaluating a panel of muscle specific promoters for promoter strength, tissue specificity and feasibility for use in the packaging size-limited rAAV genome. 2). To detarget rAAV transduction from antigen presenting cells by harnessing endogenous miRNAs for dendritic cell specific post-transcriptional transgene silencing. 3). To induce sustained systemic tolerance to immunoadhesin expression by hepatotropic rAAV8-mediated and liver-specific transduction. These studies will generate an AAV vector genome optimized to limit clearence of expressed transgenes, an objective critical to the therapeutic use of AAV vectors in many contexts.