Development of in vivo secreted antiviral entry inhibitory peptides for the treatment of HIV-infection

此前，我们已经开发了一种新的基因治疗策略来治疗HIV感染。抗病毒基因产物（maC46/M87o）是一种膜锚定的抗病毒C肽，能有效抑制HIV-1的进入。然而，患者的整体抗病毒疗效取决于m870基因保护细胞的强大选择性优势，因为最初只有一小部分潜在的HIV靶细胞可以进行基因修饰。在一项I期临床试验中，10例患者显示了自体m870基因修饰T淋巴细胞移植的安全性和可行性。然而，基因修饰细胞的比例仍低于辅助性T细胞总数的1%，不足以达到整体抗病毒效果（van Lunzen et al., 2007）。在本项目中，我们希望通过两种策略来克服这种基因治疗方法的局限性。首先，我们将改进我们目前的体内分泌抗病毒进入抑制（iSAVE）肽，即使在低水平的基因修饰细胞中也已经显示出强大的体外抗病毒活性。iSAVE肽来源于HIV-2EHO的C七重复序列，并被发现在膜融合水平上抑制HIV-1， HlV-2和SIV的进入。为了进一步完善系统，我们将最大限度地提高分泌水平，同时将C肽的潜在免疫原性降到最低。其次，这种最佳的iSAVE肽将在体内系统应用后，通过支持高水平基因表达的载体系统表达。潜在的靶组织是肝脏，甚至更有利的是造血或淋巴组织。这种可直接注射的载体将使这种基因治疗方法得到广泛应用。为此，我们将选择造血和淋巴组织特异性AAV靶向载体（H. b<s:1> ning），并将其与D. v. Laer实验室建立的人源化小鼠HIV感染模型中的嗜肝性AAV血清8型载体进行比较。

Previously, we have developed a novel gene therapeutic strategy for the treatment of HIV infection. The antiviral gene product (maC46/M87o) is a membrane-anchored antiviral C peptide, which inhibits entry of HIV-1 with great efficacy. However, overall antiviral efficacy in patients depends on a robust selective advantage of the M87o gene-protected cells as initially only a small fraction of potential target cells for HIV can be genetically modified. In a phase I clinical trial, the safety and feasibility of the transfer of autologous M87o gene-modified T lymphocytes was shown in 10 patients. The fraction of gene-modified cells, however, remained below 1% of the T helper population, to low to achieve an overall antiviral effect (van Lunzen et al., 2007). In the proposed project, we want to overcome the limitations of this gene therapy approach by two strategies. Firstly, we will improve our current version of the in vivo secreted antiviral entry inhibitory (iSAVE) peptide that already showed potent in vitro antiviral activity even with low levels of gene-modified cells. The iSAVE peptide is derived from the C heptad repeat of HIV-2EHO and was found to inhibit entry at the level of membrane fusion for HIV-1, HlV-2 and SIV. To further improve the system, we will maximize the level of secretion, while the potential immunogenicity of the C peptide will be minimized. Secondly, this optimal iSAVE peptide will be expressed from a vector system that supports high level gene expression following systemic in vivo application. Potential target tissues are liver or even more advantageous the hematopoietic or lymphatic tissue. Such a directly injectable vector would allow broad application of this gene therapy approach. For this purpose we will select hematopoietic and lymphatic tissue-specific AAV targeting vectors (H. Büning), and compare these to the hepatotropic AAV serotype 8 vectors with regard to antiviral efficacy in the humanized mouse models for HIV infecfion established in the laboratory of D. v. Laer.