Development of a one-time gene therapy for age-related macular degeneration targeting CD146

新生血管性年龄相关性黄斑变性（湿性AMD）是一种导致中央视力丧失的多因素疾病。它是西方老年人失明的主要原因，并造成了重大的临床和经济负担。抗vegf药物如阿非利塞普、贝伐单抗、雷尼单抗和最近的布卢珠单抗在临床实践中的应用显著改善了湿性AMD的预后。然而，目前还没有治愈湿性AMD的方法，现有的治疗方法不能防止萎缩和疤痕形成的发展。因此，必须寻找新的治疗方案，以解决这一毁灭性威胁视力的疾病中未得到满足的医疗需求。在这里，我们提出了一种基于新的治疗靶点（CD146，基因名称：MCAM）的一次性基因治疗AMD的发展。与以往直接抑制生长因子VEGFA的方法不同，我们的方法同时靶向促血管生成和促炎症细胞粘附分子CD146，以抑制amd相关视力丧失发病机制中至关重要的多种信号通路。自己的初步结果表明，在湿性AMD的激光诱导脉络膜新生（CNV）小鼠模型中，CD146的抑制减少了渗漏脉络膜血管的形成和病变形成的程度。之前，我们生成了针对小鼠和人CD146的重组抗体片段（fab）变体，并证实了与人CD146细胞外结构域的高亲和力结合。在本项目中，我们将进一步对重组抗cd146 fab进行表征，并在相关体外模型和湿性AMD的CNV小鼠模型中测试其有效性。然后，最有希望的变体将被生产为aav矢量化版本，并再次测试体外和体内功效。在另一种方法中，我们将生成并测试表达CRISPR-Cas9和sgRNAs的AAV载体，靶向小鼠Mcam基因灭活。我们的首要目标是为一种针对湿性AMD的新型、一次性基因治疗方法提供临床前概念证明，并为未来的临床转化选择最佳候选药物。

Neovascular age-related macular degeneration (wet AMD) is a multifactorial disease leading to loss of central vision. It is the leading cause for blindness in the elderly western population and poses a significant clinical and economic burden. Implementation of anti-VEGF drugs like aflibercept, bevacizumab, ranibizumab and more recently brolucizumab in clinical practice has dramatically improved the prognosis of wet AMD. However, currently there is no cure for wet AMD and available treatments cannot prevent the development of atrophy and scar formation. Thus, it is mandatory to find new treatment options to address the unmet medical need in this devastating sight threatening disease. Here, we propose the development of a one-time gene therapy treatment of AMD based on a novel therapeutic target (CD146; gene name: MCAM). Different from previous approaches that directly inhibit the growth factor VEGFA, our approach targets both the proangiogenic and proinflammatory cell adhesion molecule CD146 to inhibit multiple signaling pathways crucially involved in the pathogenesis of AMD-related vision loss. Own preliminary results suggest that inhibition of CD146 reduces the formation of leaky choroidal blood vessels and the extent of lesion formation in the laser-induced choroidal neovascularization (CNV) mouse model of wet AMD. Previously, we generated variants of recombinant antibody fragments (Fabs) directed against mouse and human CD146 and confirmed high affinity binding to the extracellular domain of human CD146. In this project, we will further characterize the recombinant anti-CD146 Fabs and test dem for efficacy in relevant in vitro models and in the CNV mouse model of wet AMD. The most promising variants will then be produced as AAV-vectorized versions and tested again for in vitro and in vivo efficacy. In an alternative approach, we will generate and test AAV vectors expressing CRISPR-Cas9 and sgRNAs targeting the mouse Mcam gene for inactivation. Our overarching goal is to provide preclinical proof of concept for a novel, one-time gene therapy approach against wet AMD and to select an optimal candidate for future clinical translation.