Blocking vascular permeability by targeting CD148; a novel approach for treating retinopathy

背景：糖尿病视网膜病变是由高血糖引起的糖尿病并发症，高血糖导致眼后血管通透，导致血液和液体渗漏。随着病情的发展，血管生成导致更多有缺陷的血管的形成，使病情恶化。糖尿病视网膜病变影响高达80%的糖尿病患者，这些患者患有糖尿病20年或更长时间。虽然良好的血糖水平管理可以显著减轻病情的严重程度，当治疗是必要的，手术和药物为基础的治疗使用。对于这种情况，通常每月通过玻璃体内注射抗vegf药物。然而，这种方法的使用和疗效存在一些问题，尤其是患者对这种治疗无反应的程度很高（>45%），以及生产和施用这些药物的费用。因此，有必要确定新的治疗靶点，以改善或提供现有治疗的替代方案。研究问题：我们已经确定了一种新的途径，当受到刺激时，它会阻止血管通透性对已知的这种反应的诱导剂（如VEGFA和缓激素）的反应。该途径通过刺激内皮细胞表面的蛋白酪氨酸磷酸酶受体（CD148）起作用。我们已经确定了一种与CD148相互作用并阻断血管通透性的肽QM107，但是我们的初步数据表明，来自CD148细胞外结构域的重组蛋白在抑制血管通透性和血管生成反应方面具有更有效的作用。本项目的目的是进一步探索这一点，并在此过程中回答以下问题：1)实现抑制血管通透性所需的CD148最小肽序列是什么？2)是否比QM107更有效？3) CD148衍生肽是否也能阻断血管生成？4)这种肽在糖尿病视网膜病变的体内模型中起作用吗？技术：该项目将包括Whiteford实验室常规使用的广泛技术。将使用分子生物学和蛋白质纯化方法，并结合一些生物信息学。渗透性测定，如Myles测定，将在体内使用，作为一种新的方法来可视化小鼠眼睛中的血管渗漏。此外，基于细胞的血管生成和血管通透性测定也将被采用。

Background: Diabetic retinopathy is a complication of diabetes caused by high blood sugar which causes permeabilisation of blood vessels at the back of the eye resulting in the leakage of blood and fluids. As the condition progresses angiogenesis occurs leading to the formation of more defective blood vessels exacerbating the condition. Diabetic retinopathy affects up to 80% of all diabetes patients who have had the disease for 20 years or more. Although good management of blood sugar levels can significantly alleviate the severity of this condition, when treatment is necessary both surgical and drug based therapies are used. The administration of anti-VEGF drugs via intra-vitreal injection on a monthly basis is commonly prescribed for this condition. However, there are several concerns with the use and efficacy of this approach, not least the high level of patient non-response to this therapy (>45%) and the expense of producing and administering these drugs. There is therefore a need to identify novel therapeutic targets with the aim of improving or offering an alternative to existing treatments.Research Question: We have identified a novel pathway which when stimulated blocks vascular permeability responses to known inducers of this response such as VEGFA and bradykinin. The pathway acts through the stimulation of a protein tyrosine phosphatase receptor (CD148) on the surface of endothelial cells. We have already identified a peptide QM107 which interacts with CD148 and blocks vascular permeability, however our preliminary data would suggest that recombinant proteins derived from the extracellular domain of CD148 have an even more efficacious effect in inhibiting vascular permeability and angiogenic responses. The purpose of this project is to explore this further and in so doing answer the following questions:1) What is the minimum peptide sequence derived from CD148 required to achieve inhibition of vascular permeability?2) Is this more effective than QM107?3) Can the CD148 derived peptide also block angiogenesis?4) Does this peptide work in an in vivo model of diabetic retinopathy?Techniques: This project will encompass a broad range of techniques which are routinely used in the Whiteford lab. Molecular Biology and protein purification approaches will be used, with some bioinformatics. Permeability assays such as the Myles assay will be used in vivo, as will a novel approach for visualisation vessel leakage in murine eyes. In addition cell based assays for angiogenesis and vascular permeability will also be employed.