Novel Therapy for Diabetic PAD Monitored With Dual Isotope Multimodality Imaging

 描述（由申请人提供）：症状性外周动脉疾病（PAD）是糖尿病中的一种致残性疾病，可导致腿部溃疡、截肢和死亡。目前没有有效的药物治疗有症状的PAD，只有手术血运重建和基于介入导管的方法，这些方法可能失败，导致肢体丧失。开发治疗PAD的新疗法和对疗法的成像反应代表了未满足的需求。晚期糖基化终产物（AGEs）是由葡萄糖与蛋白质的非酶结合产生的。AGEs结合受体（RECEPTOR，RECEPTOR）启动通路，在加速糖尿病血管疾病的发展和进展中发挥重要作用，并通过抑制肢体缺血的血管生成反应。我们开发了一种用于成像的单克隆抗β-淀粉样蛋白抗体，并在糖尿病和高血糖小鼠和猪中显示β-淀粉样蛋白在后肢血管组织中弥漫性表达。我们还使用成像技术证实，敲除海马可以恢复对股动脉结扎造成的组织缺氧的正常反应。我们假设，如果我们的抗体是一种阻断抗体，它可能具有作为PAD治疗剂的潜力，因此在血管平滑肌细胞的细胞培养物中进行了验证性实验，以记录阻断特性。使用通过免疫组织学证实的成像，我们显示，与安慰剂处理的小鼠相比，在用抗体相对于盐水预处理的股动脉结扎的糖尿病小鼠中，在抗体处理的小鼠的缺血后肢中，在5天时血管生成更大，并且在24天时血流改善。在这项资助申请中，我们提出了一项大型动物治疗试验，使用双同位素多模态成像来记录对治疗的反应。专门饲养的糖尿病尤卡坦小型猪将接受抗体或非免疫IgG 2个月。一个月后，将在这些猪加上额外的年龄匹配和体重匹配的非糖尿病尤卡坦小型猪中进行基于导管的单侧股动脉闭塞（FAO）。在FAO 201 Tl后24小时（急性损伤）和28天（愈合），将进行混合SPECT/CT成像，用于肢体骨骼肌灌注和CT血管造影，用于大血管解剖。在FAO后第7天，将用99 mTc scVEGF-PEG-DOTA（scV/Tc）进行混合SPECT/CT成像，99 mTc scVEGF-PEG-DOTA是靶向VEGF受体1和2的新型探针，并在小鼠中的初步实验中显示以追踪肢体缺血中的血管生成。区域后肢肌肉灌注将从早期和晚期时间点的铊-201摄取定量，并且区域肢体血管生成从第7天的scV/Tc探针摄取定量。我们预期，与安慰剂处理的猪相比，在抗体处理的猪中，从第1天到第28天灌注缺陷的减少将更大，并且这种变化的幅度将与第7天scV/Tc的定量摄取和第28天的侧支血管评分相关。所提出的工作的结果有可能作为进一步开发抗体和这种双同位素多模态成像方法的理由，以进行临床试验。

 DESCRIPTION (provided by applicant): Symptomatic peripheral arterial disease (PAD) is a disabling condition in diabetes that can lead to leg ulcers, amputation, and death. There are currently no effective drug therapies for symptomatic PAD leaving surgical revascularization and interventional catheter based approaches and these can fail leading to limb loss. Developing novel therapies to treat PAD and to image response to therapy represents an unmet need. Advanced Glycation End products (AGEs) are produced by the nonenzymatic binding of glucose to proteins. AGEs bind a receptor (RAGE) to initiate pathways that play important roles in accelerating the development and progression of vascular disease in diabetes and by inhibiting the angiogenic response to limb ischemia. We developed a monoclonal anti-RAGE antibody for imaging and have shown in diabetic and hyperlipidemic mice and pigs that RAGE is expressed diffusely in vascular tissue of hindlimbs. We have also used imaging to confirm that knocking out RAGE restores the normal response to tissue hypoxia imposed by femoral artery ligation. We hypothesized that if our antibody is a blocking antibody it may have potential as a therapeutic agent for PAD and consequently performed confirmatory experiments in cell culture of vascular smooth muscle cells to document blocking properties. Using imaging confirmed by immunohistology we showed in diabetic mice with femoral artery ligation pretreated with antibody vs. saline greater angiogenesis at 5 days and improved blood flow at 24 days in the ischemic hind limb of antibody treated mice compared to placebo treated mice. In this grant application we propose a large animal treatment trial using dual isotope multimodality imaging to document response to therapy. Purpose bred diabetic Yucatan minipigs will receive either antibody or non-immune IgG for 2 months. After one month, catheter based unilateral femoral artery occlusion (FAO) will be performed in these pigs plus additional age matched and weight matched non-diabetic Yucatan minipigs. At 24 h (acute insult) and 28 days (healing) after FAO 201Tl hybrid SPECT/CT imaging will be performed for limb skeletal muscle perfusion and CT angiography for large vessel anatomy. At day 7 after FAO hybrid SPECT/CT imaging will be performed with 99mTc scVEGF-PEG-DOTA (scV/Tc) a novel probe that targets VEGF receptors 1 and 2 and shown in preliminary experiments in mice to track angiogenesis in limb ischemia. Regional hindlimb muscle perfusion will be quantified from uptake of thallium-201 for early and late time points and regional limb angiogenesis from uptake of scV/Tc probe for day 7. We expect that the reduction in perfusion defects from day 1 to 28 will be greater in the antibody treated compared to placebo treated pigs and the magnitude of this change will relate to the quantitative uptake of scV/Tc at day 7 and to the collateral vessel score at day 28. The result of the proposed work has the potential to serve as justification for further development of the antibody and this dual isotope multimodality imaging approach towards a clinical trial.