Submicron ultrasound contrast agents as diagnostic agents and therapeutic vehicles in type 1 diabetes

项目摘要 1型糖尿病（T1D）的特征在于胰岛中自身反应性T细胞的浸润，导致糖尿病的发生。 产生胰岛素的β细胞的自身免疫性破坏和糖尿病。启动自身免疫和实质性 β细胞损失可在症状发作前开始数年开始。因此，迫切需要发展 针对T1D这一通常漫长的症状前阶段的诊断和治疗干预。 在针对症状前T1D的治疗的临床试验中已经证明了有限的成功。 虽然抗CD3已显示出前景，但抗CD3仅在研究队列的一个亚组中预防糖尿病发作， 不是特异性针对与β细胞抗原（如胰岛素）反应的T细胞。这种方法 在临床前研究中获得实质性牵引力的是使用肽治疗剂来提供对 自身反应性T细胞靶向的抗原。已经显示在小鼠中施用胰岛素肽治疗剂 通过扩增胰岛素反应性调节性T细胞来预防糖尿病发作，这种细胞具有抗炎作用， 对适当的免疫耐受和调节至关重要。尽管胰岛素肽具有治疗潜力， 在不同的组中显示出混合的结果，并且仅通过外科手术有效地施用 植入输液泵。因此，通过靶向递送和 与诊断结合是有保证的。这可以通过超声造影剂（UCA）来实现， 其是可以使用对比增强超声（CEUS）可视化的小的充气气泡， 安全，易于配制，并获得临床批准。一种新型的亚微米“纳米气泡”超声造影剂， 先前的工作已经证明了纳米气泡在小鼠胰岛中的积累增强， 症状前T1D是炎症相关微血管通透性的结果。我的总体目标是 开发和应用亚微米UCA，将治疗剂特异性靶向疾病部位， 跟踪治疗对T1D进展的影响。我假设亚微米UCA可以应用于 两者都预测治疗诱导的疾病预防和作为靶向肽递送的载体。我将研究 这通过两个具体的目标：我的目标1，我将预测治疗诱导的疾病预防使用亚微米UCA， 使用CEUS检测治疗干预后亚微米UCA的胰岛积累的变化。在 目的2，我将应用UCA作为治疗性肽递送载体。初步数据表明，肽可以 掺入到纳米气泡中，并且纳米气泡可以将肽靶向胰岛。我将描述 对肽细胞摄取的纳米泡消融表征肽-纳米泡胰岛外渗的动力学， 并评估肽-纳米泡治疗的免疫学和疾病修饰作用。开发代理 其允许治疗性肽在胰岛中积累、增强治疗功效和疾病- 反向预测诊断可以作为T1D预防的主要进展。

PROJECT SUMMARY Type 1 diabetes (T1D) is characterized by infiltration of autoreactive T cells in pancreatic islets, leading to autoimmune destruction of insulin-producing beta cells and diabetes. Initiation of autoimmunity and substantial beta cell loss may begin years prior to symptomatic onset. Therefore, there is a crucial need to develop diagnostics and therapeutic interventions directed towards this often-lengthy presymptomatic phase of T1D. Limited success has been demonstrated in clinical trials for therapeutics directed towards presymptomatic T1D. While anti-CD3 has shown promise, anti-CD3 only prevented diabetes onset in a subset of the study cohort and is not directed specifically against T cells reactive to beta cell antigens, such as insulin. An approach that has gained substantial traction in preclinical studies is the usage of peptide therapeutics to provide tolerance towards antigens targeted by autoreactive T cells. Administration of insulin peptide therapeutics in mice has been shown to prevent diabetes onset by expanding insulin-reactive regulatory T cells, which are anti-inflammatory and are essential for proper immune tolerance and regulation. Despite their therapeutic potential, insulin peptides have shown mixed results amongst different groups and have only been effectively administered via surgical implantation of an infusion pump. Therefore, optimizing therapeutic efficacy through targeted delivery and incorporation with diagnostics is warranted. This could be accomplished with ultrasound contrast agents (UCAs), which are small gas-filled bubbles that can be visualized using contrast enhanced ultrasound (CEUS) and are safe, easy to formulate, and clinically approved. A novel, submicron, ‘nanobubble’ ultrasound contrast agent has been developed and prior work has demonstrated enhanced accumulation of nanobubbles in islets of mice with presymptomatic T1D as a result of inflammation-associated microvascular permeability. My overall goal is to develop and apply submicron UCAs to both target therapeutic agents specifically to the disease site and track the effect of therapeutics on T1D progression. I hypothesize that submicron UCAs can be applied to both predict therapeutic induced disease prevention and as vehicles for targeted peptide delivery. I will examine this via two specific aims: I aim 1, I will predict therapeutic-induced disease prevention using submicron UCAs, using CEUS to detect changes in islet accumulation of submicron UCAs following therapeutic intervention. In aim 2, I will apply UCAs as therapeutic peptide delivery vehicles. Preliminary data indicates that peptide can be incorporated into nanobubbles and nanobubbles can target peptide to islets. I will characterize effect of nanobubble ablation on peptide cellular uptake characterize dynamics of peptide-nanobubble islet extravasation, and assess immunological and disease-modifying effects of peptide-nanobubble treatment. Developing an agent that allows for accumulation of therapeutic peptides in islets, enhanced therapeutic efficacy, and disease- reversal-predicting diagnostics can serve as a major advancement in T1D prevention.