Development and validation of an intraoperative imaging agent for the peripheral nervous system

项目概要/摘要 2000万美国人患有由创伤和医疗障碍引起的外周神经损伤， 一系列潜在的使人衰弱的副作用在四分之一的病例中，患者将手术视为 他们神经损伤的根本原因特别是在肿瘤切除术或创伤性损伤后， 并且可见性差会挑战外科医生精确定位和保护周围神经的能力。直觉， 如果可以使用外源性对比剂突出神经， 剂然而，在临床实践中，目前的护理标准-视觉检查和触诊-仍然是 不变 外科医生无法在手术过程中识别神经代表了巨大的未满足的临床需求， 建议在本申请中解决。我们将开发和验证一种翻译荧光成像剂 对于周围神经系统来说。为了准备临床试验，我们将在小鼠和大型动物中证明， 该试剂用于人类是安全的。具体地说，这个建议集中在Hsp 1a上，这是我们分离的一种肽， 来自秘鲁狼蛛的毒液，秘鲁种，我们发现它对 人Nav1.7。已经显示Nav1.7在人外周神经组织中高度表达。我们认为 荧光热休克蛋白1a将具有巨大的价值，作为一种可注射的，手术中的指导， 该试剂将为外科医生提供额外的造影剂，减少医源性损伤， 发病率 为了实现这一目标，我们收集了三个具体目标（SA），每个目标都将探索一个独特的目标 显示荧光Nav1.7示踪剂可以应用于临床。在SA 1中，并由我们的 为了验证原理数据，我们将合成荧光标记的Hsp 1a衍生物库（n = 20），其特征在于 不同的荧光团、连接点和连接体长度。对于所有衍生物，我们将测定IC 50值 使用膜片钳电生理学对Nav1.7进行检测;在SA 2中，将对药代动力学和体内性能进行检测。 使用表达Nav1.7的荷瘤无胸腺裸鼠和Nav1.7缺陷Scn 9atm 1Dgen/J小鼠测定。 对于最有希望的示踪剂（n = 3，基于IC 50和信噪比），我们将测试最小可检测浓度。 猪群中的剂量。在SA 3中，我们将询问最有前途的Hsp 1a的药效学特性 缩氨酸我们将确定小鼠的最大耐受剂量，同时监测心血管和 呼吸功能对于主要候选药物，我们将进行2种GLP毒性研究（小鼠和犬）。如果 如果成功，目前的研究方案和产生的数据将构成临床I期试验的基础 Hsp 1a在口腔癌中的作用

Project Summary/Abstract Twenty million Americans suffer from peripheral nerve injury caused by trauma and medical disorders, resulting in a broad spectrum of potentially debilitating side effects. In one out of four cases, patients identify surgery as the root cause of their nerve injury. Particularly during tumor resections or after traumatic injuries, tissue distortion and poor visibility can challenge a surgeon's ability to precisely locate and preserve peripheral nerves. Intuitively, surgical outcomes would improve tremendously if nerves could be highlighted using an exogeneous contrast agent. In clinical practice, however, the current standard of care – visual examination and palpation – remains unchanged. The inability of surgeons to identify nerves during surgery represents an immense unmet clinical need, which we propose to address within this application. We will develop and validate a translational fluorescent imaging agent for the peripheral nervous system. In preparation for a clinical trial, we will show in mice and large animals that the agent is safe for use in humans. Specifically, this proposal centers on Hsp1a, a peptide which we isolated from the venom of a Peruvian tarantula, Homoeomma Spec. Peru, and which we found to be highly specific for human Nav1.7. It has been shown that Nav1.7 is highly expressed in human peripheral nerve tissue. We believe that a fluorescent Hsp1a will have tremendous value as an injectable, intraoperative guide during surgery, and that the agent will provide surgeons with additional contrast, reducing iatrogenic injury and therefore surgical morbidity. To achieve this goal, we have assembled three specific aims (SAs), each of which will explore a distinct goal toward showing that fluorescent Nav1.7 tracers can be translated to the clinic. In SA1, and supported by our proof-of-principle data, we will synthesize a library of fluorescently labeled Hsp1a derivatives (n = 20), featuring different fluorophores, attachment points and linker lengths. For all derivatives, we will determine the IC50 values against Nav1.7 using patch-clamp electrophysiology; in SA2, pharmacokinetics and in vivo performance will be determined using Nav1.7 expressing, tumor-bearing athymic nude mice and Nav1.7 deficient Scn9atm1Dgen/J mice. For the most promising tracers (n = 3, based on IC50 and signal/noise ratios), we will test the minimum detectable dose in cohorts of pigs. In SA3, we will interrogate the pharmacodynamic properties of the most promising Hsp1a peptides. We will determine the maximum tolerated doses in mice while monitoring cardiovascular and respiratory function. For the lead candidate, we will perform a 2-species GLP toxicity study (mice and dogs). If successful, the current research proposal and generated data will form the foundation of a clinical phase I trial with Hsp1a in oral cancer.