Glycine Receptor Expression in Sensory Afferents to Modulate Pain Signaling

描述（由申请人提供）：慢性疼痛是一个主要的健康问题，影响8000万美国人在他们的生活中的某个时候。目前的药物治疗不是长期有效的，这导致了基因治疗方法的开发和测试。我们和其他人已经证明，单纯疱疹病毒1型（HSV）为基础的载体可以提供高度有效的疼痛调节转基因的感觉神经元在体内接种外周组织。这种方法的一个主要优点是疼痛组织可以通过局部载体递送特异性靶向。我们相信，通过靶向负责慢性疼痛的特定神经元类型，可以进一步扩展这一优势，从而使基因转移能够针对特定类型的疼痛，如炎症性或神经性疼痛，同时减少有害的副作用。我们最近表明，HSV介导的转移和甘氨酸受体11亚基（GlyR 11）的长期表达可用于控制外源性甘氨酸给药的传入沉默的时间和持续时间。基于这些发现和我们最近在创建高效、完全重靶向的HSV载体方面的成功，我们假设我们可以选择性地沉默负责神经性和炎性疼痛的初级传入神经的不同亚群，从而提供损伤特异性疼痛缓解。这些研究将使我们能够确定是否相同或不同的传入人群的基础炎症性和神经性疼痛，并提供了合理的基础上，为未来发展的HSV为基础的基因转移载体，旨在限制镇痛相关的初级传入。我们预期这些研究将（i）提供一种新的平台技术，使我们能够选择性地表达旨在调节感觉传入亚群功能的转基因，这种策略可以很容易地扩展到其他类型的感觉神经障碍;（ii）开发感觉传入亚型的初始功能和物理图谱，其在沉默后将阻断不同的持续性疼痛状况，从而提供针对性疼痛控制所需的基本信息;（iii）识别功能上已经改变以响应疼痛刺激的传入，提供关于神经纤维可塑性的进一步信息;以及（iv）识别与感觉神经元的不适当群体的沉默（例如改变的本体感受）相关的潜在风险。为了实现这些目标，我们提出了一系列相互关联的实验，如3个具体目标所述。在目的1中，HSV载体的感染性将被重新定向到选择性地粘附（a）A2纤维、（B）肽能纤维和（c）非肽能C纤维。在目标2中，我们将使用转基因启动子用转录靶向补充转导重靶向，当与转导靶向组合时，转基因启动子将微调沉默特异性。该组合策略旨在创建较大的转导靶向组内的感觉纤维亚群的初始精细地图，这些组代表对不同疼痛刺激的反应的关键传入。在目标3中，通过外周接种炎性和神经性疼痛的动物模型将重靶向HSV载体引入DRG中，并在甘氨酸诱导的GlyR 11介导的沉默后建立镇痛功效和副作用特征。

DESCRIPTION (provided by applicant): Chronic pain is a major health concern affecting 80 million Americans at some time in their lives. Current pharmacotherapies are not effective long-term, which has led to the development and testing of gene therapy approaches. We and others have demonstrated that herpes simplex virus type 1 (HSV) based vectors can deliver highly effective pain-modulating transgenes to sensory neurons in vivo following inoculation of peripheral tissue. One major advantage of this approach is that painful tissue can be specifically targeted by local vector delivery. We believe that this advantage can be further extended by targeting specific neuron types responsible for chronic pain, thus enabling gene transfer to be tailored to specific types of pain, such as inflammatory or neuropathic pain, while simultaneously reducing deleterious side effects. We have recently shown that HSV-mediated transfer and long-term expression of the glycine receptor 11 subunit (GlyR11) can be used to control the timing and duration of afferent silencing with exogenous administration of glycine. Based on these findings and our recent success in the creation of highly efficient, fully retargeted HSV vectors, we hypothesize that we can selectively silence distinct subpopulations of primary afferents responsible for neuropathic and inflammatory pain, therefore providing injury specific pain relief. These studies will enable us to determine whether the same or different afferent populations underlie inflammatory and neuropathic pain and provide the rational basis for the future development of HSV-based gene transfer vectors designed to restrict analgesia to the relevant primary afferents. We anticipate that these studies will (i) provide a novel platform technology that will allow us to selectively express transgenes designed to modulate the function of sensory afferent subpopulations, a strategy that can be readily extended to other types of sensory nerve disorders; (ii) develop initial functional and physical maps of sensory afferent subtypes that upon silencing will block different persistent pain conditions, thereby providing essential information needed for targeted pain control; (iii) identify afferents that have been functionally altered to respond to painful stimuli providing further information on nerve fiber plasticity; and, (iv) identify potential risks associated with silencing of an inappropriate population of sensory neurons (e.g. altered proprioception). To achieve these goals, we have proposed a series of interrelated experiments described in 3 Specific Aims. In Aim 1, the infectivity of HSV vectors will be retargeted to selectively transduce (a) A2 fibers, (b) peptidergic and (c) nonpeptidergic C fibers. In Aim 2, we will complement transductional retargeting with transcriptional targeting using transgene promoters that will, when combined with transductional targeting, fine tune silencing specificity. The combination strategy is intended to create initial fine maps of subpopulations of sensory fibers within the larger transductionally targeted groups representing critical afferents for the response to different painful stimuli. In Aim 3, the retargeted HSV vectors will be introduced into the DRG by peripheral inoculation of animal models of inflammatory and neuropathic pain and the analgesic efficacy and side effect profiles will be established following glycine-induced GlyR11-mediated silencing.