Cellular events in heritable peripheral neuropathies

描述（由申请方提供）：遗传性脱髓鞘神经病，包括Charcot-Marie-Tooth病1A型（CMT1A），占导致肌肉萎缩和功能障碍的周围神经疾病的很大一部分。外周髓磷脂蛋白22（Peripheral myelin protein 22，PMP 22）是雪旺细胞内的疏水性膜蛋白，其异常表达与大多数CMT1A病例相关。在大多数脱髓鞘神经病患者中，PMP 22基因是重复的，而在一小部分CMT1A和Dejerine-Sottas综合征中，PMP 22中存在单个氨基酸取代。对神经病患者的神经活检的研究揭示了PMP22在雪旺细胞胞质溶胶内的异常保留，以及缺乏正确的髓鞘蛋白表达。为了了解PMP22相关神经病变的亚细胞发病机制，我们已经表征了PMP22的翻译后加工，并发现神经病小鼠的许旺细胞内蛋白质的降解减慢和异常细胞内积累，包括点突变体Trembler J和PMP22过表达模型。由于胞质PMP22仅在神经病小鼠而非正常小鼠的神经组织中检测到，因此PMP22的异常细胞内积累可能有助于疾病发病机制。事实上，在压倒泛素-蛋白酶体途径后，PMP 22的胞质聚集体形成并募集必需的许旺细胞分子，包括分子伴侣和髓磷脂蛋白，其改变细胞的蛋白质平衡。在允许的条件下，从新生儿神经分离的许旺细胞有能力通过分子伴侣和自噬辅助的机制清除这些异常的胞质蛋白质聚集体。在本项目的当前周期中，我们刺激了来自Trembler J和PMP 22过表达小鼠的样品中的伴侣蛋白和自噬反应，并发现可以抑制PMP 22的异常胞质聚集并改善髓鞘产生。此外，我们已经证明，这些途径的饮食刺激对这些神经病小鼠有益。这些原理验证实验的成功为在神经病小鼠中推进特定药理学治疗范例奠定了基础，并评估了神经肌肉功能，神经形态学和相关亚细胞机制的治疗结果。该项目的总体目标是确定分子伴侣和自噬蛋白降解的药理学增强是否可以减缓或停止年轻小鼠神经病变的进展，并研究晚期疾病阶段的样品对这种方法的反应。我们将使用经鉴定的已知小分子来刺激年轻神经病小鼠和晚期疾病状态小鼠的离体样品中的分子伴侣和自噬途径。这些研究将确定通过刺激雪旺细胞内的蛋白质稳态机制来改善PMP 22的亚细胞加工是否可以为CMT1A和相关神经病变的治疗提供可行的方法。公共卫生相关性：遗传性神经病患者的治疗选择有限。我们已经确定蛋白伴侣和自噬蛋白水解作为治疗腓骨肌萎缩症的潜在靶点。我们现在将测试增强这些通路的小分子在神经病小鼠模型中减弱神经变性进展的功效。

DESCRIPTION (provided by applicant): Heritable demyelinating neuropathies, including Charcot-Marie-Tooth disease type 1A (CMT1A), account for a significant portion of peripheral nerve disorders leading to muscle atrophy and functional impairment. Peripheral myelin protein 22 (PMP22) is a hydrophobic integral membrane protein within Schwann cells, whose abnormal expression is associated with the majority of CMT1A cases. In most patients with demyelinating neuropathy, the PMP22 gene is duplicated, while in a smaller fraction of CMT1A and in Dejerine-Sottas Syndrome, single amino acid substitutions in PMP22 are present. Studies of nerve biopsies from neuropathic patients revealed abnormal retention of PMP22 within the Schwann cell cytosol, and the lack of correct myelin protein expression. To gain understanding into the subcellular pathogenesis of PMP22- associated neuropathies, we have characterized the posttranslational processing of PMP22 and found slowed degradation and abnormal intracellular accumulation of the protein within Schwann cells from neuropathic mice, including the point mutant Trembler J and the PMP22 overexpressor models. Since cytosolic PMP22 is only detected in nerve tissue from neuropathic and not normal mice, the abnormal intracellular accumulation of PMP22 likely contributes to the disease pathogenesis. Indeed, upon overwhelming the ubiquitin-proteasome pathway, cytosolic aggregates of PMP22 form and recruit essential Schwann cell molecules, including chaperones and myelin proteins, which alter the protein balance of the cell. Under permissive conditions, Schwann cells isolated from neonatal nerves have the ability to clear these abnormal cytosolic protein aggregates by a mechanism that is assisted by chaperones and autophagy. During the current cycle of this project, we stimulated the chaperone and autophagic responses within samples from Trembler J and PMP22 overexpressor mice, and found that the abnormal cytosolic aggregation of PMP22 can be suppressed and myelin production improved. Furthermore, we have shown that dietary stimulation of these pathways has proven beneficial to these neuropathic mice. The success of these proof-of-principle experiments sets the stage to move forward with specific pharmacologic treatment paradigms in neuropathic mice, and evaluate treatment outcome on neuromuscular function, nerve morphology and associated subcellular mechanisms. The overall aim of this project is to determine if pharmacologic enhancement of chaperones and autophagic protein degradation can slow or halt the progression of the neuropathy in young mice, and to investigate the response of samples from advanced disease stages to this approach. We will use pharmacologically characterized, known small molecules to stimulate the chaperone and autophagy pathways in young neuropathic mice and in ex vivo samples from advanced disease state mice. These studies will determine if improving the subcellular processing of PMP22 by stimulation of protein homeostatic mechanisms within Schwann cells could provide a viable approach for therapy in CMT1A and related neuropathies. PUBLIC HEALTH RELEVANCE: Treatment options for patients with hereditary neuropathies are limited. We have identified protein chaperones and autophagy proteolysis as potential targets for treating Charcot-Marie- Tooth disease. We will now test the efficacy of small molecules that enhance these pathways to attenuate the progression of neurodegeneration in neuropathic mouse models.