Promoting recovery after spinal cord injury with a targeted small molecule

描述(由申请人提供)： 为了脊髓损伤后的全面功能恢复，必须将受影响神经元和髓鞘少突胶质细胞的死亡降至最低。我们以前曾报道，神经营养素受体p75在脊髓损伤后少突胶质细胞的凋亡中起作用[7]。至于在体内结合和激活p75凋亡级联的配体，我们已经确定了proNGF[8]，这是一种新的配体形式，选择性地与p75结合[9]，其表达在损伤后上调。在这里，我们的目标是通过从一开始就阻断p75的激活来最大限度地减少损伤后少突胶质细胞的损失，在那里我们干扰proNGF启动p75介导的死亡的能力。作为一种翻译应用，应用的目标是开发一种有效的、微创的治疗策略，并在人体临床试验中获得成功的可能性很高。在这项工作中，我们开发了一种非肽小分子LM11A-31(C31)，它不仅可以在体外阻断proNGF介导的少突胶质细胞凋亡，而且当通过静脉/PO途径或IP注射时，它还可以跨越血脑屏障，提供巨大的治疗潜力。C31特别适合用于中枢神经系统的治疗，因为在大脑中检测到的化合物与血浆中检测到的化合物的比率大于5.0。因此，我们建议确定C31是否能有效地促进脊髓损伤后的功能恢复，并在没有明显细胞毒性的情况下确定最大改善的最佳剂量浓度。我们的初步数据表明，p75是参与脊髓损伤后细胞色素C释放的受体之一。细胞色素C释放到胞浆中代表线粒体稳态失调，这是许多病理疾病的标志。我们发现C31抑制挫伤小鼠细胞色素C的释放，这表明C31在体内确实以p75为靶点。我们的初步数据还表明，C31在正常成年小鼠中以每天100毫克/公斤的剂量长期给药4个月，以及单次剂量高达2000毫克/公斤时是没有毒性的。即使在遭受挫伤的小鼠中，C31在10、25和100 mg/kg的剂量下也没有毒性，每天两次，ip给药28-42天。因此，我们建议在全面试验中测试C31的疗效，以此作为促进受损小鼠功能恢复的一种方式。在这一目标下，我们建议确定C31的最佳剂量，以促进运动行为和协调性，在较长时间内没有任何毒性，或在疼痛感觉方面有显著好处。一旦确定了最佳剂量，我们将评估在该浓度下少突胶质细胞的存活率是否增加。

DESCRIPTION (provided by applicant): For comprehensive functional recovery after spinal cord injury, one must minimize the death of affected neurons and myelinating oligodendrocytes. We have previously reported that a neurotrophin receptor, p75, plays a role in apoptosis of oligodendrocytes after spinal cord injury [7]. As for the ligand that binds and activates the apoptotic cascade of p75 in vivo, we have identified proNGF [8], a novel form of the ligand, which binds p75 selectively [9], and whose expression is upregulated consequent to the injury. Here, our goal is to minimize the loss of oligodendrocytes after injury by blocking p75 activation from the outset, where we disrupt the ability of proNGF to initiate p75-mediated death. As a translational application, an applied goal is to develop an effective and minimally invasive therapeutic strategy with a high likelihood of success in human clinical trials. In this endeavor, we have developed a non-peptide, small molecule, LM11A-31 (C31), that not only blocks proNGF-mediated oligodendrocyte apoptosis in vitro, but also crosses the blood brain barrier when delivered by IV/PO routes or IP injection, providing enormous therapeutic potential. C31 is especially well suited for CNS therapeutics, since the ratio of the compound detected in the brain vs. plasma is greater than 5.0. We thus propose to determine whether C31 is effective at improving functional recovery after spinal cord injury, and determine the optimal dose concentration in terms of maximal improvement without obvious cytotoxicity. Our preliminary data suggest that p75 is one of the receptors involved in cytochrome C release after spinal cord injury. Cytochrome C release into the cytosol represents dysregulation of mitochondrial homeostasis, a hallmark of many pathological conditions. We found that C31 inhibits cytochrome C release that is induced by contusion injury in mice, suggesting that C31 indeed targets p75 in vivo when delivered by IP injection. Our preliminary data also suggest that C31 is not toxic when administered for an extended period of time at 100 mg/kg/day for 4 months and at single dosages of up to 2000 mg/kg in normal adult mice. Even in mice that have undergone contusion injury, C31 is not toxic at 10, 25, and 100 mg/kg, when delivered by IP for 28-42 days, twice daily. We therefore propose to test the efficacy of C31 in a full-scale trial as a way to promote functional recovery of injured mice. Under this goal, we propose to determine the optimal dosage of C31 that promotes motor behavior and coordination without any toxicity for an extended period or significant benefit in pain sensation. Once the optimum dose is determined, we will assess whether oligodendrocyte survival has increased at that concentration.