Injectable biomaterial depots to manipulate scar and foster axon growth after SCI

描述(申请人提供)：改善脊髓损伤(SCI)后预后的一个重要策略是实现跨损伤的轴突再生，以达到功能神经目标。各种分子具有促进轴突再生的潜力，但无法通过血脑屏障并在许多中枢神经系统(CNS)区域表现出活性，因此需要局部给药来达到疗效，同时避免副作用。延长的但临时的分娩是 需要的。这种分娩缺乏临床上可翻译的方法。我们的目标是开发功能化的二嵌段共聚多肽水凝胶(DCH)作为完全合成的生物材料，这种材料可以容易和安全地注射到SCI损伤部位及其附近，为操纵局部细胞和刺激轴突再生到健康组织中的多个分子的持续局部释放提供储存库。我们以前的工作证明了DCH输送生长因子的安全性和有效性，这些生长因子在中枢神经系统几毫米的距离内产生可预测的效果。新的初步数据显示：(1)脊髓损伤后2天注射的DCH库能够同时输送多种生长因子，刺激整个脊髓损伤核心的感觉和固有脊髓纤维的实质性再生。我们发现，这些再生的轴突沿着新上调的层粘连蛋白表达的细胞追踪，并且这种再生可以通过同时传递扰断层粘连蛋白-整合素结合的功能阻断抗体来阻止。(2)在转基因小鼠中，当DCH转导多种生长因子，并通过缺失STAT3来减轻胶质瘢痕时，轴突从病变核心再生到远端胶质瘢痕。新的数据还表明，DCH可以传递像JSI这样的疏水小分子，JSI可以抑制STAT3并减少疤痕的形成，其方式与我们的转基因小鼠相当。(3)当将多个DCH储存库同时放置到病变核心和远端健康组织中时，我们发现相当大的轴突再生到包含有存活的Neun阳性神经元的健康组织区域。建议的工作将建立在这些初步发现的基础上，并使用脊髓损伤后注射的DCH库来同时输送不同类型的分子(包括多种蛋白质生长因子、抗体和操纵基因表达的疏水小分子)，以便：(I)操纵疤痕和病变核心的细胞以支持轴突再生，(Ii)直接刺激和引导轴突再生进入、穿过病变和进入健康组织，(Iii)剖析不同分子或分子组合刺激轴突再生的细胞和分子机制，以及(Iv)测试到达健康组织的再生的固有脊髓神经元是否能够接触那里的神经元，并能够形成改善运动功能的中继连接。这种DCH库方法将为脊髓损伤后细胞和分子机制的实验研究提供一个强大的工具，并将促进为潜在的临床翻译寻找合适的分子所需的广泛的试验和错误测试。此外，因为DCH是完全合成的生物材料，所以有 DCH的临床翻译也具有在脊髓损伤中使用的现实潜力。

DESCRIPTION (provided by applicant): An important strategy for improving outcome after spinal cord injury (SCI) is to achieve axon regrowth across lesions to reach functional neural targets. Various molecules have the potential to foster axon regrowth but cannot pass the blood brain barrier and exhibit activity in many central nervous system (CNS) regions, necessitating local delivery to achieve efficacy while avoiding side effects. Prolonged but temporary delivery is needed. Clinically translatable methods for such delivery are lacking. Our goal is to develop functionalized diblock copolypeptide hydrogels (DCH) as fully synthetic biomaterials that that can easily and safely be injected into, and near, SCI lesions to provide depots for sustained local release of multiple molecules that manipulate local cells and stimulate axons to regrow into healthy tissue. Our previous works demonstrates DCH safety and efficacy to deliver growth factors that exert predictable effects over distances of several mm in CNS. New preliminary data show that: (1) DCH depots injected 2 days after SCI are able to simultaneously deliver multiple growth factors that stimulate substantive regrowth of both sensory and propriospinal fibers throughout the SCI lesion core. We find that these regrowing axons track along cells with newly upregulated laminin expression, and that regrowth can be blocked by simultaneous delivery of function-blocking antibodies that disrupt laminin-integrin binding. (2) When DCH delivery of multiple growth factors is combined with attenuation of glial scar by deletion of STAT3 in transgenic mice, axons regrow beyond the lesion core into the distal glial scar. New data also show DCH can deliver hydrophobic small molecules like JSI, which inhibits STAT3 and attenuates scar formation in a manner comparable to our transgenic mice. (3) When multiple DCH depots are placed into both the lesion core and distal healthy tissue, we find considerable axon regrowth into healthy tissue areas that contains viable NeuN-positive neurons. The work proposed will build on these preliminary findings and use DCH depots injected after SCI to simultaneously deliver different types of molecules (including multiple protein growth factors, antibodies and small hydrophobic molecules that manipulate gene expression) in order to: (i) manipulate cells in scar and lesion core to enable and support axon regrowth, (ii) directly stimulate and guide axon regrowth into, through and beyond lesions into healthy tissue, (iii) dissect cellular and molecular mechanisms that underlie the axon regrowth stimulated by different molecules or combinations of molecules, and (iv) test whether regrowing propriospinal neurons that reach healthy tissue are able to contact neurons there and are able to form relay connections that improve locomotor function. This DCH depot approach will provide a powerful tool for the experimental investigation of cellular and molecular mechanisms after SCI, and will facilitate the extensive trial and error testing needed to identify appropriate molecules for potential clinical translation. In addition, because DCH are fully synthetic biomaterials, there is also a realistic potential for clinical translation of DCH for use in SCI.