Targeted Delivery of S100B Inhibitory Peptide to SCA1 Mouse Cerebellum

描述（由申请人提供）：脊髓小脑共济失调-1（SCA 1）是一种破坏性神经系统疾病，由共济失调蛋白-1基因中的CAG重复扩增引起。多聚谷氨酰胺扩增的突变型共济失调蛋白-1主要靶向小脑的浦肯野细胞（PC）。SCA 1中PC变性的确切机制知之甚少，但已知在SCA 1 PC中观察到的最早形态学变化是含有Bergmann胶质（BG）蛋白（尤其是S100 B）的细胞质空泡的发育。这些空泡是有毒的，并改变PC形态，并可能形成响应于PC的应激信号。此外，BG或空泡释放的S100 B可调节突变型共济失调蛋白-1的Akt磷酸化。已知Akt稳定突变型共济失调蛋白-1以在PC的细胞核中聚集。该研究的目的是用治疗肽（TP）靶向SCA 1 PC，这将影响共济失调蛋白-1的聚集和毒性。虽然TP被设计为抑制特定的分子相互作用，但其体内功效受到不良药代动力学参数的限制。为了改善其药代动力学和生物分布，TP将与基于热响应多肽的载体融合。这种多肽可以通过施加局部热疗靶向Tg小鼠的小脑。热响应多肽的氨基酸序列基于弹性蛋白样多肽（ELP）生物聚合物，其在低于生理温度的水溶液中可溶，但当温度升高到39 ° C以上时聚集。将细胞穿透肽（CPP）缀合至ELP以增强多肽穿过血脑屏障（BBB）的递送并促进细胞进入。将与S100 B相互作用的治疗肽TRTK-12缀合至CPP-ELP。该TP将通过鼻内（IN）或腹膜内（IP）施用，并且通过应用局部高热，将证明这些基因工程化的多肽可以靶向患病部位并改善SCA 1小鼠模型中的运动协调和小脑病理。具体目标1：确定向小脑递送Bac-ELP或SynB 1-ELP IN或IP的可行性。将在有和没有高热的SCA 1小鼠的小脑、其他脑区域和组织中测量放射性标记的Bac/SynB 1-ELP的血浆动力学和体内分布。具体目标2：评价具体目标1中确定的连接至最佳CPP-ELP的TRTK-12的疗效。预计TP治疗将改善运动协调和PC病理学，并将为这种新技术的治疗潜力提供临床前支持。虽然SCA 1小鼠将被用作拟议工作中的模型，但这种方法也可以应用于ELP融合TP对许多其他CNS疾病的热靶向。因此，成功完成拟议的研究可能会产生重大影响，不仅推动这项技术进入临床试验，而且还提供了治疗和管理其他CNS疾病的强大工具。 公共卫生相关性：脊髓小脑性共济失调-1（SCA 1）是一种毁灭性的神经系统疾病，由共济失调蛋白-1基因中的CAG重复扩增引起，导致蛋白质水平上的多聚谷氨酰胺扩增，以及浦肯野细胞以及脑干中的一些其他细胞类型的变性。目前，没有特异性治疗方法来延迟或阻止SCA 1的进展。我们计划使用一种非常独特的策略，通过鼻内或静脉内途径将候选治疗肽热靶向递送至小脑，目的是开发用于治疗SCA 1和其他共济失调的有用的新策略。

DESCRIPTION (provided by applicant): Spinocerebellar ataxia-1 (SCA1) is a devastating neurological disorder, resulting from CAG repeat expansion in the ataxin-1 gene. The polyglutamine expanded mutant ataxin-1 primarily targets Purkinje cells (PCs) of the cerebellum. The exact mechanism of PC degeneration in SCA1 is poorly understood, but it is known that the earliest morphologic change seen in SCA1 PCs is the development of cytoplasmic vacuoles that contain Bergmann glial (BG) proteins, especially S100B. These vacuoles are toxic and alter PC morphology, and may be forming in response to a stress signal of PCs. Furthermore, S100B released by BG or from the vacuoles may modulate Akt phosphorylation of mutant ataxin-1. Akt is known to stabilize mutant ataxin-1 to aggregate in the nucleus of PCs. The objective of the proposed study is to target SCA1 PCs with therapeutic peptide (TP) that will influence ataxin-1 aggregation and toxicity. Though TPs are designed to inhibit specific molecular interactions, their efficacy in vivo is limited by poor pharmacokinetic parameters. To improve their pharmacokinetics and bio-distribution, TP will be fused to a thermally responsive polypeptide-based carrier. This polypeptide can be targeted to the cerebellum of Tg mice by applying local hyperthermia. The amino acid sequence of the thermally responsive polypeptide is based on elastin-like polypeptide (ELP) biopolymers, which are soluble in aqueous solution below physiological temperature, but aggregate when the temperature is raised above 39oC. A cell-penetrating peptide (CPP) is conjugated to the ELP to enhance delivery of the polypeptide across the blood brain barrier (BBB) and to facilitate cell entry. To the CPP-ELP, therapeutic peptide TRTK-12, which interacts with S100B will be conjugated. This TP will be administered intranasally (IN) or intraperitoneally (IP) and, by applying local hyperthermia, it will be demonstrated that these genetically engineered polypeptides can be targeted to a diseased site and improve motor coordination and cerebellar pathology in a SCA1 mouse model. This project will address the following specific aims: Specific Aim 1: Determine feasibility of delivering Bac-ELP or SynB1-ELP IN or IP to the cerebellum. The plasma kinetics and the in vivo distribution of radio-labeled Bac/SynB1-ELP will be measured in the cerebellum, other brain regions and tissues in a SCA1 mouse with and without hyperthermia. Specific Aim 2: Evaluate the therapeutic efficacy TRTK-12 attached to the optimal CPP-ELP as determined in Specific Aim 1. It is anticipated that TP treatment will result in improved motor coordination and PC pathology and will provide preclinical support for the therapeutic potential of this novel technology. Though the SCA1 mouse will be used as a model in the proposed work, this approach could also be applied to thermal targeting of ELP-fused TPs to many other CNS disorders. Therefore, successful completion of the proposed research may have a significant impact, by not only propelling this technology into clinical trials, but also providing a powerful tool to treat and manage other CNS diseases. PUBLIC HEALTH RELEVANCE: Spinocerebellar ataxia-1 (SCA1) is a devastating neurological disease resulting from CAG repeat expansion in the ataxin-1 gene, leading to polyglutamine expansion at the protein level, and degeneration of Purkinje cells as well as some other cell types in the brainstem. Currently, there is no specific treatment to delay or halt the progression of SCA1. We plan to use a very unique strategy of thermally targeted delivery of candidate therapeutic peptides to the cerebellum via intranasal or intravenous routes with the goal of developing useful new strategies for treating SCA1 and other ataxias.