Neural Tract-tracing Nucleic Acid Carriers

描述（申请人提供）：“神经道示踪核酸载体”现有转染试剂在体外成熟神经元中表现很差，不适合在体内使用。这个一期项目将测试创新表面技术公司（ISurTec）正在开发的一种新型核酸载体的性能，用于体外和体内神经元的普遍转染。该技术是基于平台的，由用于高效非病毒基因和siRNA转染的聚合物纳米颗粒组成，通过表面修饰化学掩盖作为神经束示踪剂。选择神经通道示踪表面化学作为载体，是因为通道示踪剂已经多次证明了有效的试剂摄取和逆行颗粒运输，这被认为是神经元转染的主要障碍。由于神经元过程的高度延长，逆行转运对于转染具有基因结构的成熟神经元尤为重要。所提出的载体还包括用于可定制的靶向配体附着的表面官能团。本研究项目的目标是优化试剂配方，使其在成熟哺乳动物神经元中的转染效率大于50%，通过配体附着证明神经元亚型靶向能力，并在动物模型中可预测地改变中枢神经系统回路的功能特性。这项技术的成功发展将使利用分子结构对完整神经系统中的神经可塑性和回路功能进行有针对性的调节/观察成为可能。通过结合纳米颗粒神经束示踪剂的基本特征，期望这种核酸载体具有前所未有的转染效率和成熟神经元的靶向能力。高效转染和精确体内靶向能力的结合将成为分子神经科学与系统、行为和临床前神经科学领域之间的桥梁，通过使当今使用的日益强大的分子生物学工具广泛应用于神经科学研究，极大地增加了我们理解和操纵神经可塑性和大脑功能的手段。最终，这一策略有望导致更有效的治疗各种神经系统疾病和失调，包括帕金森病、脑损伤和慢性药物成瘾。

DESCRIPTION (provided by applicant): "Neural Tract-tracing Nucleic Acid Carriers" Existing transfection reagents perform very poorly in mature neurons in vitro, and are not suitable for in vivo use. This Phase I project will test the performance of a novel nucleic acid carrier under development at Innovative Surface Technologies, Inc. (ISurTec) for universal transfection of neurons in vitro and in vivo. This technology is platform-based and consists of a polymeric nanoparticle for high-efficiency non-viral gene and siRNA transfection, chemically masked as a neural tract-tracer through surface modification. Neural tract-tracing surface chemistry was chosen for the carrier because the tract-tracers have repeatedly demonstrated efficient reagent uptake and retrograde particle transport, which are believed to be the major barriers to neuronal transfection. Retrograde transport is particularly important for transfection of mature neurons with gene constructs, due to the highly elongated nature of neuronal processes. The proposed carrier also includes surface functional groups for customizable attachment of targeting ligands. The objectives of this research project are to optimize the reagent formulation to achieve a transfection efficiency of greater than 50% in mature mammalian neurons, demonstrate neuronal sub-type targeting capability through ligand attachment, and predictably modify the functional properties of a CNS circuit in an animal model. Successful development of this technology will enable the targeted regulation/observation of neuroplasticity and circuit function in the intact nervous system using molecular constructs. An unprecedented level of transfection efficiency and targeting capability for mature neurons is expected from this nucleic acid carrier by incorporating the essential features of nanoparticulate neural tract-tracers. A combination of high efficiency transfection and precise in vivo targeting capability would serve as a bridge between the field of molecular neurosciences and the systems, behavioral, and preclinical neurosciences, greatly increasing our means to understand and manipulate neuroplasticity and brain function, by enabling the increasingly powerful molecular- biological tools in use today to be broadly applied to neuroscience research. Ultimately, this strategy is expected to lead to more effective therapeutic treatments for a variety of neurological diseases and disorders, including Parkinson's disease, brain injury, and chronic drug addiction. PUBLIC HEALTH RELEVANCE: Neurological diseases and disorders, ranging from Parkinson's disease to chronic drug addiction, are extremely difficult to treat due to the complexity and unique challenges of the brain, its neurons, and its circuits. Moreover, neurons are highly resistant to nucleic acid transfection, which hinders the application of today's increasingly powerful molecular-biological tools in neuroscience research. In this Phase I proposal we will test a novel nanoparticle technology designed to significantly increase our understanding of nervous system function through the genetic regulation of brain circuits, and enable more effective treatments for variety of ailments afflicting the human nervous system.