Engineering functionalized diamond nanoparticles for intracellular delivery of nucleic acids

我的研究项目的目标是设计一种新型纳米材料，可以将核酸携带到哺乳动物细胞中。基因传递是近二十年来一个重要的研究领域，但主流的基因传递纳米材料、脂质和聚合物在稳定性、效率和毒性方面存在局限性。我的研究项目侧重于使用固体核纳米颗粒，如纳米金刚石，来克服这些障碍。纳米金刚石具有均匀的尺寸分布和可用于化学修饰的大表面积。它们在基因传递所需的剂量下是无毒的。我的研究策略是用化学方法将氨基酸连接到纳米金刚石上，以促进它们与核酸的相互作用。该计划的新颖之处在于合成和表征纳米颗粒与生物活性分子、氨基酸结合的方法，以及阐明它们与哺乳动物细胞、器官和整个身体的有益和有害相互作用。短期研究目标建立在我之前的工作：设计自组装脂质和金刚石基纳米颗粒。为了实现我们的目标，我们用氨基酸修饰纳米金刚石的表面，从而产生可结合核酸的功能化纳米颗粒。我们将评估这些功能化纳米颗粒将核酸输送到模型细胞中的能力。各种功能化纳米金刚石的生物分布将被监测。将建立结构/结构-传递效率/毒性之间的关系。这将导致选择具有最佳特性的候选载体（改进的结合特性，细胞摄取和高活性，无毒性或最小毒性，以及在生物环境中的稳定性）将核酸输送到细胞中。为了研究输送系统的结构，将使用高分辨率显微镜、流式细胞术和基于同步加速器的技术。将这些纳米颗粒的表面特性与输送效率相关联，将有助于绘制通用和特定载体的输送机制。从长远来看，我们将探索如何将纳米金刚石颗粒化学功能化的基本知识扩展到其他固体核和软纳米颗粒，建立核酸的通用传递原理。生物分布与表面性质的相关性将被用作预测其他具有类似表面性质的人造纳米颗粒的生物命运的模型。该研究项目将推进功能化金刚石核纳米颗粒作为核酸输送系统的使用和安全性方面的知识。优化这些输送系统的特性将有利于纳米技术（一个价值12亿美元的全球市场），最终导致人类和动物的基因治疗。

The objective of my research program is to engineer novel nanomaterials that carry nucleic acids into mammalian cells. Gene delivery has been a significant research area for the past two decades, but the mainstream gene delivery nanomaterials, lipids and polymers pose limitations with respect to their stability, efficiency and toxicity. My research program focuses on the use of solid--core nanoparticles, such as nanodiamonds, to overcome these barriers. Nanodiamonds have a uniform size distribution and a large surface area available for chemical modifications. They are nontoxic in doses necessary for gene delivery. My research strategy is to chemically link amino acids to the nanodiamond in order to facilitate their interaction with nucleic acids. The novelty of this program lies in the approaches to the synthesis and characterization of nanoparticles conjugated with bioactive molecules, amino acids and elucidation of their beneficial and detrimental interactions with mammalian cells, organs and whole body. The short--term research objectives build on my previous work: design of self--assembling lipid--based and diamond--based nanoparticles. To achieve our goal, we decorate the surface of nanodiamond with amino acids resulting in functionalized nanoparticles amenable to binding nucleic acids. We will assess the ability of these functionalized nanoparticles to deliver nucleic acids into model cells. The biodistribution of various functionalized nanodiamonds will be monitored. Relationships between structure/architecture - delivery efficiency/toxicity will be established. This will lead to the selection of candidate vectors with optimal properties (improved binding properties, cellular uptake and high activity with no or minimal toxicity, and stability in biological environment) for delivering nucleic acids into cells. To study the architecture of the delivery systems, high--resolution microscopy, flow cytometry and synchrotron-based techniques will be used. Correlating the surface properties of these nanoparticles with the delivery efficiency will allow for mapping universal and carrier--specific delivery mechanisms. Long term, we will explore how the basic knowledge on chemical functionalization of the nanodiamond particles could be expanded to other solid--core and soft nanoparticles, establishing universal delivery principles for nucleic acids. Correlation of biodistribution to surface properties will be used as a model for predicting the biological fate of other man-made nanoparticles with similar surface properties. This research program will advance knowledge of the use and safety of the functionalized diamond--core nanoparticles as delivery systems for nucleic acids. Optimization of the characteristics of these delivery systems will benefit nanotechnology (a global market of $1.2B), eventually leading to gene therapy in humans and animals.