Intracellular delivery and targeting of nanoparticles

描述（由申请人提供） 摘要：纳米粒子具有重要的生物医学应用，从用基因疗法治疗人类疾病到用荧光探针了解基本细胞功能。现在可以合成具有几乎任何性质或大小的纳米颗粒，将它们引导到特定的细胞，并将它们功能化以靶向细胞内位置，但是将纳米颗粒穿过质膜以到达它们的目标仍然是一个挑战。我们最近展示了第一种将半导体纳米粒子（称为量子点（QD））同时非侵入性递送到多个细胞的胞质溶胶的方法。递送需要芘丁酸盐与阳离子肽组合，用于QD与质膜的直接相互作用。量子点穿过质膜的能力为将其他纳米颗粒递送到活细胞提供了令人兴奋的可能性。该研究计划的第一个目标是确定芘丁酸酯介导的递送的分子和细胞机制，并将其扩展到其他纳米颗粒，最终目标是用所选的纳米颗粒靶向特定的细胞内位点。新的成像方法，包括单粒子跟踪荧光显微镜，将用于探测纳米粒子的运动及其与芘丁酸盐的相互作用，因为它移动通过质膜和细胞质。由于芘丁酸酯介导的递送可能不适用于所有纳米颗粒和所有应用，我们还将努力开发一套基于充分表征的内体摄取途径的胞质递送和靶向方法。这两种递送方法将与旨在优化递送和最小化破坏的纳米颗粒的细胞反应研究结合进行。在这项研究的过程中，量子点，这是足够明亮的成像在单粒子水平，将被用来探测细胞运输的基本问题，包括通过拥挤的细胞环境，囊泡介导的运输扩散，核靶向。 公共卫生相关性：这项研究解决了使用纳米技术造福人类健康的一个根本挑战;将纳米颗粒输送到活细胞中。两种传递方法，易位和内吞作用，将结合细胞反应的研究进行探索，以获得最佳传递和最小的破坏。这些研究将使用一种模块化的功能化方案，可以扩展到各种各样的纳米颗粒，在许多不同的学科中产生广泛的影响，包括癌症治疗，基因递送和细胞成像。

DESCRIPTION (Provided by the applicant) Abstract: Nanoparticles have important biomedical applications ranging from the treatment of human disease with gene therapy to understanding basic cellular functions with fluorescent probes. It is now possible to synthesize nanoparticles with nearly any property or size, direct them to specific cells, and functionalize them to target intracellular locations, but delivering nanoparticles across the plasma membrane to reach their targets remains a challenge. We have recently demonstrated the first method for non-invasive delivery of semiconductor nanoparticles, known as quantum dots (QDs), to the cytosol of multiple cells simultaneously. Delivery requires pyrenebutyrate in combination with a cationic peptide for direct interaction of the QD with the plasma membrane. The ability of QDs to cross the plasma membrane offers exciting possibilities for the delivery of other nanoparticles to living cells. The first goal of this research program is to determine the molecular and cellular mechanism of pyrenebutyrate-mediated delivery and extend it to other nanoparticles with the ultimate goal of targeting specific intracellular sites with nanoparticles of choice. Novel imaging methods, including single particle tracking fluorescence microscopy, will be used to probe the motion of the nanoparticle and its interaction with pyrenebutyrate as it moves across the plasma membrane and through the cytosol. As pyrenebutyrate-mediated delivery may not be suitable for all nanoparticles and all applications, we will also work to develop a suite of cytosolic delivery and targeting methods that are based on the well-characterized endosomal uptake pathways. Both delivery methods will be carried out in conjunction with studies of cellular response to nanoparticles that aim to optimize delivery and minimize disruption. In the course of this research, QDs, which are sufficiently bright for imaging at the single particle level, will be used to probe fundamental questions of cellular transport including diffusion through the crowded cellular environment, vesicle-mediated transport, and nuclear targeting. Public Health Relevance: This research addresses a fundamental challenge in the use of nanotechnology for the benefit of human health; the delivery of nanoparticles into living cells. Two delivery methods, translocation and endocytosis, will be explored in conjunction with studies of cellular response for optimal delivery and minimal disruption. These studies will use a modular scheme for functionalization that can be extended to a wide variety of nanoparticles for broad impact in many different disciplines including cancer therapy, gene delivery, and cellular imaging.