Department of Biological Sciences, University of North Carolina at Charlotte, Charlotte, NC, USA.
Nanoscale Science Program, Department of Chemistry, University of North Carolina at Charlotte, Charlotte, NC 28223, USA.
Nucleic Acids Res. 2020 Nov 18;48(20):11785-11798. doi: 10.1093/nar/gkaa908.
Nucleic acid nanoparticles (NANPs) have become powerful new platforms as therapeutic and diagnostic tools due to the innate biological ability of nucleic acids to identify target molecules or silence genes involved in disease pathways. However, the clinical application of NANPs has been limited by factors such as chemical instability, inefficient intracellular delivery, and the triggering of detrimental inflammatory responses following innate immune recognition of nucleic acids. Here, we have studied the effects of altering the chemical composition of a circumscribed panel of NANPs that share the same connectivity, shape, size, charge and sequences. We show that replacing RNA strands with either DNA or chemical analogs increases the enzymatic and thermodynamic stability of NANPs. Furthermore, we have found that such composition changes affect delivery efficiency and determine subcellular localization, effects that could permit the targeted delivery of NANP-based therapeutics and diagnostics. Importantly, we have determined that altering NANP composition can dictate the degree and mechanisms by which cell immune responses are initiated. While RNA NANPs trigger both TLR7 and RIG-I mediated cytokine and interferon production, DNA NANPs stimulate minimal immune activation. Importantly, incorporation of 2'F modifications abrogates RNA NANP activation of TLR7 but permits RIG-I dependent immune responses. Furthermore, 2'F modifications of DNA NANPs significantly enhances RIG-I mediated production of both proinflammatory cytokines and interferons. Collectively this indicates that off-target effects may be reduced and/or desirable immune responses evoked based upon NANPs modifications. Together, our studies show that NANP composition provides a simple way of controlling the immunostimulatory potential, and physicochemical and delivery characteristics, of such platforms.
核酸纳米颗粒（NANPs）已成为治疗和诊断工具的强大新平台，因为核酸具有识别靶分子或沉默疾病途径中基因的固有生物学能力。然而，由于化学不稳定性、细胞内递呈效率低下以及核酸被先天免疫系统识别后引发有害炎症反应等因素，NANPs 的临床应用受到限制。在这里，我们研究了改变具有相同连接性、形状、大小、电荷和序列的一组限定的 NANPs 的化学组成的影响。我们表明，用 DNA 或化学类似物替代 RNA 链可提高 NANP 的酶和热力学稳定性。此外，我们发现这种组成变化会影响递呈效率并决定亚细胞定位，这些效果可以允许基于 NANP 的治疗剂和诊断剂的靶向递呈。重要的是，我们已经确定改变 NANP 的组成可以决定细胞免疫反应的启动程度和机制。虽然 RNA NANPs 触发 TLR7 和 RIG-I 介导的细胞因子和干扰素的产生，但 DNA NANPs 刺激最小的免疫激活。重要的是，2'F 修饰可消除 RNA NANP 对 TLR7 的激活，但允许 RIG-I 依赖的免疫反应。此外，DNA NANPs 的 2'F 修饰可显著增强 RIG-I 介导的促炎细胞因子和干扰素的产生。总的来说，这表明基于 NANPs 修饰可以减少脱靶效应并/或诱发所需的免疫反应。总之，我们的研究表明，NANP 的组成提供了一种简单的方法来控制这些平台的免疫刺激性、理化性质和递呈特征。
