Department of Chemistry , Northwestern University , Evanston , Illinois 60208 , United States.
Department of Surgery , Northwestern Feinberg School of Medicine , Chicago , Illinois 60611 , United States.
Bioconjug Chem. 2019 Mar 20;30(3):944-951. doi: 10.1021/acs.bioconjchem.9b00047. Epub 2019 Mar 4.
Liposomal spherical nucleic acids (LSNAs) are a class of nanomaterial used broadly for biomedical applications. Their intrinsic capacity to rapidly enter cells and engage cell surface and intracellular ligands stems from their unique three-dimensional architecture, which consists of densely packed and uniformly oriented oligonucleotides on the surface of a liposomal core. Such structures are promising for therapeutics because they can carry chemical cargo within the lipid core in addition to the nucleic acids that define them, in principle enabling delivery of multiple signals to a single cell. On the basis of these traits, we have designed novel dual-targeting LSNAs that deliver a nucleic acid specific for TLR9 inhibition and a small molecule (TAK-242) that inhibits TLR4. Toll-like receptors (TLRs) play a large role in pathogen recognition and disease initiation, and TLR subtypes are differentially located within the lipid membranes of the cell surface and within intracellular endosomes. Oftentimes, in acute or chronic inflammatory conditions, multiple TLRs are activated, leading to stimulation of distinct, and sometimes overlapping, downstream pathways. As such, these inflammatory conditions may respond to attenuation of more than one initiating receptor. We show that dual targeting LSNAs, comprised of unilamellar liposomal cores, the INH-18 oligonucleotide sequence, and TAK-242 robustly inhibit TLR-9 and TLR-4 respectively, in engineered TLR reporter cells and primary mouse peritoneal macrophages. Importantly, the LSNAs exhibit up to a 10- and a 1000-fold increase, respectively, in TLR inhibition compared to the linear sequence and TAK-242 alone. Moreover, the timing of delivery is shown to be a critical factor in effecting TLR-inhibition, with near-complete TLR-4 inhibition occurring when cells were pretreated with SNAs for 4 h prior to stimulation. The most pronounced effect observed from this approach is the benefit of delivering the small molecule within the SNA via the receptor-mediated internalization pathway common to SNAs.
脂质体球形核酸（LSNAs）是一类广泛应用于生物医学的纳米材料。它们具有快速进入细胞并与细胞表面和细胞内配体结合的内在能力，这源于它们独特的三维结构，该结构由脂质体核心表面上密集排列且均匀取向的寡核苷酸组成。由于这些结构可以在脂质核心内携带化学货物以及定义它们的核酸，因此对于治疗具有很大的应用前景，原则上可以将多个信号递送到单个细胞。基于这些特性，我们设计了新型的双重靶向 LSNAs，它们可以递送达 TLR9 抑制的核酸和抑制 TLR4 的小分子（TAK-242）。Toll 样受体（TLRs）在病原体识别和疾病起始中起重要作用，并且 TLR 亚型在细胞表面的脂质膜内和细胞内内体中具有不同的位置。在急性或慢性炎症条件下，通常多个 TLR 被激活，导致不同的，有时重叠的下游途径被刺激。因此，这些炎症条件可能对一种以上起始受体的衰减有反应。我们表明，由单室脂质体核心、INH-18 寡核苷酸序列和 TAK-242 组成的双重靶向 LSNAs，在工程 TLR 报告细胞和原代小鼠腹腔巨噬细胞中分别有效地抑制 TLR-9 和 TLR-4。重要的是，与线性序列和 TAK-242 单独相比，LSNAs 的 TLR 抑制作用分别增加了 10 倍和 1000 倍。此外，递药时间被证明是影响 TLR 抑制的一个关键因素，当细胞在用 SNA 预处理 4 小时后再进行刺激时，几乎完全抑制 TLR-4。从这种方法中观察到的最显著效果是通过 SNAs 共有的受体介导的内化途径在 SNA 内递送小分子的益处。
