Department of Chemistry, University of Connecticut, Storrs, Connecticut 06269, United States.
School of Chemical Sciences, National Centre for Sensor Research, Dublin City University, Glasvenin, Dublin D09 W6Y4, Ireland.
ACS Appl Mater Interfaces. 2024 Aug 21;16(33):43400-43415. doi: 10.1021/acsami.4c09894. Epub 2024 Aug 12.
Nucleic Acid Nanocapsules (NANs) are nucleic acid nanostructures that radially display oligonucleotides on the surface of cross-linked surfactant micelles. Their chemical makeup affords the stimuli-responsive release of therapeutically active DNA-surfactant conjugates into the cells. While NANs have so far demonstrated the effective cytosolic delivery of their nucleic acid cargo, as seen indirectly by their gene regulation capabilities, there remain gaps in the molecular understanding of how this process happens. Herein, we examine the enzymatic degradation of NANs and confirm the identity of the DNA-surfactant conjugates formed by using mass spectrometry (MS). With surface enhanced (resonance) Raman spectroscopy (SE(R)RS), we also provide evidence that the energy-independent translocation of such DNA-surfactant conjugates is contingent upon their release from the NAN structure, which, when intact, otherwise buries the hydrophobic surfactant tail in its interior. Such information suggests a critical role of the surfactant in the lipid disruption capability of the DNA surfactant conjugates generated from degradation of the NANs. Using NANs made with different tail lengths (C and C), we show that this mechanism likely holds true despite significant differences in the physical properties (i.e., critical micelle concentration (CMC), surfactants per micelle, ) of the resultant particles (C and C NANs). While the total cellular uptake efficiencies of C and C NANs are similar, there were differences observed in their cellular distribution and localized trafficking, even after ensuring that the total concentration of DNA was the same for both particles. Ultimately, C NANs appeared less diffuse within cells and colocalized less with lysosomes over time, achieving more significant knockdown of the target gene investigated, suggesting more effective endosomal escape. These differences indicate that the surfactant assembly and disassembly properties, including the number of surfactants per particle and the CMC can have important implications for the cellular delivery efficacy of DNA micelles and surfactant conjugates.
核酸纳米胶囊（NANs）是一种核酸纳米结构，其表面交联的表面活性剂胶束上呈现寡核苷酸。它们的化学组成赋予了治疗有效 DNA-表面活性剂缀合物在细胞内刺激响应释放的能力。尽管 NANs 迄今为止已经证明了其核酸货物的有效细胞质递送，正如其基因调控能力所间接表明的那样，但对于这一过程发生的分子理解仍然存在差距。在这里，我们研究了 NANs 的酶降解，并使用质谱（MS）证实了形成的 DNA-表面活性剂缀合物的身份。通过表面增强（共振）拉曼光谱（SE（R）RS），我们还提供了证据，证明这种能量独立的 DNA-表面活性剂缀合物的易位取决于它们从 NAN 结构中的释放，当 NAN 结构完整时，否则将疏水性表面活性剂尾部埋藏在其内部。这些信息表明，在 NAN 降解产生的 DNA 表面活性剂缀合物的脂质破坏能力中，表面活性剂起着关键作用。使用具有不同尾部长度（C 和 C）的 NANs，我们表明，尽管所得颗粒（C 和 C NANs）的物理性质（即临界胶束浓度（CMC）、每个胶束的表面活性剂）存在显著差异，但这种机制可能仍然成立。虽然 C 和 C NANs 的总细胞摄取效率相似，但观察到它们在细胞内分布和局部运输方面存在差异，即使确保两种颗粒的总 DNA 浓度相同也是如此。最终，C NANs 在细胞内的扩散程度较低，与溶酶体的共定位较少，随着时间的推移，研究的靶基因的敲低更为显著，表明内体逃逸更为有效。这些差异表明，表面活性剂的组装和拆卸特性，包括每个颗粒的表面活性剂数量和 CMC，可能对 DNA 胶束和表面活性剂缀合物的细胞递送效果有重要影响。
