Gene delivery is critical for the development of nucleic acid-based therapies against a range of severe diseases. The conception of non-viral (semi)synthetic vectors with low cytotoxicity and virus-like efficiency is gathering a lot of efforts, but it represents a fantastic challenge still far from accomplishment. Carbohydrate-based scaffolds offer interesting features towards this end, such as easy availability, relatively cheap cost, tuning properties and a good biocompatibility. The lack of analytical methods providing quantitative and qualitative data on their binding properties with oligonucleotides (DNA/RNA), with a minimal time and sample consumption, represents a limitation for these channels. Here, we attempted to fill the gap by hyphenation of capillary electrophoresis with mass spectrometry (CE-MS). This coupling strategy allows discriminating free and complexed DNA oligomers with cationic cyclodextrins (CDs), determining the stoichiometry where the highest observed is always DNAn: n/3(CD), and unambiguously assigning the partners through m/z detection. Very reliable data were obtained with migration time within 5.5 (standard deviation < 0.5%) and 25 min (standard deviation < 1.1%) for UV and MS detection, respectively. Furthermore, varying the nitrogen/phosphorus ratio (N/P), key parameters relating to the thermodynamics e.g. the micro and macroscopic dissociation constants Kd and KD, respectively (both in low μM range) and the Gibbs free energy ΔG (-16.3 to -26.9 kJ mol-1), and also the cooperativity as Hill number (nH between 0.98 and 15.75) of the supramolecular process can be delineated, providing a unique tool for the high throughput screening and selection of efficient gene delivery carriers.

中文翻译：

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通过毛细管电泳与质谱联用揭示聚阳离子环糊精与 DNA 寡聚体的协同结合

基因递送对于开发针对一系列严重疾病的基于核酸的疗法至关重要。具有低细胞毒性和病毒样效率的非病毒（半）合成载体的构想正在聚集很多努力，但它仍然是一个远未实现的巨大挑战。基于碳水化合物的支架为此提供了有趣的特性，例如容易获得、相对便宜的成本、可调节的特性和良好的生物相容性。缺乏分析方法以最少的时间和样品消耗提供有关其与寡核苷酸 (DNA/RNA) 结合特性的定量和定性数据，这代表了这些通道的局限性。在这里，我们试图通过毛细管电泳与质谱 (CE-MS) 的连字符来填补空白。这种耦合策略允许区分游离和复合的 DNA 寡聚物与阳离子环糊精 (CD)，确定观察到的最高值始终是 DNAn: n/3(CD) 的化学计量，并通过 m/z 检测明确分配伙伴。对于 UV 和 MS 检测，迁移时间分别在 5.5 分钟（标准偏差 < 0.5%）和 25 分钟（标准偏差 < 1.1%）内，获得了非常可靠的数据。此外，改变氮/磷比 (N/P)、与热力学相关的关键参数，例如分别是微观和宏观解离常数 Kd 和 KD（均在低 μM 范围内）和吉布斯自由能 ΔG（-16.3 至 - 26.9 kJ mol-1)，以及超分子过程的希尔数（nH ​​介于 0.98 和 15.75 之间）的协同性，