The recent and rapid increase in the discovery of new RNA therapeutics has created the perfect terrain to explore an increasing number of novel targets. In particular, antisense oligonucleotides (ASOs) have long held the promise of an accelerated and effective drug design compared to other RNA-based therapeutics. Although ASOs in silico design has advanced distinctively in the past years, especially thanks to the several predictive frameworks for RNA folding, it is somehow limited by the wide approximation of calculating sequence affinity based on RNA–RNA/DNA sequences. None of the ASO modifications are taken into consideration, losing hybridization information particularly fundamental to ASOs that elicit their function through RNase H1-mediated mechanisms. Here we present an inexpensive and enhanced biophysical screening strategy to investigate the affinity of ASOs for their target RNA using several biophysical techniques such as high throughput differential scanning fluorimetry (DSF), circular dichroism (CD), isothermal calorimetry (ITC), surface plasmon resonance (SPR) and small-angle X-ray scattering (SAXS).

中文翻译：

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增强的生物物理筛选策略，用于研究 ASO 与其靶标 RNA 的亲和力

最近，新 RNA 疗法的发现迅速增加，为探索越来越多的新靶点创造了完美的领域。特别是，与其他基于 RNA 的疗法相比，反义寡核苷酸 (ASO) 长期以来一直有望加速且有效的药物设计。尽管计算机设计中的ASO在过去几年中取得了显着的进步，特别是由于 RNA 折叠的几个预测框架，但它在某种程度上受到基于 RNA-RNA/DNA 序列计算序列亲和力的广泛近似的限制。没有考虑任何 ASO 修饰，从而丢失了对 ASO 特别重要的杂交信息，这些信息通过 RNase H1 介导的机制引发其功能。在这里，我们提出了一种廉价且增强的生物物理筛选策略，使用多种生物物理技术研究 ASO 与其靶标 RNA 的亲和力，例如高通量差示扫描荧光测定法 (DSF)、圆二色性 (CD)、等温量热法 (ITC)、表面等离子共振(SPR) 和小角 X 射线散射 (SAXS)。