多价相互作用常常使配体在靶分子上表现出更有效的结合性能。一般而言，可以基于靶分子的化学结构信息和传统的SELEX技术获得的单价适配体来进行后组装，构建多价适配体。但是，许多靶分子可能没有已知的匹配适配体，因此，通过全新进化作为有针对性选择高亲和力的多价适配体的替代策略将是非常有用的。在这里，受抗体与抗原之间多价相互作用的优越性的启发，我们报告了一种基于预组织的DNA框架库的直接SELEX策略，用于选择对上皮细胞黏附分子（EpCAM）的“类抗体多价适配体”（Amap），作为模型靶蛋白。Amap通过两个适配体部分同时结合EpCAM展现出相对较好的结合亲和力，这一点已经通过亲和力分析和分子建模得到证实。此外，Amap与EpCAM之间的动态相互作用在单分子水平上也可以通过磁力镊子直接可视化。Amap适配体对EpCAM阳性癌细胞的很好结合亲和力也得到了验证，这提示他们的Amap-SELEX策略有潜力成为多价适配体全新进化的一条新路。©2023 Wiley-VCH GmbH.

Multivalent interactions can often endow ligands with more efficient binding performance toward target molecules. Generally speaking, a multivalent aptamer can be constructed via post-assembly based on chemical structural information of target molecules and pre-identified monovalent aptamers derived from traditional systematic evolution of ligands by exponential enrichment (SELEX) technology. However, many target molecules may not have known matched aptamer partners, thus a de novo evolution will be highly desired as an alternative strategy for directed selection of a high-avidity, multivalent aptamer. Here, inspired by the superiority of multivalent interactions between antibodies and antigens, a direct SELEX strategy with a preorganized DNA framework library for an "Antibody-mimicking multivalent aptamer" (Amap) selection to epithelial cell adhesion molecule (EpCAM), a model target protein is reported. The Amap presents a relatively good binding affinity through both aptamer moieties concurrently binding to EpCAM, which has been confirmed by affinity analysis and molecular modeling. Furthermore, dynamic interactions between Amap and EpCAM are directly visualized by magnetic tweezers at the single-molecule level. A nice binding affinity of Amap to EpCAM-positive cancer cells has also been verified, which hints that their Amap-SELEX strategy has the potential to be a new route for de novo evolution of multivalent aptamers.© 2023 Wiley-VCH GmbH.