Design and Synthesis of Actin-Targeting 10-Phenoxy Cytochalasan Analogues: Balancing Cytotoxicity and Migrastatic Activity

Cytochalasans are
actin polymerization inhibitors with potent migrastatic
activity, but their potential therapeutic use is limited by their
cytotoxicity. Here, we describe a modular late-stage approach that
introduces unprecedented 10-phenoxy substituents into the cytochalasan
scaffold via a Mitsunobu reaction. A series of ten 10-phenoxycytochalasan
analogues was synthesized and evaluated for actin polymerization inhibition,
migrastatic activity, and cytotoxicity (BLM, MRC-5, and HaCaT). At
10 μM concentration, several 7-hydroxy-10-phenoxycytochalasans
(12a,d–g) significantly inhibited
actin polymerization in vitro and showed migrastatic effects in a
spheroid invasion assay. Para-substituents of the phenoxy group modulated
cytotoxicity without compromising actin polymerization inhibition
or migrastatic activity. In contrast, lipophilic ortho-substituents
predicted by molecular docking to enhance actin binding failed to
manifest migrastatic activity, underscoring the limitations of the
molecular docking with this type of compounds. These findings demonstrate
that migrastatic and cytotoxic effects can be decoupled in cytochalasan
analogues and highlight 10-phenoxy substitution as a promising strategy
toward noncytotoxic migrastatic agents.

细胞松弛素类（Cytochalasans）是一类具有强效细胞迁移抑制活性的肌动蛋白聚合抑制剂，但其治疗应用潜力受限于细胞毒性。本文报道一种模块化后期官能团化合成策略，可通过光延反应（Mitsunobu reaction）将前所未有的10-位苯氧基取代基引入细胞松弛素骨架。本研究合成了一系列共10种10-苯氧基细胞松弛素类似物，并针对肌动蛋白聚合抑制活性、细胞迁移抑制活性以及BLM、MRC-5、HaCaT三种细胞系的细胞毒性进行了评价。在10 μM浓度下，多款7-羟基-10-苯氧基细胞松弛素类化合物（12a、d–g）可在体外显著抑制肌动蛋白聚合，并在球体侵袭实验中表现出细胞迁移抑制活性。苯氧基基团的对位取代基可在不影响肌动蛋白聚合抑制活性与细胞迁移抑制活性的前提下调节细胞毒性。与之相反，经分子对接预测可增强肌动蛋白结合能力的亲脂性邻位取代基，却未能体现细胞迁移抑制活性，这凸显了针对此类化合物开展分子对接研究的局限性。本研究表明，可在细胞松弛素类似物中实现细胞迁移抑制活性与细胞毒性的解耦，并凸显10-苯氧基取代作为开发无细胞毒性迁移抑制剂的极具前景的策略。