Gap junctions (GJs) are dynamic structures composed of hexamers of connexins (Cxs), a class of transmembrane proteins enabling channel-mediated direct intercellular communication through cell-cell diffusion of ions and small metabolites. In defined conditions, Cxs also work as hemichannels allowing exchanges between the cytoplasm and the extracellular medium. The most common GJ channel is formed by connexin 43 (Cx43) and plays an important role in physiological and pathological processes in excitable tissues, such as heart and brain. Hence, Cx43 has been largely envisioned as a new therapeutic target in cancer, neurological and psychiatric indications, or cardiovascular diseases. Identifying new pharmacological inhibitors of Cx43 GJs with different mechanisms of action and from diverse chemical classes is thus highly challenging. We present here a high-content screening method, based on the evaluation of fluorescent dye transfer rates between adjacent cells to monitor the function of GJs in U251 glioblastoma cells expressing high levels of Cx43. This assay was validated using well-described pharmacological GJ inhibitors such as mefloquine. The method was adapted to screen a library of 1,280 Food and Drug Administration- and European Medicines Agency-approved drugs that led to the selection of both known and new inhibitors of GJ channel function. We further focused on a specific class of microtubule-targeting agents, confirming that a proper tubulin network is required for functional Cx43 GJ channels.

中文翻译：

---

高内涵筛选确定了连接蛋白43间隙连接的新抑制剂。

间隙连接（GJs）是由连接蛋白（Cxs）的六聚体组成的动态结构，连接蛋白是一类跨膜蛋白，可通过离子和小代谢物的细胞间扩散，实现通道介导的直接细胞间通讯。在确定的条件下，Cxs还可以作为半通道，允许细胞质与细胞外培养基之间进行交换。最常见的GJ通道是由连接蛋白43（Cx43）形成的，在兴奋性组织（如心脏和大脑）的生理和病理过程中起着重要作用。因此，在癌症，神经和精神病适应症或心血管疾病中，Cx43被广泛设想为新的治疗靶标。因此，鉴定具有不同作用机理和来自不同化学类别的Cx43 GJs的新药理学抑制剂具有很高的挑战性。我们在此提出了一种高含量的筛选方法，该方法基于对相邻细胞之间荧光染料转移速率的评估，以监控表达高水平Cx43的U251胶质母细胞瘤细胞中GJ的功能。使用充分描述的药理GJ抑制剂（如甲氟喹）验证了该测定法。该方法适用于筛选包含1,280种食品和药物管理局以及欧洲药品管理局批准的药物的文库，从而可以选择已知和新的GJ通道功能抑制剂。我们进一步专注于特定类别的微管靶向剂，确认功能性Cx43 GJ通道需要适当的微管蛋白网络。基于对相邻细胞之间荧光染料转移速率的评估，以监测表达高水平Cx43的U251胶质母细胞瘤细胞中GJ的功能。使用充分描述的药理GJ抑制剂（如甲氟喹）验证了该测定法。该方法适用于筛选1,280种由美国食品药品监督管理局和欧洲药品管理局批准的药物的文库，从而可以选择已知和新的GJ通道功能抑制剂。我们进一步专注于特定类别的微管靶向剂，确认功能性Cx43 GJ通道需要适当的微管蛋白网络。基于对相邻细胞之间荧光染料转移速率的评估，以监测表达高水平Cx43的U251胶质母细胞瘤细胞中GJ的功能。使用充分描述的药理GJ抑制剂（如甲氟喹）验证了该测定法。该方法适用于筛选1,280种由美国食品药品监督管理局和欧洲药品管理局批准的药物的文库，从而可以选择已知和新的GJ通道功能抑制剂。我们进一步专注于特定类别的微管靶向剂，确认功能性Cx43 GJ通道需要适当的微管蛋白网络。280食品和药物管理局（Food and Drug Administration）和欧洲药品管理局（European Medicines Agency）批准的药物导致选择了已知和新的GJ通道功能抑制剂。我们进一步专注于特定类别的微管靶向剂，确认功能性Cx43 GJ通道需要适当的微管蛋白网络。280食品和药物管理局（Food and Drug Administration）和欧洲药品管理局（European Medicines Agency）批准的药物导致选择了已知和新型GJ通道功能抑制剂。我们进一步专注于特定类别的微管靶向剂，确认功能性Cx43 GJ通道需要适当的微管蛋白网络。