Aldehyde dehydrogenases (ALDHs) catalyze the oxidation of an aldehyde to a carboxylic acid and are implicated in the etiology of numerous diseases. However, despite their importance, imaging ALDH activity in cells is challenging due to a lack of fluorescent imaging probes. In this report, we present a new family of fluorescent probes composed of an oligothiophene flanked by an aldehyde and an electron donor, termed thiophene-bridged aldehydes (TBAs), which can image ALDH activity in cells. The TBAs image ALDH activity via a fluorescence sensing mechanism based on the modulation of intramolecular charge transfer (ICT) and this enables the TBAs and their ALDH-mediated oxidized products, thiophene-bridged carboxylates (TBCs), to have distinguishable fluorescence spectra. Herein, we show that the TBAs can image ALDH activity in cells via fluorescence microscopy, flow cytometry, and in a plate reader. Using TBA we were able to develop a cell-based high throughput assay for ALDH inhibitors, for the first time, and screened a large, 1460-entry electrophile library against A549 cells. We identified α,β-substituted acrylamides as potent electrophile fragments that can inhibit ALDH activity in cells. These inhibitors sensitized drug-resistant glioblastoma cells to the FDA approved anti-cancer drug, temozolomide. The TBAs have the potential to make the analysis of ALDH activity in cells routinely possible given their ability to spectrally distinguish between an aldehyde and a carboxylic acid.

中文翻译：

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噻吩桥接醛（TBA）通过调节分子内电荷转移来成像细胞中的ALDH活性

醛脱氢酶（ALDHs）催化醛氧化为羧酸，并与多种疾病的病因有关。然而，尽管它们很重要，但由于缺少荧光成像探针，在细胞中对ALDH活性进行成像仍具有挑战性。在这份报告中，我们提出了一个新的荧光探针家族，该探针由一个寡聚噻吩和一个侧翼醛与一个电子供体组成，该噻吩被称为噻吩-桥连醛（TBA），可以成像细胞中的ALDH活性。TBAs通过以下方式显示ALDH活性一种基于分子内电荷转移（ICT）调制的荧光传感机制，这使得TBA及其ALDH介导的氧化产物，噻吩桥接的羧酸盐（TBC）具有独特的荧光光谱。在这里，我们显示TBA可以通过以下方式对细胞中的ALDH活性进行成像荧光显微镜，流式细胞仪和酶标仪。使用TBA，我们能够首次针对ALDH抑制剂开发基于细胞的高通量测定方法，并针对A549细胞筛选了一个大型的1460条目亲电试剂库。我们确定了α，β-取代的丙烯酰胺为有效的亲电子片段，可以抑制细胞中ALDH的活性。这些抑制剂使耐药胶质母细胞瘤细胞对FDA批准的抗癌药物替莫唑胺敏感。考虑到它们能够在光谱上区分醛和羧酸，TBA具有常规分析细胞中ALDH活性的潜力。