Novel use of fluorescent glucose analogues to identify a new class of triazine-based insulin mimetics possessing useful secondary effects
Novel use of fluorescent glucose analogues to identify a new class of triazine-based insulin mimetics possessing useful secondary effects
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DOI：
10.1039/c0mb00089b
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发表时间：
2011-01-01
影响因子：
--
通讯作者：
Williams, Darren R.
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文献类型：
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作者：
Jung, Da-Woon;Ha, Hyung-Ho;Williams, Darren R.
迫切需要发现通过模仿胰岛素(胰岛素模拟物)的作用来有效治疗糖尿病的新化合物。在细胞中研究抗糖尿病药物的传统方法不利于筛选化学文库来鉴定胰岛素模拟物。2-(N-(7-硝基-2-氧杂-1，3-二唑-4-基)氨基)-2-脱氧葡萄糖(2-NBDG)和6-NBDG是可用于筛选的葡萄糖的荧光类似物。然而，目前还没有关于使用这些类似物来识别新的胰岛素模拟物的公开数据。我们开发了一种基于6-NBDG的筛选系统，使用3T3-L1脂肪细胞在96孔培养板中进行筛选。在该筛选系统中，6-NBDG比2-NBDG产生更大的信号。3T3-L1脂肪细胞摄取6-NBDG对胰岛素、已知的胰岛素模拟物、葡萄糖转运抑制剂和胰岛素增敏化合物敏感。为了验证我们的筛选系统，我们筛选了一个包含576个标记的三嗪小分子的化学文库。筛选结果与商业化的基于酶的葡萄糖测定结果一致。两种葡萄糖摄取诱导剂被证明是非细胞毒性的，并且通过它们抑制肾上腺素刺激的脂肪细胞释放的游离脂肪酸而被确认为胰岛素模拟化合物。这些新的胰岛素模拟物的作用浓度明显低于两种被广泛研究的胰岛素模拟物锌(II)络合物和钒化合物，还显示出对内皮细胞功能(糖尿病继发性并发症的关键决定因素)的新的有益影响。使用6-NBDG的新胰岛素模拟物的发现证实了该探针在基于荧光标记的葡萄糖类似物摄取的大规模细胞筛选系统的开发中的使用。这项研究应该有助于开发新的战略，以发现抗击糖尿病日益流行的新药和药物靶点。
There is an urgent need to discover new compounds that effectively treat diabetes by mimicking the action of insulin (insulin mimetics). Traditional approaches to studying anti-diabetic agents in cells are inconvenient for screening chemical libraries to identify insulin mimetics. 2-(N-(7-Nitrobenz-2-oxa-1,3-diazol-4-yl) amino)-2-deoxyglucose (2-NBDG) and 6-NBDG are fluorescent analogues of glucose that could be employed in screening. However, there are no published data about the use of these analogues to identify new insulin mimetics. We have developed a screening system based on 6-NBDG using 3T3-L1 adipocytes in a 96-well culture plate format. 6-NBDG was found to produce a larger signal than 2-NBDG in this screening system. 6-NBDG uptake in 3T3-L1 adipocytes was sensitive to insulin, known insulin mimetics, inhibitors of glucose transport and insulin-sensitizing compounds. To validate our screening system, a chemical library of 576 tagged, triazine-based small molecules was screened. The screening results were identical to that obtained from a commercial enzyme-based glucose assay. Two inducers of glucose uptake were shown to be non-cytotoxic and confirmed as insulin mimetic compounds by their inhibition of epinephrine-stimulated free fatty acid release from adipocytes. These novel insulin mimetics functioned at a markedly lower concentration than two widely studied insulin mimetics, zinc(II) complexes and vanadium compounds, and also showed novel, beneficial effects on endothelial cell function (a key determinant of secondary complications in diabetes). The discovery of new insulin mimetics using 6-NBDG validates the use of this probe in the development of large-scale, cell-based screening systems based on the uptake of fluorescent-tagged glucose analogues. This research should aid the development of novel strategies to discover new drugs and drug targets for combating the increasing prevalence of diabetes.