Expanding the Redox Biology Toolbox

摘要 谷胱甘肽（GSH）是真核细胞中含量最丰富的非蛋白巯基。GSH维持细胞氧化还原 体内平衡，并参与许多生物过程。这些重要功能是动态的 受细胞内GSH浓度和分布的调节。目前，细胞内 GSH来源于细胞裂解物或GSH-S-转移酶（GST）依赖性探针。这些方法， 然而，不能提供关于GSH浓度变化的实时动态的信息。我们报道 第一个基于可逆反应的GSH探针（ThiolQuant绿色），可以进行单点定量， 在活细胞中的GSH水平，并开发了第二代GSH探针-RealThiol（RT）-允许实时 在活细胞中监测GSH动力学。我们提交了关于这些GSH探针的专利（W 02016025382），并且已经 与全球超过70个氧化还原生物学实验室共享RealThiol探针。此外，癌症科 美国国家癌症研究所（NCI）的治疗和诊断实验室支持有兴趣应用我们的GSH 探针在其药物筛选测定中的应用以及所需的RT探针的克量。这方面的首要目标是 第一阶段项目是开发探针，可以监测真实的- 时间与亚细胞特异性，并提供一个完整的工具集，研究谷胱甘肽相关的氧化还原生物学。用于相位 II商业化，我们将专注于基础和临床研究。我们将开发GSH相关的检测试剂盒， 生物学，如铁性下垂。我们还将在临床样本中应用我们的GSH探针，以帮助识别患有 耐药/耐辐射肿瘤。

Abstract Glutathione (GSH) is the most abundant non-protein thiol in eukaryotic cells. GSH maintains cellular redox homeostasis and is involved in numerous biological processes. These important functions are dynamically regulated by the intracellular concentration and distribution of GSH. Currently, the concentration of intracellular GSH is derived from either cell lysates or GSH-S-transferase (GST) dependent probes. These approaches, however, cannot provide information about the real-time dynamics of GSH concentration changes. We reported the first reversible reaction-based GSH probe (ThiolQuant Green) that can perform single-point quantification of GSH levels in living cells and developed the 2nd generation GSH probe—RealThiol (RT)—that allows real-time monitoring of GSH dynamics in living cells. We filed a patent on these GSH probes (WO2016025382) and have shared the RealThiol probe with >70 redox biology labs around the world. Additionally, the Division of Cancer Treatment & Diagnosis Laboratory Support at National Cancer Institute (NCI) is interested in applying our GSH probes in their drug screening assays and requested gram quantity of RT probes. The overarching goal for this Phase I project is to develop probes that can monitor the dynamics of the concentration change of GSH in real- time with subcellular specificity and to provide a complete toolset to study GSH-related redox biology. For Phase II commercialization, we will focus on both basic and clinical research. We will develop assay kits for GSH-related biology, such as ferroptosis. We will also apply our GSH probes in clinical samples to help identify patients with drug/radiation-resistant tumors.