Advanced GST Proteomics for Early Stage Organ-Specific Toxicity Screening. Phase

产品说明：目前用于检测器官毒性的生物标志物通常不够灵敏，无法检测急性器官损伤的早期阶段，即在减少或消除毒素暴露可以防止器官损伤或疾病过程进展的阶段，它们也不能充分区分对一个器官系统与另一个器官系统的损伤。例如，血液中的转氨酶（ALT和AST）水平是检测肝损伤的广泛接受的实践。然而，这些酶通常在低水平肝损伤的血液中检测不到，并且在检测到它们之前需要相对长的毒素暴露。细胞溶质谷胱甘肽转移酶（GST）家族的酶提供了作为器官损伤的生物标志物的更可靠的替代物，因为它们表现出该用途所需的许多特征，即组织特异性定位、以低毒性水平（高灵敏度）释放到血液中以及高细胞内浓度。GST蛋白家族由几个类别（例如A、M、P）组成，其中一些类别含有多种同种型。GST类及其亚型在组织分布中表现出显著差异。例如，GSTA 1 -1和A2-2是在肝脏中发现的主要GST酶，而GSTA 3 -3主要在类固醇生成组织中表达，而GSTA 4 -4在已检查的所有组织中表达。此外，GST P1 -1是主要存在于血小板中的GST。因此，测量血液中特定GST类别和同种型的水平将是对特定器官或组织损伤的有价值的指标。然而，目前可用的GST抗体通常不能区分GST类别的不同同种型，并且在某些情况下，特异性不足以区分GST类别。检测的可用性 特异性GST类别及其同种型，以及多种GST同种型的多重组，将使得对由于外源性暴露引起的器官损伤的分辨率和灵敏度高得多。最近几个额外的胞质GST类（S，O和Z）的鉴定提供了机会，进一步扩大GST检测的应用，毒理学测试与特定的组织相关的特定类别和亚型的免疫测定的发展。在第1阶段，我们成功地开发了针对人GST A亚型的特异性抗体和免疫测定，并证明了它们在啮齿动物毒性测试和人尿液中GST亚型的非侵入性监测中的实用性。2期计划包括（a）开发针对其他人GST类别及其同种型的高度特异性抗体（B）开发针对这些生物标志物的超灵敏免疫测定，（c）开发用于一组GST免疫测定的复杂、快速、易于使用的多重平台，（d）确定该GST蛋白质组学组用于动物模型中的临床前器官特异性毒性测试的效用，和（e）确定GST蛋白质组学组用于人类临床毒性研究的效用，包括与经历化疗的患者中的传统毒性生物标志物的比较。

DESCRIPTION: Biomarkers currently employed for the detection of organ toxicity are often not sensitive enough to detect the early stages of acute organ damage, i.e. at a stage when reducing or eliminating exposure to a toxin could prevent progression of organ damage or a disease process, nor do they adequately discriminate damage to one organ system vs. another. For example, the level of aminotransferases (ALT and AST) in blood is a widely accepted practice for detecting liver damage. However, these enzymes are typically not detectable in blood at low levels of liver damage and require a relatively long exposure to a toxin before they are detected. The cytosolic glutathione transferase (GST) family of enzymes offer a more reliable alternative as a biomarker for organ damage as they exhibit many of the required characteristics for that use, i.e. tissue specific localization, release into the blood at low leves of toxicity (high sensitivity), and a high intracellular concentration. The GST protein family is comprised of several classes (e.g. A, M, P) with some classes containing multiple isoforms. The GST classes and their isoforms exhibit marked differences in tissue distribution. For example, GSTA1-1 and A2-2 are the predominant GST enzymes found in the liver, whereas GSTA3-3 is mainly expressed in steroidogenic tissues and GSTA4-4 is expressed in all tissues that have been examined. Additionally, GSTP1-1 is the GST predominantly found in platelets. Therefore, measuring the level of specific GST classes and isoforms in blood would be a valuable indicator of damage to a particular organ or tissue. However, currently available antibodies for GSTs are often not capable of distinguishing among the different isoforms of a GST class and, in some cases, are insufficiently specific to distinguish among GST classes. The availability of assays for specific GST classes and their isoforms, and multiplex panels for many GST isoforms, would enable a much greater degree of resolution and sensitivity to organ damage due to xenobiotic exposure. Recent identification of several additional cytosolic GST classes (S, O and Z) provides the opportunity to further expand the applications of GST assays for toxicology testing with the development of immunoassays that are specific for given classes and isoforms associated with specific tissues. In Phase 1 we succeeded in developing antibodies and immunoassays specific for human GST A subforms, and demonstrated their utility for rodent toxicity testing and for non-invasive monitoring of GST isoforms in human urine. Phase 2 plans include (a) the development of highly specific antibodies for additional human GST Classes and their isoforms (b) development of ultrasensitive immunoassays for these biomarkers, (c) development of a sophisticated, rapid, easy to use multiplex platform for a panel of GST immunoassays, (d) determination of the utility of this GST proteomics panel for pre-clinical organ-specific toxicity testing in animal models, and (e) determination of the utility of a GST proteomics panel for human clinical toxicity studies, including comparison to traditional toxicity biomarkers in patients undergoing chemotherapy.