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

描述：目前用于器官毒性检测的生物标志物通常不够敏感，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进展或疾病过程的阶段，也不能充分区分一个器官系统与另一个器官系统的损伤。例如，血液中转氨酶（ALT和AST）的水平被广泛接受用于检测肝损伤。然而，这些酶通常在低水平肝损伤的血液中检测不到，并且需要相对长时间的毒素暴露才能被检测到。胞质谷胱甘肽转移酶（GST）家族作为器官损伤的生物标志物提供了更可靠的选择，因为它们表现出许多用于该用途所需的特征，即组织特异性定位，以低毒性（高敏感性）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个同种异构体。GST类及其同种异构体在组织分布上表现出显著差异。例如，GSTA1-1和A2-2是肝脏中发现的主要GST酶，而GSTA3-3主要表达于类固醇组织，而GSTA4-4在所有已检测的组织中都表达。此外，GSTP1-1是主要存在于血小板中的GST。因此，测量血液中特定GST类别和同种异构体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抗体通常无法区分GST类的不同同种异构体，并且在某些情况下，特异性不足，无法区分GST类。的可用性

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.