PROSTAGLANDIN H SYNTHASE-2 PURIFICATION AND IMMUNOASSAY

前列腺素H合成酶(PGHS)，它催化第一反应在 前列腺素和血栓烷的生物合成，在 在炎症、生殖和可能的致癌过程中起作用。一秒钟 该酶的异构体(PGHS-2)受差异调节，并具有 与PGHS-1相比有显著不同的性质。 该项目的长期目标是开发用于大型 人PGHS-1和PGHS-2的规模化生产和定量。方法 为了大规模生产人的PGHS，将利用培养的 转导PGHS基因的细胞。可获得大量的 纯化的、具有代谢活性的人PGHS亚型将允许 加速开发非类固醇抗炎药(NSAIDs) 与PGHS-1相比，它们对PGHS-2具有选择性。的可用性 区分PGHS-1和PGHS-2以及PGHS的免疫分析 启动子/报告基因结构，将允许差异定量 这两种酶在血小板、白细胞和其他组织中的水平 临床标本，将加快辨证分型的研究 PGHS亚型在不同疾病状态下的表达。这些分子 探针可以用来监测服用低剂量的人的PGHS水平 阿司匹林的抗血栓和/或抗癌研究进展 活动。 建议的商业应用：重组人PGHS-1和PGHS-2将 向制药公司推销，用于选择性筛查非类固醇抗炎药 对于PGHS-2。简便检测PGHS-1和PGHS-2的mRNA和蛋白水平， 而基因的启动子/报告基因构建是 对临床标本的评估，并可能被证明是无价的工具 用于监测特异性肿瘤的病理和药物治疗 疾病状态包括血栓形成、炎症和癌症。

Prostaglandin H synthase (PGHS), which catalyzes the first reaction in the biosynthesis of prostaglandins and thromboxanes, plays a significant role in inflammation, reproduction, and possibly carcinogenesis. A second isoform of this enzyme (PGHS-2) is differentially regulated and has significantly different properties compared with those of PGHS-1. The long term objective of this project is to develop methods for large scale production and quantification of human PGHS-1 and PGHS-2. Methods for large scale production of human PGHS will be developed using cultured cells transfected with PGHS cDNA. Availability of large quantities of purified, metabolically active human PGHS isoforms will permit accelerated development of non-steroidal anti-inflammatory drugs (NSAIDs) that are selective for PGHS-2 relative to PGHS-1. Availability of immunoassays that distinguish PGHS-1 and PGHS-2, as well as PGHS promoter/reporter gene constructs, will permit differential quantitation of the levels of these two enzymes in platelets, leukocytes, and other clinical specimens and will accelerate research on the differential expression of PGHS isoforms in different disease states. These molecular probes can be used to monitor PGHS levels in individuals taking low dose aspirin based on its reported antithrombotic and/or anticancer activities. PROPOSED COMMERCIAL APPLICATION: Recombinant human PGHS-1 and PGHS-2 will be marketed to pharmaceutical companies for screening of NSAIDs selective for PGHS-2. Facile assays for PGHS-1 and PGHS-2 mRNA and protein levels, and promoter/reporter gene constructs of the genes are needed for evaluation of clinical specimens, and may prove to be invaluable tools for monitoring the pathology and pharmacological treatment of specific disease states including thrombosis, inflammation and cancer.