Kaur Rejbinder, Sloan Lisa A, Blanchard Andy D, Smith Janet L, Churcher Ian, Wayne Gareth J, Ludbrook Steven B
1GlaxoSmithKline Medicines Research Centre, Stevenage, Hertfordshire, UK.
J Biomol Screen. 2013 Dec;18(10):1223-33. doi: 10.1177/1087057113500073. Epub 2013 Aug 27.
Mast cells are unique hematopoietic cells that are richly distributed in the skin and mucosal surfaces of the respiratory and gastrointestinal tract. They play a key role in allergic inflammation by releasing a cocktail of granular constituents, including histamine, serine proteases, and various eicosanoids and cytokines. As such, a number of drugs target either inhibition of mast cell degranulation or the products of degranulation. To identify potential novel drugs and mechanisms in mast cell biology, assays were developed to identify inhibitors of mast cell degranulation and activation in a phenotypic screen. Due to the challenges associated with obtaining primary mast cells, cord blood-derived mononuclear cells were reproducibly differentiated to mast cells and assays developed to monitor tryptase release and prostaglandin D2 generation. The tryptase assay was particularly sensitive, requiring only 500 cells per data point, which permitted a set of approximately 12,000 compounds to be screened robustly and cost-effectively. Active compounds were tested for concomitant inhibition of prostaglandin D2 generation. This study demonstrates the robustness and effectiveness of this approach in the identification of potential novel compounds and mechanisms targeting mast cell-driven inflammation, to enable innovative drug discovery efforts to be prosecuted.
肥大细胞是独特的造血细胞，大量分布于皮肤以及呼吸道和胃肠道的黏膜表面。它们通过释放包括组胺、丝氨酸蛋白酶、各种类花生酸和细胞因子在内的一系列颗粒成分，在过敏性炎症中发挥关键作用。因此，许多药物的靶点是抑制肥大细胞脱颗粒或脱颗粒产物。为了确定肥大细胞生物学中的潜在新药和机制，开发了一些检测方法，用于在表型筛选中鉴定肥大细胞脱颗粒和激活的抑制剂。由于获取原代肥大细胞存在挑战，脐血来源的单核细胞可重复性地分化为肥大细胞，并开发了检测方法来监测类胰蛋白酶释放和前列腺素D2生成。类胰蛋白酶检测特别灵敏，每个数据点仅需500个细胞，这使得能够以强大且经济高效的方式筛选约12000种化合物。对活性化合物进行了前列腺素D2生成的伴随抑制测试。这项研究证明了该方法在鉴定针对肥大细胞驱动炎症的潜在新化合物和机制方面的稳健性和有效性，从而能够推进创新药物研发工作。
