High Throughput Screening Assay for IP7K inositol pyrophosphate kinases

 描述（由申请人提供）：这项拟议的合作是在查佩尔山的北卡罗来纳州大学综合化学生物学和药物发现中心（CICBDD）与NIEHS研究三角公园的Stephen Shears博士之间进行的。我们建议开发可以应用于高通量筛选的方法，以鉴定抑制功能特异性小分子激酶对的先导化学“探针”，所述功能特异性小分子激酶被称为IP7 Ks（a.k.a. PPIP5K）。这些细胞信号酶（IP7 K1/PPIP 5 K1和IP7 K2/PPIP 5 K2）合成二磷酸化肌醇磷酸：二磷酸肌醇五磷酸（1-IP7）和双二磷酸肌醇四磷酸（IP 8）。由IP7 Ks催化的这些反应通过激活蛋白激酶AKT促进细胞增殖。此外，在病毒感染期间，控制促炎性干扰素转录的分子途径被IP7 K激活。这些相同的途径响应于代谢性炎症（例如糖尿病和肥胖症的特征性炎症）激活干扰素转录。因此，IP7 K是新的抗癌和抗炎疗法的潜在药理学靶标。改变IP7 Ks活性的探针的开发也为研究人员提供了识别和研究IP7 Ks其他功能的机会。因此，本申请的重点是开发和验证适用于高通量筛选的测定方法，伴随正交和细胞内测定，以筛选化学文库，目的是发现IP7 K活性的新的细胞渗透性修饰剂。

 DESCRIPTION (provided by applicant): This proposed collaboration is between the Center for Integrative Chemical Biology and Drug Discovery (CICBDD) at the University of North Carolina, Chapel Hill, and Dr. Stephen Shears at NIEHS, Research Triangle Park. We propose to develop the methodology that can applied to a high throughput screen to identify lead chemical "probes" to inhibit a functionally-specialized pair of small molecule kinases known as IP7Ks (a.k.a. PPIP5Ks). These cell-signaling enzymes (IP7K1/PPIP5K1 and IP7K2/PPIP5K2), synthesize diphosphorylated inositol phosphates: the diphosphoinositol pentakisphosphate, 1-IP7, and bis-diphosphoinositol tetrakisphosphate, IP8. These reactions catalyzed by IP7Ks promote cell proliferation through activation of the protein kinase, AKT. Moreover, molecular pathways controlling pro-inflammatory interferon transcription are activated by IP7Ks during viral infection These same pathways activate interferon transcription in response to metabolic inflammation, such as that which characterizes diabetes and obesity. Thus, IP7Ks are potential pharmacological targets for new anti-cancer and anti- inflammatory therapies. The development of probes that alter the activities of IP7Ks also offers opportunities for researchers to identify and study additional functions of the IP7Ks. Thus, the focus of this application is to develop and validate assay methods applicable to high throughput screening, with accompanying orthogonal and in-cell assays, to screen chemical libraries with the aim of discovering novel, cell- permeant modifiers of IP7K activity.