Characterization of ENTPD1 as a drug target in PXE

ENTPD1 作为 PXE 药物靶标的表征

• 批准号: 10790152
• 负责人: Koen van de Wetering
• 金额: $ 37.75万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-20 至 2025-09-19
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10790152
• 关键词: ATP-Binding Cassette Transporters; Affect; Alleles; Animal Model; Arteries; Binding; Biological Availability; Biological Models; Blindness; Blood Circulation; Blood Vessels; Cardiovascular system; Cells; Characteristics; Chronic Kidney Failure; Circulation; Clinical; Complex; Connective and Soft Tissue; Cutaneous; Data; Deposition; Development; Diabetes Mellitus; Diagnosis; Diphosphates; Disadvantaged; Disease; Disease Progression; Dose; Drug Targeting; Environment; Extracellular Space; Extrahepatic; Eye; Eye Development; Factor V; Family; Functional disorder; Gastrointestinal Hemorrhage; Generations; Genes; Genetic; Goals; Half-Life; Hepatic; Hepatic Tissue; Hepatocyte; Heritability; Homeostasis; Human; Hypertension; In Vitro; Individual; Inherited; Injections; Intermittent Claudication; Liver; Mediating; Metabolism; Morbidity - disease rate; Mus; Mutation; Myocardial Infarction; Nucleotides; Oral Administration; Patients; Pharmacological Treatment; Plasma; Production; Progressive Disease; Proteins; Pseudoxanthoma Elasticum; Regimen; Replacement Therapy; Research; Residual state; Retina; Role; Skin; Solid; Stroke; Supplementation; Testing; Therapeutic; Transgenic Organisms; Visual Acuity; calcification; calcium phosphate; ecto-nucleotidase; effective therapy; extracellular; high risk; in vivo; in vivo Model; inhibitor; inorganic phosphate; member; mineralization; mouse model; novel therapeutic intervention; pharmacologic; plasma cell membrane glycoprotein PC-1; pre-clinical; prevent; prototype; tripolyphosphate

This research will focus on pseudoxanthoma elasticum (PXE), the prototype of heritable ectopic mineralization disorders. Aberrant calcium phosphate deposition in PXE results in characteristic cutaneous findings which signify the potential for development of ocular complications leading to loss of visual acuity and blindness, as well as cardiovascular involvement, including nephrogenic hypertension, intermittent claudication, bleeding from gastrointestinal arteries, early myocardial infarct and stroke. There is no effective treatment for PXE and the disease therefore progresses after diagnosis. PXE is caused by mutations in the ABCC6 gene which encodes ABCC6, an efflux transporter expressed primarily in the liver. Our group elucidated the pathophysiology underlying PXE, by showing that ABCC6 in hepatocytes mediates release of ATP into the blood circulation. Released ATP is extracellularly converted by the ecto-nucleotidase ENPP1 into 1 molecule of AMP and 1 molecule of the potent mineralization inhibitor inorganic pyrophosphate (PPi). PXE patients bearing defective ABCC6 have low PPi plasma concentrations, explaining their progressive calcification of soft connective tissues. Plasma PPi homeostasis is maintained through a complex network of proteins involved in ATP release, and ecto-nucleotidases metabolizing the released nucleotides. ENPP1 is crucial in this respect, as its activity underlies all PPi present in plasma. The ~40% of residual PPi present in plasma of PXE patients is formed out of ATP released by cells independent of ABCC6. Extracellular ATP is also subject to metabolism by other ecto- nucleotidases and the most important competitor of ENPP1 in the circulation is ENTPD1, which converts ATP into 1 molecule of AMP and 2 molecules of inorganic phosphate. Importantly, ENPP1 and ENTPD1 have opposing effects on extracellular PPi homeostasis and, consequently, ectopic calcification. In this study we propose to test the hypothesis that inhibition of ENTPD1 enhances extracellular PPi formation and prevents ectopic calcification in PXE patients We have developed two Specific Aims to test our hypothesis: (1) Thoroughly characterize how competition between ENPP1 and ENTPD1 affects extracellular PPi homeostasis in vitro and (2) Determine the effect of ENTPD1 on plasma PPi concentrations and ectopic calcification in Abcc6-/- mice in vivo. These plans are based on solid preliminary data, and they take advantage of a well-characterized mouse model developed and characterized in our department – the Abcc6-/- mouse –, which recapitulates the clinical, histopathologic, ultrastructural and genetic features of PXE in humans. Collectively, our state-of-the-art studies utilizing both in vitro and in vivo model systems are expected to provide critical preclinical information of the therapeutic potential of ENTPD1 inhibition to restore plasma PPi levels in PXE, with subsequent inhibition of ectopic calcification. Our findings are expected to not only inform development of pharmacologic treatments for PXE, but also for other ectopic calcification disorders, for which no effective or specific therapy is currently available.

本研究将集中在弹性假黄瘤（PXE），遗传性异位矿化的原型 紊乱磷酸钙异常沉积导致特征性皮肤表现， 表明有可能发生导致视力丧失和失明的眼部并发症， 以及心血管受累，包括肾源性高血压、间歇性跛行、 胃肠道动脉，早期心肌梗死和中风。目前还没有有效的治疗方法， 因此，疾病在诊断后继续发展。PXE是由ABCC 6基因突变引起的，该基因编码 ABCC 6，主要在肝脏中表达的外排转运蛋白。我们小组阐明了 通过显示肝细胞中的ABCC 6介导ATP释放到血液循环中，研究了潜在的PXE。 释放的ATP在细胞外被外核苷酸酶ENPP 1转化为1分子AMP和1分子ATP。 有效的矿化抑制剂无机焦磷酸盐（PPi）分子。携带缺陷型PXE患者 ABCC 6的PPi血浆浓度较低，这解释了其软结缔组织的进行性钙化。 血浆PPi稳态通过参与ATP释放的复杂蛋白质网络来维持，并且 代谢释放的核苷酸的外核苷酸酶。ENPP 1在这方面至关重要，因为它的活动 是血浆中所有PPi的基础。存在于PXE患者血浆中的约40%的残留PPi形成于 ATP的释放不依赖于ABCC 6。细胞外ATP也受到其他外- 在循环中ENPP 1的最重要竞争者是ENTPD 1，它将ATP 转化为1分子AMP和2分子无机磷酸盐。重要的是，ENPP 1和ENTPD 1具有 对细胞外PPi稳态的相反影响，并因此导致异位钙化。本研究 提出检验抑制ENTPD 1增强细胞外PPi形成的假设， 预防PXE患者的异位钙化我们制定了两个特定目标来验证我们的假设： (1)彻底表征ENPP 1和ENTPD 1之间的竞争如何影响细胞外PPi （2）确定ENTPD 1对血浆PPi浓度和异位妊娠的影响。 在Abcc 6-/-小鼠体内的钙化。这些计划是基于坚实的初步数据，他们利用 在我们部门开发和表征的良好表征的小鼠模型-Abcc 6-/-小鼠-中， 其概括了人类PXE的临床、组织病理学、超微结构和遗传学特征。 总的来说，我们利用体外和体内模型系统的最新研究有望提供 ENTPD 1抑制恢复血浆PPi水平的治疗潜力的关键临床前信息， PXE，随后抑制异位钙化。我们的发现不仅可以为发展提供信息， 的药物治疗，但也用于其他异位钙化疾病，对于这些疾病没有有效或 目前已有特效疗法。