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.

这项研究将重点关注弹性假黄瘤（PXE），它是可遗传的异位矿化的原型 失调。 PXE 中异常的磷酸钙沉积会导致特征性皮肤表现， 表示有可能发生眼部并发症，导致视力丧失和失明，如 以及心血管受累，包括肾源性高血压、间歇性跛行、出血 胃肠动脉、早期心肌梗塞和中风。 PXE 尚无有效治疗方法， 因此，疾病在诊断后会进展。 PXE 是由 ABCC6 基因突变引起的，该基因编码 ABCC6，一种主要在肝脏中表达的外排转运蛋白。我们小组阐明了病理生理学 PXE 的基础，通过显示肝细胞中的 ABCC6 介导 ATP 释放到血液循环中。 释放的 ATP 在细胞外被核酸外切酶 ENPP1 转化为 1 分子 AMP 和 1 分子 有效矿化抑制剂无机焦磷酸盐 (PPi) 的分子。 PXE 患者轴承有缺陷 ABCC6 的 PPi 血浆浓度较低，这解释了它们的软结缔组织进行性钙化。 血浆 PPi 稳态是通过参与 ATP 释放的复杂蛋白质网络来维持的，并且 外切核苷酸酶代谢释放的核苷酸。 ENPP1 在这方面至关重要，因为它的活性 是血浆中所有 PPi 的基础。 PXE 患者血浆中约 40% 的残留 PPi 被形成 细胞释放的 ATP 独立于 ABCC6。细胞外 ATP 也受到其他胞外物质的代谢 循环中的核酸酶和 ENPP1 最重要的竞争对手是 ENTPD1，它可以转化 ATP 分解为1分子AMP和2分子无机磷酸盐。重要的是，ENPP1 和 ENTPD1 有 对细胞外 PPi 稳态的相反作用，从而导致异位钙化。在这项研究中我们 提议检验抑制 ENTPD1 会增强细胞外 PPi 形成的假设 预防 PXE 患者的异位钙化 我们制定了两个具体目标来检验我们的假设： (1) 全面表征ENPP1和ENTPD1之间的竞争如何影响细胞外PPi (2) 确定 ENTPD1 对血浆 PPi 浓度和异位的影响 Abcc6-/- 小鼠体内钙化。这些计划基于可靠的初步数据，并且利用了 我们部门开发和表征的良好表征的小鼠模型 - Abcc6-/- 小鼠 -， 它概括了人类 PXE 的临床、组织病理学、超微结构和遗传特征。 总的来说，我们利用体外和体内模型系统的最先进的研究预计将提供 ENTPD1 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。我们的研究结果不仅有望为发展提供信息 PXE 的药物治疗方法，以及其他异位钙化疾病的药物治疗方法，但目前尚无有效或有效的方法 目前已有具体治疗方法。