Enzyme Therapy for PXE: Breaking the Barrier of Ectopic Calcification

摘要 弹性纤维假黄瘤（PXE）是一种异位钙化的遗传性疾病， 由于结缔组织中羟基磷灰石晶体沉积导致死亡。虽然ABCC 6被鉴定为 作为20年前PXE的致病基因，其发病机制最近才被揭示， 目前对于病理性钙化仍然没有有效的或特异性的治疗。我们已经开发出了 这些小鼠为PXE的临床前开发提供了平台， 治疗这种目前难以治愈的疾病。PXE的一个重要病理特征是 无机焦磷酸盐（PPi）的循环水平，一个关键的内源性钙化抑制剂。因此 我们在此提出的研究目标是在临床前研究中使用我们的小鼠模型来开发安全且 通过使细胞外PPi水平正常化来预防PXE异位钙化的有效治疗。 我们已经确定ENPP 1和TNAP蛋白作为PPi稳态的关键调节因子。ENPP 1和TNAP 在维持细胞外PPi浓度方面具有相反的作用，前者产生PPi，后者产生PPi。 水解PPi。我们已经产生了重组ENPP 1酶生物制剂，我们强大的初步数据 证明这种治疗性生物制剂在PXE小鼠模型中升高血浆PPi水平，并且其循环 半衰期可以通过TNAP的药理学抑制来延长。根据这些发现和已知的 ENPP 1和TNAP的酶活性，我们建议调节血浆PPi，或者使用重组 ENPP 1酶、TNAP抑制剂或两种方法的组合代表了一种创新策略， 预防由于PPi缺乏而引起的异位钙化。为了验证这个假设，我们 我建议利用遗传学和药理学方法来确定抑制TNAP的机制， 延长来自ENPP 1酶补充的PPi的血浆半衰期，并随后预防和/或 减少了PXE小鼠模型中的异位钙化。我们的团队拥有必要的研究专长， ENPP 1-PPi-TNAP轴和合适的小鼠模型来完成这些研究。 总的来说，我们预计，拟议的研究将提供关键的翻译信息， 临床前方法，将允许开发新的治疗异位钙化的患者， PXE。如果成功的话，我们的发现将广泛地推进异位钙化的临床治疗， 缺乏在其他遗传性和获得性疾病中异位钙化的发展中起重要作用。

ABSTRACT Pseudoxanthoma elasticum (PXE) is a genetic disorder of ectopic calcification with considerable morbidity and mortality due to deposition of hydroxyapatite crystals in the connective tissues. Though ABCC6 was identified as the causative gene for PXE 20 years ago, the disease mechanism was just recently unveiled and there is currently still no effective or specific treatment for the pathologic calcification. We have previously developed and characterized mouse models for PXE, and these mice provide the platform for preclinical development of therapeutics for this currently intractable condition. A critical pathological characteristic in PXE is the reduction in circulating levels of inorganic pyrophosphate (PPi), a key endogenous inhibitor of calcification. Therefore, the goal of the research we propose herein is to use our mouse models in preclinical studies to develop safe and effective treatments that can prevent ectopic calcification in PXE by normalization of extracellular PPi levels. We have identified ENPP1 and TNAP proteins as key regulators of PPi homeostasis. ENPP1 and TNAP have opposing actions in maintaining extracellular PPi concentrations, the former generating PPi and the latter hydrolyzing PPi. We have generated a recombinant ENPP1 enzyme biologic and our strong preliminary data demonstrate that this therapeutic biologic raised plasma PPi levels in a mouse model of PXE, and its circulating half-life can be extended by pharmacologic inhibition of TNAP. Based upon these findings and the known enzymatic activities of ENPP1 and TNAP, we propose that modulation of plasma PPi, either using a recombinant ENPP1 enzyme, TNAP inhibitors, or a combination of both approaches, represents an innovative strategy to prevent the ectopic calcification that arises as a consequence of PPi deficiency. To test this hypothesis, we propose to utilize genetic and pharmacologic approaches to define mechanisms by which inhibition of TNAP extends the plasma half-life of PPi from ENPP1 enzyme supplementation, and subsequently prevents and/or diminishes the ectopic calcification in a mouse model of PXE. Our team has the requisite research expertise in the ENPP1-PPi-TNAP axis and appropriate mouse models to complete these studies. Collectively, we anticipate that the proposed studies will provide critical translational information from preclinical approaches that will allow development of novel treatments for ectopic calcification in patients with PXE. If successful, our findings will advance clinical management of ectopic calcification broadly, as PPi deficiency plays an important role in development of ectopic calcification in other genetic and acquired disorders.