Lead Optimization of Novel Inhibitors of Tissue Non-specific Alkaline Phosphatase

描述(由申请人提供)：本R01申请名为“组织非特异性碱性磷酸酶新型抑制剂的先导优化”，是对PAR-12-060“发现体内化学探针验证命中的征集”的响应。内侧血管钙化(MVC)是婴幼儿全身性动脉钙化(GACI)的主要发病和死亡原因，也是川崎病(KD)、慢性肾脏病(CKD)以及糖尿病、肥胖和老龄化等心血管疾病恶化的主要原因。矿化抑制剂胞外无机焦磷酸(Eppi)与其在这些条件下的动态平衡之间的相关性已被明确确立。Eppi水平的降低与组织非特异性碱性磷酸酶(TNAP)表达水平的升高有关，组织非特异性碱性磷酸酶可使Eppi水解性，从而消除其在MVC打击的组织中的抑制作用。例如，我们最近观察到血管平滑肌细胞(VSMC)和尿毒症主动脉中TNAP的上调，表明TNAP活性过高是Eppi缺乏和中膜钙化的重要原因。我们假设，在全身给药后，有效的小分子tnap抑制剂可能会导致tnap活性降低，导致局部eppi增加到 预防或改善血管钙化。因此，我们提出了一种策略，通过降低小分子TNAP抑制剂的TNAP活性来提高Eppi水平。我们最近报道了芳基磺酰胺衍生的TNAP抑制剂的表征和初步优化。这些化合物通过非竞争性机制发挥作用，已经准备好进行全面的化学优化，为体内概念验证研究提供准备的先导化合物。因此，我们的具体目标是：1)设计和合成具有体内口服活性的优化的TNAP抑制剂；2)在相关的体外试验中评估TNAP抑制剂的效力和选择性；3)使用体外ADME/T和体内药代动力学(PK)试验评价新型小分子TNAP抑制剂；以及4)在啮齿动物体外和体内血管钙化模型中表征领先的TNAP抑制剂探针。生成的TNAP抑制剂将提供强大的工具来测试通过调节TNAP活性来提高Eppi水平以防止MVC的假设，同时为未来开发治疗MVC引起的疾病家族的新型药物奠定基础。

DESCRIPTION (provided by applicant): This R01 application entitled "Lead Optimization of Novel Inhibitors of Tissue Non-specific Alkaline Phosphatase" is in response to PAR-12-060 "Solicitation of Validated Hits for the Discovery of in vivo Chemical Probes". Medial vascular calcification (MVC) is the major cause of morbidity and mortality in generalized arterial calcification of infancy (GACI), a severe childhood disease, and contributes to cardiovascular deterioration in Kawasaki disease (KD), chronic kidney disease (CKD), as well as in diabetes, obesity and aging. The relevance of the mineralization inhibitor extracellular inorganic pyrophosphate (ePPi) and its homeostasis in these conditions has been clearly established. Reduced levels of ePPi have been linked to elevated expression levels of tissue non-specific alkaline phosphatase (TNAP), which hydrolizes ePPi, thus eliminating its inhibitory effect in tissue stricken by MVC. For example, we have recently observed an upregulation of TNAP in vascular smooth muscle cells (VSMC) and also in uremic aortas, suggesting that excessive TNAP activity is an important cause of ePPi deficiency and medial calcification. We hypothesized that potent small molecule TNAP inhibitors are likely to cause a reduction in TNAP activity following systemic administration, resulting in an increase in the local amount of ePPi to prevent or ameliorate vascular calcification. Therefore, we propose a strategy for increasing ePPi levels by reducing TNAP activity with small molecule TNAP inhibitors. We recently reported the characterization and preliminary optimization of arylsulfonamide-derived inhibitors of TNAP. These compounds, which function via an uncompetitive mechanism, are ready for full-scale chemistry optimization to provide lead compounds ready for in vivo proof- of-concept studies. Therefore our Specific Aims are: 1) Design and synthesize optimized TNAP inhibitors that are orally active in vivo; 2) Assess the potency and selectivity of TNAP inhibitors in relevan in vitro assays; 3) Evaluate novel small molecule TNAP inhibitors using in vitro ADME/T and in vivo pharmacokinetic (PK) assays; and 4) Characterize lead TNAP inhibitor probes in rodent ex vivo and in vivo models of vascular calcification. The TNAP inhibitors generated will provide powerful tools for testing the hypothesis that enhancing ePPi levels by modulating TNAP activity protects against MVC, while laying a foundation for future development of a novel class of medications for the treatment of the family of diseases caused by MVC.