Optimization of Novel Small Molecules to Antagonize FGF-23

FGF-23是一种骨源性磷酸盐和维生素D调节激素，可激活FGFR/α-Klotho二元 肾脏中的复合物。循环FGF-23增加导致X连锁（XLH）和常染色体隐性遗传（ARH） 低磷酸盐血症以及其它遗传性和获得性低磷酸盐血症病症。适应性增长 在FGF-23中，也维持慢性肾病（CKD）中磷酸盐和维生素D稳态， 心血管（CV）死亡率增加。KRN 23，最近批准的FGF-23阻断抗体， 治疗XLH，受到半衰期长、需要全身给药、剂量滴定困难和潜在的 可能过度抑制FGF-23。由于潜在的毒性，KRN 23未被批准用于治疗升高的FGF-23 在CKD中。有机会开发可滴定的，口服生物可利用的，短效的小分子， 明显抑制FGF-23与FGFR/α-Klotho复合物的结合。我们的核心假设是一个小分子 FGF-23拮抗剂可以开发出更灵活的剂量滴定和更短的半衰期，这将是预治疗。 延迟治疗低磷酸盐血症疾病，并可能扩大治疗适应症，以防止FGF-2。 23例CKD介导的CV并发症。使用计算的，基于结构的高通量筛选，我们确定- 确定了治疗先导化合物MD-3和几种类似物。我们的目标是将原型线索发展成 用于治疗由FGF-23过量引起的病症的临床前候选药物。我们的具体目标是： 1)优化新型FGF-23拮抗剂的效力。我们将阐明构效关系 (SAR)小分子FGF-23拮抗剂，以增加其效力。IC 50 < 500的几种化合物 与在Aim 2中检查的可药用性的FGF阻断抗体相比，ηM和%最大响应> 75%。（二） 进行体外吸收、分布、代谢和排泄（ADME）、药代动力学（PK）和 毒性筛查我们将进行早期体外ADME筛选，以确定符合最佳阈值的化合物。 奥尔兹（溶解度> 10 µM，在人肝微粒体中的稳定性，t1/2> 30 min，渗透性1.0 1 E-6cm/s， 蛋白质结合（血浆，人）< 98%），随后进行体外毒性/安全性筛选， (IC50 5种主要同工酶> 10 µM）、遗传毒性（艾姆斯）、心脏毒性（hERG结合IC 50> 10 µM）、细胞毒性（hERG结合IC 50> 10 µM）。 毒性（HepG 2，IC 50> 100 µM）和脱靶效应。通过体外毒性筛选的化合物将 推进到体内PK/暴露曲线（t1/2> 60 min）。3)在临床前模型中测试FGF-23拮抗剂 FGF-23过量。我们选择了2至3种符合最佳药物样性质的FGF-23拮抗剂，并测试了它们的活性。 治疗Hyp和Dmp 1缺失的XLH和ARH临床前小鼠模型的能力。我们还将确定FGF-23是否 拮抗剂可滴定以预防CKD小鼠模型中的CV并发症，而不诱导高磷酸化。 贫血我们的预期结果是鉴定具有优势的小分子FGF-23拮抗剂 超过了现有的生物制剂。我们的影响将是确定适合开发成新疗法的化合物 遗传性低磷酸盐血症疾病，并可能是一种新的方法，拮抗FGF-23在CKD。

FGF-23 is a bone-derived phosphate and vitamin D regulating hormone that activates FGFR/α-Klotho binary complexes in the kidney. Increased circulating FGF-23 causes X-linked (XLH) and autosomal recessive (ARH) hypophosphatemia, as well as other hereditary and acquired hypophosphatemic disorders. Adaptive increases in FGF-23 also maintain phosphate and vitamin D homeostasis in chronic kidney disease (CKD) and is associ- ated with increased cardiovascular (CV) mortality. KRN23, a recently approved FGF-23 blocking antibody for treating XLH, is limited by a long half-life, need for systemic administration, difficult dose titration, and the poten- tial to over-suppress FGF-23. Due to potential toxicity, KRN23 is not approved for treatment of elevated FGF-23 in CKD. There is an opportunity to develop titratable, orally bioavailable, short-acting small molecules that re- versibly inhibit FGF-23 binding to FGFR/α-Klotho complexes. Our central hypothesis is that a small molecule FGF-23 antagonist can be developed with a more flexible dose-titration and shorter half-life that will be the pre- ferred treatment of hypophosphatemic disorders, and may expand the therapeutic indication to preventing FGF- 23 mediated CV complications in CKD. Using a computational, structure-based high-throughput screen, we iden- tified a therapeutic lead compound, MD-3 and several analogs. Our goal is to develop prototypic leads into a preclinical drug candidate for the treatment of disorders caused by FGF-23 excess. Our Specific Aims are to: 1) Optimize the potency of novel FGF-23 antagonists. We will elucidate the structure-activity relationship (SAR) of small molecule FGF-23 antagonists to increase their potency. Several compounds with an IC50 < 500 ηM and % max response > 75% compared to an FGF-blocking antibody examined for druggability in Aim 2. 2) Perform in vitro absorption, distribution, metabolism and excretion (ADME), pharmacokinetic (PK) and toxicity screens. We will perform early in-vitro ADME screens to identify compounds that meet optimal thresh- olds (solubility > 10 µM, stability in human liver microsomes with a t ½ > 30 min, permeability 1.0 1E-6cm/s, protein binding (plasma, human) < 98%), followed by in-vitro toxicity/safety screens that evaluate CYP inhibition (IC50 > 10 µM for 5 major isozymes), genotoxicity (AMES), cardiac toxicity (hERG binding IC50 > 10 µM), cyto- toxicity (HepG2, IC50 > 100 µM), and off-target effects. Compounds that pass the in-vitro toxicity screen will advance to in vivo PK/exposure profiling (t ½ > 60 min). 3) Test FGF-23 antagonists in pre-clinical models of FGF-23 excess. We select 2 to 3 FGF-23 antagonists that meet the optimal drug-like properties and test their ability to treat Hyp and Dmp1 null pre-clinical mouse models of XLH and ARH. We will also determine if FGF-23 antagonists can be titrated to prevent CV complications in mouse models of CKD without inducing hyperphos- phatemia. Our expected outcomes are identification of small molecule FGF-23 antagonists with advantages over current biologicals. Our impact will be to identify compounds suitable for development into novel treatments of hereditary hypophosphatemic disorders, and possibly a new approach to antagonize FGF-23 in CKD.