Hemoglobin Modifiers for Sickle Cell Disease Therapy

描述（由申请人提供）：我们的目标是设计和建立镰状细胞病（SCD）的新型治疗药物，即抑制初始镰状血红蛋白（HbS）的药物。 聚合和随后的病理生理学。当脱氧时，HbS聚合成长的、刚性的和不溶性的纤维，导致红细胞（RBC）镰刀状，这一过程由于HbS对氧的不寻常的低亲和力而恶化，导致氧的过早释放。基于几个证据-我们的初步数据，其他人的研究，以及我们的先导化合物5-HMF（Hb的变构效应物，AEH）最近完成的I/II期临床试验的结果-我们假设AEH不仅可以防止HbS聚合，还可以减轻几种继发性镰状化相关的病理事件，包括炎症，氧化应激/损伤，RBC溶血和疼痛。我们也有初步证据表明，我们的下一代AEH（INN和TD系列）表现出增强的效力和改善的体外作用持续时间。这些AEH通过一种新的作用机制发挥作用，即，除了增加Hb对氧的亲和力之外，还使HbS聚合物接触不稳定;提供积极的协同效应。 我们建议通过进一步研究来自INN和TD系列的候选药物以及5-HMF和INN-312的衍生物的药理学性质来测试我们的假设，重点关注二级SCD途径，以及使用我们的模型系统的潜在HbS聚合问题。具体目标是：1.新型血红蛋白变构效应物的设计与合成。我们将修改我们的母体化合物和合成衍生物，提高疗效和延长半衰期。我们还将根据需要合成保护活性醛功能部分免受醛脱氢酶（ALDH）介导的代谢的产品。2.研究新型AEH的体外功能、抗镰状化和细胞毒性活性。我们将研究AEH的体外抗镰状化/功能活性（RBC镰状化试验、P50分析、HbS溶解度和Hb加合物形成），并仔细监测不良反应。3.测定体内/体外PK/PD特性、结合和代谢，并评价AEH在SCD Berkeley小鼠中的临床前疗效。我们将表明，血清白蛋白结合和/或代谢ALDH在红细胞或肝细胞质不太可能产生不利影响，在体内药理活性。我们将使用SCD转基因小鼠的Berkeley小鼠模型证明AEH显示出有效的药理作用，增加小鼠的短期和长期存活率。我们还将研究其潜在的有益作用，例如，溶血、炎症、内皮损伤的改善以及在该模型中观察到的SCD病理生理学的总体逆转。4.确定AEH和Hb之间的原子相互作用。X射线晶体学将被用来验证我们的假设，AEH的效力是直接依赖于他们的能力，结合血红蛋白与他们的吡啶基取代基对表面的血红蛋白分子。这些结构将提供有价值的见解，以帮助指导合理的修改，以获得更好的药理学特性。

DESCRIPTION (provided by applicant): Our goal is to design and establish novel therapeutic agents for sickle cell disease (SCD), namely drugs that inhibit the initial sickle hemoglobin (HbS) polymerization and the subsequent pathophysiology. When deoxygenated, HbS polymerizes into long, rigid, and insoluble fibers causing red blood cells (RBCs) to sickle, a process worsened by the unusual low affinity of HbS for oxygen, resulting in premature release of oxygen. Based on several evidence--our preliminary data, studies by others, and the results of a recently completed phase I/II clinical testing of our lead compound, 5-HMF (an allosteric effector of Hb, AEH)-we hypothesize that AEHs, not only prevent HbS polymerization, but also mitigates several secondary sickling related pathological events that include inflammation, oxidative stress/damage, RBC hemolysis, and pain. We also have preliminary evidence that our next generation AEHs (INN- and TD-series) exhibit enhanced potency and improved in-vitro duration of action. These AEHs act via a novel mechanism of action, i.e., destabilize HbS polymer contacts, in addition to increasing Hb affinity for oxygen; providing positive synergistic effects. We propose to test our hypothesis by further investigating candidate drugs from the INN- and TD-series, as well as derivatives of 5-HMF and INN-312 for their pharmacologic properties, focusing on the secondary SCD pathways, as well as the underlying HbS polymerization problem using our model systems. The specific aims are: 1. Design and synthesis of novel allosteric effectors of hemoglobin (AEHs). We will modify our parent compounds and synthesize derivatives with enhanced efficacy and prolonged half-lives. We will also synthesize prod rugs to protect the active aldehyde functional moiety from aldehyde dehydrogenase (ALDH)- mediated metabolism as necessary. 2. Investigate in-vitro functional, ant sickling, and cytotoxicity activities of novel AEHs. We will investigate the AEHs for their in-vitro ant sickling/functional activities (RBC sickling tests, P50 analyses, HbS solubility, and Hb adduct formation), and carefully monitor for adverse effects. 3. Determine in-vivo/in-vitro PK/PD properties, binding and metabolism and evaluate preclinical efficacy of AEHs in SCD Berkeley mice. We will show that serum albumin binding and/or metabolism by ALDH in RBC or hepatic cytosol are not likely to adversely affect in-vivo pharmacologic activity. We will demonstrate using a Berkeley mouse model of SCD transgenic mice that AEHs show potent pharmacologic effects, increase short- and long-term survival rates of mice. We will also study their potential beneficial effects, e.g. amelioration of hemolysis, inflammation, endothelial damage, and overall reversal of the SCD pathophysiology observed in this model. 4. Determine the atomic interactions between AEHs and Hb. X-ray crystallography will be used to validate our hypothesis that AEH potency is directly dependent upon their abilities to bind Hb with their pyridyl substituents toward the surface of the Hb molecule. The structures would provide valuable insight to help guide rational modifications for better pharmacologic properties.