Targeted Identification of Dual Acting Antisickling Agents for Sickle Cell Disease Therapy

我们的R61/R33提案的总体目标是设计和研究新型镰状细胞治疗剂 在某些实施方案中，本发明涉及抑制和/或不稳定初始低氧诱导的血红蛋白（Hb）聚合的疾病（SCD），从而 减少随后的继发性病理生理学，具有最小的毒性倾向。当缺氧时， 镰状血红蛋白聚合成长的、刚性的和不溶性的纤维，导致红细胞（RBC）呈镰状。该提案 利用调查小组的互补专业知识、承诺和经验;以及 令人信服的初步数据，以测试新的合成分子，直接不稳定的假设， 除了增加Hb对O2的亲和力之外，聚合物的形成将为SCD提供上级治疗选择。 具体目标是：1.设计、合成并进行体外功能和生物学评价。 新的香草醛衍生物。我们以前已经建立了新的抗锯齿衍生物，表现出显着的 与母体化合物相比的药理学改进。这一目标进一步利用了一个复杂的和知情的 衍生化先导化合物以增加直接聚合物去稳定化以及降低 化合物与Hb结合的化学计量。我们将对这些化合物进行一系列筛选试验， 评价体外功能和生物学特性，包括Hb修饰、细胞镰状化抑制、 血红蛋白的亲和力为O2，和X射线晶体学，以阐明其原子与血红蛋白的相互作用。目标1研究将 鉴定上级候选物并告知进一步的结构修饰以增强随后目标的效力 R61阶段的2项研究。2：建立Hb结合动力学、体外代谢和ADME，以及 所选化合物的初步体内药效学性质。在R61的结论中， 在此阶段，我们将对目标1中的选定候选人进行进一步的验证研究。具体来说，我们将 表征其分配到RBC隔室中的效率和Hb结合动力学，体外代谢 和ADME/安全性特征，以及初步体内药效学特征。计划研究的结果 将确定并验证上级先导分子，以便在目标3（R33）中进一步开发 阶段）。3：进行体内功能和生物学研究，以确定有前景的先导化合物， 发展目标3（R33）将侧重于野生型和SCD小鼠的体内PK/PD疗效研究。我们将 进行优化先导分子的放大合成，优化配方，并正式进行详细说明- 体内PK/PD和疗效研究，包括Hb体内修饰为非镰状化、高亲和力变体; 在常氧和缺氧下减少循环镰状细胞;改善溶血，炎症， 内皮损伤;以及在该模型中观察到的SCD病理生理学的总体逆转。小说 预期化合物在降低的剂量下表现出增强的功效。我们将与加速器合作 从我们现有的候选人名单中剔除我们将获得所需的成本匹配资金， 执行此阶段并将潜在客户推进到开发阶段所需的成本。

Our overall objective of this R61/R33 proposal is to design and study novel therapeutic agents for sickle cell disease (SCD) that inhibit and/or destabilize the initial, hypoxia-induced hemoglobin (Hb) polymerization, thereby reducing the subsequent secondary pathophysiology, with minimal liability for toxicity. When deoxygenated, sickle Hb polymerizes into long, rigid, and insoluble fibers, causing red blood cells (RBCs) to sickle. The proposal leverages the complementary expertise, commitment, and experience of the investigative team; and a compelling body of preliminary data to test the hypothesis that novel synthetic molecules that directly destabilize polymer formation, in addition to increasing Hb affinity for O2, will provide a superior therapeutic option for SCD. The specific aims are: 1: Design, synthesize and conduct in-vitro functional and biological assessment of novel vanillin derivatives. We have previously established novel antisickling derivatives that exhibit significant pharmacologic improvement over their parent compounds. This aim further utilizes an intricate and informed strategy to derivatize lead compounds to increase direct polymer destabilization, as well as decrease stoichiometry of compound binding to Hb. We will subject the compounds to a battery of screening assays to evaluate in-vitro functional and biological properties, to include Hb modification, inhibition of cell sickling, effect on Hb affinity for O2, and X-ray crystallography to elucidate their atomic interactions with Hb. Aim 1 studies will identify superior candidates and inform further structural modifications to enhance potency for subsequent Aim 2 studies of the R61 phase. 2: Establish Hb binding kinetics, in-vitro metabolism and ADME, and preliminary in-vivo pharmacodynamics properties of select compounds. In this concluding Aim of the R61 phase, we will conduct further validation studies on select candidates from Aim 1. Specifically, we will characterize their efficiency of partitioning into the RBC compartment and Hb binding kinetics, in-vitro metabolic and ADME/safety profiles, and preliminary in-vivo pharmacodynamic profiles. Results from the planned studies will firmly and conclusively identify and validate superior lead molecule(s) for further development in Aim 3 (R33 Phase). 3: Conduct in-vivo functional and biological studies to establish promising lead compounds for development. Aim 3 (R33) will focus on in-vivo PK/PD efficacy studies in wild-type and SCD mice. We will conduct scale-up synthesis of optimized lead molecules, optimize formulation, and formally conduct detailed in- vivo PK/PD and efficacy studies that include in-vivo modification of Hb to the non-sickling, high affinity variant; reduction in circulating sickled cells under normoxia and hypoxia; amelioration of hemolysis, inflammation, endothelial damage; and overall reversal of the SCD pathophysiology observed in this model. The novel compounds are expected to exhibit enhanced efficacy at reduced doses. We will collaborate with accelerator partners from our current list of potential candidates. We will obtain required cost-matching funds to defray the costs required to execute this phase and advance the lead(s) into the development phase.