Bioengineering a novel therapeutic protein complex to minimize the effects of medical device induced hemolysis

生物工程新型治疗性蛋白质复合物可最大程度地减少医疗设备引起的溶血的影响

• 批准号: 10380296
• 负责人: Paul Werner Buehler
• 金额: $ 70.2万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-01-01 至 2025-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10380296
• 关键词: Acute Renal Failure with Renal Papillary Necrosis; Affinity; Animal Model; Animals; Attenuated; Binding; Binding Proteins; Binding Sites; Biological Assay; Biological Markers; Biomedical Engineering; Blood Circulation; Brain; Cardiac; Cardiac Output; Cardiac Surgery procedures; Cardiopulmonary Bypass; Cells; Cessation of life; Chemicals; Clinical; Complex; Complication; Cytolysis; Data; Dimerization; Dose; Drug or chemical Tissue Distribution; Effectiveness; Engineering; Erythrocytes; Extracorporeal Membrane Oxygenation; Fractionation; Functional disorder; Goals; Hamsters; Haptoglobins; Health Care Costs; Heart; Heme; Heme Iron; Hemeproteins; Hemoglobin; Hemoglobin concentration result; Hemolysis; Hemopexin; Hospitalization; Hour; Human; In Vitro; Individual; Injury to Kidney; Intestines; Iron; Kidney; Kinetics; Lead; Length of Stay; Ligands; Lung; Medical; Medical Device; Methods; Microvascular Permeability; Modeling; Monitor; Morbidity - disease rate; Operative Surgical Procedures; Organ; Outcome; Paste substance; Patients; Plasma; Procedures; Process; Proteins; Proteolysis; Public Health; Pump; Rattus; Renal function; Risk; Sampling; Secure; Self-Help Devices; Severities; Supportive care; System; Testing; Therapeutic; Therapeutic Intervention; Therapeutic Uses; Toxic effect; Toxin; Transferrin; Urine; Ventricular; apohemoglobin; base; blood pump; cell injury; clinically relevant; cost; cost estimate; design; dimer; experience; haptoglobin-hemoglobin complex; heart function; heme a; hemodynamics; improved; in vivo; individual patient; instrument; large scale production; lung injury; macrophage; monocyte; mortality; mortality risk; novel; novel therapeutics; organ injury; prevent; prospective; protein complex; proteostasis; pulmonary function; response; urinary

PROJECT SUMMARY Cost estimates for CNS, pulmonary, cardiac, and renal complications following medical assist devices requiring blood pumps, for example on pump cardiopulmonary bypass (CPB) is estimated at $80 million per individual states in the US over a ten-year period. Because of the need for extended CPB (> 4hours) use in complex surgeries, hemolysis, and the subsequent release of hemoglobin (Hb) into the circulation – which is known to be multi-organ toxic and vasoactive – is a relevant contributor to more intensive management of patients. To date, no effective strategy to remove toxic by-products of Hb are available. The burden caused by kidney injury alone accounts for approximately $9 billion/year, 300,000 deaths/year and an average increase in hospital stay of 3.5 days/patient This project focuses on understanding the contribution of hemolysis, Hb and heme associated with secondary end organ injury following extended on pump cardiopulmonary bypass (typically lasting up to 4 hours). To understand medical device related red blood cell lysis in this setting, we propose a strategy that is focused on sequentially understanding the contributions of Hb and heme and a therapeutic strategy to attenuate the end organ pathophysiology of these toxins. We will prospectively determine the levels of Hb, heme and iron as well as the concentrations of each toxin in plasma and urine of cardiac surgery patients. Simultaneously we will determine the concentrations of Hb, heme and iron binding and clearance proteins, haptoglobin, hemopexin and transferrin, respectively. Further we will determine plasma and urinary markers of end organ injury. We will use this data to construct a biokinetic model that determines the limits of Hb toxin concentrations that associate with end organ injury markers. The goal of this effort will be to define the need and timing for therapeutic interventions. To this end we have bioengineered a novel Hb, heme and protein scavenger based on our extensive experience with studying Hb toxicity. The novel protein construct is prepared by generating apo-Hb in multi-step process, while simultaneously isolating haptoglobin from Cohn fractionation paste IV. Finally, the two proteins are complexed and further purified to generate the apo-Hb-haptoglobin complex. This novel protein construct binds heme in high heme exposure states, secures iron in the heme ligand and safely clears the complex to monocytes and macrophages. Alternatively, in high Hb exposures apo-Hb exchanges binding sites on haptoglobin clearing Hb dimers to monocytes and macrophages, while released apo-Hb dimers are degraded by proteolysis and harmlessly cleared from circulation. To test the effect of the novel complex we have planed a range of proof-of-concept studies in animal models of on pump CPB to define the ability of apo- Hb-haptoglobin dosing to prevent end organ injury.

项目概要 医疗辅助设备使用后中枢神经系统、肺部、心脏和肾脏并发症的成本估算 需要血泵，例如泵体外循环 (CPB)，估计每台需要 8000 万美元 十年来美国各个州的情况。由于需要延长 CPB（> 4 小时）使用 复杂的手术、溶血以及随后将血红蛋白 (Hb) 释放到循环中 – 这是 已知具有多器官毒性和血管活性——是更强化管理的相关因素 患者。迄今为止，尚无有效的策略来去除 Hb 的有毒副产物。造成的负担 仅肾损伤每年就造成约 90 亿美元的损失，每年有 300,000 人死亡，平均 每个患者的住院时间增加 3.5 天 该项目的重点是了解 长期使用泵后与继发性终末器官损伤相关的溶血、血红蛋白和血红素 体外循环（通常持续长达 4 小时）。了解医疗器械相关红细胞 在这种情况下，我们提出了一种策略，重点是依次理解 血红蛋白和血红素以及减弱这些毒素的终末器官病理生理学的治疗策略。我们将 前瞻性测定 Hb、血红素和铁的水平以及每种毒素的浓度 心脏手术患者的血浆和尿液。同时我们将测定 分别为 Hb、血红素和铁结合和清除蛋白、触珠蛋白、血红素结合蛋白和转铁蛋白。 此外，我们将确定终末器官损伤的血浆和尿液标志物。我们将使用这些数据来构建 确定与终末器官损伤相关的 Hb 毒素浓度极限的生物动力学模型 标记。这项工作的目标是确定治疗干预的必要性和时机。对此 最后，我们根据丰富的经验，通过生物工程设计了一种新型 Hb、血红素和蛋白质清除剂 研究 Hb 毒性。通过多步过程生成apo-Hb来制备新型蛋白质构建体， 同时从 Cohn 分级分离糊 IV 中分离触珠蛋白。最后，这两种蛋白质是 复合并进一步纯化以生成apo-Hb-触珠蛋白复合物。这种新颖的蛋白质结构 在高血红素暴露状态下结合血红素，确保血红素配体中的铁，并安全地清除复合物 单核细胞和巨噬细胞。或者，在高 Hb 暴露中，apo-Hb 交换结合位点 结合珠蛋白将 Hb 二聚体清除为单核细胞和巨噬细胞，同时释放的 apo-Hb 二聚体被降解 通过蛋白水解并从循环中无害地清除。为了测试小说复合体的效果，我们有 计划在泵体外循环动物模型中进行一系列概念验证研究，以确定脱氧核糖核酸的能力 Hb-触珠蛋白剂量可预防终末器官损伤。