Bioengineering of Novel Synthetic Lipid-Peptide Lung Surfactants

描述(由申请人提供)：这项多学科生物工程研究伙伴关系(BRP)基金研究的是新型全合成脂质/多肽肺表面活性物质的分子生物工程、合成、表面活性和肺功效。还研究了使用合成表面活性剂来促进外源DNA输送到患有肺损伤的动物，以用于未来的基因治疗或多种药物治疗应用。BRP的主要目标是开发全合成的肺表面活性物质，与现有的动物来源或合成的临床表面活性药物相比，具有最大的活性、抗抑性、稳定性、纯度和生产经济性，用于治疗新生儿呼吸窘迫综合征(NRDS)、急性肺损伤(ALI)和急性呼吸窘迫综合征(ARDS)。在三所大学：罗切斯特大学(初级院校)、洛杉矶生物医学研究所/港湾-加州大学洛杉矶分校医学中心和圭尔夫大学，组建了一支经验丰富、协作的BRP团队，拥有生物工程、物理、化学、生物和医学方面的专业知识。BRP研究的合成肽包括含有人表面活性蛋白(SP)-B的关键分子特征的化合物，SP-B在天然表面活性物质中具有关键的功能活性。与人SP-C/SP-A相关的多肽和脂肽也被研究，并被生物工程改造为相对于天然载脂蛋白在分子稳定性方面具有优势。BRP还检测了两种类型的脂质：仿照天然表面活性物质的合成脂质(L)和具有增强吸附和扩散以及抵抗炎性肺损伤降解能力的新型磷脂酶抗性脂(RL)(ALI/ARDS)。目标1和目标2基于有希望的初始活性化合物的初步数据，研究合成肽/脂肽和新型RL化合物的生物工程、蛋白质组学、合成、纯化、分子生物物理和放大。目的3基于体外表面活性评价(脉动气泡、吸附、Wilhelmy平衡、圈养气泡)和动物肺活度研究，探讨合成表面活性剂中脂/肽组成的优化。所研究的动物模型包括：(I)FDA认可的用于评估当前临床上直接用于NRDS早产儿的临床表面活性物质的大鼠肺机械模型；(Ii)气管内滴注脂多糖(LPS)导致的在体ALI/ARDS小鼠；(3)严重高氧暴露导致的在体ALI/ARDS兔；以及(4)呼吸机治疗的肺表面活性物质缺乏和活体灌洗诱导的ALI/ARDS兔。目的研究合成表面活性物质的剪切粘度、肺组织分布和细胞毒性，以及它们在促进DNA肺转运到脂多糖小鼠中的作用，为未来ALI/ARDS的基因或多药治疗奠定基础。这项多学科的BRP拨款将加强对多肽和脂类分子行为的科学理解，同时利用工程、化学、物理、生物学、生理学和医学的原理和方法来生物工程合成具有最大活性、抗抑制性、肺功效和生产经济性的肺表面活性物质。公共卫生相关性：这项BRP研究将开发和生产具有最大活性和抗抑制性的新型全合成肺表面活性物质，用于未来治疗严重和普遍的人类肺部疾病，包括急性呼吸衰竭。生物工程表面活性剂将比现有的合成表面活性剂药物更具活性和抗抑制性，与现有的动物源性临床表面活性剂相比，在活性、抵抗力、纯度、重复性、稳定性和生产经济性方面也将具有显著的潜在优势。具体的治疗应用不仅包括早产儿的新生儿呼吸窘迫综合征(NRDS)，还包括临床急性肺损伤(ALI)和急性呼吸窘迫综合征(ARDS)，这些疾病每年在美国和世界各地影响数十万各个年龄段(从婴儿到成人)的患者。赠款中增加的研究与公众健康进一步相关，不仅检查了合成表面活性剂在改善呼吸衰竭方面的主要活性，还检查了它们促进DNA在肺部输送的能力，以用于未来治疗ALI/ARDS和其他肺部疾病的基因疗法或多种药物疗法。

DESCRIPTION (provided by applicant): This multidisciplinary bioengineering research partnership (BRP) grant studies the molecular bioengineering, synthesis, surface activity, and pulmonary efficacy of novel fully-synthetic lipid/peptide lung surfactants. Also studied is the use of synthetic surfactants to facilitate the delivery of exogenous DNA to animals with lung injury for future gene therapy or multi-drug therapy applications. The primary goal of the BRP is to develop fully-synthetic lung surfactants with maximal activity, inhibition resistance, stability, purity, and production economy compared to existing animal-derived or synthetic clinical surfactant drugs for treating the neonatal respiratory distress syndrome (NRDS), acute lung injury (ALI), and the acute respiratory distress syndrome (ARDS). A highly-experienced, collaborative BRP team with expertise in bioengineering, physics, chemistry, biology, and medicine is assembled at three universities: the University of Rochester (primary institution), LA Biomedical Research Institute/Harbor-UCLA Medical Center, and the University of Guelph. Synthetic peptides studied in the BRP include compounds incorporating key molecular features of human surfactant protein (SP)-B, which has crucial functional activity in native surfactant. Peptides and lipopeptides related to human SP-C/SP-A are also studied and are bioengineered to have advantages in molecular stability relative to native apoproteins. The BRP also examines two types of lipids: synthetic lipids (L) modeled after those in native surfactant, and novel phospholipase-resistant lipids (RL) having enhanced adsorption and spreading plus the ability to resist degradation in inflammatory lung injury (ALI/ARDS). Aims 1 and 2 study the bioengineering, proteomics, synthesis, purification, molecular biophysics, and scale-up of synthetic peptides/lipopeptides and novel RL compounds based on promising preliminary data on initial active compounds. Aim 3 investigates the optimization of lipid/peptide composition in synthetic surfactants based on surface activity assessments in vitro (pulsating bubble, adsorption, Wilhelmy balance, captive bubble) and pulmonary activity studies in animals. Animal models studied include: (i) an excised lavaged rat lung mechanical model that is FDA-accepted for evaluating current clinical surfactant drugs for direct use in premature infants with NRDS; (ii) mice with ALI/ARDS in vivo from intratracheal instillation of lipopolysaccharide (LPS); (3) rabbits with ALI/ARDS in vivo from severe hyperoxic-exposure; and (4) ventilated rabbits with surfactant-deficiency and ALI/ARDS induced by in vivo lavage. Aim 4 studies the shear viscosity, pulmonary distribution, and cytotoxicity of instilled synthetic surfactants, as well as their utility in facilitating the pulmonary delivery of DNA to LPS-mice for future applications involving gene- or multi-drug therapies for ALI/ARDS. This multidisciplinary BRP grant will enhance scientific understanding about the molecular behavior of peptides and lipids while using principles and methods of engineering, chemistry, physics, biology, physiology and medicine to bioengineer synthetic lung surfactants having maximum activity, inhibition resistance, pulmonary efficacy, and production economy. PUBLIC HEALTH RELEVANCE: This BRP research will develop and produce novel fully-synthetic lung surfactants having maximum activity and inhibition resistance for future use in treating severe and prevalent human pulmonary diseases involving acute respiratory failure. BRP surfactants will be bioengineered to be more active and inhibition-resistant than current synthetic surfactant drugs, and will also have significant potential advantages in activity, resistance, purity, reproducibility, stability, and production economy compared to existing animal-derived clinical surfactants. Specific therapeutic applications include not only the neonatal respiratory distress syndrome (NRDS) in premature infants, but also clinical acute lung injury (ALI) and the acute respiratory distress syndrome (ARDS) that affect hundreds of thousands of patients of all ages (infants to adults) each year in the United States and around the world. Added studies in the grant have further relevance to public health by examining synthetic surfactants not only for their primary activity in improving respiratory failure, but also for their ability to facilitate the pulmonary delivery of DNA for future gene therapy or multi-drug therapy approaches for treating ALI/ARDS and other lung diseases.