In Vitro Model of the Human Alveolar Epithelium

描述（由申请人提供）： 远端气道具有多种特征，这使得它们成为全身递送治疗剂的有吸引力的部位。这些包括非常薄的上皮、大的表面积、避免肝脏首过代谢和广泛的血管化。因此，肺部递送越来越多地被认为是候选药物的替代方案，候选药物在胃肠道中不稳定或吸收不良（例如蛋白质或肽）。据估计，数百种生物工程蛋白质和肽目前已经上市或正在进行临床研究。此外，即使是小分子药物也可以受益于通过肺泡上皮可能实现的几乎瞬时的递送。因此，一个强大的，市售的肺泡转运和毒性的体外模型可以显着有助于新的治疗产品的开发。然而，目前并不存在这样的模式。目前的拨款提案的目标是通过开发和生产一种商业化的人肺泡上皮细胞体外模型来满足这一未满足的需求。在进行更昂贵、耗时的体内动物研究之前，该模型将是一种有用的筛选/研究工具，从而减少了所进行的动物研究的数量和范围。除了临床前药物开发期间的运输和毒理学应用外，该模型还可用于与环境毒理学相关的应用（例如石棉，柴油机尾气，香烟烟雾，臭氧等）。或与人类肺部疾病如哮喘相关的基础研究应用。将在微孔膜插入物上培养正常人小气道上皮细胞（SAEC），以产生人肺泡上皮的体内样模型。在第I阶段，将通过包括光学显微镜、透射电子显微镜、跨上皮电阻、calveolin-1、细胞粘附分子和细胞粘附分子的方法在适当的细胞结构和功能方面表征模型。（AT1标志物），P-糖蛋白（AT 1标记物），表面活性蛋白-C（AT2标记物）和肽转运蛋白PEPT 2（AT2标记）表达，凝集素结合（AT 1和AT 2特异性），对各种分子量的模型亲水性药物、肽和P-糖蛋白底物的渗透性。将在II期研究中根据ICCVAM指南对模型进行长期批间重现性和多实验室验证。

DESCRIPTION (provided by applicant): The distal airways possess a variety of features, which make them an attractive site for systemic delivery of therapeutics. These include a very thin epithelium, large surface area, avoidance of hepatic first pass metabolism and extensive vascularization. Pulmonary delivery is therefore being increasingly considered as an alternative for drug candidates, which are either unstable or poorly absorbed in the gastrointestinal tract (e.g. protein or peptides). It is estimated that hundreds of bioengineered proteins and peptides are currently either already on the market or are undergoing clinical investigation. Furthermore, even small molecule drugs can benefit from the nearly instantaneous delivery potentially achievable through the alveolar epithelium. Thus, a robust, commercially available in vitro model of alveolar transport and toxicity could significantly contribute to the development of new therapeutic products. However, such a model does not currently exist. The goal of the present grant proposal is to fulfill this unmet need by developing and producing a commercial in vitro model of the human alveolar epithelium. This model will be a useful screening/research tool prior to more expensive, time-consuming in vivo animal studies, reducing the number and scope of animals studies conducted. In addition to transport and toxicology applications during preclinical drug development, the model will find utility for applications related to environmental toxicology (e.g. asbestos, diesel exhaust, cigarette smoke, ozone, etc.) or basic research applications related to human lung diseases such as asthma. Normal human small airway epithelial cells (SAEC) will be cultured on microporous membrane inserts to produce an in vivo-like model of the human alveolar epithelium. During Phase I, the model will be characterized in terms of appropriate cell structure and function by methods including light microscopy, transmission electron microscopy, transepithelial electrical resistance, calveolin-1 (AT1 marker), P-glycoprotein (AT 1 marker), surfactant protein-C (AT2 marker) and peptide transporter PEPT2 (AT2 marker) expression, lectin binding (AT1 and AT2 specific), permeability to model hydrophilic drugs of various molecular weights, peptides and P-glycoprotein substrates. Longer-term lot-to-lot reproducibility and multi-laboratory validation of the model according to ICCVAM guidelines will be conducted in Phase II studies.