Towards a Miniaturized Human Liver Array for High-throughput Screening

描述（由申请人提供）：本申请涉及广泛的挑战领域（11）再生医学和特定的挑战主题11-EB-104：活体人体组织微阵列，旨在生成“复杂但模块化、硬化、标准化、简化并针对传统动物模型进行验证的器官型平台”。“动物研究被证明不足以预测人类肝脏的反应，主要是由于肝功能的显着物种特异性差异。因此，在过去的三十年中，已经开发了大量的体外人类肝脏模型来补充动物试验。在目前可用的几种肝脏模型中，那些利用原代肝细胞的模型在其预测体内观察到的各种人类反应的潜力与其在各种培养形式中使用的简单性之间取得了良好的平衡。然而，众所周知，原代肝细胞难以在常规模型中维持，因为它们的表型功能在从肝脏的天然微环境分离后几小时内显示出急剧下降。事实上，这些模型中的不稳定肝细胞已被证明是临床结果的不良预测因子。我们利用微加工技术和组织工程技术开发了一种具有精确的微观细胞结构和最佳基质相互作用的人肝模型，该模型在体外显示表型稳定性数周，而在常规培养中仅需数小时。在这里，我们建议进一步开发和优化这些微型人类肝脏培养物，并将其与小型化策略和分析技术相结合，以实现具有成本效益的高通量筛选（HTS）应用。由于药物性肝损伤（DILI）是急性肝衰竭和药物高损耗率的主要原因，我们将优化我们的小型化人肝脏，特别是用于基因型特异性和临床相关药物处置和偶联DILI的体外筛选。我们在这里开发的技术可能会在开发几类治疗化合物（药物，生物制剂），评估环境毒物的处置和损伤潜力，肝脏生理学和疾病的基础研究以及肝病的个性化医学中找到广泛的用途。在未来，微技术与组织工程的持续结合可能会刺激其他组织模型的发展，并将其整合到所谓的“芯片上的人类”中。 公共卫生相关性：该项目中提出的研究旨在开发用于高通量筛选应用的小型化人类肝脏微阵列，特别是用于评估药物处置和药物诱导的肝损伤，这对患者，监管机构和制药/生物技术行业来说是一个严峻的挑战。在未来，我们的小型化人类肝脏HTS系统可能会在药物开发管道中更早地消除有问题的化合物，以减少患者接触不安全药物。我们在这里开发的技术也可能在评估环境毒物的损伤潜力，基础研究和肝病患者的个性化医疗中找到实用性。

DESCRIPTION (provided by applicant): This application addresses broad Challenge Area (11) Regenerative Medicine, and specific Challenge topic, 11-EB-104: Living Human Tissue Microarrays, aimed at generating "organotypic platforms that are complex yet modular, hardened, standardized, simplified, and validated against traditional animal models." Animal studies are proving to be insufficient for predicting human liver responses primarily due to significant species-specific differences in liver functions. Therefore, a plethora of in vitro human liver models have been developed over the last three decades to supplement testing on animals. Of the several liver models currently available, those utilizing primary hepatocytes strike a good balance between their potential to predict the diverse human responses seen in vivo and their simplicity of use in various culture formats. However, primary hepatocytes are notoriously difficult to maintain in conventional models as their phenotypic functions display a precipitous decline within a few hours after isolation from the native microenvironment of the liver. Indeed, unstable hepatocytes in these models have been shown to be poor predictors of clinical outcomes. We have utilized microfabrication technologies and tissue engineering techniques to develop a human liver model with precise microscale cytoarchitecture and optimal stromal interactions that displays phenotypic stability for several weeks in vitro as compared to a few hours in conventional cultures. Here, we propose to further develop and optimize these microscale human liver cultures and couple them with miniaturization strategies and assay technologies for cost-effective high-throughput screening (HTS) applications. Since drug-induced liver injury (DILI) is a leading cause of acute liver failures and the high attrition rate of pharmaceuticals, we will optimize our miniaturized human livers specifically for the in vitro screening of genotype-specific and clinically- relevant drug disposition and coupled DILI. The technologies we develop here may find broad utility in the development of several classes of therapeutic compounds (drugs, biologics), in evaluating the disposition and injury potential of environmental toxicants, in fundamental investigations of liver physiology and disease, and in personalized medicine for liver disease. In the future, continued combination of microtechnology with tissue engineering may spur the development of other tissue models and their integration into the so-called 'human- on-a-chip'. PUBLIC HEALTH RELEVANCE: The studies proposed in this project are aimed towards developing a miniaturized human liver microarray for high-throughput screening applications, specifically for evaluating drug disposition and drug- induced liver injury, a serious challenge for patients, regulatory agencies and the pharmaceutical/biotech industry. In the future, our miniaturized human liver HTS system may eliminate problematic compounds much earlier in the drug development pipeline towards reducing patient exposure to unsafe drugs. The technologies we develop here may also find utility in assessing the injury potential of environmental toxicants, in basic research, and in personalized medicine for patients with liver disease.