Applying a Human Liver Microphysiology System to Develop Therapeutic Strategies for Non-Alcoholic Fatty Liver Disease (NAFLD)

我们建议在定量系统药理学方法中应用四种互补技术来 建立人类非酒精性脂肪性肝病(NAFLD)的实验模型，这是增长最快的 疾病，并使用该模型来测试新的治疗策略：1)实施血管化的肝腺泡 人肝细胞微生理系统(VLAMPS)的实验研究 总结早期NAFLD表型的模型，并作为实验测试新疗法的平台；2) 基于我们在计算和系统生物学方面的经验，我们将使用来自Normal和 NAFLD患者推断疾病进展的途径，以确定潜在的分子蛋白靶点 都在推断的路径中，并使用我们的潜在因素建模方法和3D相似性模型来 确定在统计上与这些途径中的目标相互作用的药物；3)我们将使用我们的高效 结合基因编辑产生IPSC来源的成熟肝细胞以整合疾病的过程 将IPSC肝细胞(有条件地获得/丧失功能)植入vLAMPS，开始测试患者 具体疗法；以及4)应用表型药物筛选技术。 NAFLD包括从单纯性脂肪变性(NAFL)到更严重的非脂肪肝的一系列肝脏损害 酒精性脂肪性肝炎(NASH)、肝硬变和肝细胞癌(HCC)。肝硬变和肝细胞癌 对肝脏的进行性损害已成为肝脏移植的第三大常见原因。这个 NAFLD的发病机制是复杂的，并被相当大的个体差异所混淆 疾病易感性、进展和并发症，提示有必要采取针对患者的方法。 研究已经确定了NAFLD相关的基因特征和单核苷酸多态性(SNPs)。在……里面 尤其是在NAFLD中下调的SIRT1基因，已被确定为 脂肪生成、糖异生、内质网应激、脂肪酸氧化、尿素循环和抗氧化反应 肝细胞。PNPLA3基因中存在一个单核苷酸多态现象 与肝脏脂肪变性、纤维化、肝硬变和肝细胞癌有关。然而，在以下方面仍然存在重大差距 我们对NAFLD发病机制的认识。例如，尽管它与NAFLD密切相关，但 PNPLA3变异体的功能意义尚不清楚。机械论解释中的一个主要限制 PNPLA3在NAFLD中的作用在于其表达的种间差异和组织特异性 分布，表明有必要建立人体细胞模型。 这些技术和方法的这种结合有望产生新的发展战略 重新调整用途和新的治疗方法，有可能减缓或阻止早期NAFLD向 晚期，危及生命的阶段。

We propose to apply four complementary technologies in a Quantitative Systems Pharmacology approach to create a human experimental model of non-alcoholic fatty liver disease (NAFLD), the most rapidly growing disease, and to use the model to test novel therapeutic strategies:1) Implement a vascularized, liver acinus microphysiological system (vLAMPS) constructed with human patient-derived, liver cells, as an experimental model to recapitulate early NAFLD phenotypes and as a platform to experimentally test novel therapeutics; 2) Building on our experience in computational and systems biology, we will use RNAseq data from normal and NAFLD patients to infer pathways of disease progression, to identify the potential molecular protein targets that are in the inferred pathways, and to use our latent factor modeling approach and 3D similarity models to identify drugs that statistically interact with the targets in these pathways; 3) We will employ our highly efficient processes for generating mature iPSC-derived hepatocytes combined with gene editing to incorporate disease engineered iPSC hepatocytes (conditional gain/loss of function) into the vLAMPS to begin testing patient specific therapies; and 4) Apply phenotypic drug screening technologies. NAFLD encompasses a spectrum of liver damage ranging from simple steatosis (NAFL) to more serious non- alcoholic steatohepatitis (NASH), cirrhosis and hepatocellular carcinoma (HCC). Cirrhosis and HCC resulting from progressive damage to the liver have become the third most common causes of liver transplants. The disease pathogenesis of NAFLD is complex and confounded by the considerable inter-individual differences in disease susceptibility, progression and complications, suggesting the need for a patient specific approach. Studies have identified NAFLD associated gene signatures and single nucleotide polymorphisms (SNPs). In particular, the SIRT1 gene that is downregulated in NAFLD, has been identified as a key regulator of lipogenesis, gluconeogenesis, ER stress, fatty acid oxidation, urea cycle and the antioxidant response in hepatocytes. A SNP in the patatin-like phospho-lipase domain-containing 3 (PNPLA3) gene is strongly associated with hepatic steatosis, fibrosis, cirrhosis, and HCC. However, there continues to be major gaps in our understanding of the pathogenesis of NAFLD. For example, despite its strong association with NAFLD, the functional significance of the PNPLA3 variant is unknown. A major limitation in the elucidation of a mechanistic role of PNPLA3 in NAFLD has been the interspecies differences in its expression and tissue-specific distribution, suggesting the need for human cell models. This combination of the technologies and approaches is expected to lead to new strategies for development of repurposed and new therapeutics with the potential to slow or halt the progression of early NAFLD to the more advanced, life threatening stages.