Clearing intrahepatic lipids to aid in toxicity testing

清除肝内脂质以帮助毒性测试

• 批准号: 7537640
• 负责人: JOSEPH C. RUIZ
• 金额: $ 13.16万
• 依托单位: VESTA THERAPEUTICS, INC.
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-07-01 至 2010-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7537640
• 关键词: Acute; Address; Aftercare; Agonist; Alanine; Albumins; Animals; Aspartate Transaminase; Bezafibrate; Bile Acids; Biological Assay; Biological Preservation; Cell model; Cell physiology; Cells; Cellular Morphology; Characteristics; Cholic Acid; Cholic Acids; Choline; Clinical; Clinical Research; Condition; Cryopreservation; Cytochrome P450; Development; Enzymes; Excision; Excretory function; Fatty Acids; Fatty acid glycerol esters; GW0742; Gene Activation; Genes; Genetic Transcription; Goals; Hepatic; Hepatocyte; Housing; Human; Invasive; Lipids; Liver; Measures; Metabolism; Methodology; Modeling; Nuclear Receptors; Numbers; Obesity; Organ; Organ Procurements; Pan Genus; Pathway interactions; Patients; Peroxisome Proliferator-Activated Receptors; Pharmaceutical Preparations; Pharmacologic Substance; Physiological; Positioning Attribute; Production; Property; Proteins; Protocols documentation; Public Health; Purpose; Rate; Research; Research Personnel; Source; Staging; Technology; Testing; Therapeutic; Time; Toxic effect; Toxicity Tests; Transcriptional Activation; Triglycerides; United States; Urea; Very low density lipoprotein; absorption; cell transformation; density; drug development; drug discovery; fatty acid biosynthesis; fatty acid metabolism; human study; improved; improved functioning; intrahepatic; lipid biosynthesis; lipid metabolism; neoplastic cell; novel; oxidation; programs; success

DESCRIPTION (provided by applicant): Vesta Therapeutics is one of the leading procurers of non-transplantable human livers in the United States and has developed proprietary technologies for the isolation, cryopreservation, and regenerative application of isolated primary hepatocytes. As Vesta processes cells for our research programs, a significant number of surplus mature hepatocytes are collected, [which are made available to academic, clinical and pharmaceutical researchers. Primary human hepatocytes are valuable and crucial models used by pharmaceutical companies to test the absorption, distribution, metabolism, excretion (ADME) and toxicity (T) properties of novel drug candidates. However,] with the increasing rates of obesity in the United States, a high percentage of the non-transplantable organs offered by Organ Procurement Organizations [for research purposes] are steatotic (fatty), which adds an uncontrollable layer of variability to the toxicity of potential test compounds. [As a consequence, pharmaceutical companies are reluctant to utilize steatotic hepatocytes, and are forced to use non-physiological animal and tumor cell models for drug development.] Research over the last few years has shown that a number of factors that target lipid biosynthesis, oxidation, or intrahepatic export can reverse steatosis in animals and in human patients. [However, the feasibility of applying these methodologies to primary human hepatocytes for the purpose of generating a standardized supply of non-steatotic hepatocytes has not been explored.] Vesta is uniquely positioned to assess these technologies given our consistent supply of steatotic livers and our extensive cryopreserved stocks of steatotic hepatocytes. We propose to decrease steatosis levels in human hepatocytes using bile acids, PPAR agonists, and choline singly or in combination, which have been shown to reverse steatosis in animal and human studies. We will document the optimal conditions to clear lipids, the time course of lipid clearing, the stability of clearance, and the functional characteristics of "lipid-free" hepatocytes. In the latter stages of the proposed studies, we will apply these protocols to isolate hepatocytes from livers containing levels of steatosis that typically preclude their use in drug discovery applications. Our overall goal is to determine the least "invasive" protocol to clear hepatocytes of excess lipids to address the acute shortage of primary human hepatocytes for academic, clinical, and pharmaceutical applications. PUBLIC HEALTH RELEVANCE: In the application "Clearing intrahepatic lipids to aid in toxicity testing," studies are aimed at examining the feasibility of targeting three pathways known to regulate lipid (fat) metabolism with the aim of reducing the fat levels in human hepatocytes isolated from donated livers. Vesta Therapeutics is one of the leading procurers of non-transplantable livers in the United States and has developed proprietary technologies for the isolation and preservation of primary hepatocytes from these livers. Currently, pharmaceutical companies are reluctant to utilize hepatocytes with high levels of fat, which add an uncontrollable layer of variability to the toxicity of test compounds. Devising strategies to eliminate fat from freshly isolated or cryopreserved hepatocytes would provide extremely valuable technologies for the full utilization of limited number of livers donated each year. Thus, if successful, this source of "fat-free" hepatocytes could benefit many goals of public health.

描述（由申请人提供）：Vesta Therapeutics 是美国不可移植人类肝脏的主要采购商之一，并开发了用于分离原代肝细胞的分离、冷冻保存和再生应用的专有技术。当 Vesta 为我们的研究项目处理细胞时，收集了大量剩余的成熟肝细胞，[将其提供给学术、临床和药物研究人员。原代人肝细胞是制药公司用来测试新候选药物的吸收、分布、代谢、排泄 (ADME) 和毒性 (T) 特性的宝贵且重要的模型。然而，]随着美国肥胖率的不断增加，器官采购组织[用于研究目的]提供的不可移植器官中有很大一部分是脂肪变性的（脂肪），这给器官的毒性增加了一层无法控制的变异性。潜在的测试化合物。 [因此，制药公司不愿意利用脂肪变性肝细胞，而被迫使用非生理动物和肿瘤细胞模型进行药物开发。]过去几年的研究表明，许多针对脂质生物合成的因素，氧化或肝内输出可以逆转动物和人类患者的脂肪变性。 [然而，尚未探索将这些方法应用于原代人类肝细胞以产生标准化供应的非脂肪肝细胞的可行性。]鉴于我们持续供应脂肪肝并且我们广泛的研究，Vesta 在评估这些技术方面处于独特的地位。冷冻保存的脂肪变性肝细胞库存。我们建议单独或联合使用胆汁酸、PPAR 激动剂和胆碱来降低人类肝细胞的脂肪变性水平，动物和人类研究已证明这些药物可以逆转脂肪变性。我们将记录清除脂质的最佳条件、脂质清除的时间过程、清除的稳定性以及“无脂质”肝细胞的功能特征。在拟议研究的后期阶段，我们将应用这些协议从含有脂肪变性水平的肝脏中分离出肝细胞，这些脂肪变性水平通常会妨碍它们在药物发现应用中的使用。我们的总体目标是确定“侵入性”最小的方案来清除肝细胞中的过量脂质，以解决学术、临床和制药应用中原代人肝细胞的严重短缺问题。公共卫生相关性： 在“清除肝内脂质以帮助毒性测试”的应用中，研究旨在检查以已知调节脂质（脂肪）代谢的三种途径为目标的可行性，目的是降低从捐赠肝脏中分离出的人类肝细胞中的脂肪水平。 Vesta Therapeutics 是美国不可移植肝脏的领先采购商之一，并开发了用于从这些肝脏中分离和保存原代肝细胞的专有技术。目前，制药公司不愿意使用高脂肪水平的肝细胞，这给测试化合物的毒性增加了不可控制的变异性。制定从新鲜分离或冷冻保存的肝细胞中消除脂肪的策略将为充分利用每年捐赠的有限数量的肝脏提供极其有价值的技术。因此，如果成功，这种“无脂肪”肝细胞来源可以有利于公共卫生的许多目标。