Enhanced production of human hepatocytes from livers declined for transplant

肝脏产生的人类肝细胞产量因移植而下降

• 批准号: 9140604
• 负责人: MARK G CLEMENS
• 金额: $ 36.61万
• 依托单位: HEPATOSYS, INC.
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-07-17 至 2018-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9140604
• 关键词: Address; Animal Model; Biomedical Engineering; Cardiac Death; Cell Line; Cells; Clinical Trials; Collaborations; Coupled; Cryopreservation; Development; Drug Approval; Failure; Goals; Gold; Health; Health Care Costs; Hepatocyte; Hepatotoxicity; Hour; Human; Incubated; Investments; Ischemia; Left; Legal patent; Liver; Marketing; Membrane; Metabolism; Methods; Modification; Organ; Organ Preservation; Outcome; Perfusion; Pharmaceutical Preparations; Phase; Preclinical Drug Evaluation; Procedures; Process; Production; Rattus; Research; Research Personnel; Resuscitation; Series; Site; Source; System; Technology; Testing; Time; Toxicity Tests; Translating; Transplantation; Warm Ischemia; Xenobiotics; base; commercialization; cost; design; drug development; drug testing; efficacy testing; experience; flexibility; functional restoration; improved; improved functioning; innovation; innovative technologies; liver injury; novel therapeutics; oxidation; pre-clinical; preclinical study; screening; species difference

 DESCRIPTION (provided by applicant): Liver toxicity is a major barrier to the FDA approval of many new drugs. Moreover, since the liver is the major site of metabolism of most xenobiotics, it is essential to know how the liver metabolizes a potential new drug. Because of the many species differences between humans and commonly used animal models, testing on human hepatocytes remains the gold standard for drug screening. However, because the source of these hepatocytes is usually from livers that have been declined for use in transplantation, typically because of extended ischemic time, the supply of high quality cells is severely limited. The long-term goal of this project is to reliably improve the availability and viability of primary human hepatocytes isolated from non-transplantable Donation after Cardiac Death (DCD) livers through the modification of a hypothermic machine perfusion (HMP) resuscitation system that we have developed to restore function to ischemia damaged DCD livers for use in transplantation. Human hepatocytes are typically isolated from the pool of non-transplantable DCD organs which has been estimated to be as high as 40% of the current donor pool. However, inconsistent yield and quality of the isolated hepatocytes from these donors has led to limited availability resulting in higher costs for these cells. Major factors contributing to this inconsistency are the extended periods of warm ischemia experienced by these organs and the preservation process (simple cold storage) which does not resuscitate these organs. The HepatoSys solution (HS) coupled with HMP was developed to restore depleted energy stores, maintain membrane stability, and minimize oxidation damage resulting in a resuscitated liver. Our results indicate that our process can be used to successfully isolate high quality hepatocytes from otherwise unusable livers. In addition, we will add further commercial value to the technology by testing prolonged HMP of livers to provide greater flexibility in making fresh cells available, and the ability of additional oxygenated cold storage of hepatocytes in the HS to improve the quality of freshly isolated and cryopreserved hepatocytes. To achieve these goals, we propose the following aims. Specific Aim 1: Test the efficacy of prolonged HMP of livers with the HepatoSys solution on yield, viability, plateability and function of fresh hepatocytes isolated from rat DCD livers. Test the effect of up to 48 hours HMP followed by hepatocyte isolation on yield, viability, plateability and function of freshly plated isolated hepatocytes from DCD rats. Specific Aim 2: Test the efficacy of additional oxygenated cold storage (CS) of fresh hepatocytes after isolation in the HepatoSys solution on yield, viability, plateability and functio from DCD rat livers Test the ability of additional CS of isolated hepatocytes in the HS from DCD rats to maintain or improve the viability, plateability of freshly isolated rat hepatocytes. Specifc Aim 3: Test the efficacy of optimized conditions identified in aims 1 and 2 on yield, viability, plateability and function of freshly isolated hepatocytes from a small group of human DCD livers. Completion of these specific aims will provide a proof of concept that this technology has a potentially commercially valuable method to obtain viable, plateable and functional hepatocytes from livers currently not usable while providing greater flexibility in making freshly isolated hepatocytes available which will allow TRL and HepatoSys to increase market share. A Phase II application will seek to add further commercial value to the technology by expanding the pool to even poorer quality donor livers and improving the quality of cryopreserved hepatocytes.

 描述（由申请人提供）：肝毒性是 FDA 批准许多新药的主要障碍。此外，由于肝脏是大多数外源物质代谢的主要场所，因此了解肝脏如何代谢潜在的新药至关重要。由于人类和常用动物模型之间存在许多物种差异，对人类肝细胞的测试仍然是药物筛选的金标准。然而，由于这些肝细胞的来源通常来自已被拒绝用于移植的肝脏（通常是由于缺血时间延长），因此高质量细胞的供应受到严重限制。该项目的长期目标是可靠地提高主要资源的可用性和生存能力 通过修改低温机器灌注 (HMP) 复苏系统，从心源性死亡 (DCD) 肝脏的不可移植捐赠中分离出人肝细胞，我们开发了该系统，以恢复缺血受损的 DCD 肝脏的功能，用于移植。人类肝细胞通常是从不可移植的 DCD 器官库中分离出来的，据估计该器官库占当前供体库的 40%。然而，从这些供体中分离出的肝细胞的产量和质量不一致，导致可用性有限，从而导致这些细胞的成本更高。造成这种不一致的主要因素是这些器官经历的长时间的热缺血以及不能使这些器官复苏的保存过程（简单的冷藏）。 HepatoSys 解决方案 (HS) 与 HMP 相结合，旨在恢复耗尽的能量储存、维持膜稳定性并最大限度地减少氧化损伤，从而实现肝脏复苏。我们的结果表明，我们的工艺可用于成功地从无法使用的肝脏中分离出高质量的肝细胞。此外，我们将通过测试肝脏的延长 HMP 来进一步增加该技术的商业价值，以在提供新鲜细胞方面提供更大的灵活性，以及​​在 HS 中额外充氧冷藏肝细胞的能力，以提高新鲜分离和冷冻保存的肝细胞的质量。为了实现这些目标，我们提出以下目标。具体目标 1：测试使用 HepatoSys 溶液对肝脏进行延长 HMP 对分离的新鲜肝细胞的产量、活力、平板性和功能的功效 来自大鼠 DCD 肝脏。 测试长达 48 小时的 HMP，然后进行肝细胞分离对 DCD 大鼠新鲜接种的分离肝细胞的产量、活力、可接种性和功能的影响。具体目标 2：测试 HepatoSys 溶液中分离后的新鲜肝细胞额外充氧冷储存 (CS) 对 DCD 大鼠肝脏产量、活力、可铺板性和功能的功效 测试 DCD 大鼠 HS 中分离的肝细胞额外充氧冷储存 (CS) 维持或改善新鲜分离的大鼠肝细胞的活力、可铺板性的能力。具体目标 3：测试目标 1 和 2 中确定的优化条件对一小部分人 DCD 肝脏中新鲜分离的肝细胞的产量、活力、可铺板性和功能的功效。这些具体目标的完成将证明该技术具有潜在商业价值的方法，可以从目前无法使用的肝脏中获取可行的、可电镀的和功能性的肝细胞，同时在提供新鲜分离的肝细胞方面提供更大的灵活性，这将使 TRL 和 HepatoSys 能够增加市场份额。 II 期应用将寻求通过将库扩大到质量较差的供体肝脏并提高冷冻肝细胞的质量来进一步增加该技术的商业价值。