Hemodynamic Co-Culture Liver Model for Drug Discovery and Assessment

药物开发人员迫切需要实验室中更好的工具来改善临床试验中90%的药物失败率。尽管在替代动物物种中进行了广泛的临床前试验，但肝毒性仍然是药物失败的主要原因。HemoShear开发用于目标ID的人类替代技术，并在后期发现和早期临床前阶段验证和筛选化合物的安全性和有效性。HemoShear开发了一种人体血管替代技术，用于识别和验证新靶点以及筛选和选择最佳化合物以进行进一步开发（正在审查的美国专利60/879，710）。我们的人体血管替代器械独特地模拟了动脉粥样硬化早期阶段的血管解剖结构（含有内皮细胞和平滑肌细胞的共培养物，由多孔膜隔开）和血液动力学环境。该设备能够研究人类动脉粥样硬化的细胞和分子机制，并识别新的生物标志物和转录途径，以开发改进的药物疗法。基于细胞的替代模型在药物开发过程中越来越多地使用，以提供更准确的人体反应预测。该I期SBIR应用提出使用HemoShear的专有平台技术开发肝脏的血流动力学共培养模型，该平台技术将代表一种更上级的系统，可用于筛选药物的肝毒性潜力，确定肝毒性机制并确定新的治疗靶点。众所周知，用于研究药物疗效和毒性的静态单培养肝细胞模型不能预测体内反应，并且由于固有的分化表型损失，在药物开发过程中代表了用于靶点鉴定和验证的无效模型。像血管系统一样，重建对正常肝功能重要的解剖和生理特征对于有效的离体模型是必要的。例如，肝脏在体内对内源性底物以及外源性因子的生物学响应取决于内皮和上皮的不同细胞类型之间的直接和间接“串扰”。此外，它们的存活和基因表达谱在很大程度上取决于局部血流动力学。因此，本提案的总体目标是开发模拟体内生理学和血流动力学的大鼠肝脏共培养替代模型。该模型将由共培养的窦状隙内皮细胞（SEC）和肝细胞组成，其中SEC暴露于窦状隙流体血液动力学，离体重建体内细胞表型。如果成功，该系统将使制药公司能够更好地了解与当前单一培养静态系统相比，药物疗效和临床前安全性/毒性的具体作用机制。此外，该I期SBIR的成功将导致II期应用，以开发更先进的肝毒性和炎性疾病的人类模型，并纳入其他细胞类型，例如枯否细胞和星状细胞。 公共卫生相关性：药物开发人员迫切需要更好的实验室工具来改善临床试验中90%的药物失败率。尽管在替代动物物种中进行了广泛的临床前试验，但肝毒性仍然是药物失败的主要原因。HemoShear开发用于目标ID的人类替代技术，并在后期发现和早期临床前阶段验证和筛选化合物的安全性和有效性。该I期SBIR申请提出使用HemoShear的专有平台技术开发肝脏的血液动力学大鼠窦状隙内皮细胞和肝细胞共培养模型;这将代表筛选肝毒性药物潜力、确定肝毒性机制和识别治疗新靶点的远为上级的系统。这一I期SBIR的成功将导致II期应用，以开发更先进的肝毒性和炎症性疾病的人类模型，包括其他细胞类型，例如枯否细胞和星状细胞。

DESCRIPTION (provided by applicant): Drug developers desperately need better tools in the laboratory to improve the 90% failure rate of drugs in clinical trials. Liver toxicity is still the leading cause of drug failure, despite extensive preclinical testing in surrogate animal species. HemoShear develops human surrogate technologies for target ID and validation and screening of compounds for safety and efficacy in the late discovery and early pre-clinical stages. HemoShear has developed a human vascular surrogate technology for identification and validation of novel targets and for screening and selection of optimal compounds for further development (US Patent 60/879,710 under review). Our human vascular surrogate device uniquely mimics the vascular anatomy (co- culture containing endothelial and smooth muscle cells, separated by a porous membrane) and hemodynamic environment during the early stages of atherosclerosis. The device enables investigation of the cellular and molecular mechanisms of human atherosclerosis and the identification of novel biomarkers and transcriptional pathways for development of improved drug therapies. Cell-based surrogate models are being used increasingly during drug development to provide more accurate predictions of human responses. This Phase I SBIR application proposes to develop a hemodynamic co-culture model of the liver using HemoShear's proprietary platform technology that will represent a far superior system with which to screen drug potential for hepatotoxicity, determine mechanisms of liver toxicity and identify novel targets for therapy. It is widely known that static, monoculture hepatocyte models utilized to study drug efficacy and toxicity are not predictive of the in vivo response, and represent ineffective models for target identification and validation in the drug development process because of the inherent loss of differentiated phenotype. Like the vasculature, recreating anatomical and physiological features important to normal liver function is necessary for an effective ex vivo model. For example, the biological response of the liver in vivo to both endogenous substrates as well as exogenous factors is dependent on the direct and indirect 'cross-talk' between the different cell types of the endothelium and epithelium. In addition, their survival and gene expression profiles are largely dependent on the local hemodynamics. Thus, the overall goal of this proposal is to develop a rat liver co-culture surrogate model that mimics in vivo physiology and hemodynamics. The model will consist of sinusoidal endothelial cells (SECs) and hepatocytes in co-culture where the SECs are exposed to sinusoid fluid hemodynamics, recreating in vivo cell phenotypes, ex vivo. If successful, this system will allow pharmaceutical companies to better understand the specific mechanism-of-action for drug efficacy and pre-clinical safety/toxicity compared to current monoculture, static systems. Additionally, success of this Phase I SBIR will lead to a Phase II application to develop a more advanced human model of hepatotoxicity and inflammatory disease, incorporating additional cell types, e.g. Kupffer and stellate cells. PUBLIC HEALTH RELEVANCE: Drug developers desperately need better tools in the laboratory to improve the 90% failure rate of drugs in clinical trials. Liver toxicity is still the leading cause of drug failure, despite extensive preclinical testing in surrogate animal species. HemoShear develops human surrogate technologies for target ID and validation and screening of compounds for safety and efficacy in the late discovery and early pre-clinical stages. This Phase I SBIR application proposes to develop a hemodynamic rat sinusoidal endothelial cell and hepatocyte co-culture model of the liver using HemoShear's proprietary platform technology; which will represent a far superior system to screen drug potential for hepatotoxicity, determine mechanisms of liver toxicity and identify novel targets for therapy. Success of this Phase I SBIR will lead to a Phase II application to develop a more advanced human model of hepatotoxicity and inflammatory disease, incorporating additional cell types, e.g. Kupffer and stellate cells.