TeamChip for High-Throughput, Predictive Human Metabolism and Toxicology: Phase I

描述(由申请人提供)：Solidus Biosciences，Inc.与伦斯勒理工学院合作，将专注于进一步开发、验证其专利“转基因酶和代谢芯片”(或TeamChip)并将其商业化，用于系统候选药物的高通量分析以及化学新陈代谢和毒理学。TeamChip正在开发，以模拟人类肝脏的首过代谢，并预测酶特异性肝毒性。在TeamChip上表达不同代谢酶组合的人类细胞库将通过使用病毒递送系统将代谢基因导入封装在塑料芯片上的三维(3D)矩阵(小至60nL)的人类细胞中来制备。因此，目标化合物与人体单个代谢酶或人体肝脏或其他器官类型中的酶组合的反应性可以与对早期候选药物和环境化学品的预测人体毒性评估相称的速度进行评估和量化。这一第二阶段STTR方案的具体目标/里程碑是：1.构建携带代谢酶基因的重组腺病毒(例如，具有代表性的CYP450亚型和结合代谢酶)，利用荧光/发光底物在Hep3B细胞单层上进行基因转染，并通过Western印迹检测不同水平的腺病毒酶的表达。2.制备含有表达多种代谢酶组合的Hep3B细胞的TeamChip，并鉴定其差异表达影响细胞对模型化合物的反应的代谢基因。3.优化重组腺病毒和转染腺病毒的芯片上冷冻保存方案。用冷冻保存的TeamChips演示代谢诱导的毒性，并将结果与未冷冻的同行进行比较。可以用来快速评估大量化合物毒性的体外技术仍然有限。安全性评价的一个关键组成部分是化学品(如候选药物和环境化学毒物)的代谢和毒理学，它反映了化学品被人体代谢酶代谢的敏感性以及母体化合物及其代谢物的毒性。目前的化学安全评估方法成本高、耗时长，而且使用大量化合物和大量动物。因此，TeamChip作为一种安全评估工具有很大的潜力和机会，可以用来评估特定的代谢酶是否以及如何导致候选药物和化学毒物的毒性。这种能力还可以用来预测不同个体在药物和化学代谢以及毒性方面的差异。 与公共健康相关：药物发现过程是一项投资密集型、高风险的工作，导致有效和安全的药物产量较低；在总体上预测候选药物的代谢命运和预测候选药物在人体内的反应性方面存在严重缺乏信息，这是一个令人困惑的问题。为开发Solidus Bioscience的TeamChip技术而拟议的第二阶段STTR项目对公众健康具有重要意义，它为制药研究人员提供了预测候选药物在体内代谢所需的体外信息，从而有助于决定提出哪些化合物用于先导优化和更好、更安全的药物的最终开发。此外，这项研究还涉及工业和环境化学品的安全和使用方面的优先事项。

DESCRIPTION (provided by applicant): Solidus Biosciences, Inc. in partnership with Rensselaer Polytechnic Institute, will focus on further development, validation, and commercialization of its proprietary "Transfected Enzyme and Metabolism Chip" (or TeamChip) for high-throughput analysis of systematic drug candidate and chemical metabolism and toxicology. The TeamChip is being developed to mimic first-pass metabolism of the human liver and to predict enzyme-specific hepatotoxicity. A library of human cells expressing different combinations of metabolic enzymes on the TeamChip will be prepared by transfecting metabolic genes using a viral delivery system into human cells encapsulated in three-dimensional (3D) matrices (as small as 60 nL) arrayed on a plastic chip. Thus, the reactivity of target compounds with individual human metabolic enzymes or combinations of enzymes in the human liver or other organ types can be assessed and quantified at speeds commensurate with predictive human toxicity assessment of early stage drug candidates and environmental chemicals. The specific aims/milestones of this Phase II STTR proposal are to: 1. Construct recombinant adenoviruses that carry genes for metabolic enzymes from a human liver cDNA library (e.g., representative CYP450 isoforms and conjugative metabolic enzymes), demonstrate gene transfection on monolayers of Hep3B cells using fluorogenic/luminescent substrates, and measure different levels of adenoviral enzyme expression by Western blot assays. 2. Prepare the TeamChip containing Hep3B cells expressing various combinations of metabolic enzymes and identify metabolic genes whose differential expression affects the cellular response to model compounds. 3. Optimize on-chip cryopreservation protocols for recombinant adenoviruses and transfected Hep3B cells. Demonstrate metabolism-induced toxicity with cryopreserved TeamChips and compare the results with non- frozen counterparts. In vitro technologies that can be used to quickly assess large numbers of compounds for toxicity remain limited. A critical component of safety evaluation is metabolism and toxicology of chemicals (e.g., drug candidates and environmental chemical toxicants), which reflects the susceptibility of chemicals to be metabolized by human metabolic enzymes and the toxicity of parent compounds and their metabolites. Current approaches to chemical safety assessment are costly, time consuming, and use large amounts of compound and large numbers of animals. Thus, there is great potential and opportunity to apply the TeamChip as a safety assessment tool that can be used to evaluate whether and how specific metabolic enzymes contribute to the toxicity of drug candidates and chemical toxicants. This capability may also be used to predict differences among individuals in drug and chemical metabolism and toxicity. PUBLIC HEALTH RELEVANCE: The drug discovery process is an investment-intensive, high-risk endeavor that results in low yields of effective and safe drugs; a problem that is confounded by the significant lack of information that exists in predicting the metabolic fate of drug candidates, in general, and in predicting the reactivity of drug candidates in the human body. The proposed Phase II STTR project for the development of Solidus Bioscience's TeamChip technology has significant relevance to public health by providing pharmaceutical researchers with in vitro information needed to predict the in vivo metabolism of drug candidates, and thus help to decide which compounds are brought forward for lead optimization and the ultimate development of better and safer drugs. Furthermore, this research is relevant to the prioritization of industrial and environmental chemicals in terms of their safety and use.