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

描述（由申请人提供）：Solidus Biosciences，Inc.该公司将与伦斯勒理工学院合作，致力于进一步开发、验证和商业化其专有“转染酶和代谢芯片”（或TeamChip），用于系统性候选药物和化学代谢及毒理学的高通量分析。正在开发的TeamChip模拟人类肝脏的首过代谢，并预测酶特异性肝毒性。将通过使用病毒递送系统将代谢基因转染到封装在排列在塑料芯片上的三维（3D）基质（小至60 nL）中的人细胞中，制备在TeamChip上表达不同代谢酶组合的人细胞文库。因此，可以以与早期候选药物和环境化学品的预测性人体毒性评估相当的速度评估和定量目标化合物与人体肝脏或其他器官类型中的单个人体代谢酶或酶组合的反应性。 第二阶段STTR提案的具体目标/里程碑是：1.构建携带来自人肝cDNA文库的代谢酶基因的重组腺病毒（例如，代表性的CYP 450同种型和接合代谢酶），证明了使用荧光/发光底物在Hep 3B细胞单层上的基因转染，并通过Western印迹测定测量不同水平的腺病毒酶表达。2.制备含有表达各种代谢酶组合的Hep 3B细胞的TeamChip，并鉴定其差异表达影响对模型化合物的细胞应答的代谢基因。3.优化重组腺病毒和转染Hep 3B细胞的芯片冷冻保存方案。用冷冻保存的TeamChips证明代谢诱导的毒性，并将结果与非冷冻对应物进行比较。 可用于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的代谢和毒理学（例如，候选药物和环境化学毒物），这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。目前的化学品安全评估方法成本高、耗时长，并且使用大量化合物和大量动物。因此，有很大的潜力和机会，将TeamChip作为一种安全性评估工具，可用于评估特定的代谢酶是否以及如何影响候选药物和化学毒物的毒性。这种能力也可用于预测个体之间在药物和化学代谢和毒性方面的差异。 公共卫生相关性：药物发现过程是一个投资密集型，高风险的奋进，导致低产量的有效和安全的药物;一个问题，是混淆的显着缺乏信息，存在于预测代谢的候选药物的命运，一般来说，并在预测候选药物在人体中的反应。拟议的用于开发Solidus Bioscience的TeamChip技术的第二阶段STTR项目与公共卫生具有重要的相关性，为制药研究人员提供预测候选药物体内代谢所需的体外信息，从而有助于决定哪些化合物被提出用于先导优化和最终开发更好，更安全的药物。此外，这项研究与工业和环境化学品在安全和使用方面的优先次序有关。

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.