DDT-BMQ-000109, Qualification of biomarkers for in vitro developmental toxicity screening in a human system

项目总结/摘要 Stemina Biomarker Discovery，Inc.（Stemina）开发了一种基于体外人类多能干细胞的 devTOX quickPredict（devTOXqP）测定，该测定使用鸟氨酸与胱氨酸的生物标志物比率来预测 化合物在广泛的暴露范围内有可能引起发育毒性。这个目标 U 01合作协议项目是鉴定devTOXqP检测的鸟氨酸与胱氨酸（o/c）的代谢物比例 通过药物评价和研究中心（CDER）生物标志物鉴定计划（BQP）。 Stemina已接受意向书（LOI），将devTOXqP o/c比值作为安全性生物标志物， 使用人诱导多能干细胞（iPS）在体外检测人发育毒性潜力， 作为证据权重评估的一部分， 国际技术协调理事会最近发布的ICH S5（R3）指导原则中描述了 人用药品（ICH）要求。在目标1中，我们计划进行符合目的的分析 超高效液相色谱-高分辨质谱法的方法学验证研究 （UPLC-HRMS）方法，用于测量o/c比值，以评估精密度、灵敏度、专属性、稀释度 UPLC-HRMS分析的线性、再进样重现性、提取回收率和样品残留 法我们还计划测量供试品浓度并表征细胞中供试化合物的稳定性 用于测定生物标志物相关性的试验化合物子集的培养基（目的5）。 将进行前瞻性确认研究，以：（1）确定实验室内重复性， o/c比的再现性，（2）基于以下指标评价多种人iPS细胞系中o/c比的可靠性： 预测一致性和对化合物暴露的反应的相关性，以及（3）评估 用于预测广泛范围的药物化合物的发育毒性潜力的O/C比 (Aims 2、3和6）。最后，我们计划与Stemina的分销合作伙伴建立转移计划，以评估 devTOXqP标准操作规程的实验室间可转移性和可靠性 鉴定计划的实验室间部分（目标4）。Stemina的合作伙伴将为其提供财政支持 资格计划的一部分。这项资助中提出的研究，以及Stemina分发的实物支持， 合作伙伴，将提供完成资格鉴定计划中所述研究所需的数据（一次 已批准）并提交完整的资格认证包。虽然体外测定如devTOXqP将永远不会 当单独使用时，这些测定法可以完全取代动物，从而减少在动物中测试的化合物的数量。 该测定法还可以代替在某些类别的化合物中当用作第二物质时对第二物质的需要。 S5（R3）指南中描述的证据权重方法的一部分。devTOXqP测定的确认 用于发育毒性评估将减少动物使用，并提供基于人类的评估， 发育毒性，最终导致更安全的药物和更少的出生缺陷，从化学品暴露在子宫内。

Project Summary/Abstract Stemina Biomarker Discovery, Inc. (Stemina) has developed an in vitro human pluripotent stem cell-based assay, devTOX quickPredict (devTOXqP), that uses a biomarker ratio of ornithine to cystine to predict if a compound has the potential to cause developmental toxicity over a wide range of exposures. The goal of this U01 cooperative agreement project is to qualify the devTOXqP assay’s metabolite ratio of ornithine to cystine (o/c ratio) through the Center for Drug Evaluation and Research (CDER) Biomarker Qualification Program (BQP). Stemina has an accepted Letter of Intent (LOI) to qualify the devTOXqP o/c ratio as a safety biomarker for detecting human developmental toxicity potential in vitro using human induced pluripotent stem (iPS) cells at the nonclinical stage of drug development for small molecule drugs as part of a weight-of-evidence assessment as described in the ICH S5(R3) guideline recently issued by the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH). In Aim 1, we plan to conduct a fit-for-purpose analytical method validation study for the ultra-performance liquid chromatography-high resolution mass spectrometry (UPLC-HRMS) method used for measuring the o/c ratio to assess the precision, sensitivity, specificity, dilution linearity, reinjection reproducibility, extraction recovery, and sample carryover of the UPLC-HRMS analytical method. We also plan to measure test article concentrations and characterize test compound stability in the cell culture medium for a subset of the test compounds used in determining the relevance of the biomarker (Aim 5). A prospective qualification study will be conducted to: (1) determine the within-laboratory repeatability and reproducibility of the o/c ratio, (2) evaluate the reliability of the o/c ratio in multiple human iPS cell lines based on prediction concordance and the correlation of response to compound exposure, and (3) assess the relevance of the o/c ratio for predicting developmental toxicity potential across a broad range of pharmaceutical compounds (Aims 2, 3 and 6). Finally, we plan to establish a transfer plan with Stemina’s distribution partner to assess between-laboratory transferability and reliability of the of the devTOXqP standard operating procedures for the inter-laboratory portion of the Qualification Plan (Aim 4). Stemina’s partner will provide financial support for its part of the Qualification Plan. The studies proposed in this grant, and the in-kind support of Stemina’s distribution partner, will provide the necessary data for completing the studies described in the Qualification Plan (once approved) and submitting the Full Qualification Package. While in vitro assays such as devTOXqP will never entirely replace animals when used alone, these assays can reduce the number of compounds tested in animals. The assay may also replace the need for a second species in certain categories of compounds when used as part of a weight of evidence approach as described in the S5 (R3) guideline. Qualification of the devTOXqP assay for developmental toxicity assessment will reduce animal use and provide a human-based assessment of developmental toxicity, ultimately leading to safer drugs and fewer birth defects from chemical exposure in utero.