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

项目摘要/摘要 Stemina Biomarker Discovery，Inc.(Stemina)已经开发出一种基于体外人类多能干细胞的 检测，DevTOX快速预测(DevTOXqP)，使用鸟氨酸和胱氨酸的生物标志物比率来预测 化合物有可能在广泛的暴露中造成发育毒性。这样做的目的是 U01合作协议项目是鉴定DevTOXqP方法的鸟氨酸和胱氨酸的代谢物比率(o/c 比例)通过药物评估和研究中心(CDER)生物标记物资格认证计划(BQP)。 Stemina有一份被接受的意向书(LOI)，以证明devTOXqP o/c比率是安全的生物标志物 人诱导多能干细胞(IPS)体外检测人类发育毒性潜能 作为证据权重评估的一部分的小分子药物开发的非临床阶段，如 在国际技术协调理事会最近发布的ICH S5(R3)指南中进行了描述 《人用药品要求》(ICH)。在目标1中，我们计划进行因地制宜的分析 超高效液相色谱-高分辨质谱法的方法验证研究 (UPLC-HRMS)用于测量O/C比率以评估精密度、敏感性、特异性、稀释度 UPLC-HRMS分析的线性、再进样重现性、萃取回收率和样品携带 方法。我们还计划测量测试物品的浓度并表征测试化合物在细胞中的稳定性 用于确定生物标记物相关性的测试化合物子集的培养介质(目标5)。 将进行一项前瞻性资格研究，以：(1)确定实验室内的重复性和 O/C比率的重复性，(2)评估O/C比率在多个人iPS细胞系中的可靠性 预测一致性和对化合物暴露反应的相关性，以及(3)评估 预测多种药物化合物的发育毒性潜力的O/C比 (目标2、3和6)。最后，我们计划与Stemina的分销合作伙伴制定一项转让计划，以评估 DevTOXqP标准操作程序的实验室间可转换性和可靠性 资格认证计划的实验室间部分(目标4)。Stemina的合作伙伴将为其 资质计划的一部分。这项拨款中提出的研究，以及对Stemina分布的实物支持 合作伙伴将提供完成资质计划中描述的学习所需的数据(一旦 已获批准)，并提交完整的资质包。而在体外测试，如devTOXqP永远不会 完全取代动物单独使用时，这些检测方法可以减少在动物身上测试的化合物的数量。 在某些类别的化合物中，当用作 S5(R3)指南中描述的证据权重方法的一部分。DevTOXqP试剂盒的鉴定 因为发育毒性评估将减少动物的使用，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较少因在子宫内接触化学物质而导致的出生缺陷。

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.