Acute Toxicity in Renewing Tissues and In Vitro Data

更新组织的急性毒性和体外数据

• 批准号: 6410514
• 负责人: RALPH E PARCHMENT
• 金额: $ 17.57万
• 依托单位: WAYNE STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-09-01 至 2002-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6410514
• 关键词: 6 thioguanine; 6 thiopurine; albumins; alpha 1 acid glycoprotein; analog; blood disorder diagnosis; diagnosis design /evaluation; diagnostic tests; epidermal growth factor; fibroblast growth factor; fluorouracil; gastrointestinal disorder diagnosis; goblet cells; immunocytochemistry; insulin; insulinlike growth factor; intestinal mucosa; intestine disorder; laboratory mouse; neutropenia; p53 gene /protein; platelet derived growth factor; small intestines; technology /technique development; toxicology; transferrin; transforming growth factors

A very successful field of in vitro toxicology is predicting reversible neutropenia caused by acute cytotoxic drug exposure. Recent success was built on research in 1950-1980 by experimental hematologists and cancer pharmacologists who discovered hematopoietic progenitors for the different myeloid blood cells; developed methods to grow progenitors as clonal colonies of morphologically recognizable blood cells and quantify the number of functional progenitors in a sample; demonstrated that these "colony forming units" (CFUs) are targets of myelosuppressive cytoytoxic drugs in vivo and in vitro; and isolated and eventually cloned recombinant cytokines for completely defined and optimized cell culture media that stimiuated colony formation by progenitors from particular myeloid lineages. Our laboratory applied this knowledge successfully to develop in vitro hematotoxicology, i.e., predicting the nature of acute, reversible drug-induced neutropenia from the drug's in vitro toxicity to the CFUs. We published in vitro - clinical correlations for pyrazoloacridine and camptothecins. This research also discovered important principles and non-obvious maesurements to make that are critical for correctly predicting clinical outcome. Three in vitro prediction models for risk of severe neutropenia (CTC v2.0 Grade 3-4) have been published that define a population's risk of severe neutropenia as a function of dose or systemic exposure, rather than hazard classification ("myelosuppressive or not'j. One model was selected as the subject of the current validation study by the European Centre for Validation of Altemative Methods (ECVAM), in which our laboratory is the North American participant. The hypotheses of this R21 application are (1) damage to gastrointestinal mucosa from acute cytotoxic exposure is conceptually identical to neutropenia and (2) the modeling and quality control concepts that made in vitro hematotoxicology successful will also be successful when applied to predict the acute dose/exposure that will cause reversible, severe mucosal damage (manifested clinically as stomatitis and mucosifs). GI progenitors (called "transit cells") and stem cells, identified in mouse and rat, have been demonstrated to be a target of cytotoxic drugs and radiation. Mucosal populations can be grown in culture with retention of stem cell and progenitor function. The Specific Aims of this R21 build on these discoveries and combine them with the assay development strategy used in hematotoxicology to develop an optimized in vitro clonogenic assay for "CFU-Gl" progenitors that correctly predicts the relative risk of drug toxicity to GI mucosa versus CFU-GM. A subsequent R33 application will confirm similar drug exposure - CFU-GI survival curves in vitro and in vivo, perform a pre- validation study of the test, and adapt the test's configuration for application in the various stages of the drug development pathway with respect to through-put and cost.

体外毒理学的一个非常成功的领域是预测急性细胞毒药物暴露引起的可逆性中性粒细胞减少症。最近的成功建立在实验血液学家和癌症药理学家在1950-1980年的研究基础上，他们发现了不同髓系血细胞的造血祖细胞；开发了将祖细胞作为形态可识别的血细胞克隆进行培养的方法，并量化了样本中具有功能的祖细胞的数量；证明了这些“克隆形成单位”(CFU)在体内和体外是骨髓抑制细胞毒性药物的靶点；并分离并最终克隆了重组细胞因子，用于完全定义和优化的细胞培养基能刺激特定髓系祖细胞的克隆形成。我们的实验室成功地应用这一知识发展了体外血液毒理学，即根据药物对CFU的体外毒性来预测药物引起的急性、可逆性中性粒细胞减少的性质。我们发表了吡唑吖啶和喜树碱的体外-临床相关性。这项研究还发现了对正确预测临床结果至关重要的重要原则和不明显的衡量标准。已经发表了三种严重中性粒细胞减少症风险的体外预测模型(CTC v2.0 3-4级)，它们将人群严重中性粒细胞减少症的风险定义为剂量或全身暴露的函数，而不是危险分类(“骨髓抑制与否”)。其中一个模型被欧洲替代方法验证中心(ECVAM)选为当前验证研究的对象，我们的实验室是北美的参与者。R21应用的假设是：(1)急性细胞毒性暴露对胃肠粘膜的损害在概念上与中性粒细胞减少症相同，(2)使体外血液毒理学成功的建模和质量控制概念在应用于预测将导致可逆的严重粘膜损害(临床表现为口腔炎和粘液)的急性剂量/暴露时也将成功。在小鼠和大鼠中发现的GI前体细胞(称为“转运细胞”)和干细胞已被证明是细胞毒药物和辐射的靶标。在保留干细胞和祖细胞功能的情况下，粘膜群体可以在培养中生长。R21的具体目标建立在这些发现的基础上，并将它们与血液毒理学中使用的检测开发策略相结合，开发了一种针对“CFU-G1”祖细胞的优化体外克隆形成试验，该方法正确地预测了药物对胃肠道粘膜毒性与CFU-GM的相对风险。随后的R33应用将确认类似的药物暴露-CFU-GI在体外和体内的存活曲线，执行测试的预验证研究，并调整测试的配置，以适应在药物开发途径的不同阶段应用的产量和成本。