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: 6522842
• 负责人: RALPH E PARCHMENT
• 金额: $ 16.37万
• 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 2004-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6522842
• 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性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1950-1980年实验血液学家和癌症药理学家的研究基础上，他们发现了不同骨髓血细胞的造血祖细胞;开发了将祖细胞作为形态学可识别的血细胞的克隆集落生长的方法，并量化了样品中功能祖细胞的数量;证明这些“集落形成单位”（CFU）是体内和体外骨髓抑制性细胞毒性药物的靶点;分离并最终克隆重组细胞因子，用于完全确定和优化的细胞培养基，其刺激来自特定骨髓谱系的祖细胞的集落形成。我们的实验室成功地应用这一知识开发了体外血液毒理学，即，根据药物对CFU的体外毒性预测急性、可逆性药物诱导的中性粒细胞减少症的性质。我们发表了吡唑并吖啶和喜树碱的体外临床相关性。这项研究还发现了重要的原则和非明显的方法，这些原则和方法对于正确预测临床结果至关重要。严重中性粒细胞减少症风险的三种体外预测模型（CTC v2.0 3-4级），将人群的重度中性粒细胞减少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”）。欧洲替代方法验证中心（ECVAM）选择一个模型作为当前验证研究的主题，我们的实验室是北美的参与者。该R21申请的假设是：（1）急性细胞毒性暴露对胃肠道粘膜的损伤在概念上与中性粒细胞减少症相同;（2）当用于预测将导致可逆性重度粘膜损伤（临床表现为口腔炎和粘膜炎）的急性剂量/暴露时，使体外血液毒理学成功的建模和质量控制概念也将成功。在小鼠和大鼠中鉴定的GI祖细胞（称为“转运细胞”）和干细胞已被证明是细胞毒性药物和辐射的靶点。粘液细胞群可以在保留干细胞和祖细胞功能的培养物中生长。该R21的具体目的建立在这些发现的基础上，并将其与血液毒理学中使用的测定开发策略相结合，以开发用于“CFU-Gl”祖细胞的优化的体外克隆形成测定，其正确预测药物对GI粘膜的毒性相对于CFU-GM的相对风险。随后的R33应用将确认体外和体内相似的药物暴露- CFU-GI存活曲线，进行测试的预验证研究，并调整测试配置，以在药物开发途径的各个阶段应用，考虑到吞吐量和成本。