Planar culture of gastrointestinal stem cells for screening pharmaceuticals for adverse event risk

胃肠道干细胞平面培养用于筛选药物不良事件风险

基本信息

批准号：
10482465
负责人：
Christopher Eldridge Sims
金额：
$ 25.66万
依托单位：
ALTIS BIOSYSTEMS, INC.
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-07-01 至 2023-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10482465
关键词：
3-Dimensional Address Adherent Culture Adverse event Animal Model Animals Apical Area Ascending colon Benchmarking Biological Assay Biomimetics Cell Culture Techniques Cell Proliferation Cell Survival Cell physiology Cells Cellular Structures Cessation of life Characteristics Clinical Clinical Trials Collaborations Collection Colon Data Descending colon Detection Development Diarrhea Disease Dose Dose-Limiting Drug Evaluation Drug Screening Duodenum Electrical Resistance Engineering Epithelial Exposure to Gastrointestinal Physiology Gastrointestinal tract structure Goals Human In Vitro Industry Inflammation Intestines Laboratories Lead Literature Manufacturer Name Measurement Measures Membrane Microinjections Modality Modeling Monitor Oncology Organ Donor Organ Transplantation Organoids Performance Pharmaceutical Preparations Pharmacologic Substance Pharmacology Phase Phenotype Physiology Population Property Rattus Reproducibility Risk S phase Sampling Screening procedure Side Small Intestines Source Stains Surface Technology Testing Therapeutic Therapeutic Agents Therapeutic Effect Tissue Donors Tissues Toxic effect Transformed Cell Line Transverse colon Tumor Cell Line Tumor-Derived Ulcer Variant adverse event risk base biobank cell type cost effective drug testing gastrointestinal gastrointestinal epithelium gastrointestinal system high risk ileum in vitro Assay in vitro Model in vivo indexing intestinal epithelium jejunum manufacturing process monolayer novel novel therapeutics phase III trial preclinical safety preclinical study prevent programs regenerative cell repaired research clinical testing scaffold screening side effect stem stem cell differentiation stem cell population stem cell proliferation stem cells stemness success therapeutic candidate transcriptomics

项目摘要

Project Summary GI side effects, such as ulcers and diarrhea, represent the most common source of adverse events for pharmaceuticals. GI stem cells are responsible for repairing and replenishing GI epithelium, and pharmaceutical inhibition of these functions likely contribute to adverse event risk. Currently there are no high-throughput and cost-effective means of screening candidate therapeutics for effects on GI stem cells. Animal models are fraught with confounds (e.g., rats generally do not exhibit diarrhea until death is imminent) and current in vitro models like the Caco-2 tumor cell line do not include a normal stem cell population. While 3D-organoid cultures have a stem cell component, access to the apical aspect of the monolayer for compound exposure is not possible without low-throughput microinjection. Altis Biosystems, Inc. has developed a proprietary culture platform, RepliGut® Planar, enabling primary human GI cells to form an epithelium for drug screening. In preliminary efforts, we initiated development of a GI stem cell-specific platform called RepliGut® StemScreen to address the unmet need for high-throughput, cost-effective GI stem cell screening, including a range of assays to measure properties that might lead to adverse events. These include assays for proliferation using S-phase staining and assessing downstream differentiation and ability to establish barrier function via non-destructive transepithelial electric resistance (TEER) analysis. A panel of pharmaceutical agents associated with clinically adverse GI side effects, tested in a classic EdU assay using StemScreen planar cultures, demonstrated dose-dependent inhibition of stem cell proliferation and prevented stem cell differentiation into a mature epithelium with barrier function. During Phase I we will: 1) characterize monolayer proliferation properties of donor cells in the Altis biobank(initially transverse colon stem cells will be evaluated, with other regions tested in Phase II) from three donors and 2) test a selection of therapeutic agents with known GI adverse event risks for effects on stem cell proliferation, impacting differentiation and the formation of a functional epithelial barrier (indexed as TEER). Demonstrating that the StemScreen platform provides an in vitro screening tool capable of predicting a drug’s risk for GI adverse events in Phase I will enable deeper characterization of stemness, stem cells from other GI regions (e.g., ascending and descending colon; small intestine regions including jejunum, etc.), and stem cell phenotypes. In Phase 2, we plan externally validate the platform in collaboration with leading industry laboratories prior to commercial launch.

项目概要胃肠道副作用，例如溃疡和腹泻，是最常见的不良事件来源。胃肠道干细胞负责修复和补充胃肠道上皮和药物。这些功能的抑制可能会导致不良事件风险，目前还没有高通量和高通量的研究。筛选候选疗法对胃肠道干细胞影响的经济有效的方法是充满挑战的。存在混淆（例如，大鼠通常在死亡临近之前不会表现出腹泻）和当前的体外模型像 Caco-2 肿瘤细胞系不包含正常干细胞群，而 3D 类器官培养物则包含正常干细胞群。干细胞成分，不可能进入单层的顶端进行化合物暴露无需低通量显微注射 Altis Biosystems, Inc. 开发了一种专有的培养平台， RepliGut® Planar，使原代人类胃肠道细胞能够形成用于药物筛选的上皮细胞。为了解决这一问题，我们启动了名为 RepliGut® StemScreen 的胃肠道干细胞特异性平台的开发对高通量、具有成本效益的胃肠道干细胞筛查的需求未得到满足，包括一系列检测方法可能导致不良事件的特性包括使用 S 期染色进行增殖分析和评估下游分化和通过非破坏性跨上皮建立屏障功能的能力电阻（TEER）分析。一组与临床不良胃肠道副作用相关的药剂，经过经典测试使用 StemScreen 平面培养物进行的 EdU 测定表明，干细胞增殖具有剂量依赖性抑制作用并阻止干细胞分化为具有屏障功能的成熟上皮。在第一阶段，我们将：1) 表征 Altis 生物库中供体细胞的单层增殖特性（最初是横结肠干细胞）将评估细胞，并在第二阶段测试来自三个捐赠者的其他区域，以及2）测试选择的具有已知胃肠道不良事件风险的治疗药物会影响干细胞增殖，影响分化和功能性上皮屏障的形成（索引为 TEER）。 StemScreen 平台提供了一种体外筛选工具，能够预测药物发生胃肠道不良事件的风险第一阶段将能够更深入地表征干性，来自其他胃肠道区域的干细胞（例如，上行干细胞）和降结肠；小肠区域，包括空肠等）和干细胞表型。我们计划在商业化之前与领先的行业实验室合作对该平台进行外部验证发射。