Expressing bacterial bioluminescence in human cell lines: Engineering autobiolumi

描述（申请人提供）：在人细胞系中表达细菌生物发光：工程自动生物发光报告细胞筛选内分泌干扰物化学品项目摘要这个小企业创新研究第一阶段项目提出工程自动生物发光人类细胞系，用于改善筛选显示内分泌干扰物活性的化学品，以解决国家环境健康科学研究所（NIEHS）的问题。要求提供工具，以改进与内分泌活性化学品接触有关的接触评估。可能来自塑料、农药、药品和个人护理产品的内分泌干扰物化学品（EDCs）正在接受审查，因为它们可能对人类和野生动物造成不良健康影响，包括癌症发病率增加、精子数量减少、女性青春期提前和生殖器官功能改变。化学品制造商和进口商必须在美国环保署内分泌干扰物筛选计划（EDSP）的主持下测试其化学品的内分泌毒性效应。由于有87，000种化学品可能需要测试，一级EDC筛查检测被认为太慢且成本太高，无法满足当前的需求，EPA的任务是将其EDSP计划过渡到更快，更具成本效益的筛查技术。为了满足这一需求，490 BioTech提出设计一种具有细菌来源的生物发光报告基因构建体的人类细胞系，以改善对内分泌活性化学物质的监测。目前市场上应用于EDC筛选的生物发光报告细胞依赖于萤火虫荧光素酶基因构建体，该基因构建体必须提供有底物以激活其光发射响应，从而导致潜在毒理学相互作用的仅少量信息的单个时间点快照。我们提出的无底物，自生物发光报告细胞系将 响应于内分泌活性化合物而连续且真实的发光，从而在报道细胞与目标化学暴露相互作用和反应时，在报道细胞的寿命期间提供不间断的视觉数据流。此外，使用人类细胞作为传感平台提供了关于生物利用度和化学品对个体人类健康的真实影响的更准确和真实的信息。有超过500个合同实验室在87，00种化学品的潜在库存中以1，000美元/次的平均成本进行一级筛选测定，并且EPA要求将更快/更好/更便宜的技术整合到一级筛选计划中，我们相信我们拥有能够显著影响化学品/药物筛选市场的产品，特别是，提高我们对内分泌活性化学品的认识，因为它们与公共健康和消费者安全有关。

DESCRIPTION (provided by applicant): Expressing bacterial bioluminescence in human cell lines: Engineering autobioluminescent reporter cells to screen for endocrine disruptor chemicals Project Summary This Small Business Innovation Research Phase I project proposes to engineer an autobioluminescent human cell line for improved screening of chemicals displaying endocrine disruptor activity to address the National Institute of Environmental Health Sciences (NIEHS) request for tools for improved exposure assessment related to endocrine active chemical exposures. Endocrine disruptor chemicals (EDCs) potentially originating from plastics, pesticides, and pharmaceutical and personal care products are under scrutiny due to implications that they cause adverse health effects in humans and wildlife, including increased cancer rates, lowered sperm counts, early puberty in females, and altered functions of reproductive organs. Chemical manufacturers and importers are being mandated to test their chemicals for endocrine toxicity effects under the auspices of the U.S. EPA Endocrine Disruptor Screening Program (EDSP). With 87,000 chemicals potentially requiring testing, the battery of Tier 1 EDC screening assays have been recognized as being too slow and too costly to meet current demands, and the EPA has been tasked with transitioning its EDSP program towards faster and more cost-effective screening technologies. To address this need, 490 BioTech proposes to engineer a human cell line endowed with a bacterially derived bioluminescent reporter gene construct for improved monitoring of endocrine active chemicals. The current market of bioluminescent reporter cells being applied towards EDC screening rely upon a firefly luciferase gene construct that must be provided with a substrate to activate its light emission response, resulting in only marginally informative single time point snapshots of potential toxicological interactions. Our proposed substrate-free, autobioluminescent reporter cell line will emit light continuously and in real time in response to endocrine active compounds, thus providing an uninterrupted stream of visual data over the lifetime of the reporter cell as it interacts and reacts to target chemical exposure. Further, using a human cell as the sensing platform provides more accurate and realistic information in regards to bioavailability and a chemical's true effect on individual human health. With over 500 contract laboratories performing Tier 1 screening assays at an average cost of $1,000/assay over a potential inventory of 87,00 chemicals, and with the EPA mandating the integration of faster/better/cheaper technologies into the Tier 1 screening scheme, we believe we possess a product capable of significantly impacting the chemical/drug screening market and, here in particular, advancing our understanding of endocrine active chemicals as they pertain to public health and consumer safety.