SBIR-STTR Award

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.

Public Health Relevance Statement:


Public Health Relevance:
Certain hormonally active chemicals, collectively referred to as endocrine disruptor chemicals (EDCs), mimic or antagonize the actions of naturally occurring estrogens and are believed to result in adverse developmental, reproductive, neurological, and immune effects in both humans and wildlife. The chemical, pesticide, personal care products, plastics, and nutritional supplements industries have been tasked with screening their products for EDCs. Using a novel autobioluminescent human cell line, 490 BioTech proposes to advance current EDC screening assays with a reporter cell technology that surveys EDC activity continuously and in real time to deliver more data faster and more cost effectively for improved safeguarding of public health.

Project Terms:
Address; Androgens; Bacterial Luciferases; Behavior; Binding (Molecular Function); Biological Assay; Biological Availability; Bioluminescence; Breast Cancer Cell; Cancer cell line; Cell Line; Cells; Characteristics; Chemical Exposure; Chemicals; Complex; consumer product; Contracts; cost; cost effective; Data; design; Development; Dietary Supplements; Endocrine; Endocrine Disruptors; Endocrine system; Engineering; Environmental Health; Environmental Pollution; Equipment and supply inventories; Estradiol; Estrogen Antagonists; Estrogens; European Union; Evaluation; Exposure to; Faslodex(ICI 182,780); Female; Fireflies; Firefly Luciferases; Health; high throughput screening; Homeostasis; Hormones; Human; Human Cell Line; Human Engineering; Immune; improved; In Vitro; in vivo; Individual; Industry; Kinetics; Laboratories; Light; light emission; Link; Luc Gene; Maintenance; Malignant Neoplasms; Manufacturer Name; Marketing; meetings; Methods; Monitor; National Institute of Environmental Health Sciences; National Toxicology Program; Neurologic; novel; Organ; Output; Performance; Pesticides; Pharmaceutical Cares; Phase; Plastics; Positioning Attribute; Preclinical Drug Evaluation; programs; Puberty; public health medicine (field); public health relevance; Reaction; Reaction Time; Reporter; Reporter Genes; Reproduction; reproductive; reproductive function; research and development; response; Response Elements; Safety; Scheme; screening; Self Care; Side; Small Business Innovation Research Grant; Specificity; Sperm Count Procedure; stable cell line; Stream; Surveys; T47D; Technology; Testing; Time; tool; Toxic effect; United States Environmental Protection Agency; Visual

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 II project proposes to build upon our successful Phase I demonstration of autonomous endocrine disruptor chemical (EDC) detection using human cell lines to engineer a novel Tier 1 screening assay for the low cost, high-throughput detection of estrogenically- and androgenically-active compounds across multiple human tissue types to address the National Institute of Environmental Health Sciences (NIEHS) request for Improved Test Systems for Prioritization and Safety Evaluation. Current Tier 1 EDC screening approaches require the use of non-human cell lines that can obscure bioavailability data, employ radioactive materials that require dedicated use areas and specially trained personnel, or rely upon the use of expensive analytical equipment that prevents high-throughput testing. Furthermore, following this initial Tier 1 screen, those chemicals that putatively express endocrine disrupting activities must still be screened against Tier 2 animal models to validate the results. Currently, there are over 500 contract testing service companies in the U.S. alone that perform Tier 1 style assays for the chemical, pesticide, and personal care products industries at an average cost of $1,000 per assay. However, as the number of potential EDCs requiring testing expands, and with many states now adopting legislation to begin screening all municipal water supplies for EDCs, market growth within this sector is expected to increase at an annual rate of 13.5% to approach $36 billion by 2016, clearly demonstrating the need for new assays that can reduce screening costs, increase screening throughput, and provide more relevant human bioavailability data without necessitating the use of animal models. The EDC-responsive, substrate-free, autobioluminescent reporter cell lines developed by 490 BioTech under our Phase I efforts demonstrated the ability to detect EDCs similarly to existing Tier 1 screening assays while employing a human cell culture-based model that significantly reduced the cost and personnel effort of testing while simultaneously providing an uninterrupted stream of visual data over the lifetime of the reporter cell as it interacted with and reacted to EDC treatment. In this proposal, we will leverage this technology to develop fully self-contained, human cell-based, high- throughput screening assays that provide more accurate and realistic information in regards to EDC bioavailability and effects on human health. With application of these assays reducing the cost of Tier 1 EDC screening over 100-fold to less than $10 per compound, we believe we possess a product capable of significantly impacting the EDC screening market and advancing our understanding of novel compound endocrine disruptor activity as it pertains to public health and consumer safety.

Public Health Relevance Statement:


Public Health Relevance:
In response to public and scientific concerns that chemical exposure can interfere with the delicate balance of the endocrine system to trigger adverse health effects in humans and animals, government agencies worldwide have implemented comprehensive screening programs to characterize tens of thousands of compounds for endocrine disruptor activity. Unfortunately, the complex nature and high cost of the screens currently used for these efforts have thus far only been able to generate data on a limited number of chemicals, resulting in an extensive, untested backlog that is hindering our understanding of how endocrine disruption impacts human and ecosystem health. In this Phase II R&D effort, 490 BioTech proposes to implement a novel endocrine disruptor screening strategy based upon a `humanized' bacterial luciferase genetic construct that links endocrine disruptor activity to autonomous bioluminescent light emission to provide increased data workflows at a greater speed and with less cost than any existing endocrine disruptor assay.

NIH Spending Category:
Bioengineering; Estrogen

Project Terms:
Address; Adopted; Affect; Androgens; Animal Model; Animals; Architecture; Area; Bacterial Luciferases; base; Biological Assay; Biological Availability; Bioluminescence; Cell Culture Techniques; Cell Line; Cells; Chemical Exposure; Chemicals; Complex; Contracts; Cosmetics; cost; Data; Detection; Development; Dietary Supplements; early onset; Ecosystem; Endocrine; Endocrine disruption; Endocrine Disruptors; Endocrine system; Engineering; Ensure; Equilibrium; Equipment; Estrogens; Evaluation; Genes; Genetic; Genetic Engineering; Genital system; Government Agencies; Growth; Health; Hela Cells; high throughput screening; Histocompatibility Testing; Human; Human Cell Line; Human Resources; human tissue; improved; Individual; Industry; Laboratories; Life; light emission; Link; malignant breast neoplasm; Malignant neoplasm of ovary; Malignant neoplasm of prostate; Malignant neoplasm of testis; Manufacturer Name; Marketing; Measures; Mediating; meetings; Metabolic Diseases; Methods; Modeling; National Institute of Environmental Health Sciences; Nature; novel; Organism; Output; Pesticides; Phase; Phenotype; Plastics; prevent; Price; Procedures; programs; Property; Protocols documentation; Puberty; public health medicine (field); public health relevance; Quality Control; Radioactive; Reporter; reproductive; research and development; response; Safety; screening; Self Care; Services; Signal Transduction; Small Business Innovation Research Grant; small molecule libraries; Soaps; Speed (motion); Sperm Count Procedure; Standardization; Statutes and Laws; Steroid biosynthesis; Stream; Sunscreening Agents; System; Technology; Testing; Time; Training; Validation; Visual; Water; Water Supply