Biologically relevant screening of endocrine disruptors

Institution: Beckman Research Institute of the City of Hope
Investigator(s): Shiuan Chen, Ph.D. -
Award Cycle: 2011 (Cycle 17) Grant #: 17UB-8701 Award: $1,512,000
Award Type: SRI Request for Proposal (RFP)
Research Priorities
Etiology and Prevention>Etiology: the role of environment and lifestyle

Initial Award Abstract (2011)

Non-technical overview of the research topic and relevance to breast cancer:
Endocrine disruptors are environmental chemicals that interfere with the action and formation of steroid hormones in the body. Estrogen, an important steroid hormone, is known to play key roles in hormone dependent breast cancer. Although extensive efforts have been made by many outstanding researchers worldwide, a definitive conclusion regarding the impact of such chemicals in breast cancer has not been established. The goal of this proposed research is to develop screening assays for identifying and testing chemicals with relevance to known and suspected causes of estrogen-dependent breast cancer.

Research questions:
60% of premenopausal and 75% of postmenopausal patients have estrogen-dependent breast cancer. Estrogen receptor (ER) and aromatase are key players in this type of breast cancer. In our body, estrogen promotes breast cancer through its interaction with a receptor protein, estrogen receptor (ER) and drives the expression of growth factors that stimulate the growth of breast tumors. Aromatase is a protein that makes estrogen. In addition, ERRa is a receptor with similar activity as ER, but it can function without estrogen. Findings from this and other laboratories allow us to hypothesize that environmental chemicals will play critical roles in modulating breast cancer through these three proteins. We will develop functionally relevant assays that allow the identification of environmental chemicals contributing to breast cancer.

General methodology:
(1) We will refine a breast cancer-relevant high-throughput screening assay using a breast cancer cell line generated in our laboratory that is positive for all three important targets (ER, aromatase and ERRa) of endocrine disruptors. (2) A set of breast cancer-relevant genes will be tested for usage as markers to confirm the physiological importance of the chemicals identified from the screening assay. (3) Experiments will be performed to identify additional novel markers for environmental chemical-mediated responses in breast cancer. (4) Animal experiments will be performed to confirm the utility of the screening cell line and gene expression signature in the prediction of the effects of endocrine disruptors in our body.

Innovative elements:
A team of investigators with research experience in breast cancer translational research, estrogen biology, bioinformatics analysis, and animal experiments has been assembled at our institution to develop novel and breast cancer-relevant assays for an endocrine disruptor screen. New and potentially useful cell line and gene expression signatures will be established to be important tools to identify environmental chemicals with the potential to promote hormone-dependent breast cancer. An important collaboration with Dr. C. Teng, a scientist who is involved in the Tox21 project (a federal effort for environmental chemical screen) has been established. She will evaluate our assays in a timely manner through the Tox21 assay program at the National Toxicology Program, National Institute of Environmental Health Sciences. Collaboration between scientists at City of Hope and those at Tox21 community will facilitate the development of innovative assays to identify chemicals that can contribute to breast cancer.

Advocacy involvement:
Information about the risk of endocrine disruptors to breast cancer are often conflicting and confusing to patients and advocates. The development of breast cancer-relevant assays to help identify such environmental chemicals would be of great interest and receive support from patients and patient groups. Dr. Sandra Finestone, a member of Breast Cancer Solutions and the Executive Director of Hope Wellness Center, will serve as the advocate of this project, in the design of the laboratory studies and communicate the findings and progress of the project.

Final Report (2015)

Overview of the research topic and relevance to breast cancer:
The estrogen-responsive breast cancer is a major type of breast cancer. The major promoting factors in this type of cancer are aromatase (a protein to make estrogen) and estrogen receptor (ER) (activator of estrogen functions). It is thought that environmental chemicals bind and modify the activity of aromatase and ER would have an impact on the development of estrogen-dependent breast cancer. The goal of this project was to develop a biologically relevant high throughput screen system that allows us to identify environmental chemicals that modify these biological processes.

General progresses:
A breast cancer-relevant high-throughput screening assay using a breast cancer cell line, AroER tri-screen™, has been generated in our laboratory. The screening system was found to deliver highly reproducible results and to maintain a great signal/noise ratio between assays. Furthermore, it is positive for all three important biological targets (ER, aromatase and ERR?) of endocrine disruptors. AroER tri-screen™ has been shown to be able to screen through a large number of chemicals (endocrine disruptors) in a rapid fashion. This is the first screen system that can search chemicals that bind to aromatase and the first screen system that can screen chemicals interacting with aromatase and/or ER. The screening system has been adapted to be able to perform 1,536 screens simultaneously.

Barriers that were overcome:
The technical barriers for the validation of AroER tri-screen™ to be a high throughput assay that can screen a large number of chemicals were overcome through an extensive collaboration with investigators at the National Institute of Environmental Health Sciences (NIEHS) and the Tox21 program.

Major accomplishments:
At the City of Hope, we screened 446 drugs in the NIH Clinical Collection and revealed 106 of them that modulated ER and/or aromatase activities. Together with gene expression profile analysis, we defined the estrogen-like action of an antidepressant, paroxetine. Furthermore, the Tox21 program has approved our screening system and carried out a screen of a 10,000-compound library. A set of environmental chemicals that interact with aromatase were identified from this screen. One of the major hits is imazalil. It is registered for agricultural use in postharvest application and storage of fruits, vegetables, forage, and grain crops. In addition, our screening system was found to be useful to estimate overall estrogenic activities in biological samples, as a potential intermediate risk factor for breast cancer. These research findings resulted in three publications.

Plans for the next steps of the project:
The animal experiments will be essential to confirm how external exposure of a group of endocrine disruptors, identified from our rapid screen, promotes breast cancer development. The animal experiments are important to assess the quality of AroER tri-screen™ for rapid screen, and gene-expression profiles of the formed breast tumors will help in the prediction of the effects of endocrine disruptors in our body. Our findings will be presented to the CBCRP community.

AroER tri-screen(tm) is a novel functional assay to estimate both estrogenic and estrogen precursor activity of chemicals or biological specimens
OR14-5: Cell-Based High-Throughput Screening of Aromatase Inhibitors in Tox21 10K Library

AroER Tri-Screen Is a Biologically Relevant Assay for Endocrine Disrupting Chemicals Modulating the Activity of Aromatase and/or the Estrogen Receptor.
Periodical:Toxicological Sciences
Index Medicus: Toxicol Sci
Authors: Chen S, Zhou D, Hsin LY, Kanaya N, Wong C, Yip R, Sakamuru S, Xia M, et al.
Yr: 2014 Vol: 139 Nbr: 1 Abs: Pg:198-209

AroER tri-screenTM is a novel functional assay to estimate both estrogenic and estrogen precursor activity of chemicals or biological specimens
Periodical:Breast Cancer Research and Treatment
Index Medicus: Breast Cancer Res Treat
Authors: Kanaya N., Nguyen D.M., Lu H., Wang Y-Z., Hsin L-Y., Petreas M., Nelson D., et al
Yr: 2015 Vol: 151 Nbr: Abs: Pg:335-45