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Abstract for TestSmart -- Endocrine Disruptors

Validation Efforts for EPA's Endocrine Disruptor Screening Program

Anthony Maciorowski, Gary Timm, and James Kariya
Office of Science Coordination and Policy, Office of Prevention Pesticides and Toxic Substances, U.S. Environmental Protection, Washington, DC.

The U. S. Environmental Protection Agency's Proposed Endocrine Disruptor Screening Program (EDSP) has five major components: Sorting, Priority Setting, Tier 1 Screening, Tier 2 Testing, and Hazard Assessment. Implementation of the EDSP is proceeding along two fronts: development of priority setting tools, and the validation of the individual screens and tests. This presentation focuses on the progress the Agency has made to date in implementing the EDSP.

The Endocrine Disruptor Priority Setting Database (EDPSD) is being developed for use in establishing priorities for the first phase of Tier 1 Screening. The EDPSD employs a "compartment-based priority setting strategy" that builds upon distinct exposure and effects related information categories and criteria, as well as a category of specially-targeted priorities. Priority chemicals for screening will be selected from industrial chemicals, pesticide formulation ingredients, chemicals used in cosmetics and other consumer products, nutritional supplements, and food additives. The EDPSD stores and uses data from about 26 data "compartments" from the following categories:

  1. Specially-Targeted Priorities,
  2. Exposure-based compartments,
  3. Effects-based compartments, and
  4. Combined exposure/effects compartments.

The Agency used data characteristics to develop a default approach that ranks chemicals within compartments and assigns weights to the compartments. However, the database also has the flexibility for user-selected ranking algorithms and compartment weights. Version 2 of the database ranks chemicals that are Specially-Targeted Priorities or pesticide formulation ingredients that are in EPA's High Production Volume Challenge program. Although the system architecture for the EDPSD is nearly completed (, existing data specific to endocrine mechanisms is wholly lacking for the vast majority of chemicals.

The original strategy for priority setting included the option of developing a computer automated HighThroughput Prescreening (HTPS) assay system employing estrogen (ER) and androgen receptor (AR) transcriptional activation assays. The original concept was to develop ER/AR data for the approximately 15,000 chemicals produced in quantities greater than 10,000 lb/yr. These data were to be included in the EDPSD to supplement existing data and to assist priority setting. Several HTPS methods were developed in a feasibility study conducted by the Agency, but the specific assay systems were judged too unreliable for routine regulatory use following external scientific peer review. The Agency continues to collaborate with the Japanese Government who are also developing an HTPS system. Pending completion and review of the Japanese HTPS developmental studies, the data from the Japanese HTPS system will be included in the EDPSDB pending demonstration of its reliability. However, the agency is also pursuing development of ER and AR Quantitative Structure Activity (QSAR) models which provide computer simulations of relative binding affinity. The Agency is collaborating with the FDA National Center for Toxicological Research on a comparative molecular field analysis (CoMFA) QSAR model, and with the EPA Office of Research and Development and the University of Bourgas on a common reactivity pattern (COREPA) models. The Agency is developing receptor binding data for up to 500 chemicals necessary for refining the model predictions. Should the HTPS and QSAR approaches both prove to be feasible, both will be incorporated into the EDPSDB to assist priority setting.

The Tier 1 screens and Test s proposed for development and validation in the EDSP (background information is available here. The validation process follows that described in the Validation and Regulatory Acceptance of Toxicological Test Methods (NIEHS 1997). The key validation steps follow.

  1. Initial Protocol Development -- The development and/or review of Endocrine Disruptor Screening Program (EDSP) initial protocols based on existing information and experience (past and current research). The initial protocols will serve as the starting point for all subsequent prevalidation studies.
  2. Prevalidation Studies -- The further development and optimization of specific EDSP initial protocols through targeted investigations. The targeted investigations will be designed to address questions necessary for an optimized, transferable protocol suitable for inter-laboratory validation studies.
  3. Validation Studies -- The design and interpretation of comparative inter-laboratory studies to establish the reliability and relevance of the EDSP optimized transferable protocols. Following validation, the optimized transferable protocols will provide the basis for endocrine disruptor test guidelines for regulatory use.
  4. Preparation of EDSP Method Validation Documents for External Peer Review -- All EDSP methods must be peer reviewed prior to approval for regulatory use.

The EDSP Tier 1 screening battery consists of an estrogen receptor binding/reporter gene assay, an androgen receptor binding/reporter gene assay, a steroidogenesis assay with minced testes, a rodent 3-day uterotrophic assay, a rodent 20-day pubertal female assay with thyroid; a rodent 5 to 7 day Hershberger assay; a frog metamorphosis assay, and a fish reproductive screen gonadal recrudescence assay. The EDSP Tier 2 testing battery consists of a two-generation mammalian reproductive toxicity study or a less comprehensive test (e.g., alternative mammalian reproductive test), and avian, fish, and mysid shrimp reproduction studies. The status of the ongoing validation work is summarized below and will be further discussed.

Literature ReviewInitial Protocol DemonstrationPre-validation StudiesValidation Studies
ER/AR bindingxx
HTPS for priority settingx
QSAR Model Developmentx
Pubertal male assayxxx
Frog metamorphosis assayx
Fish Reproductive Screenx
Placental aromatase
In utero/lactation assay
Mammalian 2-genx
Fish 2-gen
Avian 2-genx
Amphibian 2-gen
Invertebrate 2-gen Tx