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Session E3: Validation and Implementation of Alternative Methods for Identifying Endocrine Disrupters and Reproductive ToxicantsChairs: Gary Timm (USA) and Tohru Inoue (Japan) E3: Practical Approaches to Testing and Validation for Possible Endocrine Disrupters with Respect to a Mechanism of Action The testing strategies for EDC's consist of three major components, such as in silico database screening system, in vitro high-throughput screening system, and in vivo screening system. These should be followed by possible definitive testing system. The formers are supposed to be potent in negative selection, whereas the latter is in positive selection. An in silico system consists of a variety of mathematical models, including the computer-oriented docking model and other 3D-QSR models, such as CoMFA. A high-throughput screening system consists of a variety of in vitro assay systems, such as yeast-based reporter gene assays, and a variety of receptor-binding and transcription activation assays using established mammalian cell lines including human cells. An in vivo screening including uterotrophic and/or Hershberger assay is supposed to be potentially useful system to cover comprehensive screening, although there are exceptions. Neither a perfect positive screening system nor a perfect negative screening system is currently available. Practical approaches are needed to cover the EDC screening issue. E3: An Alternative Strategy for the Assessment of Chemicals for Endocrine Disrupting Effects There is currently no consensus as to a satisfactory definition of an "endocrine disruptor" (ED), and no direct evidence for a causal link between exposure to such agents and the development of adverse health effects in humans. Also lacking is information regarding human exposure to suspected ED's. The myriad of possible mechanisms by which an ED could affect the human endocrine system, compounded by the absence of an adequate operational definition or evidence of a causal association and human exposure data, makes the development of a meaningful chemical-assessment strategy highly problematic. However, several countries, in association with the Organization for Economic Cooperation and Development (OECD), are pushing forward with a large-scale chemical-assessment strategy based almost exclusively on animal tests. This presentation addresses the animal welfare implications of current proposed programs, presents an alternative strategy for screening and testing chemicals for ED effects, and recommends priority targets for research funding to accelerate the development and validation of promising non-animal tests and testing strategies. E3: The U.S. Environmental Protection Agency's (EPA's) Endocrine Disrupter Screening Program (EDSP): Perspectives and Status Gary E. Timm. Office of Science Coordination and Policy (7201M), U. S. Environmental Protection Agency, Washington, DC 20460, USA. Timm.Gary@epa.gov. The speaker will discuss the philosophy underlying the EDSP and the approach taken in developing the assays for the Tier 1 screening battery and Tier 2 tests. This philosophy was expressed in the report of the Endocrine Disrupter Screening and Testing Advisory Committee and several other expert workshops that were asked to investigate how to design an endocrine screening program. The progress and challenges in validating the assays and implementation status of the EDSP will be discussed. Although many assays being validated now rely on animals, EPA hopes to develop and incorporate new technology in the EDSP that will replace animals or make better use of animals. The speaker will discuss future possibilities for reducing the number of animals used. E3: Candidate Chemicals for Validation of In Vitro Estrogen and Androgen Receptor Binding and Transcriptional Activation Assays In 1998, the U.S. Environmental Protection Agency's Endocrine Disrupter Screening and Testing Advisory Committee (EDSTAC) recommended four types of in vitro test methods for identifying possible estrogen-disrupting substances. These assays are estrogen (ER) and androgen receptor (AR) binding, and transcriptional activation of estrogen and androgen responsive genes. A comprehensive review of the literature identified more than 600 and 200 chemicals that have been tested, respectively, for ER and AR binding in numerous assays. To facilitate evaluation of the comparative usefulness of the current and future assays, a list of chemicals that should be considered for validation studies was prepared. The potency of the ER and AR binding positive substances covered seven orders of magnitude. Chemical selection criteria included the potency of the chemical, the reliability of the data, its chemical class, if the chemical is used in commerce or found in the environment, and its commercial availability. Based on these criteria, 30 ER and 30 AR positive and 5 ER and 5 AR binding negative substances were selected. The use of a standard list of chemicals in future validation studies will facilitate determination of the acceptability of in vitro and in vivo assays and test batteries for inclusion in the screening program for endocrine active substances. Supported by NIEHS Contract N01-ES-85424. E3: Comparative Performance of Estrogen Receptor (Er) Binding and Transcriptional Activation Assays for Use as In Vitro Endocrine Disrupter Screening Methods Recent studies indicate that a variety of natural and anthropogenic substances can act as hormonal agonists or antagonists. The public health concern surrounding these finds prompted U.S. legislation requiring a federal regulatory screening program that identifies potential endocrine-active substances. In vitro ER binding and transcriptional activation assays have been proposed as screening methods to identify substances that may mimic or block the activity of endogenous estrogen. As a step toward identifying assays that should be given priority for further validation studies, results from published reports on substances tested in ER binding and transcriptional activation assays were compiled and compared. This poster presents a summary of the available data on ER binding and transcriptional activation assays reported in the literature and assesses the various assays for their comparative effectiveness in identifying estrogen-active substances. Assays were prioritized for future validation studies based on their comparative performance. Major strengths and limitations of different receptor binding and transcriptional activation assays that were considered in prioritizing assays will be presented. Performance-based prioritization of assays for further validation studies should facilitate evaluation and selection of the most promising assays for an effective screening program. Supported by NIEHS Contract NO1-ES-85424. E3: Ideology Masquerading as Science: The Case of Endocrine Disrupter Testing Programs The global move to develop novel testing methods and strategies to identify suspected endocrine disrupting chemicals offers a unique opportunity to move away from traditional animal testing paradigms in this new area of regulatory concern. Regrettably, programs under development, both in the United States and internationally through the Organization for Economic Cooperation and Development (OECD), have thus far failed to consider in vitro and other non-animal test methods as more than "pre-screening" or "priority-setting" tools in a larger (animal) testing strategy. Validation efforts to date have focused almost exclusively on the modification of existing animal tests to detect "endocrine effects," with no demonstrable effort to promote international coordination or support for the development and validation of relevant non-animal test systems. The current orientation in these programs reflects ideological, rather than scientific, imperatives, and undermines the commitments of both the U.S. government and the OECD with respect to the Three R's and the minimization of animal testing.
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