Xenoendocrine disrupters-tiered screening and testing Filling key data gaps  L.E. Gray, Jr. a, , J. Ostby a , V. Wilson a , C. Lambright a , K. Bobseine a , P. Hartig a , A. Hotchkiss a,b , C. Wolf a,c , J. Furr a , M. Price b , L. Parks b , R.L. Cooper a , T.E. Stoker a , S.C. Laws a , S.J. Degitz d , K.M. Jensen d , M.D. Kahl d , J.J. Korte d , E.A. Makynen d , J.E. Tietge d , G.T. Ankley d a Reproductive Toxicology Division, NHEERL, ORD, USEPA, MD 72, Research Triangle Park, NC 27711, USA b USEPA/NCSU Cooperative Research Program and NCSU Zoology Department, MD 72, Research Triangle Park, NC 27711, USA c NCSU Department of Toxicology, MD 72, Research Triangle Park, NC 27711, USA d Midcontinent Ecology Division, NHEERL, ORD, USEPA, MD 72, Research Triangle Park, NC 27711, USA Abstract The US Environmental Protection Agency (EPA) is developing a screening and testing program for endocrine disrupting chemicals (EDCs) to detect alterations of hypothalamic /pituitary  /gonadal (HPG) function, estrogen (ER), androgen (AR) and thyroid hormone synthesis and AR and ER receptor-mediated effects in mammals and other animals. High priority chemicals would be evaluated in the Tier 1 Screening (T1S) battery and chemicals positive in T1S would then be tested (Tier 2). T1S includes in vitro ER and AR receptor binding and/or gene expression, an assessment of steroidogenesis and mammalian (rat) and nonmammalian in vivo assays (Table 1). In vivo, the uterotropic assay detects estrogens and antiestrogens, while steroidogenesis, antithyroid activity, (anti)estrogenicity and HPG function are assessed in a ‘Pubertal Female Assay’. (Anti-) androgens are detected in the Hershberger Assay (weight of AR- dependent tissues in castrate-immature-male rats). Fish and amphibian assays also are being developed. The fathead minnow assay can identify EDCs displaying several mechanisms of concern, including AR and ER receptor agonists and antagonists and inhibitors of steroid hormone synthesis. An amphibian metamorphosis assay is being developed to detect thyroid-active substances. Several alternative mammalian in vivo assays have been proposed. Of these, a short- term pubertal male rat assay appears most promising. An in utero-lactational screening protocol also is being evaluated. For Tier 2, the numbers of endocrine sensitive endpoints and offspring (F1) examined in multigenerational tests need to be expanded for EDCs. Consideration should be given to tailoring T2, based on the results of T1S. Tier 1 and 2 also  DISCLAIMER: The research described in this article has been reviewed by the National Health Environmental Effects Research Laboratory, U.S. Environmental Protection Agency, and approved for publication. Approval does not signify that the contents necessarily reflect the views and policies of the Agency nor does mention of trade names or commercial products constitute endorsement or recommendation for use.  Corresponding author. Present address: Endocrinology Branch, Reproductive Toxicology Division, RTD, NHEERL, ORD, USEPA, RTP, MD 72, Research Triangle Park, NC 27711, USA. Tel.:  /1-919-541-7750; fax:  /1-919-541-4017 E-mail address: gray.earl@epa.gov (L.E. Gray, Jr.). Toxicology 181  /182 (2002) 371  /382 www.elsevier.com/locate/toxicol 0300-483X/02/$ - see front matter # 2002 Elsevier Science Ireland Ltd. All rights reserved. PII:S0300-483X(02)00469-9