ID#

240

Location:

Exhibit Hall

Time of Presentation:

March 22, 9:30 AM

Category:

In Vitro/Alternative Animal Models

Phase I and II Results of a Validation Study to Evaluate In Vitro Cytotoxicity Assays for Estimating Rodent and Human Acute Systemic Toxicity
M.W. Paris^1,2, J.A. Strickland^1,2, W.S. Stokes¹, S. Casati³, R.R. Tice^1,2, H. Raabe⁴, C. Cao⁵, R. Clothier⁶, J. Harbell⁴, G. Mun⁴, A. Sizemore⁴, G. Moyer⁴, J. Madren-Whalley⁵, C. Krishna⁵, M. Owen⁶, N. Bourne⁶, J. Haseman⁷, P. Crockett⁸, M. Wenk⁹, M. Vallant⁷, A. Worth³.
1. NICEATM, NIEHS, RTP, NC, USA.
2. ILS, Inc., RTP, NC, USA.
3. ECVAM, Ispra, , Italy.
4. IIVS, Gaithersburg, MD, USA.
5. US Army, APG, MD, USA.
6. Univ. of Nottingham, Nottingham, , UK.
7. NIEHS, RTP, NC, USA.
8. ASI, RTP, NC, USA.
9. BioReliance, Rockville, MD, USA.

Studies have identified a correlation between in vitro cytotoxicity and acute oral toxicity. NICEATM and ECVAM initiated a three-phase multi-laboratory validation study to evaluate the usefulness of two standardized in vitro basal cytotoxicity assays for estimating rodent and human acute toxicity and the extent that they may reduce animal use. Seventy-two coded chemicals (12 from each of five acute oral hazard categories; 12 unclassified/nontoxic chemicals) will be tested in mouse 3T3 fibroblasts and in normal human epidermal keratinocytes (NHK) using neutral red (NR) uptake assays. Protocols were optimized after each of the first two phases to minimize intra- and inter-laboratory variation prior to testing 60 chemicals in Phase III. Phase Ia established the historical databases for the positive control chemical, sodium laurel sulfate (SLS), for each of three laboratories. Three chemicals were tested in Phase Ib and nine in Phase II. Technical challenges arose in Phases Ia/Ib (i.e., formation of NR dye crystals; uneven growth of NHKs; slow growth of 3T3 cells) that were resolved with Phase II protocols. Significant variation in NHK growth occurred in Phase II with various lots of media and supplements. The optimized final protocols are being tested in Phase III. Rodent oral LD50 values were estimated using prediction models based on the Registry of Cytotoxicity data and Phase I/II results. Human toxicity will be estimated using a prediction model based on data from human poisoning reports and the Multicentre Evaluation of In Vitro Cytotoxicity (MEIC). Supported by: N01-ES-35504, N01-ES-75408; EPA IAG DW-75-93893601-0; European Commission 19416-2002-04 F2ED ISP GB.