The simplified Whole Embryo Culture the throughput increase of a validated tool for developmental toxicology Marta Barenys CERETOX – Centre de Recerca en Toxicologia, Parc Científic de Barcelona IUF – Leibniz Research Institute for Environmental Medicine, Düsseldorf
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The simplified Whole Embryo Culture
the throughput increase of a validated tool for developmental toxicology
Marta Barenys
CERETOX – Centre de Recerca en Toxicologia, Parc Científic de Barcelona IUF – Leibniz Research Institute for Environmental Medicine, Düsseldorf
A. Yolk sac and blood vessels B. Allantois C. Flexion D. Heart E. Caudal neural tube F. Hind brain G. Mid brain H. Fore brain J. Otic system K. Optic system L. Olfactory system M. Branchial bars N. Maxillary process P. Mandibular process Q. Fore limb R. Hind limb
ECVAM Prediction Model 2
ICmax mal
ICNOEC TMS
IC50 3T3
I > II & III = non embryotoxic
II > I & III = weak embryotoxic
III > I & II = strong embryotoxic
Evaluation
Crown-rump length
Somite number
Total Morphological Score:
Malformations
„a demanding procedure which might be unsuitable for routine use as a first step in a large scale screening program“
European Partnership for Alternative Approaches to Animal Testing (EPAA) Questionnaire survey - Alternative methods for reproductive toxicity testing Aim: to investigate the real use of each alternative method among industries Participants: 11 large international pharmaceutical and chemical companies ECVAM DataBase service on Alternative Methods to animal experimentation (DB-ALM)
WEC: 6 companies substance related mechanistic studies screening or priority testing regulatory purposes studies on species differences (rat/rabbit) part of integrated testing strategy or stand-alone test
DB-ALM report Alternative methods for reproductive toxicity testing
39 total
21 developmental toxicity
9 really used
ZFET - Zebrafish Embryo Test EST - Embryonic Stem Cell Test WEC - Whole Embryo Culture
Advantages: Whole organism Same species than in in vivo reproductive toxicity assays High predictivity High reproducibility Disadvantages: High expertise required High cost Labor intensive Low throughput
Simplification achieved by: Statistical prediction model supported by one test concentration two structural endpoints (spinal cord – heart) one growth endpoint (n. of somites) Prediction model built with data from: 59 compounds (33 teratogens and 26 nonteratogens) +11 compounds ( 5t & 6nt) for independent validation
Aims: ↑ throughput ↓ animal use without reducing predictivity
Summary Simplified study design Simplified data analysis Prediction model does not require cytotoxicity data from a cell line No significant reduction in predictivity 50 % reduction of animal use and operational costs
Substance related mechanistical studies ECVAM protocol n.123 Simplified version Screening or priority testing