PREDICTION OF TERATOGENIC POTENCY OF VALPROATE ANALOGS USING CEREBELLAR AGGREGATION CULTURES

The aim of this study was to develop a novel in vitro system suitable for preclinical testing for developmental toxicity of drugs. An assay system consisting of primary cultures of dissociated cerebella from 6- day-old mice was chosen, since it allowed quantification of neuronal a ggregation and fasciculated neurites. A human teratogen, the antiepile ptic drug valproic acid (VPA), as well as its structural analogues, (/-)-4-en-VPA and E-2-en-VPA, with varying teratogenic activities, were tested and found to affect aggregation and fiber formation of cerebel lar neurons. Based on a dose-response study, the concentrations of com pounds causing 50% inhibition (IC50) of formation of thick and thin fi bers were determined. The lowest IC50 values were found for VPA (52 +/ - 7 and 86 +/- 11 mu M for thick and thin fibers, respectively), which in vivo caused the highest rate of exencephaly among the three compou nds tested, (+/-)-4-en-VPA exhibited intermediate values (150 +/- 30 a nd 300 +/- 40 mu M), whereas the highest IC50 values were found for E- 2-en-VPA (260 +/- 42 and 430 +/- 40 mu M). The latter compound does no t induce neural tube defects, but has been shown to have neurobehavior al effects in prenatally exposed animals. Subsequently, the purified S - and R-enantiomers of 4-yn-VPA (teratogenic and non-teratogenic, resp ectively) were tested for their effects on aggregation and fiber forma tion of the cerebellar neurons. Treatment with S-4-yn-VPA resulted in pronounced changes in numbers of aggregates and fasciculated processes compared to the cultures treated with R-4-yn-VPA, indicating that the intrinsic stereoselective potency of the enantiomers may be correlate d to the difference in their effects on cerebellar neurons in vitro. T hus, the teratogenic potency of VPA and its analogues correlated with their effects on aggregation of neural cells and formation of fascicul ated neurites in primary cultures of dissociated cerebella, indicating that the in vitro-assay system employed may be used as a pre-screenin g test for prediction of teratogenic potency of drugs. Copyright (C) 1 997 Elsevier Science Ireland Ltd.