Article thumbnail

The Embryonic Stem Cell Test as Tool to Assess Structure-Dependent Teratogenicity: The Case of Valproic Acid

By Christian Riebeling, Ralph Pirow, Klaus Becker, Roland Buesen, Daniel Eikel, Johanna Kaltenhäuser, Frauke Meyer, Heinz Nau, Birgitta Slawik, Anke Visan, Jutta Volland, Horst Spielmann, Andreas Luch and Andrea Seiler

Abstract

Teratogenicity can be predicted in vitro using the embryonic stem cell test (EST). The EST, which is based on the morphometric measurement of cardiomyocyte differentiation and cytotoxicity parameters, represents a scientifically validated method for the detection and classification of chemicals according to their teratogenic potency. Furthermore, an abbreviated protocol applying flow cytometry of intracellular marker proteins to determine differentiation into the cardiomyocyte lineage is available. Although valproic acid (VPA) is in worldwide clinical use as antiepileptic drug, it exhibits two severe side effects, i.e., teratogenicity and hepatotoxicity. These limitations have led to extensive research into derivatives of VPA. Here we chose VPA as model compound to test the applicability domain and to further evaluate the reliability of the EST. To this end, we study six closely related congeners of VPA and demonstrate that both the standard and the molecular flow cytometry-based EST are well suited to indicate differences in the teratogenic potency among VPA analogs that differ only in chirality or side chain length. Our data show that identical results can be obtained by using the standard EST or a shortened protocol based on flow cytometry of intracellular marker proteins. Both in vitro protocols enable to reliably determine differentiation of murine stem cells toward the cardiomyocyte lineage and to assess its chemical-mediated inhibition

Topics: IN VITRO Methods and Alternatives to Animals
Publisher: Oxford University Press
OAI identifier: oai:pubmedcentral.nih.gov:3061479
Provided by: PubMed Central

To submit an update or takedown request for this paper, please submit an Update/Correction/Removal Request.

Suggested articles

Citations

  1. (2009). A review of the implementation of the embryonic stem cell test (EST). The report and recommendations of an ECVAM/ReProTect Workshop.
  2. (2010). A robust procedure for comparing multiple means under heteroscedasticity in unbalanced designs.
  3. (2009). Anticonvulsants in bipolar disorders: current research and practice and future directions. Bipolar Disord.
  4. (2007). Antiepileptic drugs in migraine: from clinical aspects to cellular mechanisms. Trends Pharmacol.
  5. (2008). Assessment of the embryonic stem cell test and application and use in the pharmaceutical industry. Birth Defects Res.
  6. (2005). Bioassay analysis using
  7. (2004). Can the pharmaceutical industry reduce attrition rates?
  8. (2009). Carnitine in the treatment of valproic acid-induced toxicity.
  9. (2005). Depletion of latent HIV-1 infection in vivo: a proof-of-concept study.
  10. (2009). Embryonic stem cell test remastered: comparison between the validated EST and the new molecular FACS-EST for assessing developmental toxicity in vitro.
  11. (2008). Evaluation of the embryotoxic potency of compounds
  12. (1996). Further branching of valproate-related carboxylic acids reduces the teratogenic activity, but not the anticonvulsant effect.
  13. (2009). Histone deacetylase inhibitors as anti-neoplastic agents.
  14. (2001). Histone deacetylase is a direct target of valproic acid, a potent anticonvulsant, mood stabilizer, and teratogen.
  15. (1982). Immunochemical analysis of myosin heavy chain during avian myogenesis in vivo and in vitro.
  16. (2004). Improvement of an in vitro stem cell assay for developmental toxicity: the use of molecular endpoints in the embryonic stem cell test.
  17. (2009). In vitro developmental toxicity test detects inhibition of stem cell differentiation by silica nanoparticles.
  18. (2010). In vitro developmental toxicology assays: a review of the state of the science of rodent and zebrafish whole embryo culture and embryonic stem cell assays. Birth Defects Res.
  19. (2006). In vitro embryotoxicity testing of metals for dental use by differentiation of embryonic stem cell test.
  20. (1993). In vivo formation of the thiol conjugates of reactive metabolites of 4-ene VPA and its analog 4-pentenoic acid. Drug Metab.
  21. (2010). Metabolic activation capacity by primary hepatocytes expands the applicability of the embryonic stem cell test as alternative to experimental animal testing.
  22. Mood stabilizers in Alzheimer’s disease: symptomatic and neuroprotective rationales.
  23. (2003). New CNS-active drugs which are second-generation valproic acid: can they lead to the development of a magic bullet?
  24. (2006). Prediction of in vivo embryotoxic effect levels witha combination of in vitro studies and PBPK modelling.
  25. (1999). Prevalidation of the embryonic stem cell test (EST)—a new in vitro embryotoxicity test.
  26. (2009). R: A Language and Environment for Statistical Computing. R Foundation for Statistical Computing:
  27. (1983). Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays.
  28. (2009). Relative developmental toxicity of glycol ether alkoxy acid metabolites in the embryonic stem cell test as compared with the in vivo potency of their parent compounds.
  29. (2010). ReProGlo: a new stem cell-based reporter assay aimed to predict embryotoxic potential of drugs and chemicals.
  30. (2002). Selection of test chemicals for the ECVAM international validation study on in vitro embryotoxicity tests.
  31. (2008). Simultaneous inference in general parametric models.
  32. (2010). Statistical methods for assessing agreement between two methods of clinical measurement.
  33. (2006). Teratogenic effects mediated by inhibition of histone deacetylases: evidence from quantitative structure activity relationships of 20 valproic acid derivatives.
  34. (2002). The ECVAM international validation study on in vitro embryotoxicity tests: results of the definitive phase and evaluation of prediction models. European Centre for the Validation of Alternative Methods.
  35. (2007). The embryonic stem cell test for the early selection of pharmaceutical compounds.
  36. (1997). The embryonic stem cell test, an in vitro embryotoxicity test using two permanent mouse cell lines: 3T3 fibroblasts and embryonic stem cells.
  37. (1992). The enantiomers of the valproic acid analogue 2-n-propyl-4-pentynoic acid (4-yn-VPA): asymmetric synthesis and highly stereoselective teratogenicity in mice.
  38. (1985). The in vitro development of blastocyst-derived embryonic stem cell lines: formation of visceral yolk sac, blood islands and myocardium.
  39. (1979). The two kinds of linear discriminant functions and their relationship.
  40. (2009). The way forward in reproductive/developmental toxicity testing.
  41. (2010). Transcriptomics-based identification of developmental toxicants through their interference with cardiomyocyte differentiation of embryonic stem cells.
  42. (2010). Unexpected common mechanistic pathways for embryotoxicity of warfarin and lovastatin.
  43. (2006). Use of murine embryonic stem cells in embryotoxicity assays: the embryonic stem cell test.
  44. (2004). Validation of the embryonic stem cell test in the international ECVAM validation study on three in vitro embryotoxicity tests.
  45. (1999). Valproate: a reappraisal of its pharmacodynamic properties and mechanisms of action.
  46. (2009). Valproic acid in pregnancy: how much are we endangering the embryo and fetus?