Meeting Report: TestSmart’s DNT2

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Meeting Report: TestSmart’s DNT2

Jan van der Valk To appear in the ESTIV and NCA newsletters.

Published: December 17, 2008

TestSmart, DNT2: Creating a humane and efficient approach to developmental neurotoxicity testing.
12-14 November, 2008, Reston, VA, USA
Organised by Johns Hopkins University Center for Alternatives to Animal Testing
When you participate in a congress in a very busy time of the year far away from home you hope the visit is worth its time and money. The TestSmart DNT2 meeting in Reston, USA certainly was. The meeting excelled with clear and generally good presentations, lots of interactivity and discussions, and fair conclusions. For me, as a relative layperson in the area, there was a lot to learn.

About 80 scientists from industry, government (EPA), and academia participated in the meeting. The first day was marked by overviews and views on the different developments with respect to developmental neurotoxicity (DNT) testing. Kim Boekelheide (Brown University) set the stage by mentioning that although animal data have protected our health and safety for quite some time, this is mainly due to safety factors compensating for the lack of mechanistic information when extrapolating animal data to humans. There is now a need for multiple dose in vitro testing on different endpoints in high-throughput systems whereby cell-cell and organ interactions are taken into account.

Cynthia Bearer (Case Western Reserve University), paediatrician, put the need for DNT testing in the right perspective by summing up that in the US alone more than 5 million children are affected by some form of developmental neurotoxicological effect and that there are indications that some of the adult neural diseases, like Alzheimer’s and Parkinson’s, have their origin in the fetus.

Kevin Crofton (EPA) emphasised that there are many endpoints in DNT to be studied, that we know only a few of these and that it will be hard to finally choose a representative set of phenotypic endpoints and/or key events in the biochemical pathway. He stressed the need for data sharing and inter- and intra-laboratory comparisons of test methods.

Ellen Fritsche (University of Duesseldorf) discussed the application of human neural progenitor cells which are cultured as proliferating neurospheres. The advantage of the system is that the proliferating progenitor cells also express neural and glial markers, and can be used to study endpoints like proliferation, differentiation, migration and apoptosis.

William Mundy (EPA) covered the endpoints for neural connectivity, neurite outgrowth, synapse formation and function. He discussed the different aspects with illustrative slides and possible markers for the different processes.

Joseph Bressler (Johns Hopkins University) closed the day with an impressive overview of the remaining endpoints that also have a crucial role in neural development and function, and the different roles glial cells have in neural development. Glial cells are therefore also important targets for toxicants and should be taken into consideration when assessing chemical safety.

During the discussion many questions came up with regard to the endpoints: which of the many endpoints are relevant for DNT, which of these are realistic to measure in vitro and which ones will be used for screening purposes and which ones for making final decisions? It became clear that the area is still too young to give answers to these questions.

The second day focused on the models used to evaluate effects on the different endpoints.  Lucio Costa (University of Washington) gave an overview of the cell culture methods available to study (neurodevelopmental) neurotoxicity. Drawbacks of cell culture methods are that the target concentration is not known, compensatory mechanisms are not available and not all endpoints are covered. Co-cultures in which astrocytes, glial cells, etc, are also present might improve the test results.

Pamela Lein (Oregon Health & Science University) discussed non-mammalian models for DNT testing. Since several genes for neural developments have been conserved during evolution, simple organisms exhibit similar neural development processes as in mammals. These can be used to study DNT. Animals such as C. elegans, drosophila and zebra fish embryos were discussed as good candidates for DNT testing. In addition to direct effects on neuronal development these models may also give information on changes in behaviour.

After the introductions, two break-out groups discussed cell culture and alternative animal models for DNT testing. Several participants were invited to give a short presentation on their posters.  In the cell culture breakout group it was concluded that it is still difficult to establish what endpoints are relevant, that we need to generate data before prioritisation can be made and that similar culture conditions across laboratories should be aimed for.

In the afternoon session on data interpretation, integration and policy, Robert Kavlock (EPA) argued that a change in approach was needed: there are too many chemicals to be tested at too high a cost and we don’t have enough data for interpretation yet. He further discussed ToxCast, the US EPA chemical prioritization program in which over 300 chemicals are characterised for over 400 endpoints.

James Bus (Dow) discussed in vitro data in understanding human health effects. He particularly dealt with appropriate matching of in vitro dosimetry to human in vivo dosimetry to make a proper translation.

The last morning summed up the conclusions of the meeting and discussed the questions that still have to be solved:

  • What are we going to measure?
  • What does the data mean?
  • Which is the golden standard to compare in vitrodata with?
  • Does a positive result in a tier mean follow-up in an animal?
  • Does a negative result lead to acceptance or do we need confirmation?
  • How to link hazard to exposure and risk?
  • Do we have enough knowledge on the involved mechanisms?
  • In what way is DNT different from developmental toxicology?

My conclusion from the meeting is that we are still faced with questions that we also face in some other areas; that there is still a long way to go before we can get data from in vitro studies for which we have a relevant data interpretation model for the human risk evaluation. The best way forward, at least for the moment, to save time and money, seems to focus on the use of the simple organisms like zebra fish and C. elegans to evaluate hazard and possibly risk.

I tried to do my best to give you my impression of the meeting and hope that I have not mischaracterized any of the presentations. And I apologise to the speakers I have not covered here, mainly for the reason to keep this report reasonably short and readable. This was a very interesting and informative meeting, with good speakers and lively interactions. I look forward to DNT3 in Italy in 2009.
Jan van der Valk
Netherlands Centre for Alternatives to Animal Use