OECD FELLOWSHIP SUMMARY REPORT

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1 OECD FELLOWSHIP SUMMARY REPORT Name: Rolf Altenburger Subject: Status and research need in toxicogenomic mixture toxicity analysis Host Institution: University of Queensland, Brisbane, QLD Host Supervisor: Beate Escher Dates of Fellowship: 15. April to 14.October, 2011 Consent: I consent to my report being posted on the Co-operative Research Programme s website

2 Relevance To the Co-operative Research Programme objectives This project contributed to scientific advances, through reviewing and bridging of methodologies of currently separated fields of applied biological science, toxicogenomics and mixture toxicology. By cooperative efforts of the collaborators, mutual understanding of different approaches in different OECD countries has been fostered. To the Theme It is now widely held that the sustainable use and protection of natural resources is essential to support continued food production and quality of life for humans, domestic animals, and wildlife. Furthermore, natural resource stewardship efforts have led to systematic consideration and tiered approaches for the assessment of ecological risks from chemicals released to the environment, at many instances relying on OECD guideline protocols designed for testing of unwanted ecotoxic effects from individual chemicals. Organisms in the environment, however, rarely experience stress from individual contaminants alone but are exposed to several substances simultaneously or in sequence. The assessment of ecotoxic responses upon simultaneous mixture exposure has been the subject of several investigations and currently the EU is considering revising its entire environmental regulation to include explicit mixture assessments. In order to develop rational approaches for the assessment of combined effects that might be caused upon long-term low dose mixture exposure this project investigated approaches that offer the analysis of multivariate responses using postgenomic approaches such as differential detection of the transcriptome, proteome or metabolome of cells or tissues. To agricultural and food policy As we experience in the current days food as a worldwide traded commodity, agricultural and food policy heavily relies on scientific expert knowledge to secure and safeguard modern food production systems on an international scale. In this respect an often-overlooked issue is that of lack of systematic assessment of unintended or unwanted mixture exposure of humans, living stock and wildlife through food and feed. The US and EU chemical regulation have begun acknowledging the cumulative risk resulting from mixture exposure to pesticides and other chemicals, while in Australia efforts are made to deal with the combined stress resulting from effects of climate change (e.g. increase in water temperatures) and agricultural loads (fertilizer and herbicides) on ecosystem functioning (such as. The Great Barrier reef) During this project several meetings of scientific and other stakeholders were held to discuss current risk assessment approaches to these mixture exposure problems. The results of this fellowship project have advanced extrapolation techniques suitable for rational approaches to cumulative assessments of long-term effects from multiple contamination of natural resources or for combined stress evaluation regarding ecosystem functioning.

3 Objectives of the Fellowship The central objective of this research was to investigate the perspective for utilising models of concentration-response and combined effect for formulating quantitative relationships in toxicogenomic approaches. This would help to develop meaningful assessment tools for low dose mixture exposure situations. Major achievements 1. We reviewed the current approaches and evidence to mixture toxicity analysis from a toxicogenomic perspective. 2. Several presentations and seminars with Australian stakeholders from Universities and risk regulation were held to connect current domestic, international and EU perspective on the subject. 3. The involved research organizations ENTOX and UFZ arranged closer institutional cooperation, which already showed up in additional staff exchange in the field of chemical risk assessment. Follow-up Is a publication envisaged? Will this be in a journal or a publication? When will it appear? A first manuscript for publication has been submitted for publication as a review to the journal Environmental Science and Technology. It is entitled Mixture toxicity from a toxicogenomic perspective. A second manuscript Additive pressures of elevated sea surface temperatures and herbicides on symbiont-bearing foraminifera where the modeling approaches are applied to a combined stress experimental work from a collaborating Australian team is to be submitted following the fellowship. Further publications depend on the outcome of mixture experimentation that is planed for the home lab (UFZ) using the approaches identified as most promising for combined effect analysis of toxicogenomic data. Is your fellowship likely to be a start for collaboration between your home institution and your host? Yes. Exchange of further students and methods is planned. Especially joint ideas for integrated (human and environmental) assessment approaches will be followed up.

4 Is your research likely to result in protected intellectual property, novel products or processes? No. The results of our research remain typically within the public domain. Satisfaction Did your fellowship conform to your expectations? The fellowship was more than actually expected, as it provided ample training and exploration opportunities. A major aspect of this, is the OECD programme itself, which by its reliance on the capabilities of the selected fellows, i.e. due to its few administrative demands, really provides room for original thoughts and work. Will the OECD fellowship increase directly or indirectly your career opportunities? Please specify. My career opportunities will most probably profit indirectly as after several years of institutional demands for continuous strategic research planning, this fellowship provided the original opportunity to extent methodological knowledge to adjacent fields and from mainly techniquedriven exploration to modeling and assessment methodologies. This most probably will be a profit for application of our research in risk regulation and an added value for academic training of young scientists. International recognition of our work was also strengthened. Have you encountered any practical problems? None, which were unexpected. Please suggest any improvements in the fellowship Programme The greatest improvement I could think of is more senior scientists actually participating in it. So, my suggestions lead to opening the programmatic scope and allowing all stages of senior scientists to apply for projects. Additionally, as funding for cooperative project-oriented research initiatives with a wider international scope are still rare, this could be a future extension option. I am thinking of research projects (2-3 years with options for joint research or mutual exchange) that are comparable and competitive to the specific research projects of the EU. Advertising the Co-operative Research Programme How did you learn about it?

5 I learned of the OECD CRP through an academic evaluator of our group, who mentioned the programme. I than traced the programme on the Internet. What would you suggest to make it more "visible" Provide flyers for international conferences in the field; ask professional academic organizations and international institutions to announce the programme; let your homepage for the programme be linked to that of academic research organizations. Any issues you would like to record? I think this programme is a fantastic one and offers unique training and research opportunities to senior scientists.

6 Appendix Abstract for a manuscript entitled: Mixture toxicity revisited from a toxicogenomic perspective a review Submitted for publication to Environmental Science and Technology The advent of toxicogenomic techniques have raised large expectations that some of the central questions of mixture toxicology such as for mechanisms of low dose interactions can be provided with novel perspectives if not definite answers. After the first decade of experimental studies this review summarizes mixtures toxicity studies that address diagnostic, mechanistic or extrapolation questions. Since 2002 almost 40 studies were published with their major focus on mixture toxicity assessment by means of toxicogenomic techniques, mainly through microarray or qpcr techniques, though metabolomic and proteomic analysis of joint exposures have also been undertaken. It is now standard to explicitly state criteria for selected concentrations and provide insight into employed data transformation, and statistical treatment with respect to minimising sources of undue variation. Bioinformatic analysis of toxicogenomic data, by contrast, is still a field with diverse and rapidly evolving tools. The combined effect assessments achieved are discussed in the light of established toxicological dose-response and mixture toxicity models. For one, receptor-based assays seem to be most advanced towards establishing quantitative relationships between exposure and biological responses. Often transcriptomic responses are discussed based on the presence or absence of signals. As there are yet no consented ways of how to interpret these effects, there are ambiguous interpretations. Furthermore, mixture studies in their majority designed their experiments and compared their recorded outcomes against individual treatments i.e. focus was to retrieve signals of individual components under mixture exposure. This stands in stark contrast to our existing understanding of biological activity at the levels of chemical target interactions and apparent apical combined effects. Here models are employed to calculate expected combined effects based on information of the mixture components individual dose-response relationships. By joining these mixture effect models with toxicokinetic and -dynamic thinking we suggest a theoretical framework that may help to overcome the current limitation of providing mainly anecdotal evidence on mixture effects and progress into more hypothesis driven mixture studies. As ways forward we suggest to study and establish quantitative relationships between dose dependency and time dependency of responses. Moreover, development of data aggregation tools using bioinformatic and stress response concepts could play major roles in advancing the analysis of mixture toxicity.

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