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Use Of Genomics In Chemical Mode Of Action Analysis
Citation:
CORTON, C. Use Of Genomics In Chemical Mode Of Action Analysis. Presented at ECETOC meeting on (Eco)Toxicogenomics, Malaga, SPAIN, December 06 - 07, 2007.
Impact/Purpose:
This abstract is a summary of a talk given by Dr. Chris Corton at the ECETOC meeting on (Eco)Toxicogenomics in Malaga, Spain in Dec., 2007. The meeting focused on the potential uses of genomic data in chemical risk assessment but also dealt with additional subjects related to the generation and analysis of genomic, proteomic and metabolomic data. Dr. Corton gave a talk on the use of genomic data in mode of action risk assessment. There were a number of experts in toxicogenomics at the meeting and Dr. Corton benefited from informal discussions about genomics and risk assessment. Dr. Corton also gained a better understanding of the European system of toxicological assessment of chemicals including the potential use of genomics. One beneficial outcome of the meeting could be an American-European workgroup to evaluate case studies of the use of genomics in risk assessment.
Description:
The US-EPA has spent considerable effort in the last few years to develop guidance on the use of genomics to inform the process for determining effects on humans and ecological species. A number of workgroups within the Agency have developed consensus documents that cover a wide range of issues related to the use of genomics in risk assessment. In 2002, the EPA issued the Interim Policy on Genomics which provided initial guidance concerning how and when genomics information should be used to assess the risks of environmental contaminants under regulatory programs within the agency (1). In 2004, the document Potential Implications of Genomics for Regulatory and Risk Assessment Applications at EPA (2) identified four areas of oversight likely to be influenced by genomics data. These were (i) the prioritization of contaminants and contaminated sites for cleanup efforts, (ii) environmental monitoring at known or potentially contaminated sites, (iii) provisions in reporting chemical submissions and (iv) chemical risk assessment. The paper also identified a critical need for defining analysis and acceptance criteria the EPA would use for genomics information in understanding chemical toxicity and regulatory applications. This guidance encourages and supports continued genomics research for understanding the molecular basis of toxicity. While the guidance states that genomics data alone is currently insufficient to be used as the basis for risk assessment and management decisions, genomics data may help support arguments for mode of action in a weight-of-evidence approach for human or ecological health risk assessments in combination with all the other information the EPA considers for a particular assessment or decision. In 2007, an EPA Workgroup completed a document Interim Guidance for Microarray-Based Assays: Regulatory and Risk Assessment Applications at EPA that is currently undergoing peer-review. This document describes (i) the types of data that could be submitted to the EPA when using microarrays, (ii) approaches to quality assessment parameters of the submitted microarray data, (iii) microarray data analysis approaches and (iv) issues relevant to data management and storage for microarray data submitted to, or used by the EPA. This interim guidance also identifies future actions that are needed to better incorporate genomics information into the EPA's risk assessments as well as the regulatory decision making process. As mentioned above one of the areas identified in the 2004 guidance document impacted by genomics data is chemical risk assessment. Genomics data will complement conventional toxicological data to help identify key events necessary for a chemical to work through a defined mode of action. Comparing the transcript profiles between test species and human-derived samples will aid in determining the plausibility of the animal key events in humans. Genomics will likely one day decrease the overall uncertainty of a mode of action because of the global assessment of expression and individual variability of all genes in a genome. One of the major hurtles for optimizing usefulness of genomic data in mode of action based risk assessments has been the inability to comprehensively interpret changes in gene expression, especially those changes that directly impact key events in chemical mode of action. Currently, most investigators interpret the profiles generated essentially in the absence of comprehensive consideration of historical genomic data that may be extremely useful in determining a chemical mode of action. EPA and other organizations are developing or have developed databases to compare experimental profiles with historical data (e.g., references 3 and 4). This approach is even more powerful when coupled with the analysis of phenotypic endpoints that link the gene expression changes to toxicity. Additional tools are necessary to develop signatures for different key events or modes of action. Biochemical or genetic aproaches are available that when coupled to transcript profiling can aid in the interpretation of the gene expression changes. These can include profiling comparisons in wild-type and genetically modified mice, the use of techniques that can “knock down” gene expression (e.g., RNAi), and use of biochemical inhibitors of key events (e.g., inhibitors of oxidative stress in carcinogenesis). Additional approaches that have yielded more informative profiles have included dose-response and time course studies, comparisons between tissues and between active versus structurally similar but inactive compounds. Although the field of toxicogenomics is developing improved capabilities for interpretation of transcript profiles, considerable collaborative efforts are needed for development of databases of genomic data useful for comparison of newly generated profiles and for the establishment of a consensus on the information necessary to establish a key event in a chemical mode of action. One of the recommendations of the EPA interim guidance on genomic data is to identify and critically evaluate microarray data in a series of case studies with the goal to determine the utility of genomic analyses in risk assessment and regulatory applications. In the following example, one case study is described in which we have performed a toxicogenomic dissection of the transcript profiles after exposure to a chemical of interest to the EPA. To aid in the interpretation of the profiles we compared the profiles using one of the tools listed above (i.e., transgenic mice). This work has been recently published (5). A number of perfluorinated alkyl acids including perfluorooctanoic acid (PFOA) elicit effects similar to peroxisome proliferator chemicals (PPC) in mouse and rat liver. There is strong evidence that PPC cause many of their effects linked to liver cancer through the nuclear receptor peroxisome proliferator-activated receptor alpha (PPARalpha). To determine the role of PPARalpha in mediating PFOA transcriptional events, we compared the transcript profiles of the livers of wild-type or PPARalpha-null mice exposed to PFOA or the PPARalpha agonist WY-14,643 (WY). After 7 days of exposure, 85% or 99.7% of the genes altered by PFOA or WY exposure, respectively were dependent on PPARalpha. The PPARalpha-independent genes regulated by PFOA included those involved in lipid homeostasis and xenobiotic metabolism. Many of the lipid homeostasis genes including acyl-CoA oxidase (Acox1) were also regulated by WY in a PPARalpha-dependent manner. The increased expression of these genes in PPARalpha-null mice may be partly due to increases in PPARgamma expression upon PFOA exposure. Many of the identified xenobiotic metabolism genes are known to be under control of the nuclear receptor CAR and the transcription factor Nrf2. There was excellent correlation between the transcript profile of PPARalpha-independent PFOA genes and those of activators of CAR including phenobarbital and TCPOBOP but not those regulated by the Nrf2 activator, dithiol-3-thione. These results indicate that PFOA alters most genes in wild-type mouse liver through PPARalpha, but that a subset of genes are regulated by CAR and possibly PPARgamma in the PPARalpha-null mouse. As the expression and activation of the human counterparts of these nuclear receptors are well characterized, future research should focus on the possible involvement of these receptors in human liver through activation and regulation of genes involved in hepatocyte growth control. These studies will help to determine the human relevance of exposures to PFOA and the risk for human liver cancer. Although the PFOA case study above is just one case study of the use of genomics in chemical mode of action research, many other examples exist in the literature including those generated by EPA scientists and those in the European community. There is a great deal of uncertainty and lack of consensus as to the impact of these studies on chemical risk assessment. What is needed is the establishment of genomics working groups who can critically evaluate microarray data in a risk assessment context much the same way that pathology working groups evaluate tissue lesions. The workgroups would be useful to both regulatory agencies and the chemical industries to define the minimal criteria to establish key events in modes of action, define biomarkers of exposure and toxicological effects, refine data quality and analysis standards and highlight the potential usefulness of genomic data in defining mode of action and reducing uncertainty in risk assessment. Ample opportunities exist for US and European scientists to participate in genomics working groups to develop a global consensus on risk assessment issues of mutual interest.