Testing & Prioritizing Chemicals: Current & In-Development Predictive Tools
EPA is required to test a wide range of chemicals to determine if they are endocrine disrupting chemicals (EDCs). Traditional toxicology tests using animals are lengthy and expensive, as is field work to assess effects. As a result of these constraints EPA researchers have developed a number of streamlined tests for potential endocrine disrupting chemicals that EPA's Endocrine Disruption Screening Program (EDSP) uses for its Tier 1 testing battery. These tests range from 21 day reproductive studies on fish to cellular and biochemical in vitro (test tube) assays. But researchers examining the results of such tests face an additional challenge, interpreting the results. EPA researchers are approaching this challenge by harnessing the tools of an emerging scientific discipline, systems biology. Systems biologists develop tools to predict the effects of a biochemical perturbation - such as an EDC interfering with the action of estrogen - on other aspects of biology, such as effects on an individual organism and a population of organisms. Using such tools they can, for instance, look at the results of a one-day test for a potential EDC and predict whether that chemical is likely to affect the ability of fish to develop, survive and reproduce and whether these effects will influence the ecosystem.
EPA researchers have developed and are currently developing more tools that can be used to predict if a chemical is a potential EDC. EPA researchers are working to define and use "Adverse outcome pathways" for these predictive tools. An adverse outcome pathway is a biological process important to a human or wildlife species functioning normally. If these pathways are perturbed by a chemical or some other activity, the perturbation could lead to a negative effect which can lead to a function in the body not occurring normally. For instance if the activation of the estrogen receptor, the molecule that carries out the actions of estrogen, is blocked, this will have multiple effects on male and female fish. Among other effects, at the cellular level, it is predicted that blocking the activation of the estrogen receptor could increase the production of the egg yolk proteins by liver cells; at the level of the organ, it could impair the development of the testes; at the organismal level, it could reduce fecundity and skew the sex ratio of offspring; at the ecological level, result in declining fish populations. Such a sequence of events connects an initiating event (inhibition of the estrogen receptor) with an adverse outcome (fish population decline) which is relevant to assessing chemicals for potential risks.
Researchers have identified adverse outcome pathways for various hormonal systems and they have integrated them into a broader framework, called a Graphical Systems Model. Graphical models connect multiple hormonal systems at multiple biological levels - from molecule to cell to organism to population. For instance, EPA researchers have developed a systems model for the hormonal links between the brain and the sex organs in fish. It depicts the interaction of over 140 proteins and simple molecules, the regulation of 25 different genes and over 300 different biological processes. One way the researchers tested the model is by examining the effects on the system of disrupting aromatase, a molecule that is responsible for synthesizing estrogen. Using their model, they first predicted the effects of disrupting aromatase - these effects included reduction in the production of estrogen, reduction in the number of eggs female fish make, and disruption related hormonal systems in the brain that regulate cholesterol levels and other physiological systems. The researchers then tested their model by examining what happened in fish treated with a drug that inhibits aromatase. They found that their model was correct - the predicted effects occurred. They also observed additional effects, some of which were measured by assessing changes in which genes are turned on and off in the ovary. Using such 'genomics' tools they were then able to incorporate other effects into their model.
One advantage of these systems biology methods is that they can ultimately be used to ask informed questions about chemicals for which little is known.
Results and Impact
EPA researchers have already developed tests that are currently being used by EDSP to screen chemicals for potential EDCs. In addition to these current methods, new tests and models are being developed using the systems biology approach. Systems biology approaches such as adverse outcome pathways and graphical systems models are being used to predict if a chemical is an EDC. Such models and knowledge can leverage results from laboratory and field tests to predict the various endocrine related effects. Research efforts in this area are also useful for the EDSP, where developed tests and predictive tools are used to assess chemicals for potential endocrine disruption.