Lisa Ortego

Relevance Weighting of Tier 1 Endocrine Screening Endpoints by Rank Order

Weight of evidence (WoE) approaches are recommended for interpreting various toxicological data, but few systematic and transparent procedures exist. A hypothesis-based WoE framework was recently published focusing on the U.S. EPAs Tier 1 Endocrine Screening Battery (ESB) as an example. The framework recommends weighting each experimental endpoint according to its relevance for deciding eight hypotheses addressed by the ESB. Here we present detailed rationale for weighting the ESB endpoints according to three rank ordered categories and an interpretive process for using the rankings to reach WoE determinations. Rank 1 was assigned to in vivo endpoints that characterize the fundamental physiological actions for androgen, estrogen, and thyroid activities. Rank 1 endpoints are specific and sensitive for the hypothesis, interpretable without ancillary data, and rarely confounded by artifacts or nonspecific activity. Rank 2 endpoints are specific and interpretable for the hypothesis but less informative than Rank 1, often due to oversensitivity, inclusion of narrowly context-dependent components of the hormonal system (e.g., in vitro endpoints), or confounding by nonspecific activity. Rank 3 endpoints are relevant for the hypothesis but only corroborative of Ranks 1 and 2 endpoints. Rank 3 includes many apical in vivo endpoints that can be affected by systemic toxicity and nonhormonal activity. Although these relevance weight rankings (WREL ) necessarily involve professional judgment, their a priori derivation enhances transparency and renders WoE determinations amenable to methodological scrutiny according to basic scientific premises, characteristics that cannot be assured by processes in which the rationale for decisions is provided post hoc.

Estimating potential risks to terrestrial invertebrates and plants exposed to bisphenol A in soil amended with activated sludge biosolids.

Bisphenol A (BPA) is a high production volume substance primarily used to produce polycarbonate plastic and epoxy resins. During manufacture and use, BPA may enter wastewater treatment plants. During treatment, BPA may become adsorbed to activated sludge biosolids, which may expose soil organisms to BPA if added to soil as an amendment. To evaluate potential risks to organisms that make up the base of the terrestrial food web (i.e., invertebrates and plants) in accordance with international regulatory practice, toxicity tests were conducted with potworms (Enchytraeids) and springtails (Collembolans) in artificial soil, and six plant types using natural soil. No-observed-effect concentrations (NOEC) for potworms and springtails were equal to or greater than 100 and equal to or greater than 500 mg/kg (dry wt), respectively. The lowest organic matter-normalized NOEC among all tests (dry shoot weight of tomatoes) was 37 mg/kg-dry weight. Dividing by an assessment factor of 10, a predicted-no-effect concentration in soil (PNEC(soil)) of 3.7 mg/kg-dry weight was calculated. Following international regulatory guidance, BPA concentrations in soil hypothetically amended with biosolids were calculated using published BPA concentrations in biosolids. The upper 95th percentile BPA biosolids concentration in North America is 14.2 mg/kg-dry weight, and in Europe is 95 mg/kg-dry weight. Based on recommended biosolids application rates, predicted BPA concentrations in soil (PEC(soil)) would be 0.021 mg/kg-dry weight for North America and 0.14 mg/kg-dry weight for Europe. Hazard quotients (ratio of PEC(soil) and PNEC(soil)) for BPA were all equal to or less than 0.04. This indicates that risks to representative invertebrates and plants at the base of the terrestrial food web are low if exposed to BPA in soil amended with activated sludge biosolids.

Comparison of Four Species Sensitivity Distribution Methods to Calculate Predicted No Effect Concentrations for Bisphenol A

ABSTRACT Bisphenol A (BPA, CAS RN 80-05-7) is a high production volume chemical used as an intermediate in the production of polycarbonate plastic and epoxy resins. During its manufacture and use, some emissions to surface waters are anticipated. Chronic predicted no effect concentrations (PNECs) for aquatic systems are used to support the assessment of potential risks to aquatic organisms in receiving waters. PNECs for a compound are considered protective of populations, communities, and ecosystems. Traditionally, PNECs are derived by taking the lowest no-observed effect concentration (NOEC) from a set of toxicity studies and dividing by an assessment factor (e.g., 10 to 1000). This traditional approach is appropriate for substances with few data, but may not be necessary for substances with many valid studies. For well-studied substances, statistical approaches (i.e., development of Species Sensitivity Distribution or SSD methods) can be used to calculate a PNEC that makes use of the full distribution of available NOEC values. Bisphenol A has an extensive set of aquatic toxicity studies covering diverse taxa including algae, hydra, rotifers, mollusks, crustaceans (both benthic and pelagic), insects, annelids, fish, and amphibians. The full chronic data set was used to calculate PNEC values using four SSD methods: (1) the Hazard Concentration (HC5) approach developed by The Netherlands National Institute of Public Health and the Environment (RIVM), (2) the U.S. Environmental Protection Agencys water quality criteria procedure, (3) SigmaPlot (Systat 2000) commercial software that calculates percentile values, and (4) a distributional method consistent with that used by Environment Canada. Using these approaches, PNEC values for BPA range from 11 to 71 μ g/L. Literature studies suggest that application of an additional assessment factor is unwarranted if an SSD-based PNEC is based on chronic data. SSD-derived PNEC values and the traditionally derived PNEC value of 1.6 μ g/L are then compared to concentrations of BPA that have been measured in North American and European surface waters. Adverse risks to aquatic organisms are not anticipated from measured concentrations of BPA in North American and European surface waters.

Adult fathead minnow, Pimephales promelas, partial life-cycle reproductive and gonadal histopathology study with bisphenol A.

Bisphenol A (BPA) is an intermediate used to produce epoxy resins and polycarbonate plastics. Although BPA degrades rapidly in the environment with aquatic half-lives from 0.5 to 6 d, it can be found in aquatic systems because of widespread use. To evaluate potential effects from chronic exposure, fathead minnows were exposed for 164 d to nominal concentrations of 1, 16, 64, 160, and 640 µg/L BPA. Population-level endpoints of survival, growth, and reproduction were assessed with supplemental endpoints (e.g., vitellogenin, gonad histology), including gonad cell type assessment and quantification. No statistically significant changes in growth, gonad weight, gonadosomatic index, or reproduction variables (e.g., number of eggs and spawns, hatchability) were observed; however, there was a significant impact on male survival at 640 µg/L. Vitellogenin increased in both sexes at 64 µg/L or higher. Gonad cell type frequencies were significantly different from controls at 160 µg/L or higher in males with a slight decrease in spermatocytes compared with less mature cell types, and at 640 µg/L in females with a slight decrease in early vitellogenic cells compared with less mature cells. The decrease in spermatocytes did not correspond to a decrease in the most mature sex cell type (spermatozoa) and did not impair male fertility, as hatchability was not impacted. Overall, marginal shifts in gametogenic cell maturation were not associated with any statistically significant effects on population-relevant reproductive endpoints (growth, fecundity, and hatchability) at any concentration tested.

Recommended approaches to the scientific evaluation of ecotoxicological hazards and risks of endocrine-active substances

A SETAC Pellston Workshop® “Environmental Hazard and Risk Assessment Approaches for Endocrine-Active Substances (EHRA)” was held in February 2016 in Pensacola, Florida, USA. The primary objective of the workshop was to provide advice, based on current scientific understanding, to regulators and policy makers; the aim being to make considered, informed decisions on whether to select an ecotoxicological hazard- or a risk-based approach for regulating a given endocrinedisrupting substance (EDS) under review. The workshop additionally considered recent developments in the identification of EDS. Case studies were undertaken on 6 endocrine-active substances (EAS—not necessarily proven EDS, but substances known to interact directly with the endocrine system) that are representative of a range of perturbations of the endocrine system and considered to be data rich in relevant information at multiple biological levels of organization for 1 or more ecologically relevant taxa. The substances selected were 17α-ethinylestradiol, perchlorate, propiconazole, 17β-trenbolone, tributyltin, and vinclozolin. The 6 case studies were not comprehensive safety evaluations but provided foundations for clarifying key issues and procedures that should be considered when assessing the ecotoxicological hazards and risks of EAS and EDS. The workshop also highlighted areas of scientific uncertainty, and made specific recommendations for research and methods-development to resolve some of the identified issues. The present paper provides broad guidance for scientists in regulatory authorities, industry, and academia on issues likely to arise during the ecotoxicological hazard and risk assessment of EAS and EDS. The primary conclusion of this paper, and of the SETAC Pellston Workshop on which it is based, is that if data on environmental exposure, effects on sensitive species and life-stages, delayed effects, and effects at low concentrations are robust, initiating environmental risk assessment of EDS is scientifically sound and sufficiently reliable and protective of the environment. In the absence of such data, assessment on the basis of hazard is scientifically justified until such time as relevant new information is available.

Challenges in assigning endocrine-specific modes of action: Recommendations for researchers and regulators

As regulatory programs evaluate substances for their endocrine-disrupting properties, careful study design and data interpretation are needed to distinguish between responses that are truly endocrine specific and those that are not. This is particularly important in regulatory environments where criteria are under development to identify endocrine-disrupting properties to enable hazard-based regulation. Irrespective of these processes, most jurisdictions use the World Health Organization/International Programme on Chemical Safety definition of an endocrine disruptor, requiring that a substance is demonstrated to cause a change in endocrine function that consequently leads to an adverse effect in an intact organism. Such a definition is broad, and at its most cautious might capture many general mechanisms that would not specifically denote an endocrine disruptor. In addition, endocrine responses may be adaptive in nature, designed to maintain homeostasis rather than induce an irreversible adverse effect. The likelihood of indirect effects is increased in (eco)toxicological studies that require the use of maximum tolerated concentrations or doses, which must produce some adverse effect. The misidentification of indirect effects as truly endocrine mediated has serious consequences for prompting animal- and resource-intensive testing and regulatory consequences. To minimize the risk for misidentification, an objective and transparent weight-of-evidence procedure based on biological plausibility, essentiality, and empirical evidence of key events in an adverse outcome pathway is recommended to describe the modes of action that may be involved in toxic responses in nontarget organisms. Confounding factors such as systemic toxicity, general stress, and infection can add complexity to such an evaluation and should be considered in the weight of evidence. A recommended set of questions is proffered to help guide researchers and regulators in discerning endocrine and nonendocrine responses. Although many examples provided in this study are based on ecotoxicology, the majority of the concepts and processes are applicable to both environmental and human health assessments. Integr Environ Assess Manag 2017;13:280-292.

Distinguishing between endocrine disruption and non-specific effects on endocrine systems

ABSTRACT The endocrine system is responsible for growth, development, maintaining homeostasis and for the control of many physiological processes. Due to the integral nature of its signaling pathways, it can be difficult to distinguish endocrine‐mediated adverse effects from transient fluctuations, adaptive/compensatory responses, or adverse effects on the endocrine system that are caused by mechanisms outside the endocrine system. This is particularly true in toxicological studies that require generation of effects through the use of Maximum Tolerated Doses (or Concentrations). Endocrine‐mediated adverse effects are those that occur as a consequence of the interaction of a chemical with a specific molecular component of the endocrine system, for example, a hormone receptor. Non‐endocrine‐mediated adverse effects on the endocrine system are those that occur by other mechanisms. For example, systemic toxicity, which perturbs homeostasis and affects the general well‐being of an organism, can affect endocrine signaling. Some organs/tissues can be affected by both endocrine and non‐endocrine signals, which must be distinguished. This paper examines in vitro and in vivo endocrine endpoints that can be altered by non‐endocrine processes. It recommends an evaluation of these issues in the assessment of effects for the determination of endocrine disrupting properties of chemicals. This underscores the importance of using a formal weight of evidence (WoE) process to evaluate potential endocrine activity.

Letter to the editor (Wright-Walters et al., 2011).

We have read with interest the recent paper updating the aquatic hazard assessment of Bisphenol A (Wright-Walters et al., 2011). The paper developed a “new” predicted no effect concentration (PNEC) for Bisphenol A (BPA) of 0.06 μg/L for long term aquatic exposure to freshwater organisms. This new PNEC was calculated using the species sensitivity distribution (SSD) HC5 (lower 5th percentile hazard concentration) method of van der Hoeven (2001) with a large set of acute and chronic studies taken from published literature and unpublished sources. The new PNEC is considerably lower than other SSD-based HC5 values derived for BPA for use as a PNEC (Staples et al., 2008; EC, 2010). The HC5 values calculated by Staples et al. (2008) and EC (2010) are 18 and 7.5 μg/L, respectively. EC (2010) used a further assessment factor of 5 to arrive at a PNEC of 1.5 μg/L, while neither Staples et al. (2008) nor Wright-Walters et al. (2011) applied an assessment factor. Although different models were used, the differences between the HC5 values calculated by Staples et al. (2008), EC (2010) and Wright-Walters et al. (2011) are largely due to the use of different datasets. Wright-Walters et al. (2011) conducted literature searches to identify potentially relevant studies and also reportedly assessed the quality of each study. The resulting dataset is shown in Table 2 of the paper and includes 61 studies and 94 no observed effect concentrations (NOEC). Unfortunately, the dataset that resulted from this process and was used by Wright-Walters et al. (2011) to derive the PNEC is fraught with errors and omits a number of important studies. The result of the errors and omissions in the dataset is that the “new” PNEC calculated by Wright-Walters et al. (2011) is incorrect and should not be used to assess the hazard of BPA. The dataset assembled by Wright-Walters et al. (2011) includes values that do not exist in the specified studies. NOEC forHydra vulgaris, Gammarus pulex, and Chironomus riparius of 0.002, 0.01 and 0.078 μg/L from Pascoe et al. (2002) and Watts et al. (2001, 2003) were cited by Wright-Walters et al. (2011), but these values are not in any of the published papers. Acute data were improperly included in the chronic toxicity dataset even though valid chronic data were available. For example, the Wright-Walters et al. (2011) dataset included a 96-h NOEC of 1400 μg/L from an acute lethality study with Chironomus tentans (Sayers, 2005), and an extended acute 14-d NOEC of 3200 μg/L with zebrafish (Bayer AG, 1999). Instead, the dataset should have included the valid chronic studies with the midge C. riparius and zebrafish that do exist for these species (Watts et al., 2003; Segner et al., 2003a). Studies were improperly included that lacked sufficient documentation to score their validity using procedures based on Klimisch et al. (1997). The studies reported by Fraunhofer (2000) and Segner et al.