Slide 1 of 15 

# FIFRA Science Advisory Panel Meeting

Prioritization of EDSP
Universe of Chemicals Using Computational Toxicology Tools

Arlington, VA January 29, 2013Slide 2 of 15 1.1 Please comment on appropriateness of the overall conceptual approach for prioritizing the EDSP chemical inventory ( note : subsequent charge questions address issues associated with specific components of the prioritization approach). In your comments, please address:
The extent to which EPA’s description of the process transparently captures and describes the key technical steps of the prioritization scheme and whether there are other scientific considerations the agency should incorporate into its EDSP prioritization scheme;
The robustness of the scientific support for the overall approach and, in particular, the logical sequence of filters,
and
Whether changes to the approach could set priorities more efficiently and effectively 

# Question 1. Overall Prioritization Approach

Section 2.2; also see Section 9Slide 3 of 15 2.1 Please comment on the proposed exclusion criteria for each physicochemical property as appropriate for identifying chemicals that are untestable and/or unable to elicit systemic toxicity. Please include a discussion of the extent to which varying the range of exclusion criterion can influence confidence in identifying chemicals that are untestable and/or unable to elicit systemic effects ( i.e. , those substances that are unlikely to trigger the molecular initiating event). Please include in your

comments:

Whether there are additional chemical characteristics that should be considered ( e.g. , volatility),

Whether there are additional data sources for each parameter identified ( e.g. , ChemAxon for pKa values, EpiSuite for half life values, etc.), and how to consider multiple data sources in a weight of evidence approach. 

# Question 2. Physicochemical Properties

Section 2.2.2Slide 4 of 15 2.1 (Continued)
Whether the exclusion criteria cut offs and the confidence that application of the exclusion criteria will reasonably identify those substances that are untestable and/or unable to trigger the molecular initiating event of the ER pathway. Also please discuss options for evaluating cases where uncertainty in a chemical’s specific parameter range overlaps a p

- chem filter cut off. Please consider in your comments the following exclusion criteria:

- pK
a values <2 or >11.5;

- Hydrolysis half life of <40 days and

- Charged species of 99% or greater at pH 7 

# Question 2. Physicochemical Properties

Section 2.2.2Slide 5 of 15 3.1 Please comment on the approach used to build ESv2 through chemical testing and effect

- based category and read across methods to provide a scientifically defensible approach to predict ER binding potential for a larger number of chemical groups, particularly with regard to defining new groups from within the Mixed Organics and Mixed Phenols. Please indicate any considerations unique to ESv2 that indicate the approach used to develop ESv1 (SAP, 2009a) needs to be modified.

3.2 Building from the ESv1 training set, please comment on the level of scientific confidence that a chemical is unlikely to initiate the ER AOP, if the in vitro assay TrSet data shows no activity. Please comment on the extent to which the level of confidence may vary by chemical category. 

# Question 3. ER Expert System Development

Section 3Slide 6 of 15 The agency is proposing to use the HTP data to expand the ER expert system ( i.e. , ESv3). Therefore, it is important to understand how the HTP data are generated and interpreted for the intended use.

4.1 Given the importance of a well

- defined endpoint to determining if/how endpoint data are appropriately used, the panel is asked to comment on:

Whether the assays are sufficiently described so that others can reconstruct the assay conditions and data analysis and to what
extent additional information would be useful?

Whether EPA has described sufficiently the important experimental conditions that affect the assay measurements, and whether EPA has sufficiently discussed their potential impact on assay results and interpretation of the results? 

# Question 4. Overall Prioritization Approach

Sections 4

- 6Slide 7 of 15 The agency is proposing to use the HTP data to expand the ER expert system ( i.e. , ESv3). Therefore, it is important to understand how the HTP data are generated and interpreted for the intended use.

4.1 (Continued)

Whether the chemical library ( e.g. , purity and analysis), chemical exposure ( e.g. , solvents used, chemical dilutions), and plates and plate layouts provides information necessary for data interpretation?

The adequacy of test concentrations (maximum and minimum concentration tested) and cutoffs, and whether the concentration cutoffs affected some assays results more than others, especially in the context of false negatives. 

# Question 4. Overall Prioritization Approach

Sections 4

- 6Slide 8 of 15 The agency is proposing to use the HTP data to expand the ER expert system ( i.e. , ESv3). Therefore, it is important to understand how the HTP data are generated and interpreted for the intended use.

4.2 With respect to data interpretation, please comment on the approach for defining an active chemical ( i.e. , to initiate the ER AOP) and an inactive chemical ( i.e. , unlikely to initiate the ER AOP) compound and whether the method of data interpretation is adequately described and if the rationale for the approach is sufficiently presented. Please include in your comments:

The adequacy of the approach to generate the data plots; how and what parameters are calculated, etc.,

How the background data were used to establish the control level,

Whether the process for identifying assay interference is adequately described,

Adequacy of data normalization, outlier identification, curve fitting, background subtractions, and all other data processing and calculation techniques, and appropriateness of the statistical analyses. 

# Question 4. Overall Prioritization Approach

Sections 4

- 6Slide 9 of 15 The agency is proposing to use the HTP data to expand the ER expert system ( i.e. , ESv3). Therefore, it is important to understand how the HTP data are generated and interpreted for the intended use.

4.3 Would you recommend other considerations or approaches to analyzing
the HTP data? 

# Question 4. Overall Prioritization Approach

Sections 4

- 6Slide 10 of 15 5.1 Please comment on the selection of reference chemicals and whether they are sufficient to assess the performance of each HTP assay for ER agonists. How well do the reference chemicals represent the range of potencies needed to establish the reliability and relevance of these assays for use in EDSP chemical prioritization for the inert ingredients, fragrances, and SDWA
chemicals?

5.2 Please comment on whether the Agency’s evaluation of the performance of the HTP ER binding and activation assays has considered and accurately assessed all relevant aspects of the assays. For example, have signal

- to

- noise ratios, background subtraction and interferences been adequately characterized when describing the strengths and limitations of the assay? 

# Question 5. Performance Evaluation of the HTP ER Binding and Transcriptional Activation Assays Against a Set of
Reference Chemicals

Section 6Slide 11 of 15 5.3 Based on analyses in sections 5 and 6, please comment on the comparative performance of the 8 HTP assays for detecting ER reference agonists ( e.g. , do some assays perform differently, and if so to what extent?) Was sufficiently detailed information provided explaining the likely reasons for assay differences, when they are observed ( e.g. , cutoffs, assay interferences, background differences)? 

# Question 5. Performance Evaluation of the HTP ER Binding and Transcriptional Activation Assays Against a Set of
Reference Chemicals

Section 6Slide 12 of 15 6.1 Please comment on the agency’s assessment of how HTP in vitro assays performed with respect to expectations of performance for chemicals within chemical categories when compared with the in vitro training set data used to build the ESv1/2.

6.2 Please comment on EPA's approach to comparing the ES TrSet data with the HTP data and the interpretations regarding discordant results between the two assay approaches. Please recommend other considerations or approaches to conducting this comparative analysis?

6.3 Given that there may be varying occurrences of false negatives and false positives based on different sources of training set data, please include in your comments options for addressing these uncertainties for different categories in the context of building ESv3? 

# Question 6. Analysis of ER Expert Training Set
and HTP in vitro Assay Data

Section 7Slide 13 of 15 7.1 Please comment on the adequacy and efficiency of the category based approach to select chemicals for testing to expand the training sets in terms of covering the ~1700 chemicals not
covered in the domain of ESv2.

7.2 Based on the HTP assay performance (analysis of reference chemicals and comparative analysis with the ES TrSet ), please comment on the adequacy of the HTP data for advancing the ER expert system’s rules to cover the additional groups of chemicals

in ESv3. 

# Question 7. Category

- Based Approach and Strategic Testing to Expand the ES applicability domain with
additional in vit ro data

Section 8Slide 14 of 15 7.3 To the extent there are differences between the ES TrSet and HTP data in detecting the ability of chemicals to initiate the ER AOP, especially in the context of minimizing ‘false negatives’ for low potency compounds, please comment on the strengths and limitations of combining data from the different assays to generate training set data for building the structure based rules within the ESv3. What are the strengths and limitations of using training set data from assays that have the same or similar degree of sensitivity. 

# Question 7. Category

- Based Approach and Strategic Testing to Expand the ES applicability domain with
additional in vit ro data

Section 8Slide 15 of 15 8.1 Please provide any initial thoughts on how varying empirical and/or computational techniques could be employed to account for metabolites that may trigger the ER MIE. 

# Question 8. In Vitro Testing and Computer
Based Simulations: Addressing Active Metabolites

Section 7.4.3Slide 16 of 15 9.1 Please comment on whether the principles and concepts used to develop the ER expert system are generally applicable for any
category

- based prioritization system for other molecular initiating events (e.g., other AOPs for perturbing estrogen, androgen receptor, and thyroid hormone systems)?

9.2 What lessons have been learned from the development of the ER focused prioritization model that will facilitate more efficient and effective development of an effect

- based prioritization model for the androgen pathway.

9.3 What will be the challenges in developing a prioritization using in vitro methods and computer based simulations for the thyroid hormone system? 

# Question 9. Use of the AOP and Category

Based Testing Strategies for Other Endocrine Pathways

Section 9