Document q3e59qgpEKRw0Jyy73qZMrDqM

A chemical category-based approach for selecting and screening PFAS for toxicity and toxicokinetic testing PhD PFAS Global 2022 March 24, 2022 Phone: @epa.gov The views expressed in this presentation are those of the author and do not necessarily reflect the views or policies of the U.S. EPA 1 Overview Large numbers of PFAS whose hazard potential needs to be characterized Characterize PFAS chemical space Categories and sub-categories Chemical feature-based categories (ToxPrints) vs. OECD categories Find PFAS chemicals that can be purchased (limited subset of PFAS Universe) Select subset of these chemicals for NAM (New Approach Methods, in vitro) testing Compile legacy in vivo data Use categories and data as input for selecting chemicals for in vivo testing Work in progress 2 EPA PFAS Strategic Roadmap Accelerate public health protections by identifying PFAS categories Advance the science to assess human health and environmental risks from PFAS 3 https://www.epa.gov/system/files/documents/2021-10/pfas-roadmap_final-508.pdf Assemble a PFAS Chemical Library for NAM Work Attempted to procure ~3,000 based on chemical diversity, Agency priorities, and other considerations Obtained 480 total unique chemicals 430/480 soluble in DMSO (90%) 54/75 soluble in water (72%) (incl. only 3 DMSO insolubles) Issues with sample stability and volatility Categories initially assigned based on three approaches Buck et al., 2011 categories Markush categories (ToxPrints) OECD categories 4 Kathy Coutros, Chris Grulke, Grace Patlewicz and Ann Richard 4 PFAS Structure-based Categorization: ToxPrints Publicly available tools exist to generate & download ToxPrints e.g. ChemoTyper, CompTox Chemicals Dashboard Provides excellent coverage of PFAS chemical space Nested, hierarchical nature lends itself to creating flexible categories Can augment with computed structure properties (e.g., MW, size, etc.) ToxPrints: 729 chemical features Chemically interpretable Coverage of diverse chemistry Includes scaffolds, functional groups, chains, rings, bonding patterns, atomtypes 5 Selecting a Subset of PFAS for Tiered Toxicity and Toxicokinetic Testing (NAMs) Goals: Generate data to support development and refinement of categories for read-across Incorporate substances of interest to Agency Characterize mechanistic and toxicokinetic properties of the broader PFAS landscape Selected 150 PFAS in two phases representing 83 different categories 9 categories with > 3 members Lots of singletons 6 6 In Vitro Toxicity and Toxicokinetic Testing (NAMs) Toxicological Response Developmental Toxicity Immunotoxicity Mitochondrial Toxicity Developmental Neurotoxicity Endocrine Disruption General Toxicity Assay Zebrafish embryo assay Bioseek Diversity Plus Mitochondrial membrane potential (HepaRG) Microelectrode array assay (rat primary neurons) ACEA real-time cell proliferation assay (T47D) Attagene cis- and trans- Factorial assay (HepG2) Assay Endpoints Fertilisation, lethality, and structural defects Protein biomarkers across multiple primary cell types Mitochondrial membrane potential Neuronal electrical activity Cell proliferation Nuclear receptor and transcription factor activation High-throughput transcriptomic assay (multiple cell types) High-throughput phenotypic profiling (multiple cell types) Cellular mRNA Nuclear, endoplasmic reticulum, nucleoli, golgi, plasma membrane, cytoskeleton, and mitochondria morphology Purpose Assess potential teratogenicity Measure potential disease and immune responses Measure mitochondrial health and function Impacts on neuron function Measure ER activity Activation of key receptors and transcription factors involved in hepatotoxicity Measures changes in important biological pathways Changes in cellular organelles and general morphology Toxicokinetic Parameter Intrinsic hepatic clearance Pla7sma protein binding Assay Assay Endpoints Purpose Hepatocyte stability assay (primary Time course metabolism of parent chemical Measure metabolic breakdown by the human hepatocytes) liver Ultracentrifugation assay Fraction of chemical not bound to plasma Measure amount of free chemical in protein the blood 7 Chemical Inventories In Vitro = 99 NAM chemical passing QC & VP criteria In Vivo = chemicals with PODs in ToxValDB PFAS TSCA = non-CBI TSCA Active Inventory with structure 8 Category / Sub-Category Approach sub-category Chemical substances are placed into high-level categories based on structure Sub-categories are derived using NAM, chemical property and existing in vivo data At least one chemical substance needs in vivo data per subcategory for read-across Chemical Types Untested Existing in vivo data Proposed for new testing 9 Sub-Category Approach Similarity metric combines Chemical structures Chemical properties NAM data In vivo data Select chemical nearest centroid for testing 10 Current Status NAM data generation for PFAS 150 close to complete Initial categorization approach developed Used as input to the National PFAS Testing Strategy (October 2021) https://www.epa.gov/assessing-and-managing-chemicals-under-tsca/national-pfastesting-strategy 11 Acknowledgements 12