Potential applicability and challenges of using in pp y g g vitro - PowerPoint PPT Presentation

Potential applicability and challenges of using in pp y g g vitro and in silico methodologies in food ingredient safety assessment S Suzanne Fitzpatrick, PhD, DABT, ERT Fit t i k PhD DABT ERT CFSAN/FDA SOT Food Safety Specialty Section Webinar August 18 2016 August 18, 2016

Topics of Discussions for this Talk Topics of Discussions for this Talk • Vision of the NRC Report on Toxicity Testing in the 21 p y g Century • Tox 21 Partnership between FDA, EPA, NIEHS, and NCATS NCATS • Other Emerging “Tox 21” Methods – SAR/QSAR – SAR/QSAR – Read Across – AOPs – Organs of a Chip • New ICCVAM Activities • New NRC Report

2007 NRC - Toxicity Testing in the 21 t C 21st Century: A Vision & Strategy t A Vi i & St t • Sponsored by EPA • Use cell-based (high throughput) assays to understand how chemicals perturb normal cellular functions (i.e., toxicity pathway) ( , y p y) – Establish relationships of perturbations with “ adverse outcomes ” • Develop in vitro to in vivo extrapolation p p methods • Integrate results to predict hazard/risk Broader coverage of chemicals & endpoints Reduce cost & time of testing Reduce cost & time of testing Use fewer animals

Vision of the NRC report Vision of the NRC report • The NRC report laid out a roadmap for The NRC report laid out a roadmap for revamping toxicity testing • Focus should shift away from identification of y toxicant-induced apical endpoint effects towards an identification of a sequence of key events/modes of action as the organizing t / d f ti th i i principle for risk assessment • The use of mechanistic data will help risk • The use of mechanistic data will help risk assessors gain a better understanding of how chemicals exert their toxic effects

Vision of the NRC report Vision of the NRC report  NRC also advocated the use of adverse NRC also advocated the use of adverse outcome pathways (AOPs)as a critical aspect of predictive toxicity testing.  The AOP Framework was established to systematically collect, organize and evaluate mechanistic data and causally link them to adverse effects.

Memorandum of Understanding for Tox 21 • 5-year Memorandum of Understanding (MoU) on “ High-Throughput Screening, Toxicity Pathway Profiling, and Biological Interpretation of S i T i it P th P fili d Bi l i l I t t ti f Findings ” released on Feb 14, 2008 signed by NHGRI (F.S. Collins), NIEHS/NTP (S.H. Wilson), and EPA (G.M. Gray). • Revised 5-year MoU to add FDA signed on July 19, 2010 by NHGRI (E.D. Green), NIEHS/NTP (L.S. Birnbaum), EPA (P.T. Anastas) and FDA (J Woodcock) Anastas), and FDA (J. Woodcock). • Known informally as Tox21 for Toxicology in the 21 st Century • MOU revised July 2015. Dr. Susan Mayne, Director of the Center for Food Safety & Applied Nutrition, signed for FDA. 6

Tox21 Goals • Identify patterns of compound- induced biological response in order to: d t − characterize toxicity/disease pathways − facilitate cross-species extrapolation − model low-dose extrapolation • Prioritize compounds for more Prioritize compounds for more extensive toxicological evaluation • Develop predictive models for biological response in humans 7

Agency Points of Contact FDA – Suzanne Fitzpatrick Ph.D. FDA Suzanne Fitzpatrick Ph D NCGC/NCATS – Anton Simeonov, Ph.D. EPA/NCCT – Russell Thomas, Ph.D. NIEHS/NTP – Rick Paules, Ph.D. Assays & Pathways Chemical Selection Informatics Targeted Testing Working Group Working Group Working Group Working Group Working Group Working Group Working Group Working Group Co-Chairs Co-Chairs Co-Chairs Co-Chairs Kevin Gaido, Ph.D. (FDA) William Leister, Ph.D. (NCGC) Ruili Huang, Ph.D. (NCGC) Michael DeVito, Ph.D. (NTP) Keith Houck, Ph.D. (EPA) Richard Judson, Ph.D. (EPA) David Gerhold, Ph.D. (NCGC) Donna Mendrick, Ph.D. (FDA) Kristine Witt, M.S. (NTP) Nisha Sipes, Ph.D. (NIEHS) Timothy Shafer, Ph.D. (EPA) Ann Richard, Ph.D. (EPA) Menghang Xia Ph D (NCGC) Menghang Xia, Ph.D. (NCGC) Weida Tong, Ph.D. (FDA) Weida Tong Ph D (FDA) Robert Sprando Ph D (FDA) Robert Sprando, Ph.D. (FDA) Suramya Waidanatha,Ph.D.(NTP) Suramya Waidanatha Ph D (NTP)  Evaluate assay  Establish compound  Evaluate relevance  Identify toxicity performance pathways & libraries for qHTS of prioritization  Develop corresponding (10K, mixtures, water- schemes & prioritization assays soluble) prediction models schemes and  Establish QC  Extrapolate in vitro  Review nominated prediction models procedures for concentration to in assays and  Make all data Make all data compound identity compound identity, vivo dose i o dose prioritize for use at i iti f t publicly accessible purity, concentration, the NCGC and stability 8

Tox 21: Outcome Pathways of NAS Tox 21: Outcome Pathways of NAS Exposure Exposure Uptake-Delivery to Target Tissues Perturbation Cellular response pathway “Normal” Biological Biologic Function inputs Early cellular Adaptive changes Responses Responses Cell Adverse injury, Inability Outcomes to (e.g., mortality, regulate Reproductive Impairment) Impairment) Adverse outcome Molecular Perturbed cellular relevant to initiating event response pathway risk assessment risk assessment Toxicity Pathway NAS, 2007 Adverse Outcome Pathway

Tox Cast Inventories Tox Cast Inventories • ToxCast Phase I (293 unique cmpds) – EPA pesticidal actives w/ rich in vivo data � – PFOAs, BPA, approx 12 metabolite/parent pairs PFOA BPA 12 t b lit / t i • ToxCast Phase II (767 unique new cmpds) – EPA pesticides, high interest EPA and stakeholder inventories, data rich chemicals (EDSP, OPPT, antimicrobials, inerts, green alternatives, fragrances, h i l (EDSP OPPT ti i bi l i t lt ti f water …) – FDA CFSAN data rich, NCTR LTKB Priority 1 drugs – Toxicity reference chemicals, data ‐ rich chemicals, NTP immunotox Toxicity reference chemicals data rich chemicals NTP immunotox – 135 Donated pharma cmpds ‐‐ failed drugs w/ pre ‐ clinical or clinical tox data • ToxCast E1K (800 unique new cmpds) – Endocrine active reference cmpds, SAR predicted ER ‐ active/inactives, EDSP E d i ti f d SAR di t d ER ti /i ti EDSP cmpds • EPA’s Tox21 library (3727 unique cmpds out of current 8599 total) – Complete on ‐ hand EPA sample library used to build ToxCast inventories C l t h d EPA l lib d t b ild T C t i t i

Tox21 Robot System NCGC NCGC screened 1408 compounds (1353 unique) from NTP d 1408 d (1353 i ) f NTP and 1462 compounds (1384 unique) from EPA in 140 qHTS assays representing 77 predominantly cell ‐ based reporter gene endpoints. http://www.youtube.com/watch?v=ECloTsdD-xo 11

Current Limitations of Data for Regulatory Use • Lack of xenobiotic metabolism • Inability to screen volatile or highly hydrophillic chemicals • Limited coverage of biological targets • Lack of a pragmatic path forward for validation • Inability to confidently translate perturbations at molecular level to likely tissue and organ ‐ level effects • These are all the challenges/goals for all four agencies going forward with this program

SAR/QSAR /Q • Structure-Activity Relationships (SAR) are relationships between a compound’s chemical relationships between a compound s chemical structure and physiochemical properties and biological effects on living systems g g y • Complex computer software modeling programs have been and are being developed to predict carcinogenic and mutagenic potential using quantitative SAR or QSAR. • QSAR analysis – could be useful tool for complementing and possibly reducing the battery of genetic toxicity testing requested for battery of genetic toxicity testing requested for food contact substances

Read ‐ Across Read Across • Read-across is when the already available data of a y data-rich substance (the source) is used for a data-poor substance (the target), which is considered similar enough to the source substance to use the same data as enough to the source substance to use the same data as a basis for the safety assessment • Uses of read Across – To avoid additional animal testing – To save time and cost – To use human data if available for one compound but not – To use human data, if available for one compound but not possible to produce for another – To cover more substances with one safety assessment

Opportunities for incorporating in vitro/in silico data into read-across d t i t d it /i ili Reducing uncertainty in a read-across argument in a regulatory submission: g y • Using in vitro/in silico data to confirm the similarity in the mechanism of action within a category and/or between the “target” and “source” d/ b t th “t t” d “ ” compounds • Confirming or refuting a hypothesis that proposed • Confirming or refuting a hypothesis that proposed analogues may have “other” effects • Assessing the relative “potency” of the analogues g p y g

Understanding an AOP Provides A Basis to Inform The Use of Data for Risk Assessment & Decision Making Structure Activity In vitro In vivo R l Relationships i hi studies di studies di Key events or predictive Adverse outcome Molecular relationships spanning relevant to initiating event levels of biological risk assessment organization g Greater Toxicological Greater Risk Understanding Relevance (Quantitative AOP) (Qualitative AOP)