DESIGN AND DOSE SELECTION
FOR CHRONIC RODENT STUDIES
SOT RASS TELECON
April 9, 2008
Lorenz Rhomberg
Gradient Corporation
Dale Hattis
Clark University
Stephen Olin
ILSI Research Foundation
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Authors
Lorenz Rhomberg
Gradient Corporation
Karl Baetcke
U.S. EPA/OPP
Jerry Blancato
U.S. EPA/ORD/NERL
James Bus
Dow Chemical Company
Samuel Cohen
U. of Nebraska Medical Center
Rory Conolly
U.S. EPA/ORD/NCCT
Rakesh Dixit
MedImmune, Inc.
John Doe
Dale Hattis
Clark University
Abigail Jacobs
U.S. FDA/CDER
David Jacobson-Kram
U.S. FDA/CDER
Tom Lewandowski
Gradient Corporation
Robert Liteplo
Health Canada
Olavi Pelkonen
U. of Oulu, Finland
Jerry Rice
Georgetown University
Syngenta CTL, UK
Diana Somers
Karen Ekelman
PMRA, Canada
U.S. FDA/CVM
Penny Fenner-Crisp
ILSI RF/RSI
Paul Harvey
NICNAS, Australia
ILSI® Research Foundation
Angelo Turturro
U.S. FDA/NCTR
Webster West
U. of South Carolina
Stephen Olin
ILSI RF/RSI
Process
• Convened working group:
– International
– Multi-disciplinary
– Government, industry, academia
• Building on ILSI 1997 “Principles for the
Selection of Doses in Chronic Rodent
Bioassays”, working group:
– Examined issues that come into play in putting those
principles into practice
– Offered perspectives and insights in the context of
increasing demands on the bioassay and the growing
importance of mechanism/mode of action in the
assessment of human health risks
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Scope and Focus
• Not a new testing paradigm, but an effort to
facilitate continuing improvement in quality,
consistency and utility of bioassay data
• Recognizes the ‘yin and yang’ of design and
interpretation
• Broadly focused on dose selection but not
just MTD
• More weight on cancer bioassay but also
applicable to chronic toxicity studies
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Other Information
• Peer consultation (2005) informed
development of final document
• OECD draft guidance on dose selection
based on ILSI 1997 and 2007
• Financial support provided by EPA/OPP and
Health Canada
**********************************************************
• Citation: “Issues in the Design and Interpretation of
Chronic Toxicity and Carcinogenicity Studies in
Rodents: Approaches to Dose Selection” (2007)
Critical Reviews in Toxicology, 37(9): 729-837.
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Dose Selection Questions:
• Top Dose?
• Number of Dose Levels?
• Placement of Dose Levels?
• Numbers of Animals per Dose Level?
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Cancer vs. Noncancer Endpoints
Different dose-selection criteria
• Cancer:
• Power to Detect
• Stochastic Endpoints
• Noncancer:
• Severity Spectrum
• NOAEL / BMD Identification
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Prospective vs. Retrospective?
RETROSPECTIVE (evaluate the success
of a design by interpreting outcomes)
PROSPECTIVE (plan a design likely to
succeed in providing needed
information)
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Bioassay Objectives
• screening chemicals for identifying carcinogens or
other toxic effects;
• characterizing the dose-response curve in the
observable range;
• characterizing the dose-response curve to
facilitate low-dose extrapolation;
• defining a threshold or benchmark dose point of
departure;
• providing data on health effects at human
exposure levels;
• providing data to test hypotheses regarding mode
of action.
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Combining Objectives
• Different objectives invariably conflict
in demands on dose selection
• Illuminate conflicts by considering ideal designs for
different objectives
• No single ideal design – compromises
are necessary
• Hedging against poor choices
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A Systematic Approach to Selecting Doses
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A Systematic Approach to Selecting
Doses
3 CONTEXTS:
• scientific
• regulatory / risk mgmt
• practical
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OBJECTIVES:
A Systematic Approach to Selecting
Doses
• Haz Screening
• D-R
• Low-Dose Extrapol’n
• ID Threshold
• BMD
• Human Safety Study
• MoA
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A Systematic Approach to Selecting Doses
• PK
• MoA
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A Systematic Approach to Selecting Doses
• Top Dose
• Number of Doses
• Location of Doses
• Animal Allocation
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A Systematic Approach to Selecting Doses
Imagine Possible Outcomes
&
Evaluate Design
Performance
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A Systematic Approach to Selecting Doses
Iterate to Improve
Likelihood of Desired
Performance
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Design / Interpretation Factors
•
•
•
•
•
•
•
•
•
mortality
clinical signs
site-of-administration effects
pharmacokinetics and altered metabolism
physiological effects
nutritional effects
hormonal effects
organ-weight changes
cell proliferation and apoptosis
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Case Studies
• Formaldehyde
• Methylene Chloride
• Drug X
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Appendix:
Criteria for MTD Attainment:
Clinical Pathology and Pathology-based Endpoints
•
•
•
•
Liver
Kidney
Hematopoietic System
Reproductive System
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Looking to the Future of the Chronic
Bioassay--Potential Paradigm Shifts
• Decision-making questions to be
addressed are different
• Experimental tools available are different
• Scientific issues that can be and need to
be addressed are different
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Decision-Making Questions-From Qualitative to Quantitative
• How extensive is the cleanup that is needed
for a superfund site?
• How quantitatively “significant” are the risks
posed by occupational exposures to specific
(air levels x durations) of X?
• How large are the health prevention benefits
from defined reductions in exposures to X
achievable by specific interventions?
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New Experimental Tools as
Sources of Paradigm Shifts
• Carcinogenesis
– From quantal observations of one+ tumor per animal
after lifetime dosing to shorter term measurements of
continuous parameters (somatic mutation rates for
relevant mutations, rates of epigenetic changes and
relative growth advantages for initiated clones)
– More definitive mode of action characterizations with
genetic “knockout” animals and related manipulations
(e.g. RNAi, animals with specific genes over
expressed)
– “-omics” based assay systems applied to quantitative
elucidation of dose/time/response relationships after
identification/separation of specific relevant cell types
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Newer Tools for Newer Questions for both
Carcinogenesis and Other Health Endpoints
• Accounting for more diversity in toxic susceptibility
(by age, other interacting factors)
• Chronic cumulative modes of harm (e.g. Parkinson’s,
Alzheimer’s, obstructive lung disease conditions
where there are long term losses of functional units).
Need biomarkers of:
– Cumulative total of past damage (e.g. FEV1)
– Current rate of increase in damage (e.g. enzymes
released into the blood that indicate death of key
types of cells or structural damage)
– Dysfunction indicators (e.g. heart rate variability
changes)
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Building a Deeper Understanding of
Homeostatic Control Systems
• How exactly are the numerous set points for
different levels of biological organization set?
– Coded in the genome? If so, how?
– Developed by some sort of evolutionary learning process?
How?
• How do failures of homeostatic controls happen?
– Barker observations--chronic cumulative loss of “wetware” to
accomplish control
– Other reasons for deterioration of homeostatic system controls
with ageing
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Plot of the Incidence of Type 2 Diabetes
in Relation to Log(Mean Birth Weight)
--Data of Forsen et al., 2000
12
y = 47.05 - 11.46x R^2 = 0.960
11
% Type 2 Diabetes
10
9
8
7
6
5
4
3.2
3.3
3.4
3.5
Log(Mean Birth Wt g)
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3.6
3.7
New Paradigms for Risk Analysis-Tools for Probabilistic Analyses
• From analysis of data sets to data bases
• Quantitative structure activity relationships
specific to different modes of action require
standardized effect levels (e.g. ED50s) rather
than no-effect levels or even BMDL’s that mix
in effects of inter-animal variability and assay
uncertainty/measurement errors
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Conclusion
• The current chronic animal bioassay is perhaps best
seen as an aircraft carrier—a central anchor of a large
naval battle group including many subsidiary
vessels/experiments that both protect the carrier and
enhance the capabilities to address increasingly
diverse information needs. It remains a mainstay for
current risk assessment, but will increasingly be
supplemented, if not supplanted, with numerous other
tools involving exposures during specific life-stages
and measuring a variety of endpoints short of apical
measures of adverse health endpoints.
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