&EPA
United States .
Environmental Protection
Agency
Office of Health and
Environmental Assessment
Washington DC 2O46O
EPA/600/6-88/007 Aa
June 1988
Externaf Review Draft
Research and Development
A Cancer
Risk-Specific
Dose Estimate for
2,3,7,8-TCDD
Review
Draft
(Do Not
Cite or Quote)
NOTICE
This document is a preliminary draft. It has not been formally
released by EPA and should not at this stage be construed to
represent Agency policy. It is being circulated for comment on its
technical accuracy and policy implications.
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DRAFT EPA-600/6-88/007Aa
DO NOT QUOTE OR CITE June 1988
Review Draft
A CANCER
RISK-SPECIFIC DOSE ESTIMATE
FOR 2,3,7,8-TCDD
NOTICE
THIS DOCUMENT IS A PRELIMINARY DRAFT. It has not been formally released by the
U.S. Environmental Protection Agency and should not at this stage be construed
to represent Agency policy. It is being circulated for comments on its
technical accuracy and policy implications.
U.S. Environmental Protection Agency
Washington, D.C.
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DISCLAIMER
This document is an external draft for review purposes only and does
not constitute Agency policy. Mention of trade names or commercial products
does not constitute endorsement or recommendation for use.
11
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CONTENTS
Tables and Figures vi
Preface vii
Authors and Reviewers ix
I. OVERVIEW 1
A. NEW DATA AND METHODS 1
B. COMPARISON OF POTENCY ESTIMATES 5
C. ORGANIZATION. 7
II. ANALYSIS. . 8
A. BACKGROUND 8
1. Exposure Considerations . .9
2. Carcinogenicity of 2,3,7,8-TCDD . .9
a. Chronic Animal Studies 10
b. Epidemic logic Studies 12
3. Altered Cell Function 15
4. Weight-of-Evidence Conclusion 17
B. MECHANISMS OF CARCINOGENISIS 17
1. Hypothesis 1: 2,3,7,8-TCDD as a Direct-Acting,
Complete Carcinogen 19
a. Qualitative Considerations 19
b. Quantitative Considerations 21
2. Hypothesis 2: 2,3,7,8-TCDD as a "Pure" Promoter 22
a. Qualitative Considerations 22
b. Quantitative Considerations 26
iii
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CONTENTS (continued)
3. Hypothesis 3: Secondary or Indirect Mechanisms of
Carcinogenesis 26
a.
General Considerations 26
b. Qualitative Evaluation of 2,3,7,8-TCDD as an
Indirect or Secondary!Carcinogen. . 28
c. Quantitative Considerations 31
C. EVALUATION OF 2,3,7,8-TCDD AS|AN "ANTICARCINOGEN" 32
D. USE OF PREDICTIVE MODELS. . I . . 32
III. CONCLUSIONS • 38
A. QUALITATIVE CONSIDERATIONS. . 41
B. QUANTITATIVE CONSIDERATIONS 42
1. Selection of Models 43
a. Noel/Uncertainty Factor (Threshold Approaches) 43
b. Sielken Approach. . 44
c. M-K-V Model 44
d. LMS Model 45
2. Selection of the RsD Range 46
a. Base Analysis 46
b. Modification of Base Analysis . . : 47
REFERENCES 52
APPENDIX A: QUANTITATIVE IMPLICATIONS OF THE USE OF DIFFERENT
EXTRAPOLATION PROCEDURES FOR LOW-DOSE CANCER RISK
ESTIMATES FROM EXPOSURE TjO 2,3,7,8-TCDD
APPENDIX B: EPIDEMIOLOGIC CANCER STUDIES ON POLYCHLORINATED
DIBENZO-fi-DIOXINS, PARTICULARLY 2,3,7,8-TCDD
iv
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CONTENTS (continued)
APPENDIX C: REPRODUCTIVE AND DEVELOPMENTAL TOXICITY OF 2,3,7,8-TCDD
APPENDIX D: EPIDEMIOLOGIC DATA ON REPRODUCTION AND EXPOSURE TO
2,3,7,8-TCDD: ITS USEFULNESS IN QUANTITATIVE RISK
ASSESSMENT
APPENDIX E: IMMUNOTOXICITY OF 2,3,7,8-TCDD: REVIEW, ISSUES,
AND UNCERTAINTIES
APPENDIX F: RATIONALE FOR A HORMONE-LIKE MECHANISM OF
. 2,3,7,8-TCDD FOR USE IN RISK ASSESSMENT
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TABLE
Table 1. Promoting effect of 2,3,7,8-JTCDD on hepatocarcinogenesis
in partially hepatectomized female rats by a single dose of
diethylnitrosamine \
24
FIGURES
Figure l.;
Figure 2.
Figure 3.
Example of risk specific doses and reference
doses for 2,3,7,8-TCDD. . J
Potential secondary mechanisms of carcinogenic activity
of 2,3,7,8-TCDD
"Multiple mechanism" hypothesis for 2,3,7,8-TCDD carcinogenesis
37
40
vi
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PREFACE
Responding to both internal and external questions about the U.S.
Environmental Protection Agency's (EPA's) 1985 Health Assessment Document for
Polychlorinated Dibenzo-p.-Dioxins, the EPA Administrator asked the Assistant
Administrator of the Office of Research and Development (ORD) to re-examine the
data and methodology upon which the assessment for 2,3,7,8-tetrachlorodibenzo-
p_-dioxin (2,3,7,8-TCDD) was based in light of new data or alternative
interpretations of the older literature that have become available since 1985
and, if appropriate, to modify EPA's approach. This report, entitled "A Cancer
Risk-Specific Dose Estimate for 2,3,7,8-TCDD,"* and its appendices present the
results of that effort.
Although there are many components to any risk assessment for
2,3,7,8-TCDD, two factors have been particularly important in recent Agency
decisions, i.e., estimates of cancer potency and estimates of human exposure.
Consequently, while other issues were reviewed and are briefly discussed in the
appendices to this report, the report itself focuses on cancer potency and
*In EPA terminology, the risk specific dose (RsD) is an estimate of dose, or
exposure, that would equal the dose estimated to result in an upper-bound
estimate of incremental lifetime cancer risk, e.g., one in a million.
The reference dose (RfD)s which is referred to later in this document, is an
estimate (uncertainty spanning perhaps an order of magnitude) of a daily
exposure to the human population (including sensitive subgroups) that is
likely to be without an appreciable risk of deleterious effects during a
lifetime (U.S. EPA, 1987b).
vii
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exposure issues. This analysis is thus not a complete risk characterization
for 2,3,7,8-TCDD, but rather a re-examination of the hazard identification and
dose-response assessment for the potential human carcinogenicity of this
chemical.
An ad hoc inter-office workgroup (hereafter, the "Workgroup") prepared the
report and recommendations. While scientists outside of this group have
provided useful analyses, review, and!comment, the conclusions and
recommendations are those of the Workgroup alone. Similarly, although the
; _ . i* |
appendices to the report contain important background information on a broad
range of 2,3,7,8-TCDD issues discussed in the report, the special focus of the
report precluded use of many of these analyses in the final document. Other
major related sources include:
• the report of an EPA Workshop! on the Development of Risk Assessment
Methodologies for Tumor Promoters,
• the report on Estimating Exposure to 2,3,7,8-TCDD
- the report of the "Dioxin" Update Committee
This report, its appendices, and| the reports listed above represent an
effort by many people within EPA to grapple with the difficult scientific
issues presented by the very large bujt incomplete data base on 2,3,7,8-TCDD.
Credit for each report or appendix in| this overall effort belongs to the
individual authors.
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AUTHORS AND REVIEWERS
AUTHORS
This document, "A Cancer Risk-Specific Dose Estimate for 2,3,7,8-TCDD,"
was prepared by an interoffice Workgroup. Members of the Workgroup are as
follows:
Peter W. Preuss, Ph.D., Chairman
Director
Office of Technology Transfer and Regulatory Support
Donald G. Barnes, Ph.D.
Acting Director
Science Advisory Board
William H. Farland, Ph.D.
Director
Office of Health and Environmental Assessment
Dorothy E. Patton, Ph.D., J.D.
Executive Director
Risk Assessment Forum
Patricia A. Roberts, J.D., B.S.
Office of General Counsel
Hugh L. Spitzer, B.A.*
Office of Technology Transfer and Regulatory Support
Deborah A. Taylor, M.A., B.A.*
Office of the Administrator
*Mr. Spitzer and Ms. Taylor contributed to the report before they left EPA in
February 1988 and August 1987, respectively.
The Appendices were prepared by scientists from ORD's Office of Health and
Environmental Assessment and Office of Health Research, and by outside
consultants.
Appendix A: Steven P. Bayard, Ph.D.
Carcinogen Assessment Group
Todd W. Thorslund, Ph.D.
ICF-Clement Associates
Fairfax, VA
Appendix B: David L. Bayliss, M.S.
Carcinogen Assessment Group
ix
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Appendix C:
Appendix D:
Appendix E:
Appendix F:
Gary L. Kimmel, Ph.D.
Reproductive Effects Assessment Group
Sherry G. Selevan, Ph.D.
Reproductive Effects Assessment Group
Babasahab R. Sonawane,!Ph.D.
Reproductive Effects Assessment Group
i
Ralph J. Smialowicz, Ph.D.
Office of Health Research
Robert W. Luebke, Ph.D;.
Office of Health Research
Michael A. Gallo, Ph.D. .
Department of Environmental and Community Medicine
University of Medicine and Dentistry of New Jersey
Robert Wood Johnson Medical School
REVIEWERS
Dr. Milt Clark
U.S. Environmental Protection Agency
Region 5 |
Chicago, IL 60605 |
Mr. David H. Cleverly i
Office of Air Quality Planning and Standards
U.S. Environmental Protection Agency
Research Triangle Park, NC 27711
Mr. Barry Commoner
Queens College
Flushing, NY 11367
Mr. Michael B. Cook
Office of Drinking Water
U.S. Environmental Protection Agency
Washington, DC 20460
Dr. Diane K. Courtney!
Health Effects Research Laboratory
U.S. Environmental Protection Agency
Research Triangle Park, NC 27711
Dr. Marilyn Fingerhut[
National Institute for Occupational
Safety and Health
Cincinnati, OH 45226
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Dr. Mike Gough
Resources for the Future
Washington, DC 20036
Mr. Richard N. Hill
Office of Pesticides and Toxic Substances
U.S. Environmental Protection Agency
Washington, DC 20460
Dr. David Hoel
National Institute for Environmental
Health Sciences
Research Triangle Park, NC 27709
Dr. Vernon Houck
Centers for Disease Control
Atlanta, GA 30323
Dr. Renate Kimbrough
Regional Operations
U.S. Environmental Protection Agency
Washington, DC 20460
Dr. Gary Kimmel
Reproductive Effects Assessment Group
U.S. Environmental Protection Agency
Washington, DC 20460
Dr. Richard J. Kociba
The Dow Chemical Company
Midland, MI 48674
Mr. James C. Lamb
Office of Pesticides and Toxic Substances
U.S. Environmental Protection Agency
Washington, DC 20460
Dr. Debdas Mukerjee
Environmental Criteria and Assessment Office
U.S. Environmental Protection Agency
Cincinnati, OH 45268
Dr. Marvin Schneiderman
National Research Council
National Academy of Sciences
Washington, DC 20418
Dr. Sherry Selevan
Reproductive Effects Assessment Group
U.S. Environmental Protection Agency
Washington, DC 20460
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Dr. Ellen Silbergeldi
Environmental Defense Fund
Washington, DC 20036
Mr. Ralph Smialowiczj
Health Effects Research Laboratory
U.S. Environmental Protection Agency
Research Triangle Park, NC 27711
I
i
Dr. Bob Sonawane
Reproductive Effects Assessment Group
U.S. Environmental Protection Agency
Washington, DC 20460
xii
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I. OVERVIEW
This report re-examines the scientific basis and methods used by the U.S.
Environmental Protection Agency (EPA) for estimating the cancer potency for
2,3,7,8-tetrachlorodibenzo-p.-dioxin (2,3,7,8-TCDD) (U.S. EPA, 1985). The
object is to determine if the 1985 assessment should be modified in light of
recent data and other plausible risk assessment methods or alternative data
interpretations.
; The Analysis uses two different approaches. One examines EPA's earlier
analysis in terms of new data and recent reviews that offer scientific
information and views for re-assessing 2,3,7,8-TCDD cancer risks. The other
involves comparing EPA's 1985 assessment with that of other regulatory agencies
in this country and elsewhere. The Agency Workgroup could not reach consensus
on all issues. However, for the reasons developed below, the Workgroup
convened for this task agreed that (1) the 1985 assessment that associates a
0.006 pg/kg/day dose with a plausible upper-bound increased cancer risk of one
in a million (10"6) should be reconsidered, and (2) a majority of the group
agreed that a change to a 0.1 pg/kg/day dose as a plausible upper-bound
associated with an increased lifetime risk of one in a million is consistent
with the available data and theories, and represents a reasonable science
policy position for the Agency.
A. NEW DATA AND METHODS
Although the scientific literature is replete with studies on
2,3,7,8-TCDD, which might be brought into a comprehensive characterization of
cancer risk, most discussions and debate about quantitative risk focus on the
1
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interpretation and use of a small subset of animal and human studies.
i
Laboratory studies conclusively establish a relationship between exposure to
2,3,7,8-TCDD and cancer in test animals. There is, however, considerable
uncertainty and controversy about theimechanism by which 2,3,7,8-TCDD causes
i
cancer, an uncertainty that can strongly influence both qualitative assessments
i
and the mathematical methods used to assess cancer risk to humans. Also,
variabilities in conduct and response among studies in human populations that
may have been exposed to 2,3,7,8-TCDD--an assumption that is itself
i
uncertain—raise additional questions and obscure the overall assessment.
A question often asked is whether 2,3,7,8-TCDD is a "complete carcinogen,"
a "promoter," or whether it produces cancer by some unknown mechanism that may
functionally have elements of initiation, promotion, and progression.1 The
previous EPA assessment (U.S. EPA, 1985) analyzed 2,3,7,8-TCDD as a complete
carcinogen. Recent studies support the assertion put forth a number of years
ago that one of the major mechanisms of action for 2,3,7,8-TCDD involves the
"promotion" of carcinogenesis in cells. However, despite changes and additions
to the data base, the analysis for risk assessment is neither obvious nor
simple, and important uncertainties remain.
These considerations give rise to the issues upon which this analysis is
[
founded.
According to generally accepted theory (OSTP, 1985; U.S. EPA, 1986a), both
complete carcinogens and promoters are capable of increasing cancer incidence
in humans. Thus, the question of complete carcinogenicity versus promotion
has little effect on identifying potential human cancer hazard associated
with exposure to 2,3,7,8-TCDD. Differences, however, in mechanisms of
carcinogenesis may lead to differences in approaches to quantitative risk
assessment, with resulting differences in numerical risk estimates.
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• Is the carcinogenic mechanism of action better understood today than it
was for the previous assessment?
• Is the linearized multistage (IMS) model as employed by the EPA to
estimate the risk associated with exposure to carcinogens appropriate
for estimating risk from exposure to 2,3,7,8-TCDD?
• Are there other appropriate ways to characterize the risk associated
with exposure to 2,3,7,8-TCDD that more fully incorporate the
biological data? Are these approaches more appropriate than the IMS
model {as used by EPA) for 2,3,7,8-TCDD assessments?
- Does the choice of approach (and related assumptions) have any bearing
on a discrepancy perceived by some between the observed human cancer
. experience and human risk estimates based on animal studies?
• What is currently understood about the mechanism(s) of action of
2,3,7,8-TCDD?
• How significant are the remaining uncertainties?
This analysis identifies several reasonable approaches to estimating
2,3,7,8-TCDD cancer risk, but concludes that there do not appear to be
compelling scientific reasons for regarding any one of them as a "most
appropriate" approach. Indeed, among the several contributors to this report,
there was a diversity of viewpoints. Preferring somewhat different assumptions
and interpretive criteria, the individual contributors brought different
perspectives to the review process. Therefore, based on rationales grounded in
science and/or science policy, several different risk assessment approaches
have been considered.
The Workgroup recognizes that there is a range of cancer risk estimates
for 2,.3,7,8-TCDD (see Figure 1). Estimates at one end of the range are based
on several different linearized models and those at the other end are based on
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California
EPA
(LMS)
Relative
Half-life*
0.001-
0.004
0
Netherlands
NRC __.
Canada FHG
CD
007
i
C
PDi
o.oi
0.006 0.03
• * * U I J
I 1 1
0.0001 0.001 0.01
\
M-K-V
Model
1
0 07 0-6
*, '*'
1
N
Sta
3
Y
te
4
Canada
3
•
1
Slelkln
(MLE)
260
2.0 1 10.0
r 1
0.1 1.0
rl 1 *. _
1 1
10.0 100.0
pg/kg/day
Figure 1. Some examples of risk specific doses (10"s ) and reference doses calculated by
Individual scientists, scientific organizations, and regulatory agencies for 2,3,7,8-TCDD. Solid
lines represent conclusions reached by regulatory agencies or scientific organizations; hatched
lines represent research efforts. Some values represented as lines with a single point could
be represented as a range. The point shown is generally the lowest RsD or RfD of the range.
* Very preliminary analysis, taking into account the longer half-life of 2,3,7,8-TCDD in humans
relative to rats.
Abbreviations used in the chart are as follows:
CDC Centers for Disease Control, U.S. Public Health Service
EPA U.S. Environmental Protection Agency
FDA U.S. Food and Drug Administration
FRG Ftderal Republic of Germany !
LMS Linear multistage model
M-K-V Moolgavkar, Knudson, Venzon
MLE Maximum likelihood estimate
NRC National Research Council, Canada
i
The Una Indicated as "Canada" represents both Health and Welfare Canada and the
Province of Ontario, Canada
Source: Taken from Appendix A.
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a traditional toxicological approach.2 Based on science policy considerations
developed in section III.B.2.b.6., the Workgroup proposes that the Agency adopt
a dose of 0.1 pg/kg/day as a plausible upper-bound increased lifetime risk of
one in a million. The estimate is consistent with the available data and
theories and represents a reasonable science policy position.
B. COMPARISON OF POTENCY ESTIMATES
Plausible upper-bound cancer potency estimates for 2,3,7,8-TCDD published
by EPA, other U.S. agencies, some state agencies, foreign governments, and
individual investigators fall within a range that spans more than three orders
of magnitude. This range represents the lack of current consensus regarding
approaches to estimating levels associated with potential cancer risk.
Comparison of the various assessments has two purposes: (1) to demonstrate how
scientists, using the same data but different assumptions and/or science
policies, arriye at different risk estimates, and (2) to discuss alternative
methods to EPA's customary approach, which is based on the upper-confidence
limit (UCL) of the IMS model, for estimating human cancer risk from exposure to
2,3,7,8-TCDD. This explicit discussion of assumptions and policy choices
highlights some of the uncertainties inherent in the risk assessment process
generally, and in analyses and assessments specific to the carcinogenicity of
2,3,7,8-TCDD.
2It should be recognized that neither of these methods attempts to estimate the
"true risk" posed by exposure to a chemical. In the case of the linearized
models, because our understanding of the mechanism of carcinogenesis is so
limited, an upper limit to the risk is calculated. In its risk assessment,
using the IMS model, EPA stresses that the true risk is likely to be lower
than the "plausible upper bound," and may be zero. Likewise, the traditional
toxicological approach does not attempt to estimate risk; rather, it estimates
a lifetime daily dose likely to be without significant risk.
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Alternative approaches to estimating cancer risk are discussed in terms of
existing 2,3,7,8-TCDD risk assessments to give EPA risk assessors and risk
managers established,points of reference and to distinguish science, science
policy, and assumptions. The analysis shows that differences among the risk
assessments for 2,3,7,8-TCDD developed by various regulatory agencies are not
due to disagreements about the scientific data base per se, but rather are due
to the judgments, science policy positions, and methods used in estimating
human risk. Indeed, a major factor accounting for the differences in the
various assessments is the judgment reached on whether or not a threshold
exists for'the carcinogenic activity |of 2,3,7,8-TCDD. U.S. agencies, including
EPA, have selected the LMS model, a mathematical model based on a dose-response
function that does not have a threshold (that is, some non-zero risk can be
I „• :
calculated for all dose levels) and h|as a low-dose response characteristic that
is essentially linear. Some Canadian and European environmental agencies, as
well as some state agencies in this Country, have selected a traditional
toxicological approach based on an experimentally established
no-observed-effect-level (NOEL)3 to estimate a presumed "safe dose." While
choice of model often reflects, in part, the historical or philosophical
tradition of a particular agency, in the case of 2,3,7,8-TCDD it also reflects
important differences in the way different scientists interpret and weigh the
scientific evidence and related uncertainties.
3Dose in the chronic animal bioassay at which no Increase in tumor incidence
was observed.
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C. ORGANIZATION
The background information in Chapter II refers to a report on sources and
routes of human exposure to 2,3,7,8-TCDD, summarizes animal and human data on
the potential for human canceri, and surveys existing quantitative cancer risk
assessments for 2,3,7,8-TCDD. The section on mechanisms of carcinogenesis
evaluates 2,3,7,8-TCDD in light of several different mechanistic hypotheses,
while the remainder of the chapter focuses on mathematical approaches for risk
extrapolation. Chapter III synthesizes the range of qualitative and
quantitative considerations bearing on the human cancer risk potential of
2,3,7,8-TCDD, and explains the basis for selecting 0.1 pg/kg/day as a cancer
risk-specific dose (10~6) for this chemical.
While this report draws on EPA's 1985 Health Assessment Document (HAD)
for Polychlorinated Dibenzo-p_-Dioxins (U.S. EPA, 1985) for certain data, it
incorporates new information and alternative interpretations of the scientific
evidence.* The analysis follows EPA's Guidelines for Carcinogen Risk
Assessment which call for articulation of "major assumptions, scientific
judgments and to the extent possible, estimates of the uncertainties embodied
in the assessment. . .distinguishing clearly between fact, assumption, and
science policy" (U.S. EPA, 1986a).
40ther sources include a "Dioxin" Update Committee Report of a meeting held
July 1-2, 1986 (submitted to the Office of Pesticides and Toxic Substances
August 28, 1986; hereafter called the "Pitot Report") (U.S. EPA, 1986c), the
"Report of the EPA Workshop on the Development of Risk Assessment
Methodologies for Tumor Promoters" (hereafter called the U.S. EPA "Promoter
Workshop") (U.S. EPA, 1987c), and six issue papers developed as background
information for this reanalysis (Appendices A through F).
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II. I ANALYSIS
In this chapter data from laboratory and human studies are evaluated in
terms of the factors that influence risk assessment methodology generally and
that raise specific questions about existing cancer risk assessments for
2,3,7,8-TCDD. In place of models that appear to be based upon a dichotomous
view of carcinogens as either complete carcinogens or pure promoters,
researchers are exploring more sophisticated models which span a variety of
direct and indirect mechanisms, including a combination of several modes of
action. Qualitative answers to the questions regarding mechanism(s) by which
2,3,7,8-TCDD exerts its carcinogenicity could have a significant effect on the
quantitative estimates of risk.
This chapter addresses these questions. Section A identifies relevant
data from laboratory and human studies. Section B briefly reviews current
[
theories on mechanisms of carcinogenesis as they apply to this chemical. In
section C, several risk assessment models are reviewed in light of current data
on 2,3,7,8-TCDD and mechanistic considerations. The special question of body
burden data, particularly its meaning for the epidemiologic studies, is
reviewed in section D. The implications of these several factors for human
cancer risk from exposure to 2,3,7,8-TCDD is discussed in section E.
i
A. BACKGROUND ;
This section summarizes basic data on the effects of 2,3,7,8-TCDD in
animal, human, microbial, and in vitro studies. Section 1 outlines data on
sources and routes of exposure; section 2 summarizes the animal and human
studies that provide the foundation for most risk assessments for 2,3,7,8-TCDD;
8
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and section 3 presents relevant ancillary information. These summaries
abstract data reviewed and analyzed in the issue papers prepared for this
report (Appendices A through F) and other sources cited.
1. Exposure Considerations
A comprehensive review and analysis of human exposure to 2,3,7,8-TCDD
appears in a draft document entitled "Estimating Exposures to 2,3,7,8-TCDD"
(U.S. EPA, I987a). This report should be consulted for information on sources
of 2,3,7,8-TCDD, its movement through the environment, routes of human
exposure, and possible human doses resulting from those exposures. v
The report was prepared by scientists and engineers from the Exposure
Assessment Group, Office of Health and Environmental Assessment. The primary
purpose of the report is to provide a review and update of information related
to exposure to 2,3,7,8-TCDD that has come to light since 1984. In addition,
this report provides an illustration of the application of this information in
performing exposure assessments for 2,3,7,8-TCDD. This is accomplished by
using the information to construct several scenarios where contaminated
material may result in exposure to 2,3,7,8-TCDD, and estimating what the
exposure would be for various pathways from source to humans exposed. Sources
used as examples in this report include contaminated soil, various land
disposal situations, and municipal waste incinerators. It must be emphasized
that these scenarios are not to be interpreted as an exposure/risk assessment
for all sources of these types. This report should, however, provide a sound
starting point for many exposure assessments of 2,3,7,8-TCDD contamination.
2- Carcinogenlcltv of 2.3.7.8-TCDD
The carcinogenic potential of 2,3,7,8-TCDD for humans has been the focus
of intensive study arid debate for almost a decade, and the issue is still not
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resolved. While chronic exposure studies in laboratory animals demonstrate
that 2,3,7,8-TCDD is a potent carcinogen in rodents, epidemiologic data have
been much more difficult to interpret and more controversial. These issues are
addressed in this section and described more fully in the issue papers prepared
by Bayard (Appendix A) and Bayliss (Appendix B).
a. Chronic Animal Studies
There is general agreement that 2,3,7,8-TCDD is carcinogenic in laboratory
animals. The critical studies are from two independent laboratories and show
effects in both rats and mice. These and other experimental animal studies are
fully discussed in EPA's HAD (U.S. EPA, 1985).
In a chronic toxicity and oncogenicity study by Kociba et al. (1978),
dietary doses of approximately 0, 1,000, 10,000, and 100,000 pg/kg/day
2,3,7,8-TCDD were fed to rats for up to 2 years (1 pg = 10'12 gram, so that
100,000 pg/kg/day is equal to 0.1 ug/kg/day). In the high-dose group, both
male and female animals had significant site-specific increases in tumors. The
target organs and tumor types in male animals were squamous cell carcinomas of
the tongue, hard palate, and nasal turbinates, and adenomas of the adrenal
cortex; in female animals, the target organs and tumor types were
i
hepatocellular carcinomas and squamous cell carcinomas of the tongue, nasal
turbinates, and lung. Most investigators interpret this study as demonstrating
I
that dietary exposure to 2,3,7,8-TCDD at 100,000 pg/kg/day or greater results
i
in increased tumor Incidences in both male and female rats. If neoplastic
10
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nodules are combined with hepatocellular carcinomas, a statistically
significant response is also seen at 10,000 pg/kg/day in female rats.^
The National Toxicology Program (NTP) has also tested 2,3,7,8-TCDD for
carcinogenicity in rats and mice following administration by gavage (NTP,
1982). Rats were exposed to weekly doses of 0.0, 10,000, 50,000, and 500,000
pg/kg. The only tumors that appeared to be treatment-related were follicular
cell adenomas or carcinomas of the thyroid in male animals, and neoplastic
nodules or hepatocellular carcinomas of the liver in female animals. The
incidence of these tumors was significantly greater in the high-dose groups
than in controls, and the incidence of tumors at both sites showed a positive
dose-related trend. Under the conditions of this bioassay, the NTP concluded
that 2,3,7,8-TCDD was carcinogenic in both male and female rats.
In the NTP mouse study, male mice were exposed to weekly doses of 0.0,
10,000, 50,000, and 500,000 pg/kg, while female mice were exposed to weekly
doses of 0.0, 40,000, 200,000, and 2,000,000 pg/kg. An increased incidence of
liver tumors was also observed in the NTP study in the high-dose male mice and
in the high-dose female mice. Female mice also had an increased incidence of
follicular-cell adenomas of the thyroid. In this study, 2,3,7,8-TCDD was
carcinogenic to mice, with effective doses ranging between 500,000 and
2,000,000 pg/kg/week (0.5 and 2.0 ug/kg/week) depending on sex.
5The EPA Guidelines for Carcinogen Risk Assessment (U.S. EPA, 1986a) address
the question of combining benign and malignant lesions of identical histogenic
origin. In addition, the Agency's Risk Assessment Forum has published
guidance on the use of various pro!iferative hepatocellular lesions of the
rat in risk assessment (U.S. EPA, 1986b). In the case of 2,3,7,8-TCDD, both
would recommend combining benign and malignant lesions; however, both suggest
that when such lesions are combined, the impact of the benign lesions on the
quantitative response should be presented explicitly.
11
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On the basis of information contained in these two studies, and other
animal data described in the HAD (U.S. EPA, 1985), the Agency concluded that
i
the evidence from animal studies for a carcinogenic response induced by
L
exposure to 2,3,7,8-TCDD is "sufficient" under the weight-of-evidence system in
EPA's then Proposed Guidelines for Carcinogen Risk Assessment (U.S. EPA, 1984).
b. Epidemiologic Studies j
Several epidemiologic studies designed to evaluate cancer in humans
potentially exposed to 2,3,7,8-TCDD or substances presumed to contain
2,3,7,8-TCDD are subjects of sharp debate. Results of these studies, including
both cohort and case-control designs, iare arguably conflicting, with some
studies reporting high risks and others reporting low or no detectable risks.
i
Although most of the studies have followed standard epidemiologic
I
procedures, the conclusions of all are subject to specific limitations. Of
particular concern are uncertainties about the nature and extent of actual
exposure to 2,3,7,8-TCDD.6 In every instance, because of a lack of empirical
i
exposure data, some surrogate basis for estimating exposure has been used.
t
Studies in which human populations have been examined for carcinogenic
responses to exposure to substances containing 2,3,7,8-TCDD are reviewed in the
issue paper by Bayliss (Appendix B).
Two epidemiologic studies from Sweden (Hardell and Sandstrom, 1979;
i
Eriksson et a!., 1981) are considered by many to be critical studies in
assessing potential human cancer risk. They suggest a high cancer risk
associated with exposure to chemicals! contaminated with 2,3,7,8-TCDD. These
6Recent analyses of tissue from subjects in some of these studies suggest that
"control" and "exposed" have roughly the same levels of 2,3,7,8-TCDD in their
tissues (Hardell, 1987). The impact! of this finding on the interpretation of
the studies has not been fully assessed.
-------
studies have reported statistically significant, five- to sevenfold elevated
risks of soft tissue sarcoma (STS)7 related to occupational exposure to phenoxy
herbicides and/or chlorophenols, some of which were assumed to contain
2,3,7,8-TCDD. While some methodological questions have been raised about these
studies, it^appears that the elevated risks (at some level of exposure) are
real and,,,,should be considered in hazard evaluation.
In addition to the two Swedish studies and certain case reports, other
studies (including studies in certain U.S. populations) may give some support
to the association between exposure to 2,3,7,8-TCDD and STS (see Appendix B);
however, such an assessment is widely debated. These and other studies have
also reported associations between other forms of cancer and exposure to
2,3,7,8-TCDD or chemicals likely to be contaminated with chlorinated dibenzo-fi-
dioxins (CDDs). For example, Hardell et al. (1981) reported a statistically
significant risk of non-Hodgkin's lymphoma (NHL) in agricultural, forestry, and
woodworking employees exposed to phenoxy herbicides, chlorophenols, or both.
The relative risk ratio ranged from 4.3 to 6.0 for both classes of compounds
together as well as separately. In addition to NHL, results from these studies
have raised questions about increased risks of stomach cancer, prostate cancer,
Hodgkin's disease, and kidney cancer. Since reports of increased risks for
cancers other than STS and NHL occur sporadically throughout these studies,
they are generally considered inconclusive.
7"Soft tissue sarcomas constitute a category of rare cancers with a total
mortality rate of 1 STS per 100,000 persons per year in the United States.
The soft tissue sarcomas are malignant neoplasms of diverse histologic
subtypes, which occur throughout the body in mesenchymal connective tissue
other than bone. These malignancies are often not reported accurately on
death certificates and may not be recognized accurately by general
pathologists" (Fingerhut, 1986). The effect of such misdiagnoses could either
under- or overreport the number of STS cases in a study.
13
-------
In contrast, several studies involving populations believed to have been
exposed to 2,3,7,8-TCDD or 2,3,7,8-TCDD-containing chemicals have not shown any
significant increased incidence of cancer (Appendix B).
Studies of Vietnam veterans have been the subject of particular interest
because it is thought that some of these veterans were exposed to 2,3,7,8-TCDD
as a result of exposure to Agent Orange in Vietnam.8 Based on Agent Orange use
and potential for exposure under the conditions in Vietnam, it has been assumed
i
some subsets of the population may have been exposed to relatively high levels
of 2,3,7,8-TCDD. To date, most of these studies have shown no statistically
<• . • j
significant correlation between Vietnam service (and, therefore, possible
exposure to 2,3,7,8-TCDD) and an increased risk of cancer. For example, the
Ranch Hand study has so far reported only a limited number of deaths (six) from
cancer among the exposed group, none of which was from STS or lymphoma
(Fingerhut et al., 1984). Studies of the mortality patterns among New York
service men with and without Vietnam experience found no significant
association between cancer and serviced in Vietnam (Greenwald et al., 1984),
i
although it should be noted that a study of Massachusetts Vietnam veterans
reports a significant excess of connective tissue sarcomas compared to
non-Vietnam veterans (Kogan and Clapp, 1985).
Although the epidemiologic data afe not persuasive regarding one
interpretation over another, the high relative risks seen in the Swedish
studies are noteworthy. While an association may exist between exposure to
8Agent Orange, an herbicide widely use'd in Vietnam, was composed of equal parts
of butylesters of 2,4,5-trichlorophenpxyacetic acid (2,4,5-T) and 2,4-
dichlorophenoxyacetic acid (2,4-D). 2,3,7,8-TCDD, a contaminant in Agent
Orange, has been shown to originate from the 2,4,5-T component with levels
ranging from 0.1 to 47 ug/g (U.S. EPA, 1985).
14
-------
chemicals contaminated with 2,3,7,8-TCDD (e.g., phenoxy herbicides) and
increased incidences of cancer, the data are still too uncertain to attribute
the effects seen to 2,3,7,8-TCDD.9
In light of the above considerations, and in accordance with the Agency's
Guidelines for Carcinogen Risk Assessment (U.S. EPA, 1986a), the human evidence
supporting an association between exposure to 2,3,7,8-TCDD and cancer is
considered inadequate.
3. Altered Cell Function
Animals given 2,3,7,8-TCDD as a single dose of 300,000 to 3,000,000 pg/kg
exhibited a significant increase in a variety of enzymes responsible for the
toxification/detoxification of foreign chemicals in the liver (increased
enzymatic activity has also been found in other organs) (Poland and Glover,
1975). Sloop and Lucier (1987) have shown a statistically significant increase
in arylhydrocarbon hydroxylase (AHH) in animals exposed to 15,000 pg/kg of
2,3,7,8-TCDD in corn oil.10 In other systems it has been shown that the
toxification/detoxification process can, in some cases, yield genotoxic
intermediates when metabolizing ingested chemicals to harmless substances
One current investigation involves a National Institute for Occupational
Safety and Health (NIOSH) registry of U.S. workers who have
been employed in industries that manufactured chemicals thought to be
contaminated with 2,3,7,8-TCDD, as well as other chlorinated dibenzo-p.-
dioxins (CDDs) and chlorinated dibenzofurans (CDFs). A study of this
group of 7,000 workers, scheduled for completion in 1989, could provide
substantial, valuable, and additional information on the question of the
carcinogenic potential of 2,3,7,8-TCDD in humans.
AHH is a cytochrome P45G-mediated microsomal mono-oxygenase that metabolizes
numerous chemicals. AHH is induced by a sequence of events starting with
the binding of 2,3,7,8-TCDD to the Ah receptor located at the cellular mem-
brane. It is the translocation of the Ah receptor/2,3,7,8-TCDD complex to
5e«[]ucleus and b1nd™9 to. the DNA that leads to the transcription/induction
of AHH.
15
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suitable for excretion. 2,3,7,8-TCDDjis poorly metabolized, and therefore the
I
increased activity of these enzymes i? likely to have little effect on the
genotoxic potential of 2,3,7,8-TCDD itself. Available data suggest that
2,3,7,8-TCDD has little or no ability to act as a direct genotoxin. However,
the increase in toxification/detoxification enzymes does increase the
possibility that other exogenous chemicals will be activated to genotoxic
substances. Thus, it is possible to postulate a process whereby 2,3,7,8-TCDD,
I
through a secondary route, causes cancer through a mechanism involving
genotoxicity.
The available information on enzyme induction indicates that 2,3,7,8-TCDD
i
is one of the most potent inducers studied. Poland and Glover (1984) found
that 0.85 nmoles/kg of 2,3,7,8-TCDD ejlicited the ED50 level of AHH hydrocarbon
hydroxylase in rat liver compared to J25,500 nmoles/kg of 3-methylcholan-
threne.ll
In addition, the increased enzymatic activity elicited by 2,3,7,8-TCDD
remains elevated, near peak level, for over 30 days while the enzymatic
activity elicited by 3-methylcholanth|rene returned to normal in 8 days,
reflecting the influence of 2,3,7,8-TCDD's long half-life (see Appendix A).
Limited data using human cells in culture indicate that enzyme induction is
likely to take place in humans exposed to 2,3,7,8-TCDD (Jaiswal et al., 1985).
Additional cellular data suggest an impact of 2,3,7,8-TCDD on such diverse
I
responses as increased proliferation,! antagonism of hormone-mediated responses,
cytotoxicity, and in vitro transformation (see section B).
n3-Methylcholanthrene is used routinely by investigators as an inducer of
AHH.
16
-------
While this information 1s of interest and does signal qualitative concern
for carcinogenicity in humans exposed to 2,3,7,8-TCDD, it is not possible at
present to factor these observations into a quantitative risk assessment.
4- Weight-of-Evidence Conclusion
Based on sufficient evidence in animal studies, inadequate human evidence,
and consideration of ancillary or supportive information, 2,3,7,8-TCDD is
classified as a B2--probable human carcinogen in EPA's weight-of-evidence
scheme. The Agency reached the same conclusion in the 1985 HAD without
•: -,•'••
invoking the contribution of the ancillary data.
This classification represents a qualitative judgment as to the likelihood
that 2,3,7,8-TCDD may be a human carcinogen at some dose. It does not reflect
a calculation of potency nor does it resolve the issue of threshold versus
nonthreshold approaches to describe the carcinogenic dose-response. In its
simplest terms, this classification represents the consensus of scientific
opinion that ". . .in the absence of adequate data on humans, it is
biologically plausible and prudent to regard agents for which there is
sufficient evidence. . .of carcinogenicity in experimental animals as if they
presented a carcinogenic risk to humans." (IARC, 1987).
B. MECHANISMS OF CARCINOGENESIS
Qualitative evidence for designating a chemical as a potential human
carcinogen comes from a variety of observations—human epidemiologic studies,
chronic animal bioassays, in vitro studies, metabolic studies, and mechanism
studies. Thus, conclusions depend not on a single piece of information, but
rather, a weight-of-evidence assessment of all data bearing on whether the
chemical is or is not carcinogenic.
17
-------
At present, carcinogenesis, as a process, is hypothesized to be a
continuum of events characterized by a number of steps — some irreversible and
others reversible—that result in the [transformation of a normal cell to a
malignant one. While some of these steps have been defined by responses
observed in laboratory experiments, the actual sequence of cellular events
leading to (1) initiation of a cell, (2) clonal expansion of the population of
I
initiated cells, and (3) progression leading to malignant transformation is
i
still unknown for even a single chemical.*2 Thus, chemicals are often
i
categorized by responses seen in vivo!and in vitro studies and/or where in the
carcinogenic process they act. For example, simply stated, a "complete
carcinogen" is one that can lead to tumor formation in the absence of any other
known exogenous factor and a "promoted" is a substance that can affect the
growth and clonal expansion of a population of initiated cells, and can alter
gene expression (U.S. EPA, 1987c).
While such categories are theoretically easy to describe, in practice it
is often difficult (if not impossible! to separate carcinogens into discrete
categories based on mechanism of actibn. This process is further complicated
by the possibility of multiple carcinogenic mechanisms, direct and indirect,
occurring as a result of exposure to a single compound. In fact, especially at
low doses it is not clear that any carcinogen affects all stages; the
hypothesis of "linearity-at-low-doses" does not require this.
12It should be noted, that initiation!, promotion, and transformation may
also be made up of a series of steps.
18
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1- Hypothesis 1: 2.3,7,8-TCDD as a Direct-Acting. Complete Carcinogen
a. Qualitative Consideration
At present, carcinogens are defined operationally because of a lack of
understanding of the mechanisms by which chemicals cause cancer. Thus, obser-
vations of an increased number of rare tumors, a decrease in the time-to-tumor,
or a statistically significant increase of site-specific tumor incidence in
multiple species and strains in animal bioassays have generally been taken to
support the conclusion that the test chemical is a complete carcinogen.
Unfortunately, the bioassay gives us no information about the process leading
to the tumorigenic response. To give further support to the characterization
of a chemical as a direct-acting, complete carcinogen, i.e., one that acts
through a mechanism that involves DNA damage, the weight-of-evidence analysis
includes information from a variety of other observations. These include
short-term mutagenicity studies, irreversible binding to DNA, clastogenicity,
and other indications of genotoxicity. Positive results in some or all of
these types of tests are considered by some as necessary before a chemical can
be considered to be a complete carcinogen. Conversely, negative results in
these types of tests are considered by some as an indication that a chemical
may not be a complete carcinogen. Although some mechanisms, such as oncogene
activation, may also be considered as "direct" negative results in more
traditional genotoxicity tests, they have been used to argue against
interpretation of positive bioassays results as the consequence of direct,
complete carcinogenesis.13
13The use of the term "direct" in this context is meant to convey the notion of
a direct impact of a chemical or its metabolites on the carcinogenic process
and should not be confused with the use of the term direct (parent compound)
versus indirect (metabolite) carcinogenicity as described by some in the
scientific literature.
19
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2,3,7,8-TCDD produces tumors inlanimals in lifetime studies (Kociba et
a!., 1978; NTP, 1982). Tumors were seen at multiple sites in rats and mice of
both sexes. However, attempts to demonstrate that 2,3,7,8-TCDD is either
mutagenic or genotoxic have been mostly negative. Recent reviews by Fishbein
(1987), Giri (1987), and Shu et al. (1987) of over 20 studies have concluded
that 2,3,7,8-TCDD is probably not gehotoxic. In addition, Randerath et al.
[
(1987) demonstrated that 2,3,7,8-TCDD exposure does not result in measurable
j
DNA adducts, and, therefore, probably does not bind irreversibly to DNA.
Furthermore, Lim et al. (1987) published data that show a lack of ;
clastogenicity in animals exposed to!2,3,7,8-TCDD. Thus, the kind of
mechanistic data that would support (classification of 2,3,7,8-TCDD as a
direct-acting, complete carcinogen on the basis of genotoxicity are lacking.
i
A recent review comparing the results from short-term tests and long-term
chronic bioassays for 76 chemicals (Tehnant et al.» 1987) showed that the
i
concordance between short-term tests! and a response in a chronic bioassay was
only 60%. While highly predictive for certain classes of compounds and for
those substances that are routinely positive, this review indicates that the
lack of positive results in short-term tests is often not a predictor of the
outcome of a chronic bioassay, and, 'therefore, may not be a reliable predictor
of direct, complete carcinogenicity.
Also, Reynolds et al. (1987) have shown
that at least two chemicals that tes|t negative in mutagenicity tests conducted
by the NTP can activate oncogenes wh|ich may result in a specific irreversible
i
cellular change. This is considered by many scientists to affect control of
cell proliferation and, perhaps, to !be an important step in the carcinogenic
process. These kinds of observations suggest that as new approaches to
investigating molecular events potentially involved in the carcinogenic process
1 20
-------
become available, our understanding of how particular chemicals exert their
carcinogenic effect may change.
It is importan^ to note that while the promoting capability of
2,3,7,8-TCDD has been clearly demonstrated in the liver, the tumors observed in
one animal bioassay in the lung, soft palate, and nasal turbinates can be taken
as evidence supporting complete carcinogenicity; although, even in these
tissues,"promotion of pre-existing initiation (or complete carcinogenesjs by
unknown mechanisms that are not representative of response at the administered
dose), based on locally high levels of 2,3,7,8-TCDD found in food particles,
cannot be ruled out (Kociba, 1984). In addition, studies that use a liver
. ' '\
system to show promoting effects of 2,3,7,8-TCDD yield data suggesting that
some initiating potential cannot be totally ruled out. In sum, it is not
possible, at this time, to conclude definitively whether or not 2,3,7,8-TCDD is
acting as a complete carcinogen in some tissues.
b. Quantitative Considerations
For complete carcinogens that act by directly causing an irreversible
initiating event and then by fostering promotion and progression of the cells
to a frank tumor, no threshold would be expected on the basis of current
theory. In addition, if a carcinogen caused its effect by adding irreversibly
to a background process already underway, linearity at low doses would be
expected. The EPA has integrated the two concepts of irreversibility and
additivity in deriving the use of a plausible upper bound from the IMS model
for carcinogens as a matter of science policy. While a number of models, in
addition to the IMS model, incorporate the concept of low-dose linearity, there
is currently no biological basis for the choice of one of these alternatives
over the IMS model.
21
-------
Consequently, if the Agency considers 2,3,7,8-TCDD to be a direct-acting,
complete carcinogen, then, the use by IEPA of a plausible upper bound derived
i - '• '
from the IMS model is appropriate and Iconsistent with Agency science policy.
i ' • , - •
To the extent that the mechanism of action of 2,3,7,8-TCDD is not in accord
with the derivation of the model, the |use of the IMS model may be less
appropriate. ,
2. Hypothesis 2: 2,3,7,8-TCDD as a "Pure" Promoter
f
f ' .
a. Qualitative Considerations |
i . ' . .
Like complete carcinogens, promoters are defined operationally. Promoters
are defined as providing a certain pattern of results in initiation/promotion
tests. In theory, therefore, because "pure" promoters do not cause initiation,
one might expect a promoter to give negative results in an animal
carcinogenicity bioassay. In practice, however, a promoter could yield
positive results in such a test because of initiated cells present as a result
of "background" events that can be promoted to yield a tumorigenic response.
For most promoters, the sensitivity of the bioassay would be expected to be too
low to yield a statistically significant response based only on background
i • ,
events. Thus, under these terms, bioassay studies would allow certain
promoters to be distinguished from complete carcinogens.
In order to characterize a chemical as having promoting potential only,
I . .
additional information from other tests is generally considered to be
necessary, such as: a lack of initialing potential as measured by a lack of
- i .
genotoxicity, reversibility of the promotion response, inhibition of
cell-to-cell communication14 and/or demonstration of a dose-response having
^Several investigators (e.g., Troskoi, 1983) have suggested that many promoters
may act by inhibiting cell-to-cell Communication as measured in metabolic
cooperation or dye-transfer studies!.
. '22 ' , .
-------
threshold characteristics. It should be noted, however, that promoters do not
necessarily have to exhibit all of these characteristics, but that positive
results in these kinds of tests can help to support the categorization of a
chemical as having only promotion potential.
Currently, two in vivo systems are commonly used to study the promotional
phase of the carcinogenic process—rat liver and mouse skin. In both systems
the animal is given a very small amount of a known potent initiator followed by
a promoter; in the case of one of the controls, the order is reversed.
Assuming the test compound is a promoter only, the results of such experiments
can be expected to yield results as follows:
Test Protocol Result
initiator only no tumors
initiator followed tumors
by a promoter
promoter followed no tumors
by an initiator
promoter only no tumors
In both rat liver and in some mouse skin studies, 2,3,7,8-TCDD is a potent
promoter when tested with a known initiator (U.S. EPA 1986c; 1987c). In both
systems, however, a low incidence of tumors in animals given only 2,3,7,8-TCDD
suggests that 2,3,7,8-TCDD may have some initiating potential or that the
observed tumors are the result of potent promotion of background events or
unidentified initiators found in the animals' environment (e.g., substances in
food). The results of such a study involving diethylnitrosamine (DEN) and
2,3,7,8-TCDD in partially hepatectomized rats are illustrated in Table 1.
In addition to the in vivo studies for promotional activity, a number of
laboratories have investigated the promotion potential of 2,3,7,8-TCDD in
23
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TABLE 1 PROMOTING EFFECT OF 2,3,7,8-TCDD (TCDD) ON HEPATOCARCINOGENESIS IN PARTIALLY
TABLE 1. FEMALE RATS BY A SINGLE DOSE OF DIETHYLNITROSAMINE (DEN)
No. of enzyme-
altered foci per
No. of cubic centimeter
Group Treatment animals of liver
1 DENa 4 . ' i 309 ± 98b
2 TCDD (low dose)c 4 34 ± 17
3 TCDD (high dose)d 5 25 ± 7
4 Phenobarbital (control) 4 .56 ± 13
5 DEN + TCDD (low dose) 5 1068 ± 166
6 DEN + TCDD (high dose) 7 871 ± 66
7 DEN + phenobarbital 10 533 ± 103
a DEN given at dose of 10 mg/kg.
b Mean ± SD.
c 0.14 ug/kg/2 weeks.
d 1.40 ug/kg/2 weeks.
e Three rats exhibited neoplastic nodules in the liver.
f One rat exhibited neoplastic nodules in the liver.
Source: Pi tot et a!., 1980.
24
Mean volume No. of
of enzyme- % liver rats
altered volume with
foci (cubic occupied car-
millimeter) by foci cinoma
0.02 0.7 0
0.05 0.2 0
0.04 0.1 0
0.01 0.1 0
0.08 9.0 Oe
0.49 43.0 5f
0.15 6.0 8
-------
isolated cell systems (see, for example, Abernethy et al., 1985). The results
of these studies clearly demonstrate that 2,3,7,8-TCDD affects isolated cells
as a promoter but does not alter the cells in a manner which would suggest that
an initiating event has occurred. Hence, under the most stringent conditions
(in vitro), 2,3,7,8-TCDD may act as a "pure" promoter, but under in vivo
conditions the results are not as clear cut.
The reversibility of clonal expansion after the removal of a promoting
substance has been considered a key effect in characterizing promotional
activity. For example, Pitot et al.. (1987) showed that the size of liver foci
in rats increased in the presence of phenobarbital (a chemical generally
regarded as a "pure" promoter) and then returned to normal when phenobarbital
was removed from the diet.15 Investigation of 2,3,7,8-TCDD for reversibility
of foci formation in similar experiments has not been attempted because of its
long half-life in tissues, making results less easily interpretable.
It can also be argued that 2,3,7,8-TCDD does not meet the above criteria
for a "pure" promoter. Some tumors observed in the lung, nasal turbinate, and
hard palate may be the result of "complete carcinogenicity," although, as
discussed previously, this is not uniformly accepted. Similarly, the
reversibility of the promoter step has not been demonstrated for 2,3,7,8-TCDD.
Finally, some evidence of clastogenicity exists although the results have not
been duplicated (Green et al., 1977).
15It should be noted that re-administration of phenobarbital in this experiment
resulted in a greater than anticipated clonal expansion which could be
interpreted to mean that in addition to reversible effects, some irreversible
step(s) had occurred.
25
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b. Quantitative Considerations
A "pure" promoter that acts by facilitating clonal expansion of a
population of initiated cells without! any capacity to induce irreversible
changes in DNA may be expected to dempnstrate a threshold, theoretically,
although no quantitative framework exists which demonstrates that thresholds
must exist under all circumstances. h>r example, the Moolgavkar, Knudson, and
i
Venson (M-V-K) model, now being investigated by a number of scientists for
modeling the carcinogenicity of promoters, does not require an assumption of a
I
threshold. The theoretical basis for| the threshold response would be the
likelihood of a dose at which the netj response of clonal expansion and normal
cell death (or control of cell proliferation) would be zero. Consequently, for
such a situation an appropriate method to estimate a level of exposure that
would be unlikely to pose a significant risk may be the traditional
toxicological approach for organ-specific toxicity via the derivation of a
reference dose (RfD). This RfD is calculated from the no-observed-adverse-
effect-level (NOAEL) based on carcino|genicity noted in the bioassay, divided by
i
several uncertainty factors.
i
To the extent that the data demonstrate that 2,3,7,8-TCDD is a "pure"
[
promoter, the traditional toxicologicjal approach is arguably appropriate.
Conversely, data that indicate that 2,3,7,8-TCDD has the ability to initiate
i
the carcinogenic process or that 2,3,!7,8-TCDD may be a complete carcinogen
would make the application of the abpve method less appropriate.
3. Hypothesis 3: Secondary or Indirect Mechanisms of Carcinoaenesis
a. General Considerations
As stated earlier in this chapter, a carcinogenic response in animals may
be the result of direct interaction of the chemical (or its metabolites) under
26
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test with sensitive cellular targets. On the other hand, this response may be
the result of indirect effects or even a combination of the two.
While it is not possible to distinguish between direct and indirect
responses in animal bioassays, a variety of test data may be used to draw a
conclusion that indirect carcinogenicity may be responsible for an observed
effect. The nature of such indirect effects may be such that the chemical
affects the pool of "initiated cells" either qualitatively by making cells more
sensitive to initiating agents or quantitatively by causing increased numbers
• - ' • t
of initiated cells. On the other hand, chemicals may indirectly affect the
establishment or progression of preneoplastic lesions by making the cellular
environment more conducive to such events or by inhibiting the cellular
processes which keep tumor growth in check. It is quite plausible that
induction or inhibition of enzymes, competitive inhibition of normal feedback
mechanisms regulating cell proliferation, cytotoxicity, or effects on the
immune system could be responsible for an indirect increase in carcinogenic
response.
A wide variety of experimental data may be used to evaluate the inference
of indirect carcinogenicity. These may be studies focused on molecular
mechanisms or they may be at the level of altered organ function. The
following section discusses such experimental data on 2,3,7,8-TCDD. It should
be understood that this discussion is included to establish the plausibility of
this approach, and does not represent an indepth review of potential
mechanistic data. A number of lines of evidence require further development
before their impact on earcinogenicity can be assessed. A more detailed
discussion of several aspects of this approach can be found in Appendix F.
27
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b. Qualitative Evaluation of 2,3,7,8-TCDD as an Indirect
or Secondary Carcinogen
(1) Enhancement of Initiation
One way in which 2,3,7,8-TCDD could affect the pool of initiated cells is
by inducing enzymes that activate other endogenous and exogenous chemicals to
proximate carcinogens. The effects, of 2,3,7,8-TCDD on the induction of AHH
I
activity is well established in a number of in vitro and in vivo systems
(Poland and Knutson, 1982). This ehzyme induction could result in increased
levels of reactive intermediates frfDm other xenobiotic chemicals, and
i
ultimately increased numbers of initiated cells and increased potential
I . , . .
carcinogenicity. Further enhancement of carcinogenicity could result from the
potent promotional activity of 2,3,7,8-TCDD acting on indirectly initiated
cells. Studies have shown increased genotoxicity of chemicals such as
benzo(a)pyrene that require metaboljic activation when they are administered or
incubated with 2,3,7,8-TCDD in systems capable of responding to enzyme inducers
(Pahlman and Pelkonen, 1987). :
i
The integrity of the DNA molecjjle is generally thought to be important for
normal cellular function. Specific types of DNA damage alter DNA molecular
weight and may increase the probability of initiation. One measure of both
i
normal metabolic processing and increases in DNA damage is quantification of
single strand breaks in DNA. Such jstrand breaks may be caused by increased
levels of free radicals in actively metabolizing tissues. Direct measurements
of strand breaks have indicated increased breakage with exposure to
2,3,7,8-TCDD (Randerath et al., 198J7). Studies that examined changes in DNA
i
molecular weight after 2,3,7,8-TCDD treatment have provided conflicting
results. Molecular weight increases were observed in treated animals
28
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indicating fewer rather than Increased numbers of single strand breaks. Strand
breaks may also be associated with increases in DNA adducts either directly or
indirectly to chemical exposures. Randerath et al. (1987) found no DNA adducts
within the limits of detection of his assay in the livers of rats chronically
exposed to low doses of 2,3,7,8-TCDD, and Romkes and Safe (1987) did not find
2,3,7,8-TCDD enhancement of DNA adduct formation from endogenous steroids.
Changes in DNA repair capacity could also have an impact on the pool of
initiated cells. While few studies have measured repair parameters after
2,3,7,8-TCDD treatment, several observations can be noted. Treatment with
2,3,7,8-TCDD causes increases in 0-6-methylguanine content in DNA. This
observation may be consistent with an indirect effect on methylating capacity
or on a failure to rapidly repair these lesions. Studies to date have not
demonstrated that 2,3,7,8-TCDD causes increases in unscheduled DNA synthesis
(UDS). This observation suggests that, under the conditions of the study, DNA
damage has not increased to a level that can be measured with this type of DNA
repair. Based on recent studies by Busser and Lutz (1987) and Den Engelse et
al. (1986), effects on the repair system itself cannot be ruled out.
Another way that 2,3,7,8-TCDD could impact the pool of initiated cells
indirectly is through its ability to cause organ-specific cell proliferation,
thereby increasing the number of pre-existing initiated cells, which may
represent an increase of potential tumors. The ability of 2,3,7,8-TCDD to
cause hypertrophy and hyperplasia in several tissues where tumors arise has
been noted in chronic-toxicity studies (Kociba et al., 1978). The question of
stimulation of DNA synthesis caused by exposure to 2,3,7,8-TCDD is still an
open one (Busser and Lutz, 1987).
29
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(2) Enhancement of Carcinogenic! Progression
Currently accepted theory suggests that an aspect of the carcinogenic
i
process is the ability of chemical carcinogens to affect the progression of
preneoplastic cells or foci towards the malignant state. It is generally held
that this part of the process shows ah irreversible dedifferentiation based on
permanent genetic change. In the cas|e of 2,3,7,8-TCDD, the long half-life of
the molecule in human and animal systems allows for a constant impact over a
prolonged period of time, somewhat akin to the constitutive presence of a
regulator of cellular differentiation. While there is no evidence of permanent
genetic changes associated with 2,3,7,8-TCDD exposure, there are data to
suggest that the chemical can affect terminal differentiation and carcinogenic
transformation in vitro. Increases in "flat cell"/"XB cell" (Gierthy and
Crane, 1985) transformation support the notion of the potential to cause
dedifferentiation, and studies in "10T1/2 cells" (Abernethy et al., 1985) show
increased cellular transformation. ;
[
These activities represent a release from growth control which is
characteristic of the carcinogenic process and might allow for the appearance
of increased carcinogenic response in an indirect manner. Further support for
this role of 2,3,7,8-TCDD comes from its apparent ability to act as a hormone
agonist or antagonist (Umbreit and Gallo, 1988), without competing for sites on
certain hormone receptors. The suggestion that 2,3,7-8 TCDD may be acting
i
through cellular receptors as opposed to acting through the disruption of
membranes, as many "classical promoters" do, is further supported by evidence
that 2,3,7,8-TCDD does not disrupt cell-to-cell communication in vitro (Lincoln
et al., 1987). Disruption of cell-tb-cell communication has been suggested by
some investigators (e.g., Trosko, 1983) to be an attribute of many promoting
! 30
-------
compounds. This activity, or perhaps lack of activity, is found in cells with
and Without an inducible AHH system, so the characteristic is independent of
that event, again suggesting the necessary but not sufficient role that enzyme
induction might have in the carcinogenic process. This issue is described more
fully in the issue paper prepared by Gallo (Appendix F).
c. Quantitative Considerations
The suggestion, in qualitative terms, that 2,3,7,8-TCDD may be acting as
an indirect or secondary complete carcinogen through multiple mechanisms
affects the way that this chemical is viewed in quantitative terms. This
hypothesis is also consistent with the observed potency of 2,3,7,8-TCDD in
producing a carcinogenic response in vivo, in that the observed tumors could be
postulated to result from mechanisms such that 2,3,,7,8-TCDD: may increase the
pool of initiated cells, either directly or indirectly; is a potent promoter
based on experimental evidence in specific test systems; may be actively
involved in stimulating progression of tumors from benign to malignant; and may
be indirectly influencing both repair and surveillance capabilities in vivo.
Such activities, if viewed as adding to events that are already underway and
contributing to the background or "spontaneous" incidence rate of tumors, have
been argued as reasons for considering a nonthreshold, low-dose linear type of
dose-response curve as being applicable (OSTPj 1985; Crump and Howe, 1984).
This would imply an approach, under the Agency's current guidance, that might
be identical to that generally used for carcinogens, namely the use of the IMS
model to describe a; plausible upper bound on the risk. It would not be likely .
that such indirect effects would result in a greater response than would direct
effects, although this cannot be ruled out. These issues are discussed further
in section II:B.l.b. of this document and in Appendix F.
• ".' .•' -.'.-.'. " ".' • : : ..•. si ' " •• •..'.• . : •
-------
C. EVALUATION OF 2,3,7,8-TCDD AS AN "ANTICARCINOGEN"
Another attribute of chemical carcinogens that should be taken Into
i
account when attempting to understand their behavior is the activity termed
i
"anticarcinogenesis." In some cases, chemicals have been shown to overcome
background processes or their own effects at some doses to produce a net
negative effect on the carcinogenic
outcome of animal bioassays. Such
responses could have a profound effect on expectations of the dose-response if
anticarcinogenic effects were cancelling out carcinogenic effects at a given
dose in some tissues. The reproducible dip in response (at low doses) to below
'
background tumor levels in female rat liver (Kociba et al., 1978; NTP, 1982)
has suggested to some observers that* 2,3,7,8-TCDD may show such a phenomenon.
These observations are supported by ,more recent data which showed that
induction of liver foci were reduced at low doses of 2,3,7,8-TCDD in an
i
initiation/promotion assay (Pitot et al., 1987). Responses in other rat
tissues (i.e., uterine and breast tissue), also show decreases in background or
spontaneous tumor incidence which again support the observation that there is
an overall decrease in the carcinogenic response with some doses of
2,3,7,8-TCDD (Kociba et al., 1978).
The implications for an analysis of dose-response and the applicability
of certain models based on the observation of a potentially "anticarcinogenic"
response for 2,3,7,8-TCDD is discussed in the next section.
D. USE OF PREDICTIVE MODELS i
Scientific interest in interspecies and high- to low-dose extrapolation
has led to the development of a variety of predictive models, including models
for estimating the likelihood of hunian cancer based on data from animal
32
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studies. Selection of the appropriate model in a given set of circumstances is
difficult because of a lack of knowledge of concordant events across species
and at low doses. Selection is further complicated by the possibility of
multiple effects affecting the carcinogenic process.
Pleiotropism, i.e., action through multiple pathways, is not an uncommon
finding with molecules such as 2,3,7,8-TCDD. One has only to review the
earlier experiments on multistage mouse skin carcinogenesis of 2,3,7,8-TCDD to
see that, in some cases, it inhibited tumor formation by PAH initiators
'*'.'•<,': f
(DiGiovanni-et al., 1977; 1980; Berry et al., 1978). It must be emphasized
that the responses in multistage models are dependent on time, sequence of
administration, dose, and species. Hence, inhibition under some conditions
might have been predictable. This is contrasted with the two-stage liver model
(Pitot et al., 1980) in which it has been shown that orally administered
2,3,7,8-TCDD enhances the tumorigenic action of DEN. In subsequent experiments
at lower doses of 2,3,7,8-TCDD, a parabolic dose-response curve has been
reported in the DEN/TCDD initiation-promotion protocol (Pitot et al., 1987).
These results are not well understood, but they do not appear to be solely the
function of enhanced metabolism or Ah receptor binding (Safe et al., 1987).
Perhaps it is the result of alteration of epidermal growth factor (EGF)
receptors at low doses (Madhukar et al., 1984) which displays a commonality
with several steroid hormone receptors.
These findings are of importance to the approximation of human and animal
health risks from exposure to 2,3,7,8-TCDD and related molecules. Mathematical
modeling of physiological phenomena, especially those related to receptor
function, is often conducted using the Michaelis-Menton equation (1913) as
modified by Clark (1933) for the "classical" receptor model. The weight of
33
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evidence for the most prevalent 2,3,7J,8-TCDD effects falls into the category of
the receptor model (Poland and Knutsoh, 1982). According to recent findings,
hepatocarcinogenesis observed after exposure of animals to 2,3,7,8-TCDD is
related to estrogen levels or to the presence of functional ovaries (Goldstein
et a!., 1987), and di ethyl nitrosamine; hepatocarcinogenesis in partially
hepatectomized rats is first inhibited and then promoted by 2,3,7,8-TCDD (Pitot
et al., 1980; 1987). These findings indicate that 2,3,7,8-TCDD is not causing
its myriad of effects in liver by a simple one-step event such as binding to
the Ah receptor and subsequent induction of cytochrome PI-450.
Risk modeling for carcinogenic xenobiotics can be segregated into three
classes or types of models: (1) physiologically based pharmacokinetic (PBPK)
models in which the body is considered to be a small group of physiological
compartments (Hoel et al., 1983; Krewski et al., 1986; Bischoff, 1987); (2)
biologically motivated models of carcinogenesis (BMMC) in which the
carcinogenic process is considered to occur through a series of linked
reactions that result from two or more molecular events followed by a cellular
i
amplification by "promoter" molecule^ (Moolgavkar et al., 1987; Thorslund et
al., 1987; Krewski et al., 1987; U.S. EPA, 1987a); and (3) the IMS model of
Armitage-Doll as modified by Crump and Howe (1984) in which it is assumed that
a sequence of events occur within a single cell, some of which are
irreversible, leading to the neoplastic change (Armitage, 1985).
j
A model that appears to accommodate most of the critical components from
the biological data base on 2,3,7,8-jCDD is the BMMC model, which is generally
I
referred to as the M-V-K model (Moolgavkar .and Venzon, 1979; Moolgavkar and
L
Knudson, 1981). This model allows for several of the concepts of
initiation-promotion-progression, along with the growth-stimulating role of
34
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endogenous substrates such as hormones (Moolgavkar, 1987). Incorporation of
some of the factors necessary for the PBPK model can also be done using the
M-V-K model as expanded by Thorslund et al. (1987). These expansions of the
M-V-K model give the risk assessor a powerful tool for looking at cancer risk
mechanistically. This approach requires the use of assumptions for several
critical parameters and, thus, is testable but as yet unvalidated. This
approach is not available with the IMS model as originally proposed.
The IMS model might be accommodated if one hypothesizes that: (1) the
initiating event is the result of an indirect action of 2,3,7,8-TCDD through
modification of exogenous or endogenous compounds; (2) a population of
initiated cells exist; or (3) 2,3,7,8-TCDD acts through a variety of
mechanisms, some of which are, at low doses, additive to other processes
related to carcinogenicity arid already underway.
Recent reports have shown that 2,3,7,8-TCDD and other promoters in liver
enhance stimulation of DNA synthesis in situ, and stimulate repair of
0-6-methylguanine in liver DNA (Busser and Lutz, 1987; Den Englese et al.,
1986). Lutz et al. (1984) presented a scheme for promoter potency based on
stimulation of DNA synthesis and the assumption that cell division is a
prerequisite for several stages in the carcinogenesis process. These reports
indicate that 2,3,7,8-TCDD may act as a complete, indirect carcinogen,
including promoter activity, despite the lack of DNA binding or direct
mutagenesis. The sum of all these findings, along with the myriad of other
toxic responses, suggests a complex hypothesis for 2,3,7,8-TCDD carcinogenesis
in rodent liver, which is illustrated schematically in Figure 2. This
hypothesis can account for the dose-response data in the bioassays and the
multistage promotion experiments, as well as allow for incorporation into
35
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existing risk models, and the scheme is not inconsistent with the reports of
decreased tumor formation in some tissues. If the pathway through AHH activity
can be verified by demonstration of a net increase of reactive intermediates
after 2,3,7,8-TCDD treatment, then the IMS model can be used. The scheme also
presents several testable hypotheses that should be examined.
37
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III.
CONCLUSIONS
This re-examination of the carcinogen risk assessment for 2,3,7,8-TCDD
identifies several approaches to describing the possible carcinogenic
I
mechanisms for this chemical and reviews several different mathematical models
for estimating its carcinogenic potency. However, the enormously rich data
base on 2,3,7,8-TCDD is incomplete when it comes to answering the questions
j
posed at the beginning of the analysis. The available data permit
consideration of a broad array of possible scientific theories and science
policy approaches when reconsidering EPA's 1985 carcinogen risk assessment for
2,3,7,8-TCDD (U.S. EPA, 1985), but they do not provide a clear basis for
confidently choosing among them. The evaluation of each possible theory
results in varying degrees of confidence as to their plausibility, as discussed
i
in more detail below.
The literature review regarding 2,3,7,8-TCDD leads the Workgroup to
i
several conclusions:
• 2,3,7,8-TCDD is a potent promoter of carcinogenesis;
• the possibility that 2,3,7,8-TCDD acts as a direct-acting, complete
carcinogen cannot be eliminated;
• 2,3,7,8-TCDD may act through a secondary mechanism(s) which may affect
the carcinogenic process at|different stages; and
• 2,3,7,8-TCDD may act through a number of different mechanisms so that
the observed effects represent an integrated composite of several
mechanisms in operation. !
After considering all of the data, the Workgroup has concluded that
i
thinking about 2,3,7,8-TCDD either solely as a "promoter" or as a "complete"
carcinogen is an oversimplification! Rather, while it is clear that 2,3,7,8-
TCDD acts as a potent promoter, it may also affect other important carcinogenic
38
-------
processes, some of which may result In a linear carcinogenic response in the
low-dose region.
Schematically, this multiple mechanism hypothesis for 2,3,7,8-TCDD
carcinogenicity is graphically displayed in Figure 3. The upper curve is the
UCL of the IMS model, as applied in U.S. EPA (1985), in which the chemical is
treated as a complete carcinogen with no distinction made for the specific
contributions of initiation, promotion, or progression to the response. The
low-dose behavior that is appropriately modeled by the IMS approach for the
multiple mechanism hypothesis is depicted by the dotted line for the composite
effect, represented here by an arbitrarily placed line. However, the magnitude
of any, difference between the slopes of these two lines is uncertain.
Particularly from the available information, it is impossible to determine
whether there is any difference at all, or if the "true" difference is
negligible or substantial. The fact that the promoting behavior is taken as a
major factor in the tumor response for 2,3,7,8-TCDD could argue for a greater
rather than a lesser difference between the two slopes. On the other hand,
because the animal tumor response observed is the net result of many processes,
2,3,7,8-TCDD potency at low doses could be characterized by the LMS upper-bound
estimate, even if promotion is the predominant activity, because the linear
portion of the curve would be the composite of a number of different activities
acting in concert.
Quantitatively, it is not. easy to fit such a hypothesis into currently
available and well-accepted risk assessment models. Accordingly, the Workgroup
recommends that, until either a more appropriate model is developed or
additional data demonstrate that a currently available model is correct, as
assessment approach that recognizes the possibility of linearity at low doses
39
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R
I
S
K
UCL-LMS (1985)
- "Composite (1988)
D OS E
Figure 3. "Multiple mechanism" hypothesis for 2,3,7,8-TCDD
carcinogenesis. j
40
-------
and presents estimates in terms of plausible upper bounds would be preferable
as a interim approach.
The basis for these conclusions is set forth below. Section A summarizes
qualitative factors relating to choosing a mathematical model, while section B
reviews related quantitative considerations. Section C sets forth a rationale
for a recommended cancer risk-specific dose (RsD) for 2,3,7,8-TCDD.
A. -QUALITATIVE CONSIDERATIONS
Qualitatively, at least three classes of mechanisms of carcinogenic action
for 2,3,7,8-TCDD have been considered in this document. First, tumors found in
long-term bioassays in which 2,3,7,8-TCDD is the only known agent suggest that
this agent is, at least operationally speaking, a complete carcinogen. Second,
data from in vivo studies for promoter activity demonstrate that 2,3,7,8-TCDD
is a potent promoter of carcinogenesis with little or no demonstrated ability
to act as a direct genotoxin. Finally, an indirect impact on carcinogenic
processes is suggested by studies linking 2,3,7,8-TCDD to responses such as
enhancement of initiation, increased cell proliferation, antagonism of
hormone-mediated responses, cytotoxicity, and in vitro transformation activity.
Despite evidence supporting each potential mechanism, 2,3,7,8-TCDD does
not exhibit certain properties generally expected for each of the first two
mechanisms. For example, if 2,3,7,8-TCDD directly initiated a "complete"
carcinogenic response, genotoxicity or binding to DNA would be expected. The
absence of these effects in studies involving this chemical does not rule out
the possibility that 2,3,7,8-TCDD is a direct initiator, but it suggests that
2,3,7,8-TCDD is not a "typical" complete carcinogen. Unlike many promoting
agents, 2,3,7,8-TCDD acts at low doses. (2,3,7,8-TCDD is active at doses 1000
41
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times less than other known promoters.) Furthermore, reversibility, an
j
oft-cited characteristic of promotion, has not and cannot easily be
demonstrated because of the long half-life of 2,3,7,8-TCDD in biological
systems.
In short, it appears that 2,3,M-TCDD has characteristics of a potent
promoter, and, operationally, of a complete carcinogen. As observed
previously, the Workgroup has concluded that thinking about 2,3,7,8-TCDD either
solely as a "promoter" or as a "complete" carcinogen is an oversimplification.
Rather, 2,3,7,8-TCDD produces a broad spectrum of biological responses that
allows many hypotheses regarding the mechanism of 2,3,7,8-TCDD toxicity and
carcinogenicity. Because the available data are not adequate to absolutely
confirm or refute one or more of thjese approaches, each is considered in
evaluating potential quantitative methods for estimating the risk associated
with exposure to this chemical.
B. QUANTITATIVE CONSIDERATIONS !
I
The range of potential mechanisms suggests a range of different
quantitative approaches. If 2,3,7,8-TCDD is treated as a direct-acting,
complete carcinogen, a model incorporating linearity at low doses would be
appropriate for dose-response assessment, for the reasons mentioned in the EPA
guidelines and the OSTP Cancer Principles (U.S. EPA, 1986a; OSTP, 1985). At
the other end of the spectrum, if 2,3,7,8-TCDD is regarded solely as a
promoter, a threshold approach to quantitative assessment may be more
appropriate. And, finally, if 2,3J7,8-TCDD is regarded as an indirect
carcinogen, possibly acting by multiple mechanisms in the carcinogenic process,
some of which may display linear behavior at low doses, then a linearized model
42
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could be appropriate although the slope of the response will be uncertain. It
should be noted here that the 1985 estimate of the potency of 2,3,7,8-TCDD
(U.S. EPA, 1985) has always been characterized as a plausible upper bound to
the risk; the extent of this "overestimate" is unknown, but it may be higher
than previously thought,
An assessment of several potential approaches for estimating the
carcinogenic potency of 2,3,7,8-TCDD and related risk-specific doses is
summarized in the following section.
1. Selection of Models
a. NOEL/Uncertainty Factor (Threshold Approaches)
If 2,3,7,8-TCDD acts solely as a promoter, the traditional toxicological
approach based on a NOEL or a lowest-observed-effect-level (LOEL) may be an
appropriate risk estimation approach under certain circumstances. However,
while there is evidence that 2,3,7,8-TCDD acts as a promoter, the data suggest
that this chemical may also act through multiple mechanisms, direct or
indirect. If some of these component mechanisms were linear at low doses, then
the composite dose-response curve would be expected to be linear at low doses,
and a threshold approach would not be appropriate. Although 2,3,7,8-TCDD is a
strong promoter, biological data and statistical limitations to the power of
bioassays suggest that a threshold cannot be adequately demonstrated and may
not exist. This assessment is tempered somewhat by observations both in vivo
and in vitro suggesting "anticarcinogen" effects at low doses which could
offset the other effects of 2,3,7,8-TCDD and possibly produce a threshold. An
anticarcinogenic effect working in conjunction with a carcinogenic effect in
the same tissue might result in a net response of zero over background and
might be described as a threshold effect, in summary.
43
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Until such time that some of th|ese critical Issues are resolved, the
Workgroup concluded that it would not be prudent to adopt a threshold approach
i
for estimating human cancer risk for 2,3,7,8-TCDD.
i
I
b. Siel ken Approach
Sielken has produced a risk assessment with several provocative aspects;
e.g., generating a maximum likelihood estimate (MLE, as opposed to a UCL),
i
discarding the data obtained from thje highest dose, using time-to-tumor
information, and drawing attention to the lower-than-background response
I ' i. • . . .
observed at the lowest dose in both ^he Kociba et al. (1978) and the NTP (1982)
studies. While this type of exploratory analysis is interesting and raises
points that should be examined more Closely, the Workgroup is reluctant to
adopt this approach at this time for several reasons. For example, the use of
the MLE has traditionally been avoided due to its instability in the face of
relatively small changes in the data| (see Appendix A) and, for this reason, the
UCL has generally been preferred. Cfump (1987) provided a critical review of
i
the Sielken analysis and observed that even if "the true shape of the dose
response is a straight line connecting the background response and the response
at the mid-dose," the probability of an MLE of zero for the linear term in the
multistage model is about 1/3. He concludes that while the data are consistent
with Sielken's interpretation of a higher RsD, they are also consistent with
much lower RsDs, as displayed in the confidence limits.
c. M-K-V model
j
This model, which is based on the use of biologically-based models (see
Section II.D.), represents significant progress in carcinogen risk assessment.
Its use for estimating the carcinogenic potency of chemicals in general, and of
2,3,7,8-TCDD in particular, is viewed by this group as premature for several
44
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reasons. There are concerns about its biological bases and assumptions, the
statistical derivation and application of the model, the resulting large range
of "best" estimates (as opposed to upper-bound estimates) for which there are
no adequate criteria for selecting any estimate within the range, and the lack
of use and "experience" with other chemicals. Other concerns include
uncertainties and sensitivities about application of the model. Although this
model has many interesting features, and the Agency will continue to encourage
its development, the reasons described above and in Appendix A preclude
recommendation of its use to select an RsD at this time.
d. IMS Model
Although there are many uncertainties about the mode of action of
2,3,7,8-TCDD, data from bioassays and information on possible indirect
mechanisms provide some basis for assuming that this chemical might initiate
the carcinogenic response. If 2,3,7,8-TCDD is both an initiator and a promoter
of carcinogenesis, or if it functions in some other way as a complete
carcinogen, a model that is linear at low doses would be appropriate for
estimating an upper bound on the carcinogenic potency. If the chemical acts by
a combination of direct and indirect mechanisms, some with the characteristic
of low-dose linearity, the composite dose-response might be expected to exhibit
low-dose linearity as well, but perhaps with a lower slope than previously
estimated.
The EPA has elected to use the IMS model for cancer risk assessments, in
general, because it has a plausible biological basis, incorporates the
assumptions of nonthreshold linearity at low doses, provides a plausible upper
bound to the risk, and can be used as a "yardstick" to compare the "potency" of
one chemical with another. The EPA guidelines for the assessment of risk from
45
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carcinogens point out that other modjels can, and should, be used if biological
I
considerations dictate. In the casei of 2,3,7,8-TCDD, some other models have
been proposed (Longstreth and Hushon, 1983), but the biological basis that
would justify their use is not any stronger than with the IMS model.
Consequently, the Workgroup has concluded that the use of these alternative
models provides no additional advantages to the risk assessment of
2,3,7,8-TCDD.
In summary, the Workgroup concludes that none of the available models
adequately describe the carcinogenic; behavior of 2,3,7,8-TCDD at low doses.
Specifically:
• While there is evidence that! 2,3,7,8-TCDD acts as a promoter, there is
little evidence on which to conclude that a threshold exists. Without
a more scientific basis for such a radical departure from EPA's
traditional approach to the risk assessment for carcinogens, the
Workgroup is unwilling to adbpt a threshold approach for 2,3,7,8-TCDD.
• The innovative approaches of;Sielken and Moolgavkar, Venson, and
Knudson are interesting, butj untested. Therefore, the Workgroup
concludes that it would be imprudent to use them at this time for
2,3,7,8-TCDD.
• The available evidence suggests that reliance on the IMS model, as
traditionally used by EPA, may be less appropriate for 2,3,7,8-TCDD
than for many other chemicalis, and that the Agency's 1985 assessment
based on the IMS model may overestimate the upper bound on the risk by
some unknown amount. Howeveir, a rationale for a possible linear
behavior at low doses has been developed in this report, and the IMS
model provides a useful and familiar context which is widely used in
the Federal government when discussing risk estimates. Therefore, the
Workgroup discusses its recommendation using the IMS model as a
construct, that is, the plausible upper-bound estimate of risk and the
risk-specific dose.
2. Selection of the RsD Range
a. Base Analysis
Application of the IMS model to estimate the carcinogenic potency of
2,3,7,8-TCDD results in a range of RsDs (10~6) from 0.001 pg/kg/day to 1.2
i
P9/k§/day depending on the data used as the basis for the analysis and the
I 46
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assumptions made in the assessment. This range would be even greater if the
RfDs established in Canada, Europe, and some states in the United States were
included. Moreover, it is not clear where in the range potency estimates,
based on composite direct and indirect mechanisms, would fall. In fact, it is
possible that such estimates may fall outside of the IMS model range described
previously. The discussion in the subsequent sections attempts to narrow this
wide range and provide a bound to the RsD selected.
, b. Modification of Base Analysis
(1) Incorporation of Alternative Inferences
Incorporation of the factors used by the FDA for scaling from animals to
humans, along with use of only the tumors of the lung, hard palate, and nasal
turbinates observed in the Kociba et al. (1978) study results in an RsD (10~6)
of 1.2 pg/kg/day, the highest RsD we have developed based on the IMS model.
This analysis excludes the tumors observed in the liver because of the strong
promoter activity shown by 2,3,7,8-TCDD in this organ. However, the Workgroup
gives less weight to this point on the range because (1) it excludes 90% of the
response observed in the bioassay, and (2) there is controversy whether the
tumors of the lung, hard palate, and nasal turbinates should be considered at
all (Kociba, 1984), since they may be a localized carcinogenic response to
inhaled microscopic food particles containing 2,3,7,8-TCDD. This suggestion
gains support from the fact that these tumors were not observed in experiments
using other routes of exposure, i.e., dermal, gavage, and intraperitoneal,
while liver tumors were common to all four routes. The Workgroup concludes
that there is not, as yet, sufficient evidence to accept or reject this
hypothesis.
47
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(2) Incorporation of Relative Half-Lives
The lowest RsD in the range is based on differences in the relative
half-lives of 2,3,7,8-TCDD in rats and humans. The Workgroup gives less weight
I
to this point because of numerous uncertainties about the pharmacokinetics of
2,3,7,8-TCDD, particularly the absence of information on species differences
and rates of incorporation and absorption of 2,3,7,8-TCDD in different tissues
and species. Similarly, the rate ofjrelease of this chemical is likely to be
different from tissue to tissue and species to species. Furthermore, it is not
I
clear how 2,3,7,8-TCDD will behave in different species, particularly humans,
i
under conditions of chronic exposure^ incorporation, and release.
(3) Alternative Dose-Response Curves
The suggestion by Hoel (1987) that AHH induction may be useful for
defining the shape of the dose-response curve is interesting, but it is not
clear how the results of such a calculation would be incorporated into a final
f
risk assessment. Initially, use of AHH data to define the curve is appealing
because many data points are available, AHH induction is closely related to
many of the toxic effects observed in 2,3,7,8-TCDD, and AHH induction appears
i
to be linear at low doses. However,;there is no demonstrated correlation
between AHH induction and tumorigenicity, and it is not apparent that the shape
of the AHH dose-response curve reflects the shape of the dose-response curve
for 2,3,7,8-TCDD's carcinogenic effects at low doses.
(4) Multiple Mechanisms
Qualitatively, the concept of multiple mechanisms acting in concert, or
even opposition, is given added weight because it allows inclusion of much of
i
the body of scientific data on 2,3,7|,8-TCDD other than the standard bioassay
i
data. As discussed previously, it ils not unreasonable to assume that a
48
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composite dose-response curve may, under certain conditions, exhibit linearity
at low doses. However, quantitative application of this approach is limited
because we do not have information on the slopes of the component curves that
would be necessary to define the overall slope of the linear portion of the
curve. In general, if such multiple mechanisms could be incorporated into the
LMS methodology, the slope of the line would be lower (extent not quantifiable)
rather than higher than that derived as a plausible upper bound for direct-
acting, complete carcinogens.
(5) Different Base Assumptions
Using the same basic LMS model approach, EPA, CDC, and FDA have estimated
carcinogenic RsDs (10~5) for 2,3,7,8-TCDD of 0.006, 0.03, and 0.06 pg/kg/day,
respectively. Several different policy-based assumptions account for the
differences. For example, EPA scales from animals to humans on the basis of
relative body surface area, while FDA uses relative body weights. In addition,
both EPA and FDA used the 2,3,7,8-TCDD concentration in rat food as a surrogate
for dose, while CDC used the concentration of the chemical in rat liver as a
measure of dose. There is no obvious scientific basis for excluding any of
these values from the range.
(6) Choice of RsD
As noted previously, a majority of the Workgroup has concluded that the
1985 EPA estimate of the upper-bound potency (RsD) generated from the
application of the UCL LMS model to the Kociba et al. (1978) data is likely to
have led to an overestimate of risk (or underestimate of the risk-specific
dose). The weight of evidence indicates that a more appropriate upper-bound
estimate would be obtained by a reduction of the potency by some unquantifiable
amount. Therefore, in recommending a new RsD and indirectly suggesting a
49
-------
change in potency, the Workgroup was confronted with the question, "How great a
reduction in slope (or increase in RsD) is appropriate?"
The Workgroup concluded that tjiere is currently no definitive scientific
i
basis for an answer to that questioji. Given, however, that the question must
be answered for Agency purposes, this answer should be grounded in rational,
prudent science policy.
While some argument could be mpunted that a threshold for the
carcinogenicity of 2,3,7,8-TCDD may; in fact exist, the evidence for such a
contention is not compelling. Therefore, prudence dictates against adopting a
simple threshold approach to setting an RsD. Hence, the Workgroup recommends
against adopting a policy position similar to that of the European countries at
i
this time.
The Workgroup encourages the type of analysis generated by Si el ken and the
application of the newer M-K-V modeil to 2,3,7,8-TCDD. These fresh looks at the
problem stimulate discussion and challenge old ways of thinking. However, as
noted previously, neither of these approaches is sufficiently developed nor has
received sufficient standing in the| critical scientific community that the
Workgroup feels comfortable in recommending a potency (RsD) on this basis at
this time. ;
The Workgroup is not convinced^ that the UCL IMS model is an appropriate
model for estimating upper-bound cancer'risks associated with exposure to
2,3,7,8-TCDD. However, without a b[etter alternative the Workgroup has used the
UCL IMS model as a construct within: which to discuss the carcinogenic potency
I
or RsD of 2,3,7,8-TCDD. The scientific evidence is consistent with, and would
support, a recommended science policy position that the RsD (10~6) for
i A
2,3,7,8-TCDD be 0.1 pg/kg/day, which is associated with a qj, in the UCL IMS
50
-------
construct of 1 x W^/mg/kg/day, for the following reasons:
• the scientific data indicate that the Agency's current upper bound for
2,3,7,8-TCDD may be an overestimate;
• the scientific data do not permit an estimate of the extent of the
overestimate;
• al] of the UCL IMS RsD estimates generated by the Federal agencies are
arguably of equal scientific merit at this time;
• for strictly policy purposes, there is great benefit in Federal
agencies' adopting consistent positions in the absence of compelling
scientific information; and
• an order of magnitude estimate of the RsD (potency), as opposed to some
more precise estimate of the risk-specific dose, helps to convey the
notion that the numerical expression is only a rough estimate (the
science permits no greater accuracy).
The available scientific data can give us no clearer guidance. Some of
the research now underway holds the promise of clarifying the issue but not
resolving it totally. While a series of considerations, based on science, may
be brought to bear on the selection of the RsD, the Workgroup does not advance
them as compelling scientific support for a recommended RsD (10"^), which is,
in this case, simply a rational and prudent science policy position. The
Workgroup further recommends that the 2,3,7,8-TCDD risk-specific dose issue be
examined again regularly as new information becomes available. It is felt that
the rate of research on this chemical is so great and the fundamental questions
relating to multistage carcinogenicity and risk are so important to Agency
decision-makers that a regular re-evaluation of the science and/or science
policy underlying the selection of an RsD is appropriate.
51
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57
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