Tuesday
April 23, 1996
Li J
Part II
Environmental
Protection Agency
Proposed Guidelines for Carcinogen Risk
Assessment; Notice
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17960
Federal Register / Vol. 61, No. 79 / Tuesday, April 23, 1996 / Notices
ENVIRONMENTAL PROTECTION
AGENCY
[FRL-5460-3]
Proposed Guidelines for Carcinogen
Risk Assessment
AGENCY: Environmental Protection
Agency (EPA).
ACTION: Notice of Availability and
Opportunity to Comment on Proposed
Guidelines for Carcinogen Risk
Assessment.
SUMMARY: The U.S. Environmental
Protection Agency (EPA) is today
publishing a document entitled
Proposed Guidelines for Carcinogen
Risk Assessment (hereafter "Proposed
Guidelines"). These Proposed
Guidelines were developed as part of an
interoffice guidelines development
program by a Technical Panel of the
Risk Assessment Forum within EPA's
Office of Research and Development.
These Proposed Guidelines are a
revision of EPA's 1986 Guidelines for
Carcinogen Risk Assessment (hereafter
"1986 cancer guidelines") published on
September 24,1986 (51 FR 33992).
When final, these guidelines will
replace the 1986 guidelines.
In a future Federal Register notice,
the Agency intends to publish for
comment how it will implement the
Proposed Guidelines once they are
finalized. The plans will propose and
seek comment on how die Guidelines
will be used for Agency carcinogen risk
assessment and, in particular, will
address the impact of the Guidelines on
the Agency's existing assessments, and
any mechanisms for handling
reassessments under finalized
Guidelines.
DATES: The Proposed Guidelines are
being made available for a 120-day
public review and comment period.
Comments must be in writing and must
be postmarked by August 21,1996. See
ADDRESSES section for guidance on
submitting comments.
ADDRESSES: The Proposed Guidelines
will be made available in the following
ways:
(1) The electronic version will be
accessible on EPA's Office of Research
and Development home page on the
Internet at http://www.epa.gov/ORD
(2) 3W high-density computer
diskettes in Wordperfect 5.1 format will
be available from ORD Publications,
Technology Transfer and Support
Division, National Risk Management
Research Laboratory, Cincinnati, OH;
telephone: 513-569-7562; fax: 513-
569-7566. Please provide the EPA No.
(EPA/600/P-92/003Ca) when ordering.
(3) This notice contains the full draft
document. In addition, copies of the
draft will be available for inspection at
EPA headquarters and regional libraries,
through the U.S. Government
Depository Library program, and for
purchase from the National Technical
Information Service (NTIS), Springfield,
VA; telephone: 703-487-4650, fax: 703-
321-8547. Please provide the NTIS PB
No. (PB96-157599) ($35.00) when
ordering.
SUBMITTING COMMENTS: Comments on the
Proposed Guidelines may be mailed or
delivered to the Technical Information
Staff (8623), NCEA-WA/OSG, U.S.
Environmental Protection Agency, 401
M Street, S.W., Washington, DC 20460.
Comments should be in writing and
must be postmarked by the date
indicated. Please submit one unbound
original with pages numbered
consecutively, and three copies. For
attachments, provide an index, number
pages consecutively with the comment,
and submit an unbound original and
three copies.
Please note that all technical
comments received in response to this
notice will be placed in a public record.
For that reason, commenters should not
submit personal information (such as
medical data or home address),
Confidential Business Information, or
information protected by copyright. Due
to limited resources, acknowledgments
will not be sent.
FOR FURTHER INFORMATION CONTACT:
Technical Information Staff, Operations
and Support Group, National Center for
Environmental Assessment—
Washington Office, telephone: 202-260-
7345, Email inquiries may be sent to
cancer-guidelines@epamail.epa.gov. .
SUPPLEMENTARY INFORMATION: In 1983,
the National Academy of Sciences
(NAS)/National Research Council (NRG)
published its report entitled Risk
Assessment in the Federal Government:
Managing the Process (NRC, 1983). In
that report, the NRC recommended that
Federal regulatory agencies establish
"inference guidelines" to ensure
consistency and technical quality in risk
assessments and to ensure that the risk
assessment process was maintained as a
scientific effort separate from risk
management. The 1986 cancer
guidelines were issued on September
24,1986 (51 FR 33992). The Proposed
Guidelines published today continue
the guidelines development process.
These guidelines set forth principles
and procedures to guide EPA scientists
in the conduct of Agency cancer risk
assessments and to inform Agency
decisionmakers and the public about
these procedures.
Both the 1986 guidelines and the
current proposal contain inference
guidance in the form of default
inferences to bridge gaps in knowledge
and data. Research conducted in the
past decade has elucidated much about
the nature of carcinogenic processes and
continues to provide new information.
The intent of this proposal is to take
account of knowledge available now
and to provide flexibility for the future
in assessing data and employing default
inferences, recognizing that the
guidelines cannot always anticipate
future research findings. Because
methods and knowledge are expected to
change more rapidly than guidelines
can practicably be revised, the Agency
will update specific assessment
procedures with peer-reviewed
supplementary, technical documents as
needed. Further revision of the
guidelines themselves will take place
when extensive changes are necessary.
Since 1986, the EPA has sponsored
several workshops about revising the
cancer guidelines (U.S. EPA, 1989b,
1989c, 1994a). The Society for Risk
Analysis conducted a workshop on the
subject in connection with its 1992
annual meeting (Anderson et al., 1993).
Participants in the most recent
workshop in 1994 reviewed an earlier
version of the guidelines proposed here
and made numerous recommendations
about individual issues as well as broad
recommendations about explanations
and perspectives that should be added.
Most recently, the Committee on the
Environment and Natural Resources of
the Office of Science and Technology
Policy reviewed the guidelines at a
meeting held on August 15,1995. The
EPA appreciates the efforts of all
participants in the process and has tried
to address their recommendations in
this proposal.
In addition, the recommendations of
the NRC (1994) in Science and
Judgment in Risk Assessment have been
addressed. Responses to these
recommendations are given generally in
Appendix B as well as being embodied
in the Proposed Guidelines. Responses
that explain the major default
assumptions adopted under these
guidelines and the policy for using and
departing from these default
assumptions appear in Section 1.3.
The Science Advisory Board also will
review these Proposed Guidelines at a
meeting to be announced in a future
Federal Register notice. Following these
reviews Agency staff will prepare
summaries of the public and SAB
comments. Appropriate comments will
be incorporated, and the revised
Guidelines will be submitted to the Risk
Assessment Forum for review. The
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17961
Agency will consider comments from
tho public, the SAB, and the Risk
Assessment Forum in its
recommendations to the EPA
Administrator.
Major Changes From the 1986
Guidelines
Characterizations
Increased emphasis on providing
characterization discussions for the
hazard, dose response, and exposure
sections is part of the proposal. These
discussions will summarize the
assessments to explain the extent and
weight of evidence, major points of
interpretation and rationale, and
strengths and weaknesses of the
evidence and the analysis, and to
discuss alternative conclusions and
uncertainties that deserve serious
consideration (U.S. EPA, 1995). They
serve as starting materials for the risk
characterization process which
completes the risk assessment.
We/g/i/ng Evidence of Hazard
A major change is in the way hazard
evidence is weighed in reaching
conclusions about the human
carcinogenic potential of agents. In the
1986 cancer guidelines, tumor findings
in animals or humans were the
dominant components of decisions.
Other information about an agent's
properties, its structure-activity
relationships to other carcinogenic
agents, and its activities in studies of
carcinogenic processes was often
limited and played only a modulating
role as compared with tumor findings.
In this proposal, decisions come from
considering all of the evidence. This
change recognizes the growing
sophistication of research methods,
particularly in their ability to reveal the
modes of action of carcinogenic agents
at cellular and subcellular levels as well
as toxicokinetic and metabolic
processes. The effect of the change on
tho assessment of individual agents will
depend greatly on the availability of
now kinds of data on them in keeping
•with tho state of the art. If these new
kinds of data are not forthcoming from
public and private research on agents,
assessments under these guidelines will
not differ significantly from assessments
under former guidelines.
Weighing of the evidence includes
addressing the likelihood of human
carcinogenic effects of the agent and the
conditions under which such effects
may be expressed, as these are revealed
in the lexicological and other
biologically important features of the
agent. (Consideration of actual human
exposure and risk implications are done
separately; they are not parts of the
hazard characterization). In this respect,
the guidelines incorporate
recommendations of the NRG (1994). In
that report, the NRG recommends
expansion of the former concept of
hazard identification, which rests on
simply a finding of carcinogenic
potential, to a concept of
characterization that includes
dimensions of the expression of this
potential. For example, an agent might
be observed to be carcinogenic via
inhalation exposure and not via oral
exposure, or its carcinogenic activity
might be secondary to another toxic
effect. In addition, the consideration of
evidence includes the mode(s) of action
of the agent apparent from the available
data as a basis for approaching dose
response assessment.
Classification Descriptors
To express the weight of evidence for
carcinogenic hazard potential, the 1986
cancer guidelines provided summary
rankings for human and animal cancer
studies. These summary rankings were
integrated to place the overall evidence
in classification groups A through E,
Group A being associated with the
greatest probability of human
carcinogenicity and Group E with
evidence of noncarcinogenicity in
humans. Data other than tumor findings
played a modifying role after initial
placement of an agent into a group.
These Proposed Guidelines take a
different approach, consistent with the
change in the basic approach to
weighing evidence. No interim
classification of tumor findings followed
by modifications with other data takes
place. Instead, the conclusion reflects
the weighing of evidence in one step.
Moreover, standard descriptors of
conclusions are employed rather than
letter designations, and these are
incorporated into a brief narrative
description of their informational basis.
The narrative with descriptors replaces
the previous letter designation. The
descriptors are in three categories:
"known/likely," "cannot be
determined," or "not likely." For
instance, using a descriptor in context,
a narrative could say that an agent is
likely to be carcinogenic by inhalation
exposure and not likely to be
carcinogenic by oral exposure. The
narrative explains the kinds of evidence
available and how they fit together in
drawing conclusions, and points out
significant issues/strengths/limitations
of the data and conclusions.
Subdescriptors are used to further refine
the conclusion. The narrative also
summarizes the mode of action
information underlying a recommended
approach to dose response assessment.
In considering revision of the former
classification method, the Agency has
examined other possibilities that would
retain the use of letter and number
designation of weights of evidence. The
use of standard descriptors within a
narrative presentation is proposed for
three primary reasons. First, the
proposed method permits inclusion of
explanations of data and of their
strengths and limitations. This is more
consistent with current policy emphasis
on risk characterization. Second, it
would take a large set of individual
number or letter codes to cover
differences in the nature of contributing
information (animal, human, other),
route of exposure, mode of action, and
relative overall weight. When such a set
becomes large—10 to 30 codes—it is too
large to be a good communication
device, because people cannot
remember the definitions of the codes so
they have to be explained in narrative.
Third, it is impossible to predefine the
course of cancer research and the kinds
of data that may become available. A
flexible system is needed to
accommodate change in the underlying
data and inferences, and a system of
codes might become out of date, as has
the one in the 1986 cancer guidelines.
Dose Response Assessment
The approach to dose response
assessment calls for analysis that
follows the conclusions reached in the
hazard assessment as to potential
mode(s) of action. The assessment
begins by analyzing the empirical data
in the range of observation. When
animal studies are the basis of the
analysis, the estimation of a human
equivalent dose utilizes toxicokinetic
data, if appropriate and adequate data
are available. Otherwise, default
procedures are applied. For oral dose,
the default is to scale daily applied
doses experienced for a lifetime in
proportion to body weight raised to the
0.75 power. For inhalation dose, the
default methodology estimates
respiratory deposition of particles and
gases and estimates internal doses of
gases with different absorption
characteristics. These two defaults are a
change from the 1986 cancer guidelines
which provided a single scaling factor of
body weight raised to the 0.66 power.
Another change from the 1986
guidelines is that response data on
effects of the agent on carcinogenic
processes are analyzed (nontumor data)
in addition to data on tumor incidence.
If appropriate, the analyses of data on
tumor incidence and on precursor
effects may be combined, using
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Federal Register / Vol. 61, No. 79 / Tuesday, April 23, 1996 / Notices
precursor data to extend the dose
response curve below the tumor data.
Even if combining data is not
appropriate, study of the dose response
for effects believed to be part of the
carcinogenic influence of the agent may
assist in thinking about the relationship
of exposure and response in the range
of observation and at exposure levels
below the range of observation.
Whenever data are sufficient, a
biologically based or case-specific dose
response model is developed to relate
dose and response data in the range of
empirical observation. Otherwise, as a
standard, default procedure, a model is
used to curve-fit the data. The lower
95% confidence limit on a dose
associated with an estimated 10%
increased tumor or relevant nontumor
response (LEDio) is identified. This
generally serves as the point of
departure for extrapolating the
relationship to environmental exposure
levels of interest when the latter are
outside the range of observed data. The
environmental exposures of interest
may be measured ones or levels of risk
management interest in considering
potential exposure control options.
Other points of departure may be more
appropriate for certain data sets; as
described in the guidance, these may be
used instead of the LEDio. Additionally,
the LEDio is available for comparison
with parallel analyses of other
carcinogenic agents or of noncancer
effects of agents and for gauging and
explaining the magnitude of subsequent
extrapolation to low-dose levels. The
LEDio, rather than the EDio (the estimate
of a 10% increased response), is the
proposed standard point of departure
for two reasons. One is to permit easier
comparison with the benchmark dose
procedure for noncancer health
assessment—^-also based on the lower
limit on dose. Another is that the lower
limit, as opposed to the central estimate,
accounts for uncertainty in the
experimental data. The issue of using a
lower limit or central estimate was
discussed at a workshop held on the
benchmark procedure for noncancer
assessment (Barnes et al., 1995) and at
a workshop on a previous version of this
proposal (U.S. EPA, 1994b). The latter
workshop recommended a central
estimate; the benchmark workshop
recommended a lower limit.
The second step of dose response
assessment is extrapolation to lower
dose levels, if needed. This is based on
a biologically based or case-specific
model if supportable by substantial
data. Otherwise, default approaches are
applied that accord with the view of
mode(s) of action of the agent. These
include approaches that assume
linearity or nonlinearity of the dose
response relationship or both. The
default approach for linearity is to
extend a straight line to zero dose, zero
response. The default approach for
nonlinearity is to use a margin of
exposure analysis rather than estimating
the probability of effects at low doses.
A margin of exposure analysis explains
the biological considerations for
comparing the observed data with the
environmental exposure levels of
interest and helps in deciding on an
acceptable level of exposure in
accordance with applicable
management factors.
The use of straight line extrapolation
for a linear default is a change from the
1986 guidelines which used the
"linearized multistage" (LMS)
procedure. This change is made because
the former modeling procedure gave an
appearance of specific knowledge and
sophistication unwarranted for a
default. The proposed approach is also
more like that employed by the Food
and Drug Administration (U.S. FDA,
1987). The numerical results of the
straight line and LMS procedures are
not significantly different (Krewski et
al., 1984). The use of a margin of
exposure approach is included as a new
default procedure to accommodate cases
in which there is sufficient evidence of
a nonlinear dose response, but not
enough evidence to construct a
mathematical model for the
relationship. (The Agency will continue
to seek a modeling method to apply in
these cases. If a modeling approach is
developed, it will be subject to peer
review and public notice in the context
of a supplementary document for these
guidelines.)
The public is invited to provide
comments to be considered in EPA
decisions about the content of the final
guidelines. After the public comment
period, the EPA Science Advisory Board
will be asked to review and provide
advice on the guidelines and issues
raised in comments. EPA asks those
who respond to this notice to include
their views on the following:
(1) The proposed guidance for
characterization of hazard, including the
weight of evidence descriptors and
weight of evidence narrative which are
major features of the proposal. There are
three categories of descriptors: "known/
likely," "cannot be determined," and
"not likely" which are further refined
by subdescriptors. It is felt that these
three descriptors will satisfactorily
delineate the types of evidence bearing
on carcinogenicity as they are used with
subdescriptors in the context of a
narrative of data and rationale.
However, an issue that has been
discussed by external peer reviewers
and by EPA staff is whether the
descriptor-subdescriptor called "cannot
be determined—suggestive evidence"
should become a separate, fourth
category called "suggestive." The EPA
may choose this course in the final
guidelines and requests comment. In
considering this issue, commenters may
wish to refer not only to Sections 2.6.2.
and 2.7.2. which cover the descriptors
and narrative, but also to case study
example #6 in Section 2.6.3. and
example narrative #2 in Appendix A of
the proposal. EPA asks commenters on
this question to address the rationale
(science as well as policy) for leaving
the categories of descriptors as proposed
or making the fourth category. How
might the coverage of a "suggestive"
category be defined in order to be most
useful?
(2) The use of mode of action
information in hazard characterization
and to guide dose response assessment
is a central part of the proposed
approach to bringing new research on
carcinogenic processes to bear in
assessments of environmental agents'
(Sections 1.3.2., 2.3.2., 2.5., 3.1.). The
appropriate use of this information now
and in the future is important. EPA
requests comment on the treatment of
such information in the proposal,
including reliance on peer review as a
part of the judgmental process on its •
application.
(3) Uses of nontumor data in the dose
response assessment and the
methodological and science policy
issues posed are new to these guidelines
(Sections 1.3.2., 3.1.2.). EPA requests
comment on both issues.
(4) Dose response assessment is
proposed to be considered in two
parts—range of observed data and range
of extrapolation (Section 3.1.). The
lower 95% confidence limit on a dose
associated with a 10% response (tumor
or nontumor response) is proposed as a
default point of departure, marking the
beginning of extrapolation. This is a
parallel to the benchmark procedure for
evaluating dose-response of noncancer
health endpoints (Barnes et al., 1995).
An alternative is to use the central
estimate of a 10% response. Another
alternative is to use a 1%, instead of a
10%, response when the observed data
are tumor incidence data. Does the
generally larger sample size of tumor
effect studies support using a 1%
response as compared with using 10%
for smaller studies? Are there other
approaches for the point of departure
that might be considered?
(5) Discussions of default assumptions
and other responses to the 1994 NRC
report Science and Judgment in Risk .
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Assessment appear in Section 1.3.1. and
Appendix B of the proposal,
respectively. Comments are requested
on responses to the NRC
recommendations and how the
guidelines as a whole address them.
Dated: April 10,1996.
Carol M. Browner,
Administrator.
Contents
List of Figures
1. Introduction
1.1. Purpose and Scope of the Guidelines
1,2. Organization and Application of the
Guidelines
1.2.1. Organization
1.2.2. Application
1.3. Uso of Default Assumptions
1.3.1. Default Assumptions
1.3.2. Major Defaults
1.3.2.1. Is the Presence or Absence of
Effects Observed in a Human Population
Predictive of Effects in Another Exposed
Human Population?
1.3.2.2. Is the Presence or Absence of
Effects Observed in an Animal
Population Predictive of Effects in
Exposed Humans?
1.3,2.3. How Do Metabolic Pathways Relate
Across Species?
1.3.2.4. How Do Toxicokinetic Processes
Relate Across Species?
1.3.2.5. What Is the Correlation of the
Observed Dose Response Relationship to
the Relationship at Lower Doses?
1.4. Characterizations
2. Hazard Assessment
2.1. Overview of Hazard Assessment and
Characterization
2.1.1. Analyses of Data
2.1.2. Cross-Cutting Topics for Data
Integration
2.1.2.1. Conditions of Expression
2.1.2.2. Mode of Action
2.1.3. Presentation of Results
2.2. Analysis of Tumor Data
2.2.1. Human Data
2.2.1.1. Types of Studies
2.2.1.2. Criteria for Assessing Adequacy of
Epidemiologic Studies
2.2.1.3. Criteria for Causality
2.2.1.4. Assessment of Evidence of
Qirclnogcnicity from Human Data
2.2.2. Animal Data
2.2.2.1. Long-Term Carcinogenicity Studies
2.2.2.2. Other Studies
2.2.3. Structural Analogue Data
2.3. Analysis of Other Key Data
2.3.1. Physlcochemical Properties
2.3.2. Structure-Activity Relationships
2.3.3. Comparative Metabolism and
Toxteokinotics
2.3.4. Toxicological and Clinical Findings
2.3.5. Mode of Action-Related Endpoints
and Short-Term Tests
2.3.5.1. Direct DMA Effects
2.3.5.2. Secondary DNA Effects
2.3.5.3. Nonmutagcnic and Other Effects
2.3.5.4. Criteria for Judging Mode of Action
2.4. Biomarkcr Information
2.5. Mode of Action—Implications for
Hazard Characterization and Dose
Response
2.6. Weight of Evidence Evaluation for
Potential Human Carcinogenicity
2.6.1. Weight of Evidence Analysis
2.6.2. Descriptors for Classifying Weight of
Evidence
2.6.3. Case Study Examples
2.7. Presentation of Results
2.7.1. Technical Hazard Characterization
2.7.2. Weight of Evidence Narrative
3. Dose Response Assessment
3.1. Dose Response Relationship
3.1.1. Analysis in the Range of Observation
3.1.2. Analysis in the Range of
Extrapolation
3.1.3. Use of Toxicity Equivalence Factors
and Relative Potency Estimates
3.2. Response Data
3.3. Dose Data
3.3.1. Interspecies Adjustment of Dose
3.3.2. Toxicokinetic Analyses
3.3.3. Route-to-Route Extrapolation
3.3.4. Dose Averaging
3.4. Discussion of Uncertainties
3.5. Technical Dose Response
Characterization
4. Technical Exposure Characterization
5. Risk Characterization
5.1. Purpose
5.2. Application
5.3. Presentation of Risk Characterization
Summary
5.4. Content of Risk Characterization
Summary
Appendix A
Appendix B
Appendix C
References
List of Figures
Figure 1—1. Decisions on Dose Response
Assessment Approaches for the Range of
Extrapolation
Figure 1-2. Risk Characterization
Figure 2—1. Factors for Weighing Human
Evidence
Figure 2-2. Factors for Weighing Animal
Evidence
Figure 2-3. Factors for Weighing Other Key
Evidence
Figure 2-4. Factors for Weighing Totality of
Evidence
Figure 3-1. Graphical Presentation of Data
and Extrapolations
1. Introduction
1.1. Purpose and Scope of the
Guidelines
These guidelines revise and replace
United States Environmental Protection
Agency (EPA) Guidelines for Carcinogen
Risk Assessment published in 51 FR
33992, September 24,1986. The
guidelines provide EPA staff and
decisionmakers with guidance and
perspectives to develop and use risk
assessments: They also provide basic
information to the public about the
Agency's risk assessment methods.
The guidelines encourage both
regularity in procedures to support
consistency in scientific components of
Agency decisionmaking and innovation
to remain up-to-date in scientific
thinking. In balancing these goals, the
Agency relies on input from the general
scientific community through
established scientific peer review
processes. The guidelines incorporate
basic principles and science policies
based on evaluation of the currently
available information. As more is
discovered about carcinogenesis, the
need will arise to make appropriate
changes in risk assessment guidance.
The Agency will revise these guidelines
when extensive changes are due. In the
interim, the Agency will issue special
reports, after appropriate peer review, to
supplement and update guidance on.
single topics, (e.g., U.S. EPA, 1991b)
1.2. Organization and Application of the
Guidelines
1.2.1. Organization
Publications of the Office of Science
and Technology Policy (OSTP, 1985)
and the National Research Council
(NRC, 1983,1994) provide information
and general principles about risk
assessment. Risk assessment uses
available scientific information on the
properties of an agent * and its effects in
biological systems to provide an
evaluation of the potential for harm as
a consequence of environmental
exposure to the agent. Risk assessment
is one of the scientific analyses available
for consideration, with'Other analyses,
in decisionmaking on environmental
protection. The 1983 and 1994 NRC
documents organize risk assessment
information into four areas: hazard
identification, dose response
assessment, exposure assessment, and
risk characterization. This structure
appears in these guidelines, which
additionally emphasize characterization
of evidence and conclusions in each
part of the assessment. In particular, the
guidelines adopt the approach of the
NRC's 1994 report in adding a
dimension of characterization to the
hazard identification step. Added to the
identification of hazard is an evaluation
of the conditions under which its
expression is anticipated. The risk
assessment questions addressed in these
guidelines are:
• For hazard—Can the agent present
a carcinogenic hazard to humans, and if
so, under what circumstances?
• For dose response—At what levels
of exposure might effects occur?
• For exposure—What are the
conditions of human exposure?
• For risk—What is the character of
the risk? How well do data support
conclusions about the nature and extent
of the risk?
1 The term "agent" refers generally to any
chemical substance, mixture, or physical or
biological entity being assessed, unless otherwise
noted.
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1.2.2, Application
The guidelines apply within the
framework of policies provided by
applicable EPA statutes and do not alter
such policies. The guidelines cover
assessment of available data. They do
not imply that one kind of data or
another is prerequisite for regulatory
action concerning any agent. Risk
management applies directives of
regulatory legislation, which may
require consideration of potential risk,
or solely hazard or exposure potential,
along with social, economic, technical,
and other factors in decisionmaking.
Risk assessments support decisions, but
to maintain their integrity as
decisionmaking tools, they are not
influenced by consideration of the
social or economic consequences of
regulatory action.
Not every EPA assessment has the
same scope or depth. Agency staff often
conduct screening-level assessments for
priority-setting or separate assessments
of hazard or exposure for ranking
purposes or to decide whether to invest
resources in collecting data for a full
assessment. Moreover, a given
assessment of hazard and dose response
may be used with more than one
exposure assessment that may be
conducted separately and at different
times as the need arises in studying
environmental problems in various
media. The guidelines apply to these
various situations in appropriate detail
given the scope and depth of the
particular assessment. For example, a
screening assessment may be based
almost entirely on structure-activity
relationships and default assumptions.
As more data become available,
assessments can replace or modify
default assumptions accordingly. These
guidelines do not require that all of the
kinds of data covered here be available
for either assessment or decisionmaking.
The level of detail of an assessment is
a matter of Agency management policy
regarding the applicable decisionmaking
framework.
1.3. Use of Default Assumptions
The National Research Council, in its
1983 report on the science of risk
assessment (NRG, 1983), recognized that
default assumptions are necessarily
made in risk assessments where gaps
exist in general knowledge or in
available data for a particular agent.
These default assumptions are
inferences based on general scientific
knowledge of the phenomena in
question and are also matters of policy
concerning the appropriate way to
bridge uncertainties that concern
potential risk to human health (or, more
generally, to environmental systems)
from the agent under assessment.
EPA's 1986 guidelines for cancer risk
assessment (EPA, 1986) were developed
in response to the 1983 NRG report. The
guidelines contained a number of
default assumptions. They also
encouraged research and analysis that
would lead to new risk assessment
methods and data and anticipated that
these would replace defaults. The 1986
guidelines did not explicitly discuss
how to depart from defaults. In practice,
the agency's assessments routinely have
employed defaults and, until recently,
only occasionally departed from them.
In its 1994 report on risk assessment,
the NRG supported continued use of
default assumptions (NRG, 1994). The
NRG report thus validated a central
premise of the approach to risk
assessment that EPA had evolved in
preceding years—the making of science
policy inferences to bridge gaps in
knowledge—while at the same time
recommending that EPA develop more
systematic and transparent guidelines to
inform the public of the default
inferences EPA uses in practice. It
recommended that the EPA review and
update the 1986 guidelines in light of
evolving scientific information and
experience in practice in applying those
guidelines, and that the EPA explain the
science and policy considerations
underlying current views as to the
appropriate defaults and provide
general criteria to guide preparers and
reviewers of risks assessments in
deciding when to depart from a default.
Pursuant to this recommendation, the
following discussion presents ;
descriptions of the major defaults and
their rationales. In addition, it presents
general policy guidance on using and
departing from defaults in specific risk
assessments.
1.3.1. Default Assumptions
The 1994 NRG report contains several
recommendations regarding flexibility
and the use of default options:
• EPA should continue to regard the
use of default options as a reasonable
way to deal with uncertainty about
underlying mechanisms in selecting
methods and models for use in risk
assessment.
• EPA should explicitly identify each
use of a default option in risk
assessments.
• EPA should clearly state the
scientific and policy basis for each
default option.
• The Agency should consider
attempting to give greater formality to
its criteria for a departure from default
options in order to give greater guidance
to the public and to lessen the
possibility of ad hoc, undocumented
departures from default options that
would undercut the scientific credibility
of the Agency's risk assessments. At the
same time, the Agency should be aware
of the undesirability of having its
guidelines evolve into inflexible rules.
• EPA should continue to use the
Science Advisory Board and other
expert bodies. In particular, the Agency
should continue to make the greatest
possible use of peer review, workshops,
and other devices to ensure broad peer
and scientific participation to guarantee
that its risk assessment decisions will be
based on the best science available
through a process that allows full public
discussion and peer participation by the
scientific community.
In the 1983 report (p. 28), NAS
defined the use of "inference options"
(default options) as a means to bridge
inherent uncertainties in risk
assessment. These options exist when
the assessment encounters either
"missing or ambiguous information on a
particular substance" or "gaps in
current scientific theory." Since there is
no instance in which a set of data on an
agent or exposure is complete, all risk
assessments must use general
knowledge and policy guidance to
bridge data gaps. Animal toxicity data
are used, for example, to substitute for
human data because we do not test
human beings. The report described the
components of risk assessment in terms
of questions encountered during
analysis for which inferences must be
made. The report noted (p. 36) that
many components "* * * lack
definitive scientific answers, that the
degree of scientific consensus
concerning the best answer varies (some
are more controversial than others), and
that the inference options available for
each component differ in their degree of
conservatism. The choices encountered
in risk assessment rest, to various
degrees, on a mixture of scientific fact
and consensus, on informed scientific
judgment, and on policy determinations
(the appropriate degree of
conservatism)* * *." The report did
not note that the mix varies significantly
from case to case. For instance, a
question that arises in hazard
identification is how to use
experimental animal data when the
routes of exposure differ between
animals and humans. A spectrum of
inferences could be made, ranging from
the most conservative, or risk adverse
one that effects in animals from one
route may be seen in humans by another
route, to an intermediate, conditional
inference that such translation of effects
will be assumed if the agent is absorbed
by humans through the second route, to
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17965
a nonconservative view that no
inference is possible and the agent's
effects in animals must be tested by the
second route. The choice of an
inference, as the report observed, comes
from more than scientific thinking
alone. While the report focused mainly
on the idea of conservatism of public
health as a science policy rationale for
making the choice, it did not evaluate
other considerations. These include
such things as the matters of time and
resources and whether the analysis is
for an important decision required to he
made soon or is simply a screening or
ranking effort. For a screening analysis,
one might make several "worst case"
inferences to determine if, even under
those conditions, risk is low enough that
a problem can be eliminated from
further consideration. In the above
discussion concerning inferences about
routo-to-route extrapolation, one might
uso the most conservative one for
screening.
Those revised guidelines retain the
uso of default assumptions as
recommended in the 1994 report.
Generally, these defaults remain public
health conservative, but in some
instances, they have been modified to
reflect the evolution of scientific
knowledge since 1986.
In addition, the guidelines reflect
evaluation of experience in practice in
applying defaults and departing from
them in individual risk assessments
conducted under the 1986 guidelines.
Tho application and departure from
dofaults and the principles to be used in
those judgments have been matters of
debato among practitioners and
reviewers of risk assessments. Some
observers believe that in practice EPA
risk assessors have been too resistant to
considering departures; others question
whether proposed departures have been
adequately supported. Some cases in
which departures have been considered
have been generally accepted, while
others have been controversial. The
guidelines here are intended to be both
explicit and more flexible than in the
past concerning the basis for making
departures from defaults, recognizing
that expert judgment and peer review
aro essential elements of the process.
In response to the recommendations
of tho 1994 report, these guidelines call
for identification of the default
assumptions used within assessments
and for highlighting significant issues
about defaults within characterization
summaries of component analyses in
assessment documents. As to the use of
poor review to aid in making judgments
about applying or departing from
dofaults, wo agree with the NRG
recommendation. The Agency has long
made use of workshops, peer review of
documents and guidelines, and
consultations as well as formal peer
review by the Science Advisory Board
(SAB), hi 1994, the Administrator of
EPA published formal guidance for peer
review of EPA scientific work products
that increases the amount of peer review
for risk assessments as well as other
work, as a response to the NRC report
and to SAB recommendations (U.S.
EPA, 1994b).
The 1994 NRC report recommended
that EPA should consider adopting
principles or criteria that would give
greater formality and transparency to
decisions to depart from defaults. The
report named several possible criteria
for such principles (p. 7): "* * *
[Protecting the public health, ensuring
scientific validity, minimizing serious
errors in estimating risks, maximizing
incentives for research, creating an
orderly and predictable process, and
fostering openness and trustworthiness.
There might be additional relevant
criteria* * *." The report indicated,
however, that the committee members
had not reached consensus on a single
criterion to address the key issue of how
much certainty or proof a risk assessor
must have in order to justify departing
from a default. Appendix N of the report
contains two presentations of alternative
views held by some committee members
on this issue. One view, known as
"plausible conservatism," suggested
that departures from defaults should not
be made unless new information
improves the understanding of a
biological process to the point that '
relevant experts reach consensus that
the conservative default assumption
concerning that process is no longer
plausible. The same criterion was
recommended where the underlying
scientific mechanism is well
understood, but where a default is used
to address missing data. In this case, the
default should not be replaced with
case-specific data unless it is the
consensus of relevant experts that the
proffered data make the default
assumption no longer plausible.
Another view, known as the "maximum
use of scientific information" approach,
acknowledged that the initial choice of
defaults should be conservative but
argued that conservatism should not be
a factor in determining whether to
depart from the default in favor of an
alternate biological theory or alternate
data. According to this view, it should
not be necessary to reach expert
consensus that the default assumption
had been rendered implausible; it
should be sufficient that risk assessors
find the alternate approach more
plausible than the default.
The EPA is notadopting a list of
formal decision criteria in the sense of
a checklist based on either view. It
would not be helpful to generate a
.checklist of uniform criteria for several
reasons. First, risk assessments are
highly variable in content and purpose.
Screening assessments may be
purposely "worst case" in their default
assumptions to eliminate problems from
further investigation. Subsequent risk
assessments based on a fuller data set
can discard worst-case default
assumptions in favor of plausibly
conservative assumptions and
progressively replace or modify the
latter with data. No uniform checklist
will fit all cases. Second, a checklist
would likely become more a source of
rote discussion than of enlightenment
about the process.
Instead, these guidelines use a
combination of principles and process
in the application of and departure from
default assumptions. The guidelines
provide a framework of default
assumptions to allow risk assessment to
proceed when current scientific theory
or available case-specific data do not
provide firm answers in a particular
case, as the 1983 report outlined. Some
of the default assumptions bridge large
gaps in fundamental knowledge which
will be filled by basic research on the
causes of cancer and on other biological
processes, rather than by agent-specific
testing. Other default assumptions
bridge smaller data gaps that can
feasibly be filled for a single agent, such
as whether a metabolic pathway in test
animals is like (default) or unlike that in
humans.
The decision to use a default, or not,
is a choice considering available
information on an underlying scientific
process and agent-specific data,
depending on which kind of default it
is. Generally, if a gap in basic
understanding exists, or if agent-specific
data are missing, the default is used
without pause. If data are present, their
evaluation may reveal inadequacies that
also lead to use of the default. If data
support a plausible alternative to the
default, but no more strongly than they
support the default, both the default and
its alternative are carried through the
assessment and characterized for the
risk manager. If data support an
alternative to the default as the more
reasonable judgment, the data are used.
(This framework of choices is not
wholly applicable to screening ,
assessments. As mentioned above,
screening assessments may
appropriately use "worst case"
inferences to determine if, even under
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those conditions, risk is low enough that
a problem can be eliminated from
further consideration.)
Scientific peer review, peer
consultative workshops and similar
processes are the principal ways
determining the strength of thinking and
generally accepted views within the
scientific community about the
application of and departure from
defaults and about judgments
concerning the plausibility and
persuasiveness of data in a particular
case. The choices made are explicitly
discussed in the assessment, and if a
particular choice raises a significant
issue, it is highlighted in the risk
characterization.
The discussion of major defaults in
these guidelines together with the
explicit discussion of the choice of
inferences within the assessment and
the processes of peer review and peer
consultation will serve the several goals
stated in the 1994 report. One is to
encourage research, since results of
research efforts will be considered.
Another is to allow timely
decisionmaking, when time is a
constraint, by supporting completion of
the risk assessment using defaults as
needed. Another is to be flexible, using
new science as it develops. Finally, the
use of public processes of peer
consultation and peer review will
ensure that discipline of thought is
maintained to support trust in
assessment results.
Experience has shown that the most
difficult part of the framework of
choices is the judgment of whether a
data analysis is both biologically
plausible and persuasive as applied to
the case at hand. There is no set of rules
for making this judgment in all cases.
Two criteria that apply in these
guidelines are that the underlying
scientific principle has been generally
accepted within the scientific
community and that supportive
experiments are available that test the
application of the principle to the agent
under review. For example,
mutagenicity through reactivity with
DNA has been generally accepted as a
carcinogenic influence for many years.
This acceptance, together with evidence
of such mutagenicity in experiments on
an agent, provides plausible and
persuasive support for the inference that
mutagenicity is a mode of action for the
agent.
Judgments about plausibility and
persuasiveness of analyses vary
according to the scientific nature of the
default. An analysis of data may replace
a default or modify it. An illustration of
the former is development of EPA
science policy on the issue of the
relevance for humans of male rat kidney
neoplasia involving alpha 2u globulin .
(U.S. EPA, 1991b). The 1991 EPA policy
gives guidance on the kind of
experimental findings that demonstrate
whether the alpha 2u globulin
mechanism is present and responsible
for carcinogenicity in a particular case.
Before this policy guidance was issued,
the default assumption was that
neoplasia in question was relevant to
humans and indicated the potential for
hazard to humans. A substantial body of
data was developed by public and
private research groups as a foundation
for the view that the alpha 2u globulin-
induced response was not relevant to
humans. These studies first addressed
the alpha 2u globulin mechanism in the
rat and whether this mechanism has a
counterpart in the human being, both
were large research efforts. The resulting
data presented difficulties; some
reviewers were concerned that the
mechanism in the rat appeared to be
understood only in outline, not in
detail, and felt that the data were
insufficient to show the lack of a
counterpart mechanism in humans. It
was particularly difficult to support a
negative such as the nonexistence of a
mechanism in humans because so little
is known about what the mechanisms
are in humans. Despite these concerns,
in its 1991 policy guidance, EPA
concluded that the alpha 2u globulin-
induced response in rats should be
regarded as not relevant to humans (i.e.,
as not indicating human hazard).
One lesson in the development and
peer review of this policy is that if the
default concerns an inherently complex
biological question, large amounts of
work will be required to replace the
default A second is that addressing a
negative is difficult. A third is that
"proof in the strict sense of having laid
all reasonable doubt to rest is not
required. Instead, an alternative may
displace a default when it is generally
accepted in peer review as the most
reasonable judgment. The issue of
relevance may not always be so
difficult. It would be an experimentally
easier task, for example, to determine
whether carcinogenesis in an animal
species is due to a metabolite of the
agent in question that is not produced
in humans.
When scientific processes are
understood but case-specific data are
missing, defaults can be constructed to
be modified by experimental data, even
if data do not suffice to replace'them
entirely. For example, the approaches
adopted in these guidelines for scaling
dose from experimental animals to
humans are constructed to be either
modified or replaced as data become
available on toxicokinetic parameters
for the particular agent being assessed.
Similarly, the selection of an approach
or approaches for dose response
assessment is based on a series of
decisions that consider the nature and
adequacy of available data in choosing
among alternative modeling and default
approaches.
The 1994 NRG report notes (p. 6) that
"[a]s scientific knowledge increases, the
science policy choices made by the
Agency and Congress should have less
impact on regulatory decisionmaking.
Better data and increased understanding
of biological mechanisms should enable
risk assessments that are less dependent
on conservative default assumptions
and more accurate as predictions of
human risk." Undoubtedly, this is the
trend as scientific understanding
increases. However, some gaps in
knowledge and data will doubtless
continue to be encountered in
assessment of even data-rich cases, and
it will remain necessary for risk
assessments to continue using defaults
within the framework set forth here.
1.3.2. Major Defaults
This discussion covers the major
default assumptions commonly
employed in a cancer risk assessment
and adopted in these guidelines. They
are predominantly inferences necessary
to use data observed under empirical
conditions to estimate events and
outcomes under environmental
conditions. Several inferential issues
arise when effects seen in a
subpopulation of humans or animals are
used to qualitatively infer potential
effects in the population of
environmentally exposed humans.
Several more inferential issues arise in
extrapolating the exposure-effect
relationship observed empirically to
lower-exposure environmental
conditions. The following issues cover
the major default areas. Typically, an
issue has some subissues; they are
introduced here, but are discussed in
greater detail in subsequent sections.
• Is the presence or absence of effects
observed in a human population
predictive of effects in another exposed
human population?
• Is the presence or absence of effects
observed in an animal population
predictive of effects in exposed
humans?
• How do metabolic pathways relate
across species?
• How do toxicokinetic processes
relate across species?
• What is the correlation of the
observed dose response relationship to
the relationship at lower doses?
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1.3.2.1.7s the Presence or Absence of
Effects Observed in a Human
Population Predictive of Effects in
Anotlwr Exposed Human Population?
When cancer effects in exposed humans
are attributed to exposure to an
exogenous agent, the default assumption
is that such data are predictive of cancer
in any oilier exposed human
population. Studies either attributing
cancer effects in humans to exogenous
agents or reporting no effects are often
studios of occupau'onally exposed
humans. By sex, age, and general health,
workers are not representative of the
general population exposed
environmentally to the same agents. In
such studios there is no opportunity to
observe whether infants and children,
males, or females who are under
represented in the study, or people
whose health is not good, would
respond differently. Therefore, it is
understood that this assumption could
still underestimate the response of
certain sensitive human subpopulations,
i.e. biologically vulnerable parts of the
population may be left out of risk
assessments (NRG, 1993a, 1994).
Consequently, this is a default that does
not err on the side of public health
conservatism, as the 1994 NRG report
also recognizes.
On the one hand, If effects are seen in
a worker population, this may be in fact
indicative of heightened effects in
sensitive subpopulations. There is not
enough knowledge yet to form a basis
for any generally applicable, qualitative
inference to compensate for this
knowledge gap. In these guidelines, this
problem is loft to case-by-case analysis,
to be attended to as future research and
information on particular agents allow.
When information on a sensitive
subpopulation exists, it will be used.
The topic of variability is addressed
further in the discussion of quantitative
default assumptions about dose
response relationships below. On the
other hand, when cancer effects are not
found in an exposed human population,
this information by itself is not
generally sufficient to conclude that the
agent poses no carcinogenic hazard to
this or other populations of potentially
exposed humans. This is because
opidomiologic studies usually have low
power to detect and attribute responses
(section 2,2.1.). This may be particularly
true when extrapolating null results
from a healthy, worker population to
oilier potentially sensitive exposed
humans. Again, the problem is left to
case-by-case analysis.
1.3.2.2.7s tlw Presence or Absence of
Effects Observed in an Animal
Population Predictive of Effects in
Exposed Humans? The default
assumption is that positive effects in
animal cancer studies indicate that the
agent under study can have
carcinogenic potential in humans. Thus,
if no adequate human data are present,
positive effects in animal cancer studies
are a basis for assessing the carcinogenic
hazard to humans. This assumption is a
public health conservative policy, and it
is both appropriate and necessary given
that we do not test for carcinogenicity
in humans. The assumption is
supported by the fact that nearly all of
the agents known to cause cancer in
humans are carcinogenic in animals in
tests with adequate protocols (IARC,
1994; Tomatis et al., 1989; Huff, 1994).
Moreover, almost one-third of human
carcinogens were identified subsequent
to animal testing (Huff, 1993). Further
support is provided by research on the
molecular biology of cancer processes,
which has shown that the mechanisms
of control of cell growth and
differentiation are remarkably
homologous among species and highly
conserved in evolution. Nevertheless,
the same research tools that have
enabled recognition of the nature and
commonality of cancer processes at the
molecular level also have the power to
reveal differences and instances in
which animal responses are not relevant
to humans (Linjinsky, 1993; U.S. EPA,
199ib). Under these guidelines,
available mode of action information is
studied for its implications in both
hazard and dose response assessment
and its effect on default assumptions.
There may be instances in which the
use of an animal model would identify
a hazard in animals that is not truly a
hazard in humans (e.g., the alpha-2u-
globulin association with renal
neoplasia in male rats (U.S. EPA,
1991b)). The extent to which animal
studies may yield false positive
indications for humans is a matter of
scientific debate. To demonstrate that a
response in animals is not relevant to
any human situation, adequate data to
assess the relevancy issue must be
available.
Animal studies are conducted at high
doses in order to provide statistical
power, the highest dose being one that
is minimally toxic (maximum tolerated
dose). Consequently, the question often
arises whether a carcinogenic effect at
the highest dose may be a consequence
of cell killing with compensatory cell
replication or of general physiological
disruption, rather than inherent
carcinogenicity of the tested agent.
There is little doubt that this may
happen in some cases, but skepticism
exists among some scientists that it is a
pervasive problem (Ames and Gold,
1990; Melnick et al., 1993a; Melnick et
al., 1993b; Barrett, 1993). In light of this
question, the default assumption is that
effects seen at the highest dose tested
are appropriate for assessment, but it is
necessary that the experimental
conditions be scrutinized. If adequate
data demonstrate that the effects are
solely the result of excessive toxicity
rather than carcinogenicity of the tested
agent per se, then the effects may be
regarded as not appropriate to include
in assessment of the potential for human
carcinogenicity of the agent. This is a
matter of expert judgment, considering
all of the data available about the agent
including effects in other toxicity
studies, structure-activity relationships,
and effects on growth control and
differentiation.
When cancer effects are not found in
well-conducted animal cancer studies in
two or more appropriate species and
other information does not support the
carcinogenic potential of the agent,
these data provide a basis for
concluding that the agent is not likely
to possess human carcinogenic
potential, in the absence of human data
to the contrary. This default assumption
about lack of cancer effects is not public
health conservative. For instance, the
tested animal species may not be
predictive of effects in humans; arsenic
shows only minimal or no effect in
animals, while it is clearly positive in
humans. (Other information, such as
absence of mutagenic activity or absence
of carcinogenic activity among
structural analogues, can increase the
confidence that negative results in
animal studies indicate a lack of human
hazard.) Also, it is recognized that
animal studies (and epidemiologic
studies as well) have very low power to
detect cancer effects. Detection of a 10%
tumor incidence is generally the limit of
power with currently conducted animal
studies (with the exception of rare
tumors that are virtually markers for a
particular agent, e.g., angiosarcoma
caused by vinyl chloride).
Target organs of carcinogenesis for
agents that cause cancer in both animals
and humans are most often concordant
at one or more sites (Tomatis et al.,
1989; Huff, 1994). However,
concordance by site is not uniform. The
default assumption is that target organ
concordance is not a prerequisite for
evaluating the implications of animal
study results for humans. This is a
public health conservative science
policy. The mechanisms of control of
cell growth and differentiation are
concordant among species, but there are
marked differences among species in the
way control is managed in various
tissues. For example, in humans,
mutation of the tumor suppressor gene
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p53 is one of the most frequently
observed genetic changes in tumors.
This tumor suppressor is also observed
to be operating in some rodent tissues,
but other growth control mechanisms
predominate in rodents. Thus, an
animal response may be due to changes
in a control that are relevant to humans,
but appear in animals in a different way.
However, it is appropriate under these
guidelines to consider the influences of
route of exposure, metabolism, and,
particularly, hormonal modes of action
that may either support or not support
target organ concordance between
animals and humans. When data allow,
these influences are considered in
deciding whether the default remains
appropriate in individual instances
(NRG, 1994, p. 121). An exception to the
basic default of not assuming site
concordance exists in the context of
toxicokinetic modeling. Site
concordance is inherently assumed
when these models are used to estimate
delivered dose in humans based on
animal data.
As in the approach of the National
Toxicology Program and the
International Agency for Research on
Cancer, the default is to include benign
tumors observed in animal studies in
the assessment of animal tumor
incidence if they have the capacity to
progress to the malignancies with which
they are associated. This treats the
benign and malignant tumors as
representative of related responses to
the test agent, which is scientifically
appropriate. This is a science policy
decision that is somewhat more
conservative of public health than not
including benign tumors in the
assessment. Nonetheless, in assessing
findings from animal studies, a greater
proportion of malignancy is weighed
more heavily than a response with a
greater proportion of benign tumors.
Greater frequency of malignancy of a
particular tumor type in comparison
with other tumor responses observed in
an animal study is also a factor to be
considered in selecting the response to
be used in dose response assessment.
Benign tumors that are not observed
to progress to malignancy are assessed
on a case-by-case basis. There is a range
of possibilities for their overall
significance. They may deserve
attention because they are serious health
problems even though they are not
malignant; for instance, benign tumors
may be a health risk because of their
effect on the function of a target tissue
such as the brain. They may be
significant indicators of the need for
further testing of an agent if they are
observed in a short term test protocol,
or such an observation may add to the
overall weight of evidence if the same
agent causes malignancies in a long
term study. Knowledge of the mode of
action associated with a benign tumor
response may aid in the interpretation
of other tumor responses associated
with the same agent. In other cases,
observation of a benign tumor response
alone may have no significant health
hazard implications when other sources
of evidence show no suggestion of
carcinogenicity.
1.3.2.3. How Do Metabolic Pathways
Relate Across Species? The default
assumption is that there is a similarity
of the basic pathways of metabolism and
the occurrence of metabolites in tissues
in regard to the species-to-species
extrapolation of cancer hazard and risk.
If comparative metabolism studies were
to show no similarity between the tested
species and humans and a metabolite(s)
were the active form, there would be
less support for an inference that the
animal response(s) relates to humans. In
other cases, parameters of metabolism
may vary quantitatively between
species; this becomes part of deciding
on an appropriate human equivalent
dose based on animal studies, optimally
in the context of a toxicokinetic model.
1.3.2.4. How Do Toxicokinetic
Processes Relate Across Species? A
major issue is how to estimate human
equivalent doses in extrapolating from
animal studies. As a default for oral
exposure, a human equivalent dose is
estimated from data on another species
by an adjustment of animal oral dose by
a scaling factor of body weight to the
0.75 power. This adjustment factor is
used because it represents scaling of
metabolic rate across animals of
different size. Because the factor adjusts
for a parameter that can be improved on
and brought into more sophisticated
toxicokinetic modeling, when such data
become available, the default
assumption of 0.75 power can be refined
or replaced.
For inhalation exposure, a human
equivalent dose is estimated by default
methodologies that provide estimates of
lung deposition and of internal dose.
The methodologies can be refined to
more sophisticated forms with data on
toxicokinetic and metabolic parameters
of the specific agent. This default
assumption, like the one with oral
exposure, is selected in part because it
lays a foundation for incorporating
better data. The use of information to
improve dose estimation from applied,
to internal, to delivered dose is
encouraged, including use of
toxicokinetic modeling instead of any
default, where data are available. Health
conservatism is not an element in
choosing the default.
For a route-to-route of exposure
extrapolation, the default assumption is
that an agent that causes internal tumors
by one route of exposure will be
carcinogenic by another route if it is
absorbed by the second route to give an
internal dose. This is a qualitative
assumption and is considered to be
public health conservative. The
rationale is that for internal tumors an
internal dose is significant no matter
what the route of exposure.
Additionally, the metabolism of the
agent will be qualitatively the same for
an internal dose. The issue of
quantitative extrapolation of the dose-
response relationship from one route to
another is addressed case by case.
Quantitative extrapolation is
complicated by considerations such as
first-pass metabolism, but is
approachable with empirical data.
Adequate data are necessary to
demonstrate that an agent will act
differently by one route versus another
route of exposure. ,
1.3.2.5. What Is the Correlation of the
Observed Dose Response Relationship to
the Relationship at Lower Doses? The
overriding preferred approach is to use
a biologically based or case-specific
model for both the observed range and
extrapolation below that range when
there are sufficient data. While
biologically based models are still under
development, it is likely that they will
be used more frequently in the future.
The default procedure for the observed
range of data, when the preferred
approach cannot be used, is to use a
curve-fitting model.
In the absence of data supporting a
biologically based or case-specific
model for extrapolation outside of the
observed range, the choice of approach
is based on the view of mode of action
of the agent arrived at in the hazard
assessment. A linear default approach is
used when the mode of action
information is supportive of linearity or,
alternatively, is insufficient to support a
nonlinear mode of action. The linear
approach is used when a view of the
mode of action indicates a linear
response, for example, when a
conclusion is made that an agent
directly causes alterations in DNA, a
kind of interaction that not only
theoretically requires one reaction, but
also is likely to be additive to ongoing,
spontaneous gene mutation. Other kinds
of activity may have linear implications,
e.g., linear rate-limiting steps, that
support a linear procedure also. The
linear approach is to draw a straight line
between a point of departure from
observed data, generally, as a default,
the LED 10, and the origin (zero dose,
zero response). Other points of
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departure may be more appropriate for
certain data sets; these may be used
Instead of the LEDio. This approach is
generally considered to be public health
conservative. The LEDio is the lower
95% limit on a dose that is estimated to
cause a 10% response. This level is
chosen to account (conservatively) for
experimental variability. Additionally,
it is chosen because it rewards
experiments with better designs in
regard to number of doses and dose
spacing, since these generally will have
narrower confidence limits. It is also an
appropriate representative of the lower
end of the observed range because the
limit of detection of studies of tumor
effect is about 10%.
The linear default is thought to
generally produce an upper bound on
potential risk at low doses, e.g., a I/
100,000 to 1/1,000,000 risk; the straight
lino approach gives numerical results
about the same as a linearized
multistage procedure (Krewski et al.,
1984). This upper bound is thought to
cover the range of human variability
although, in some cases, it may not
completely do so (Bois et al., 1995). The
EPA considers the linear default to be
inherently conservative of public health,
without addition of another factor for
human variability. In any case, the size
of such a factor would be hard to
determine since a good empirical basis
on which to construct an estimate does
not currently exist. The question of
what may be the actual variability in
human sensitivity is one that the 1994
NRC report discussed as did the 1993
NRG report on pesticides in children
and infants. The NRC has recommended
research on the question, and the EPA
and other agencies have begun such
research.
When adequate data on mode of
action show that linearity is not the
most reasonable working judgment and
provldo sufficient evidence to support a
nonlinear mode of action, the default
changes to a different approach—a
margin of exposure analysis—which
assumes that nonlinearity is more
reasonable. The departure point is again
generally the LEDio- A margin of
exposure analysis compares the LEDio
with the dose associated with the
environmental exposure(s) of interest by
computing the ratio between the two.
The purpose of a margin of exposure
analysis is to provide the risk manager
with all available information on how
much reduction in risk may be
associated with reduction in exposure
from the point of departure. This is to
support the risk manager's decision as
to what constitutes an acceptable
margin of exposure, given requirements
of the statute under which the decision
is being made. There are several factors
to be considered. (For perspective, keep
in mind that a sufficient basis to support
this nonlinear procedure often will
include data on responses that are
precursors to tumor effects. This means
that the point of departure may well be
from these biological response data
rather than tumor incidence data, e.g.,
hormone levels, mitogenic effects.) One
factor to consider is the slope of the
dose response curve at the point of
departure. A steeper slope implies an
apparent greater reduction in risk as
exposure decreases. This-may support a •
smaller margin of exposure. Conversely,.
a shallow slope may support use of a
greater margin of exposure. A second
factor is the nature of the response used
in the assessment—A precursor effect or
frank toxicity or tumor response. The
latter two may support a greater margin
of exposure. A third factor is the nature
and extent of human variability in
sensitivity to the phenomenon. A fourth
factor is the agent's persistence in the
body. Greater variability or persistence
argue for greater margins of exposure. A
fifth factor is human sensitivity to the
phenomenon as compared with
experimental animals. The size of the •.
margin of exposure that is acceptable
would increase or decrease as this factor
increases or decreases. If human
variability cannot be estimated based on
data, it should be considered to be at
least 10-fold. Similarly, if comparison of
species sensitivities cannot be estimated
from available data, humans can be
considered to be 10-fold more sensitive.
If it is found that humans are less
sensitive than animals a factor that is a
fraction no smaller than Vio may be
assumed. The 10-fold factors are
moderately conservative, traditional
ones used for decades in the assessment
of toxicological effects. It should not be
assumed that the numerical factors are
the sole components for determination
of an acceptable margin of exposure.
Each case calls for individual judgment.
It should be noted that for cancer
assessment the margin of exposure
analysis begins from a point of
departure that is adjusted for
toxicokinetic differences between
species to give a human equivalent
dose. Since the traditional factor for
interspecies difference is thought to
contain a measure for toxicokinetics as
well as sensitivity to effect, the result of
beginning with a human equivalent
dose is to add some conservatism. The
ultimate judgment whether a particular
margin of exposure is acceptable is a
risk management decision under
applicable law, rather than being
inherent in the risk assessment.
Nonetheless, the risk assessor is
responsible for providing scientific
rationale to support the the decision.
When the mode of action information
indicates that the dose response may be
adequately described by both a linear
and a nonlinear approach, then the
default is to present both the linear and
margin of exposure analyses. An
assessment may use both linear and
nonlinear approaches either for
responses that are thought to result from
different modes of action or for
presenting considerations for a response
that appears to be very different at high
and low doses due to influence of
separate modes of action. Also, separate
approaches may be used for different
induced responses (i.e. tumor types)
from the same agent. These would also
be carried forward and presented in the
assessment. Figure 1—1 presents the
decision points in deciding on a dose
response approach or approaches.
BILLING CODE 6560-50-P
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Federal Register / Vol. 61, No. 79 / Tuesday, April 23, 1996 / Notices
Data to Support:
Biologically
Based or Case-
Specific Model
Linearity .
Nonlinearity
1
1 Extrapolation Used:
yes
model
no
yes
no
default--
linear
no
no
yes
default--
nonlinear
no
yes
yes
default—linear
and nonlinear
no
no
no
default--
linear
Figure 1-1. Decisions on dose response assessment approaches for
the range of extrapolation.
BILLING CODE 6560-50-C
A default assumption is made that
cumulative dose received over a
lifetime, expressed as a lifetime average
daily dose, is an appropriate measure of
dose. This assumes that a high dose of
such an agent received over a shorter
period of time is equivalent to a low
dose spread over a lifetime. This is
thought to be a relatively public health
conservative assumption and has
empirical support (Monro, 1992). An
example of effects of short-term, high
exposure that results in subsequent
cancer development is treatment of
cancer patients with certain
chemotherapeutic agents. An example
of cancer from long-term exposure to an
agent of relatively low potency is
smoking. Whether the cumulative dose
measure is exactly the correct measure
in both such instances is not certain and
should be assessed case by case and .
altered when data are available to
support another approach. Other
measures of dose that consider dose rate
and duration are appropriate, e.g., when
an agent acts by causing cell toxicity or
hormone disruption. In these cases both
agent concentration and duration are
likely to be important, because such
effects are generally observed to be
reversible at cessation of short-term
exposure.
1.4. Characterizations
The risk characterization process first
summarizes findings on hazard, dose
response, and exposure
characterizations, then develops an
integrative analysis of the whole risk
case. It ends in a nontechnical Risk
Characterization Summary. The Risk
Characterization Summary is a
presentation for risk managers who may
or may not be familiar with the
scientific details of cancer assessment. It
also provides information for other
interested readers. The initial steps in
the risk characterization process are to
make building blocks in the form of
characterizations of the assessments of
hazard, dose response, and exposure.
The individual assessments and
characterizations are then integrated to
arrive at risk estimates for exposure
scenarios of interest. There are two
reasons for individually characterizing
the hazard, dose response, and exposure
assessments. One is that they are often
done by different people than those who
do the integrative analyses. The second
is that there is very often a lapse of time
between the conduct of hazard and dose"
response analyses and the conduct of
exposure assessment and integrative
analysis. Thus, it is necessary to capture
characterizations of assessments as the
assessments are done to avoid the need
to go back and reconstruct them. Figure
1—2 shows the relationships of analyses.
The figure does not necessarily
correspond to the number of documents
involved; there may be one or several.
"Integrative analysis" is a generic term.
At EPA, the documents of various
programs that contain integrative
analyses have other names such as the
"Staff Paper" that discusses air quality
criteria issues. In the following sections,
the elements of this figure are discussed.
BILLING CODE 6560-50-P
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17971
CHARACTERIZATIONS
Hazard
Assessment
Dose Response
Assessment
Exposure
Assessment
Technical
Hazard
Characterization
Technical
Dose Response
Characterization
Technical
Exposure
Characterization
RISK
CHARACTERIZATION
SUMMARY
Integra tive
Analysis
Risk Characterization Process
Figure 1-2. Risk Characterization
DILUIKJ CODE K60-60-C
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Federal Register / Vol. 61, No. 79 / Tuesday, April 23, 1996 / Notices
2. Hazard Assessment
2.1. Overview of Hazard Assessment
and Characterization
2.1.1. Analyses of Data
The purpose of hazard assessment is
to review and evaluate data pertinent to
two questions: (1) whether an agent may
pose a carcinogenic hazard to human
beings and (2) under what
circumstances an identified hazard may
be expressed (NRG, 1994, p. 142).
Hazard assessment is composed of
analyses of a variety of data that may
range from observations of tumor
responses to analysis of structure-
activity relationships. The purpose of
the assessment is not simply to
assemble these separate evaluations; its
purpose is to construct a total case
analysis examining the biological story
the data reveal as a whole about
carcinogenic effects, mode of action,
and implications of these for human
hazard and dose response evaluation.
Weight of evidence conclusions come
from the combined strength and
coherence of inferences appropriately
drawn from all of the available
evidence. To the extent that data permit,
hazard assessment addresses the mode
of action question as both an initial step
in considering appropriate approaches
to dose response assessment and as a
part of identifying human hazard
potential.
The topics in this section include
analysis of tumor data, both animal and
human, and analysis of other key
information about properties and effects
that relate to carcinogenic potential. The
section addresses how information can
be used to evaluate potential modes of
action. It also provides guidance on
performing a weight of evidence
evaluation.
2.1.2. Cross-Cutting Topics for Data
Integration
Two topics are included in the
analysis of each kind of available data:
first, gathering information from
available data about the conditions of
expression of hazard and second,
gathering perspectives on the agent's
potential mode of action.
2.1.2.1. Conditions of Expression.
Information on the significance of the
route of exposure may be available from
human or animal studies on the agent
itself or on structural analogues. This
information may be found in studies of
the agent or analogue for toxicological
endpoints other than cancer under acute
or subchronic or chronic exposure
regimens. Studies of metabolism or
toxicokinetics of the agent similarly may
provide pertinent data.
Each kind of data is also examined for
information on conditions that affect
expression of carcinogenic effect such as
presence or absence of metabolic
pathways. If carcinogenicity is
secondary to another toxic effect, the
physiological or tissue changes that
mark the other toxicity are examined.
Comparison of metabolic processes and
toxicity processes in humans and
animals also bears on the relevance of
animal responses to human hazard.
Included in the examination are the
questions of the potential range of
human variability and whether any
special sensitivity may occur because of
age, sex, preexisting disease, or other
condition.
2.1.2.2. Mode of Action. Information
on an agent's potential mode(s) of action
is important in considering the
relevance of animal effects to
assessment of human hazard. It also
plays an important role in selecting dose
response approach(es), which are
generally either biologically based
models or case-specific models
incorporating mode of action data or
default procedures based on more
limited data that support inferences
about the likely shape of the dose
response curve.
Each kind of data may provide some
insight about mode of action and
insights are gathered from each to be
considered together as discussed in
section 2.4. In Appendix C, is a
background discussion of some of the
development of views about
carcinogenic processes.
2.1.3. Presentation of Results.
Presentation of the results of hazard
assessment follows Agency guidance as
discussed in section 2.7. The results are
presented in a technical hazard
characterization that serves as a support
to later risk characterization. It includes:
• a summary of the evaluations of
hazard data,
• the rationales for its conclusions,
and
• an explanation of the significant
strengths or limitations of the
conclusions.
Another presentation feature is the
use of a weight of evidence narrative
that includes both a conclusion about
the weight of evidence of carcinogenic
potential and a summary of the data on
which the conclusion rests. This
narrative is a brief summary that
replaces the alphanumerical
classification system used in EPA's
previous guidelines.
2.2. Analysis of Tumor Data
Evidence of carcinogenicity comes
from finding tumor increases in humans
or laboratory animals exposed to a given
agent, or from finding tumors following
exposure to structural analogues to the
compound under review. The
significance of observed or anticipated
tumor effects is evaluated in reference to
all of the other key data on the agent.
This section contains guidance for
analyzing human and animal studies to
decide whether there is an association
between exposure to an agent or a
structural analogue and occurrence of
tumors. Note that the use of the term
"tumor" here is generic, meaning
malignant neoplasms or a combination
of malignant and corresponding benign
neoplasms.
Observation of only benign neoplasias
may or may not have significance.
Benign tumors that are not observed to
progress to malignancy are assessed on
a case-by-case basis. There is a range of
possibilities for their overall
significance. They may deserve
attention because they are serious health
problems even though they are not
malignant; for instance, benign tumors
may be a health risk because of their
effect on the function of a target tissue
such as the brain. They may be
significant indicators of the need for
further testing of an agent if they are
observed in a short term test protocol,
or such an observation may add to the
overall weight of evidence if the same
agent causes malignancies in a long
term study. Knowledge of the mode of
action associated with a benign tumor
response may aid in the interpretation
of other tumor responses associated
with the same agent. In other cases,
observation of a benign tumor response
alone may have no significant health
hazard implications when other sources
of evidence show no suggestion of •
carcinogenicity.
2.2.1. Human Data
Human data may come from
epidemiologic studies or case reports.
Epidemiology is the study of the
distributions and causes of disease
within human populations. The goals of
cancer epidemiology are to identify
differences in cancer risk between
different groups in a population or
between different populations, and then
to determine the extent to which these
differences in risk can be attributed
causally to specific exposures to
exogenous or endogenous factors.
Epidemiologic data are extremely useful
in risk assessment because they provide
direct evidence that a substance
produces cancer in humans, thereby
avoiding the problem of species to
species inference. Thus, when available
human data are extensive and of good
quality, they are generally preferable
over animal data and should be given
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greater weight in hazard
characterization and dose response
assessment, although both are utilized.
Null results from a single
epidomiologic study cannot prove the
absence of carcinogenic effects because
they can arise either from being truly
negative or from inadequate statistical
power, inadequate design, imprecise
estimates, or confounding factors.
However, null results from a well-
designed and well-conducted
epidomiologic study that contains
usable exposure data can help to define
upper limits for the estimated dose of
concern for human exposure if the
overall weight of the evidence indicates
that tho agent is potentially carcinogenic
in humans.
Epidemiology can also complement
experimental evidence in corroborating
or clarifying the carcinogenic potential
of the agent in question. For example,
observations from epidemiologic studies
that elevated cancer incidence occurs at
sites corresponding to those at which
laboratory animals experience increased
tumor incidence can strengthen the
weight of evidence of human
carcinogonicity. On the other hand,
strong nonpositive epidemiologic data
alone or in conjunction with compelling
mechanistic information can lend
support to a conclusion that.animal
responses may not be predictive of a
human response. Furthermore, the
advent of biochemical or molecular
epidemiology may help improve
understanding of the mechanisms of
human corcinogenesis.
2.2.1.1, Types of Studies. The major
types of cancer epidemiologic studies
are analytical epidemiologic studies and
descriptive or correlation epidemiologic
studies. Each study type has well-
known strengths and weaknesses that
affect interpretation of study results as
summarized below (Kelsey et al., 1986;
Lilionfold and Lilienfeld, 1979; Mausner
and Kramer, 1985; Rothman, 1986).
Analytical opidemiologic studies are
most useful for identifying an
association between human exposure
and adverse health effects. Analytical
study designs include case-control
studies and cohort studies. In case-
control studios, groups of individuals
with (cases) and without (controls) a
particular disease are identified and
compared to determine differences in
exposure. In cohort studies, a group of
"exposed" and "nonexposed"
individuals are identified and studied
over lime to determine differences in
disease occurrence. Cohort studies can
either bo performed prospectively or
retrospectively from historical records.
Descriptive or correlation
opidomiologic studies (sometimes called
ecological studies) examine differences
in disease rates among populations in
relation to age, gender, race, and
differences in temporal or
environmental conditions. In general,
these studies can only identify patterns
or trends in disease occurrence over
time or in different geographical
locations but cannot ascertain the causal
agent or degree of exposure. These
studies, however, are often very useful
for generating hypotheses for further
research.
Biochemical or molecular
epidemiologic studies are studies in
which laboratory methods are
incorporated in analytical
investigations. The application of
techniques for measuring cellular and
molecular alterations due to exposure to
specific environmental agents may
allow conclusions to be drawn about the
mechanisms of carcinogenesis. The use
of biological biomarkers in
epidemiology may improve assessment
of exposure and internal dose.
Case reports describe a particular
effect in an individual or group of
individuals who were exposed to a
substance. These reports are often
anecdotal or highly selected in nature
and are of limited use for hazard
assessment. However, reports of cancer
cases can identify associations
particularly when there are unique
features such as an association with an
uncommon tumor (e.g., vinyl chloride
and angiosarcoma or diethylstilbestrol
and clear-cell carcinoma of the vagina).
2.2.1.2. Criteria for Assessing
Adequacy of Epidemiologic Studies.
Criteria for assessing the adequacy of
epidemiologic studies are well
recognized. Characteristics that are
desirable in these studies include (1),
clear articulation of study objectives or
hypothesis, (2) proper selection and
characterization of the exposed and
control groups, (3) adequate
characterization of exposure, (4)
sufficient length of follow-up for disease
occurrence, (5) valid ascertainment of
the causes of cancer morbidity and
mortality, (6) proper consideration of
bias and confounding factors, (7)
adequate sample size to detect an effect,
(8) clear, well-documented, and
appropriate methodology for data
collection and analysis, (9) adequate
response rate and methodology for
handling missing data, and (10)
complete and clear documentation of
results. Ideally, these conditions should.
be satisfied, where appropriate, but
rarely can a study meet all of them. No
single criterion determines the overall
adequacy of a study. The following
discussions highlight the major factors
included in an analysis of
epidemiologic studies.
Population Issues. The ideal
comparison would be between two
populations that differ only in exposure
to the agent in question. Because this is
seldom the case, it is important to
identify sources of bias inherent in a
study's design or data collection
methods. Bias can arise from several
sources, including noncomparability
between populations of factors such as
general health (McMichael, 1976), diet,
lifestyle, or geographic location;
differences in the way case and control
individuals recall past events;
differences in data collection that result
in unequal ascertainment of health
effects in the populations; and unequal
follow-up of individuals. Both
acceptance of studies for assessment
and judgment of their strengths or
weaknesses depend on identifying their
sources of bias and the effects on study '
results.
Exposure Issues. For epidemiologic
data to be useful in determining
whether there is an association between
health effects and exposure to an agent,
there must be adequate characterization
of exposure information. In general,
greater weight should be given to
studies with more precise and specific
exposure estimates.
Questions to address about exposure
are: What can one reliably conclude
about the level, duration, route, and , '
frequency of exposure of individuals in -
one population as compared with
another? How sensitive are study results
to uncertainties in these parameters?
Actual exposure measurements are
not available for many retrospective
studies. Therefore, surrogates are often
used to reconstruct exposure parameters
when historical measurements are not
available. These may involve attributing.
exposures to job classifications in a
workplace or to broader occupational or
geographic groupings. Use of surrogates
carries a potential for misclassification
in that individuals may be placed in the
incorrect exposure group.
Misclassification generally leads to i
reduced ability of a study to detect
differences between study and referent
populations.
When either current or historical
monitoring data are available, the
exposure evaluation includes
consideration of the error bounds of the
monitoring and analytic methods and
whether the data are from routine or
accidental exposures. The potentials for
misclassification and measurement
errors are amenable to both qualitative
and quantitative analysis. These are
essential analyses for judging a study's
results because exposure estimation is
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Federal Register / Vol. 61, No. 79 / Tuesday, April 23, 1996 / Notices
the most critical part of a retrospective
study.
Biological markers potentially offer
excellent measures of exposure (Hulka
and Margolin, 1992; Peto and Darby,
1994). Validated markers of exposure
such as alkylated hemoglobin from
exposure to ethylene oxide (van Sittert
et al., 1985) or urinary arsenic (Enterline
et al., 1987) can greatly improve
estimates of dose. Markers closely
identified with effects promise to greatly
increase the ability of studies to
distinguish real effects from bias at low
levels of relative risk between
populations (Taylor et al., 1994; Biggs et
al., 1993) and to resolve problems of
confounding risk factors.
Confounding Factors. Because
epidemiologic studies are mostly
observational, it is not possible to
guarantee the control of confounding
variables, which may affect the study
outcome. A confounding variable is a
risk factor, independent of the putative
agent, that is distributed unequally
among the exposed and unexposed
populations (e.g., smoking habits,
lifestyle). Adjustment for possible
confounding factors can occur either in
the design of the study (e.g., matching
on critical factors) or in the statistical
analysis of the results. The influence of
a potential confounding factor is limited
by the effect of the exposure of interest.
For example, a twofold effect of an
exposure requires that the confounder
effect be at least as big. The latter may
not be possible due to the presentation ,
of the data or because needed
information was not collected during
the study. In this case, indirect
comparisons may be possible. For
example, in the absence of data on
smoking status among individuals in the
study population, an examination of the
possible contribution of cigarette
smoking to increased lung cancer risk
may be based on information from other
sources such as the American Cancer
Society's longitudinal studies
(Hammand, 1966; Garfinkel and
Silverberg, 1991). The effectiveness of
adjustments contributes to the ability to
draw inferences from a study.
Different studies involving exposure
to an agent may have different
confounding factors. If consistent
increases in cancer risk are observed
across a collection of studies with
different confounding factors, the
inference that the agent under
investigation was the etiologic factor is
strengthened, even though complete
adjustment for confounding factors
cannot be made and no single study
supports a strong inference.
It also may be the case that the agent
of interest is a risk factor in conjunction
with another agent. This relationship
may be revealed in a collection of
studies such as in the case of asbestos
exposure and smoking.
Sensitivity. Sensitivity, or tha ability
of a study to detect real effects, is a
function of several factors. Greater size
of the study population(s) (sample size)
increases sensitivity, as does greater
exposure (levels and duration) of the
population members. Because of the
often long latency period in cancer
development, sensitivity also depends
on whether adequate time has elapsed
since exposure began for effects to
occur. A unique feature that can be
ascribed to the effects of a particular
agent (such as a tumor type that is seen
only rarely in the absence of the agent)
can increase sensitivity by permitting
separation of bias and confounding
factors from real effects. Similarly, a
biomarker particular to^the agent can
permit these distinctions. Statistical
reanalyses of data, particularly an
examination of different exposure
indices, can give insight on potential
exposure-response relationships. These
are all factors to explore in statistical
analysis of the data.
Statistical Considerations. The
analysis applies appropriate statistical
methods to ascertain whether or not
there is any significant association
between exposure and effects. A
description of the method or methods
should include the reasons for their
selection. Statistical analyses of the
potential effects of bias or confounding
factors are part of addressing the
significance of an association, or lack of
one, and whether a study is able to
detect any effect.
The analysis augments examination of
the results for the whole population
with exploration of the results for
groups with comparatively greater
exposure or time since first exposure.
This may support identifying an
association or establishing a dose
response trend. When studies show no
association, such exploration may apply
to determining an upper limit on
potential human risk for consideration
alongside results of animal tumor effects
studies.
Combining Statistical Evidence
Across Studies. Meta-analysis is a
means of comparing and synthesizing
studies dealing with similar health
effects and risk factors. It is intended to
introduce consistency and
comprehensiveness into what otherwise
might be a more subjective review of the
literature. When utilized appropriately,
meta-analysis can enhance
understanding of associations between
sources and their effects that may not be
apparent from examination of
epidemiologic studies individually.
Whether to conduct a meta-analysis
depends on several issues. These
include the importance of formally
examining sources of heterogeneity, the
refinement of the estimate of the
magnitude of an effect, and the need for
information beyond that provided by
individual studies or a narrative review.
Meta-analysis may not be useful in some
circumstances. These include when the
relationship between exposure and
disease is obvious without a more
formal analysis, when there are only a
few studies of the key health outcomes,
when there is insufficient information
from available studies related to disease,
risk estimate, or exposure classification,
or when there are substantial
confounding or other biases that cannot
be adjusted for in the analysis (Blair et
al., 1995; Greenland, 1987; Peto, 1992).
2.2.1.3. Criteria for Causality. A
causal interpretation is enhanced for
studies to the extent that they meet the
criteria described below. None of the
criteria is conclusive by itself, and the
only criterion that is essential is the
temporal relationship. These criteria are
modeled after those developed by
Bradford Hill in the examination of
cigarette smoking and lung cancer
(Rothman, 1986) and they need to be
interpreted in the light of all other
information on the agent being assessed.
• Temporal relationship: The
development of cancers require certain
latency periods, and while latency
periods vary, existence of such periods
is generally acknowledged. Thus, the
disease has to occur within a
biologically reasonable time after initial
exposure. This feature must be present
if causality is to be considered.
• Consistency: Associations occur in
several independent studies of a similar
exposure in different populations, or
associations occur consistently for
different subgroups in the same study.
This feature usually constitutes strong
evidence for a causal interpretation
when the same bias or confounding is
not also duplicated across studies.
• Magnitude of the association: A
causal relationship is more credible
when the risk estimate is large and
precise (narrow confidence intervals).
• Biological gradient: The risk ratio
(i.e., the ratio of the risk of disease or
death among the exposed to the risk of
the unexposed) increases with
increasing exposure or dose. A strong
dose response relationship across
several categories of exposure, latency,
and duration is supportive for causality
given that confounding is unlikely to be
correlated with exposure. The absence
of a dose response relationship,
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howovor, is not by itself evidence
against a causal relationship.
• Specificity of the association: The
likelihood of a causal interpretation is
increased if an exposure produces a
specific effect Cone or more tumor types
also found in other studies) or if a given
effect has a unique exposure.
• Biological plausibility: The
association makes sense in terms of
biological knowledge. Information is
considered from animal toxicology,
toxicokinetics, structure-activity
relationship analysis, and short-term
studios of tho agent's influence on
events in the carcinogenic process
considered.
• Coherence: The cause-and-effect
interpretation is in logical agreement
with what is known about the natural
history and biology of the disease, i.e.,
the entire body of knowledge about the
agent.
2.2.1.4. Assessment of Evidence of
Carclnogenicity from Human Data. In
tho evaluation of carcinogenicity based
on opidemiologic studies, it is necessary
to critically evaluate each study for the
confidence in findings and conclusions
as discussed under section 2.2.1.2. All
studies that are properly conducted,
whether yielding positive or null
results, or even suggesting protective
carcinogenic effects, should be
considered in assessing the totality of
tho human evidence. Although a single
study may bo indicative of a cause-effect
relationship, confidence in inferring a
causal relationship is increased when
several independent studies are
concordant in showing the association,
when tho association is strong, and
when other criteria for causality are also
met. Conclusions about the overall
evidence for carcinogenicity from
available studies in humans should be
summarized along with a discussion of
strengths or limitations of the
conclusions.
2.2.2. Animal Data
Various kinds of whole animal test
systems are currently used or are under
development for evaluating potential
carcinogenicity. Cancer studies
involving chronic exposure for most of
tho life span of an animal are generally
accepted for evaluation of tumor effects
(Tomalis etal., 1989; Rail, 1991; Allen
ot al., 1988; but see Ames and Gold,
1990). Other studies of special design
ore useful for observing formation of
proneoplnstic lesions or tumors or
investigating specific modes of action.
2.2.2.1. Long-Term Carcinogenicity
Studios. Tho objective of long-term
carclnogcnesis bioassays is to determine
tho carcinogenic potential and dose
response relationships of the test agent.
Long-term rodent studies are designed
to examine the production of tumors as
well as preneoplastic lesions and other
indications of chronic toxicity that may
provide evidence of treatment-related
effects and insights into the way the test
agent produces tumors. Current
standardized long-term studies in
rodents test at least 50 animals per sex
per dose group in each of three
treatment groups and in a concurrent
control group, usually for 18 to 24
months, depending on the rodent
species tested (OECD, 1981; U.S. EPA,
1983a; U.S. EPA, 1983b; U.S. EPA,
1983c). The high dose in long-term
studies is generally selected to provide
the maximum ability to detect
treatment-related carcinogenic effects
while not compromising the outcome of
the study due to excessive toxicity or
inducing inappropriate toxicokinetics
(e.g., overwhelming detoxification or
absorption mechanisms). The purpose
of two or more lower doses is to provide
some information on the shape of the
dose response curve. Similar protocols
have been and continue to be used by
many laboratories worldwide.
All available studies of tumor effects
in whole animals are considered, at
least preliminarily. The analysis
discards studies judged to be wholly
inadequate in protocol, conduct, or
results. Criteria for the technical
adequacy of animal carcinogenicity
studies have been published and should
be used as guidance to judge the
acceptability of individual studies (NTP,
1984; OSTP, 1985). Care is taken to
include studies that provide some
evidence bearing on carcinogenicity or
help interpret effects noted in other
studies even if they have some
limitations of protocol or conduct. Such
limited, but not wholly inadequate,
studies can contribute as their
deficiencies permit. The findings of
long-term rodent bioassays are always
interpreted in conjunction with results
of prechronic studies along with
toxicokinetic and metabolism studies
and other pertinent information, if
available. Evaluation of tumor effects
requires consideration of both biological
and statistical significance of the
findings (Haseman, 1984,1985,1990,
1995). The following sections highlight
the major issues in the evaluation of
long-term carcinogenicity studies.
Dosing issues. In order to obtain the
most relevant information from a long-
term carcinogenicity study, it is
important to require maximization of
exposure to the test material. At the
same time, there is a need for caution in
using excessive high dose levels that
would confound the interpretation of
study results to humans. The high dose
is conventionally defined as a dose that
produces some toxic effects without
either unduly affecting mortality from
effects other than cancer or producing
significant adverse effects on the
nutrition and health of the test animals
(OECD, 1981; NRC, 1993b). It should be
noted that practical upper limits have
been established to avoid the use of
excessive high doses in long-term
carcinogenicity studies (e.g., 5% of the
test substance in the feed for dietary
studies [OECD, 1981]).
Evaluating the appropriateness of the
high dose in carcinogenicity studies is
based on scientific judgment using all
available relevant information. In
general, if the test agent does not appear
to cause any specific target organ
toxicity or perturbation of physiological
function, an adequate high dose would
be a dose that causes no more than 10%
reduction of body weight gain over the
life span of the animals. On the other
hand, significant increases in mortality
from effects other than cancer is
accepted as clear evidence of frank
toxicity, which indicates that an
adequate high dose may have been
exceeded. Other signs of treatment-
related toxicity that may indicate that an
adequate high dose has been exceeded
include the following: (a) Reduction of
body weight gain of 10% or greater, (b)
significant increases in abnormal
behavioral and clinical signs, (c)
significant changes in hematology or
clinical chemistry, (d) saturation of
absorption and detoxification
mechanisms, or (e) marked changes in
organ weight, morphology, and
histopathology.
For dietary studies, weight gain
reductions should be evaluated as to
whether there is a palatability problem
or an issue with food efficiency;
certainly, the latter is a toxic
manifestation. In the case of inhalation
studies with respirable particles,
evidence of impairment of normal
clearance of particles from the lung
should be considered along with other
signs of toxicity to the respiratory
airways to determine whether the high
exposure concentration has been
appropriately selected. For dermal
studies, evidence of skin irritation may
indicate that an adequate high dose has
been reached.
Interpretation of carcinogenicity study
results is profoundly affected by
exposure conditions, especially by
inappropriate dose selection. This is
particularly important in studies that
are nonpositive for carcinogenicity,
since failure to reach a sufficient dose
reduces the sensitivity of a study. A lack
of tumorigenic responses at exposure
levels that cause significant impairment
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of animal survival may also not be
acceptable as negative findings because
of the reduced sensitivity of the study.
On the other hand, overt toxicity or
inappropriate toxicokinetics due to
excessive high doses may result in
tumor effects that are secondary to the
toxicity rather than directly attributable
to the agent.
There are several possible outcomes
regarding the study interpretation of the
significance and relevance of
tumorigenic effects associated with
exposure or dose levels below, at, or
above an adequate high dose. General
guidance is given here that should not
be taken as prescriptive; for each case,
the information at hand is evaluated and
a rationale should be given for the
position taken.
• Adequate high dose: If an adequate
high dose has been utilized, tumor
effects are judged positive or negative
depending on the presence or absence of
significant tumor incidence increases,
respectively.
• Excessive high dose: If toxicity or
mortality is excessive at the high dose,
interpretation depends on the finding of
tumors or not.
(a) Studies that show tumor effects
only at excessive doses may be
compromised and may or may not carry
weight, depending on the interpretation
in the context of other study results and
other lines of evidence. Results of such
studies, however, are generally not
considered suitable for risk
extrapolation.
(b) Studies that show tumors at lower
doses, even though the high dose is
excessive and may be discounted,
should be evaluated on their own
merits.
(c) If a study does not show an
increase in tumor incidence at a toxic
high dose and appropriately spaced
lower doses are used without such
toxicity or tumors, the study is generally
judged as negative for carcinogenicity.
• Inadequate high dose: Studies of
inadequate sensitivity where an
adequate high dose has not been
reached may be used to bound the dose
range where carcinogenic effects might
be expected.
Statistical Considerations. The main
aim of statistical evaluation is to
determine whether exposure to the test
agent is associated with an increase of
tumor development. Statistical analysis
of a long-term study should be
performed for each tumor type
separately. The incidence of benign and
malignant lesions of the same cell type,
usually within a single tissue or organ,
are considered separately and are
combined when scientifically defensible
(McConnell et al., 1986).
Trend tests and pairwise comparison
tests are the recommended tests for
determining whether chance, rather
than a treatment-related effect, is a
plausible explanation for an apparent
increase in tumor incidence. A trend
test such as the Cochran-Armitage test
(Snedecor and Cochran, 1967) asks
whether the results in all dose groups
together increase as dose increases. A
pairwise comparison test such as the
Fisher exact test (Fisher, 1932) asks
whether an incidence in one dose group
is increased over the control group. By
convention, for both tests a statistically
significant comparison is one for which
p <0.05 that the increased incidence is
due to chance. Significance in either
kind of test is sufficient to reject the
hypothesis that chance accounts for the
result. A statistically significant
response may or may not be biologically
significant and vice versa. The selection
of a significance level is a policy choice
based on a trade-off between the risks of
false positives and false negatives. A
significance level of greater or less than
5% is examined to see if it confirms
other scientific information. When the
assessment departs from a simple 5%
level, this should be highlighted in the
risk characterization. A two-tailed test
or a one-tailed test can be used. In either
case a rationale is provided.
Considerations of multiple
comparisons should also be taken into
account. Haseman (1983) analyzes
typical animal bioassays testing both
sexes of two species and concludes that,
because of multiple comparisons, a
single tumor increase for a species-sex-
site combination that is statistically
significant at the 1% level for common
tumors or 5% for rare tumors
corresponds to a 7—8% significance
level for the study as a whole.
Therefore, animal bioassays presenting
only one significant result that falls
short of the 1% level for a common
tumor may be treated with caution.
Concurrent and Historical Controls.
The standard for determining statistical
significance of tumor incidence comes
from a comparison of tumors in dosed
animals as compared with concurrent
control animals. Additional insights
about both statistical and biological
significance can come from an
examination of historical control data
(Tarone, 1982; Haseman, 1995).
Historical control data can add to the
analysis particularly by enabling
identification of uncommon tumor types
or high spontaneous incidence of a
tumor in a given animal strain.
Identification of common or uncommon
situations prompts further thought
about the meaning of the response in the
current study in context with other
observations in animal studies and with
other evidence about the carcinogenic
potential of the agent. These other
sources of information may reinforce or
weaken the significance given to the
response in the hazard assessment.
Caution should be exercised in simply
looking at the ranges of historical
responses because the range ignores
differences in survival of animals among
studies and is related to the number of
studies in the database.
In analyzing results for uncommon
tumors in a treated group that are not
statistically significant in comparison to
concurrent controls, the analyst can use
the experience of historical controls to
conclude that the result is in fact
unlikely to be due to chance. In
analyzing results for common tumors, a
different set of considerations comes
into play. Generally speaking,
statistically significant increases in
tumors should not be discounted simply
because incidence rates in the treated
groups are within the range of historical
controls or because incidence rates in
the concurrent controls are somewhat
lower than average. Random assignment
of animals to groups and proper
statistical procedures provide assurance
that statistically significant results are
unlikely to be due to chance alone.
However, caution should be used in
interpreting results that are barely
statistically significant or in which
incidence rates in concurrent controls
are unusually low in comparison with
historical controls.
In cases where there may be reason to
discount the biological relevance to
humans of increases in common animal
tumors, such considerations should be
weighed on their own merits and clearly
distinguished from statistical concerns.
When historical control data are used,
the discussion needs to address several
issues that affect comparability of
historical and concurrent control data.
Among these issues are the following:
genetic drift in the laboratory strains;
differences in pathology examination at
different times and in different
laboratories (e.g., in criteria for
evaluating lesions; variations in the
techniques for preparation or reading of
tissue samples among laboratories);
comparability of animals from different
suppliers. The most relevant historical
data come from the same laboratory and
same supplier, gathered within 2 or 3
years one way or the other of the study
under review; other data should be used
only with extreme caution.
Assessment of Evidence of
Carcinogenicity from Long-Term
Animal Studies. In general, observation
of tumor effects under different
circumstances lends support to the
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17977
significance of the findings for animal
cnrcinogonicity. Significance is a
function of the number of factors
present, and for a factor such as
malignancy, the severity of the observed
pathology. The following observations
add significance to the tumor findings:
• uncommon tumor types
• tumors at multiple sites
• tumors by more than one route of
administration
• tumors in multiple species, strains,
or both sexes
• progression of lesions from
prcnooplaslic to benign to malignant
• reduced latency of neoplastic
lesions
• motostases
• unusual magnitude of tumor
response
• proportion of malignant tumors
• doso-related increases
Those guidelines adopt the science
policy position that tumor findings in
animals indicate that an agent may
produce such effects in humans.
Moreover, the absence of tumor findings
in well-conducted, long-term animal
studios in at least two species provides
reasonable assurance that an agent may
not bo a carcinogenic concern for
humans. Each of these is a default
assumption that may be adopted, when
appropriate, after evaluation of tumor
data and other key evidence.
Site concordance of tumor effects
between animals and humans is an
issue to be considered in each case.
Thus for, there is evidence that growth
control mechanisms at the level of the
cell are homologous among mammals,
but there is no evidence that these
mechanisms are site concordant.
Moreover, agents observed to produce
tumors in both humans and animals
have produced tumors either at the
same (e.g., vinyl chloride) or different
sites (e.g., benzene) (NRG, 1994). Hence,
site concordance is not assumed a
priori. On the other hand, certain
processes with consequences for
particular tissue sites (e.g., disruption of
thyroid function) may lead to an
anticipation of site concordance.
2.2.2.2. Other Studies. Various
intermediate-term studies often use
protocols that screen for carcinogenic or
prcneoplastic effects, sometimes in a
single tissue. Some involve the
development of various proliferative
lesions, like foci of alteration in the liver
(Goldsworthy ot al.» 1986). Others use
tumor ondpoints, like the induction of
lung adenomas in the sensitive strain A
mouso (Maronpot et al., 1986) or tumor
induction in initiation-promotion
studies using various organs such as the
bladder, intestine, liver, lung, mammary
gland, and thyroid (Ito et al., 1992). In
these tests, the selected tissue is, in a
sense, the test system rather than the
whole animal. Important information
concerning the steps in the carcinogenic
process and mode of action can be
obtained from "start/stop" experiments.
In these protocols, an agent is given for
a period of time to induce particular
lesions or effects, then stopped to
evaluate the progression or reversibility
of processes (Todd, 1986; Marsman and
Popp, 1994).
Assays in genetically engineered
rodents ma}' provide insight into the
chemical and gene interactions involved
in carcinogenesis (Tennant et al.,
1995a). These mechanistically based
approaches involve activated oncogenes
that are introduced (transgenic) or
tumor suppressor genes that are deleted
(knocked-out). If appropriate genes are
selected, not only may these systems
provide information on mechanisms,
but the rodents typically show tumor
development earlier than the standard
bioassay. Transgenic mutagenesis assays
also represent a mechanistic approach
for assessing the mutagenic properties of
agents as well as developing
quantitative linkages between exposure,
internal dose, and mutation related to
tumor induction (Morrison and Ashby,
1994; Sisk et al., 1994; Hayward et al.,
1995). These systems use a stable
genomic integration of a lambda shuttle
vector that carries a lacl target gene and
a lacZ reporter gene.
The support that these studies give to
a determination of carcinogenicity rests
on then* contribution to the consistency
of other evidence about an agent. For
instance, benzoyl peroxide has promoter
activity on the skin, but the overall
evidence may be less supportive (Kraus
et al., 1995). These studies also may
contribute information about mode of
action. One needs to recognize the
limitations of these experimental
protocols such as short duration, limited
histology, lack of complete development
of tumors, or experimental
manipulation of the carcinogenic
process that may limit their contribution
to the overall assessment. Generally,
their results are appropriate as aids in
the assessment for interpreting other
lexicological evidence (e.g., rodent
chronic bioassays), especially regarding
potential modes of action. With
sufficient validation, these studies may
partially or wholly replace chronic
bioassays in the future (Tennant et al.,
1995).
2.2.3. Structural Analogue Data
For some chemical classes, there is
significant information available on the
carcinogenicity of analogues, largely in
rodent bioassays. Analogue effects are
instructive in investigating carcinogenic
potential of an agent as well as
identifying potential target organs,
exposures associated with effects, and
potential functional class effects or
modes of action. All appropriate studies
are included and analyzed, whether
indicative of a positive effect or not.
Evaluation includes tests in various
animal species, strains, and sexes; with
different routes of administration; and at
various doses, as data are available.
Confidence in conclusions is a function
of how similar the analogues are to the
agent under review in structure,
metabolism, and biological activity.
This confidence needs to be considered
to ensure a balanced position.
2.3. Analysis of Other Key Data
The physical, chemical, and structural
properties of an agent, as well as data on
endpoints that are thought to be critical
elements of the carcinogenic process,
provide valuable insights into the
likelihood of human cancer risk. The
following sections provide guidance for
analyses of these data.
2.3.1. Physicochemical Properties
Physicochemical properties affect an
agent's absorption, tissue distribution
(bioavailability), biotransformation, and
degradation in the body and are
important determinants of hazard
potential (and dose response analysis).
Properties to analyze include, but are
not limited to, the following: molecular
weight, size, and shape; valence state;
physical state (gas, liquid, solid); water
or lipid solubility, which can influence
retention and tissue distribution; and
potential for chemical degradation or
stabilization in the body.
An agent's potential for chemical
reaction with cellular components,
particularly with DNA and proteins, is
also important. The agent's molecular
size and shape, electrophilicity, and
charge distribution are considered in
order to decide whether they would
facilitate such reactions.
2.3.2. Structure-Activity Relationships
Structure-activity relationship (SAR)
analyses and models can be used to
predict molecular properties, surrogate
biological endpoints, and
carcinogenicity. Overall, these analyses
provide valuable initial information on
agents, which may strengthen or weaken
the concern for an agent's carcinogenic
potential.
Currently, SAR analysis is useful for
chemicals and metabolites that are
believed to initiate carcinogenesis
through covalent interaction with DNA
(i.e., DNA-reactive, mutagenic,
electrophilic, or proelectrophilic
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chemicals) (Ashby and Tennant, 1991).
For organic chemicals, the predictive
capability of SAR analysis combined
with other toxicity information has been
demonstrated (Ashby and Tennant,
1994). The following parameters are
useful in comparing an agent to its
structural analogues and congeners that
produce tumors and affect related
biological processes such as receptor
binding and activation, mutagenicity,
and general toxicity (Woo and Arcos,
1989):
• nature and reactivity of the
electrophilic moiety or moieties present,
• potential to form electrophilic
reactive intermediate(s) through
chemical, photochemical, or metabolic
activation,
• contribution of the carrier molecule
to which the electrophilic moiety(ies) is
attached,
• physicochemical properties (e.g.,
physical state, solubility, octanol-water
partition coefficient, half-life in aqueous
solution),
• structural and substructural features
(e.g., electronic, stearic, molecular
geometric),
• metabolic pattern (e.g., metabolic
pathways and activation and
detoxification ratio), and
• possible exposure route(s) of the
agent.
Suitable SAR analysis of non-DNA-
reactive chemicals and of DNA-reactive
chemicals that do not appear to bind
covalently to DNA requires knowledge
or postulation of the probable mode(s)
of action of closely related carcinogenic
structural analogues (e.g., receptor-
mediated, cytotoxicity-related).
Examination of the physicochemical
and biochemical properties of the agent
may then provide the rest of the
information needed in order to make an
assessment of the likelihood of the
agent's activity by that mode of action.
2.3.3. Comparative Metabolism and
Toxicokinetics
Studies of the absorption,
distribution, biotransformation, and
excretion of agents permit comparisons
among species to assist in determining
the implications of animal responses for
human hazard assessment, supporting
identification of active metabolites,
identifying changes in distribution and
metabolic pathway or pathways over a
dose range, and making comparisons
among different routes of exposure.
If extensive data are available (e.g.,
blood/tissue partition coefficients and
pertinent physiological parameters of
the species of interest), physiologically
based pharmacokinetic models can be
constructed to assist in a determination
of tissue dosimetry, species-to-species
extrapolation of dose, and route-to-route
extrapolation (Connolly and Andersen,
1991; see section 3.2.2). If it is not
contrary to available data, it is assumed
as a default that toxicokinetic and
metabolic processes are qualitatively
comparable between species. Discussion
of the defaults regarding quantitative
comparison and their modifications
appears in section 3.
The qualitative question of whether
an agent is absorbed by a particular
route of exposure is important for
weight of evidence classification
discussed in section 2.7.1. Decisions
whether route of exposure is a limiting
factor on expression of any hazard, in
that absorption does not occur by a
route, are based on studies in which
effects of the agent, or its structural
analogues, have been observed by
different routes, on physical-chemical
properties, or on toxicokinetics studies.
Adequate metabolism and
pharmacokinetic data can be applied
toward the following as data permit.
Confidence in conclusions is enhanced
when in vivo data are available.
• Identifying metabolites and reactive
intermediates of metabolism and
determining whether one or more of
these intermediates are likely to be
responsible for the observed effects.
This information on the reactive
intermediates will appropriately focus
SAR analysis, analysis of potential
modes of action, and estimation of .
internal dose in dose response
assessment (D'Souza et al., 1987;
Krewski et al., 1987).
• Identifying and comparing the
relative activities of metabolic pathways
in animals with those in humans. This
analysis can provide insights for
extrapolating results of animal studies
to humans.
• Describing anticipated distribution
within the body and possibly
identifying target organs. Use of water
solubility, molecular weight, and
structure analysis can support
qualitative inferences about anticipated
distribution and excretion. In addition,
describing whether the agent or
metabolite of concern will be excreted
rapidly or slowly or will be stored in a
particular tissue or tissues to be
mobilized later can identify issues in
comparing species and formulating dose
response assessment approaches.
• Identifying changes in
toxicokinetics and metabolic pathways
with increases in dose. These changes
may result in important differences in
disposition of the agent or its generation
of active forms of the agent between
high and low dose levels. These studies
play an important role in providing a
f physicochemical data (e.g.,
ol-water partition coefficient
rationale for dose selection in
carcinogenicity studies.
• Determining bioavailability via
different routes of exposure by
analyzing uptake processes under
various exposure conditions. This
analysis supports identification of
hazards for untested routes. In addition,
use of i
octanol-water partitio
information) can support an inference
about the likelihood of dermal
absorption (Flynn, 1990).
Irt all of these areas, attempts are
made to clarify and describe as much as
possible the variability to be expected
because of differences in species, sex,
age, and route of exposure. The analysis
takes into account the presence of
subpopulations of individuals who are
particularly vulnerable to the effects of
an agent because of toxicokinetic or
metabolic differences (genetically or
environmentally determined) (Bois et
al., 1995).
2.3.4. Toxicological and Clinical
Findings
Toxicological findings in
experimental animals and clinical
observations in humans are an
important resource to the cancer hazard
assessment. Such findings provide
information on physiological effects,
effects on enzymes, hormones, and
other important macromolecules as well
as on target organs for toxicity. Given
that the cancer process represents
defects in terminal differentiation,
growth control, and cell death,
developmental studies of agents may
provide an understanding of the activity
of an agent that carries over to cancer
assessment. Toxicity studies in animals
by different routes of administration
support comparison of absorption and
metabolism by those routes. Data on
human variability in standard clinical
tests may provide insight into the range
of human sensitivity and common
mechanisms to agents that affect the
tested parameters.
2.3.5. Mode of Action-Related
Endpoints and Short-Term Tests
A myriad of biochemical and
biological endpoints relevant to the
carcinogenic process provide important
information in determining whether a
cancer hazard exists and include, but
are not limited to, mutagenesis,
inhibition of gap junctional intercellular
communication, increased cell
proliferation, inhibition of programmed
cell death, receptor activation, and
immunosuppression. These precursor
effects are discussed below.
2.3.5.1. Direct DNA Effects. Because
cancer is the result of multiple genetic
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17979
defects in genes controlling proliferation
and tissue homeostasis (Vogelstein et
al.( 1988), the ability of an agent to affect
DNA is of obvious importance. It is well
known that many carcinogens are
olectrophiles that interact directly with
DNA, resulting in DNA damage and
adducts, and subsequent mutations
(referred to in these guidelines as direct
DNA effects) that are thought to
contribute to the carcinogenic process
(Shelby and Zoiger, 1990; Tinwell and
Ashby, 1991). Thus, studies of these
phenomena continue to be important in
the assessment of cancer hazard. The
EPA has published testing guidelines for
detecting the ability of agents to affect
DNA or chromosomes (EPA, 1991a).
Information on agents that induce
mutations in animal germ cells also
deserves attention; several human
carcinogens have been shown to be
positive in rodent tests for the induction
of genetic damage in both somatic and
germ colls (Shelby, 1995).
2.3.5.2. Secondary DNA Effects.
Similarly of interest are secondary
mechanisms that either increase
mutation rates or the number of
dividing cells. An increase in mutations
might be due to cytotoxic exposures
causing regenerative proliferation or
mitogenic influences, either of which
could result in clonal expansion of
initiated cells (Cohen and Ellwein,
1990). An agent might interfere with the
enzymes involved in DNA repair and
recombination (Barrett and Lee, 1992).
Also, programmed cell death (apoptosis)
can potentially be blocked by an agent,
thereby permitting replication of
damaged cells. For example, peroxisome
proliferators may act by suppressing
apoptosis pathways (Shulte-Hermann et
at., 1993; Bayly eta/., 1994). An agent
may also generate reactive oxygen
species that produce oxidative damage
to DNA and other important
macromolecules that become important
elements of the carcinogenic process
(Kehrer, 1993; Clayson etal, 1994;
Chang et a!., 1988). Damage to certain
critical DNA repair genes or other genes
(e.g., the p53 gene) may result in
gonomic instability, which predisposes
cells to further genetic alterations and
increases the probability of neoplastic
progression independent of any
exogenous agent (Harris and Hollstein,
1993; Lovine, 1994).
The loss or gain of chromosomes (i.e.,
anouploidy) is an effect that can result
in gonomic instability (Feoron and
Vogolstoin, 1990; Cavenee et al, 1986).
Although the relationship between
induced aneuploidy and carcinogenesis
is not completely established, several
carcinogens have been shown to induce
aneuploidy (Gibson et al., 1995; Barrett,
1992). Agents that cause aneuploidy
interfere with the normal process of
chromosome segregation and lead to
chromosomal losses, gains, or
aberrations by interacting with the
proteins (e.g., microtubules) needed for
chromosome movement.
2.3.5.3. Nonmutagenic and Other
Effects. A failure to detect DNA damage
and mutation induction in several test
systems suggests that a carcinogenic
agent may act by another mode of
action.
It is possible for an agent to alter gene
expression (transcriptional,
translational, or post-translational
modifications) by means not involving
mutations (Barrett, 1995). For example,
perturbation of DNA methylation
patterns may cause effects that
contribute to carcinogenesis (Jones,
1986; Goodman and Counts, 1993;
Holliday, 1987). Overexpression of
genes by amplification has been
observed in certain tumors (Vainio et
al., 1992). Other mechanisms may
involve cellular reprogramming through
hormonal mechanisms or receptor-
mediated mechanisms (Ashby et al.,
1994; Barrett, 1992).
Gap-junctional intercellular
communication is widely believed to
play a role in tissue and organ
development and in the maintenance of
a normal cellular phenotype within
tissues. A growing body of evidence
suggests that chemical interference with
gap-junctional intercellular
communication is a contributing factor
in tumor development; many
carcinogens have been shown to inhibit
this communication. Thus, such
information may provide useful
mechanistic data in evaluating cancer
hazard (Swierenga and Yamasaki, 1992;
Yamasaki, 1995).
Both cell death and cell proliferation
are mandatory for the maintenance of
homeostasis in normal tissue. The
balance between the two directly affects
the survival and growth of initiated
cells, as well as preneoplastic and tumor
cell populations (i.e., increase in cell
proliferation or decrease in cell death)
(Bellamy et al., 1995; Cohen and
Ellwein, 1990,1991; Cohen et al., 1991).
In studies of proliferative effects,
distinctions should be made between
mitogenesis and regenerative
proliferation (Cohen and Ellwein, 1990,
1991; Cohen et al., 1991). In applying
information from studies on cell
proliferation and apoptosis to risk
assessment, it is important to identify
the tissues and target cells involved, to
measure effects in both normal and
neoplastic tissue, to distinguish between
apoptosis and necrosis, and to
determine the dose that affects these
processes.
2.3.5.4. Criteria for Judging Mode of
Action. Criteria that are applicable for
judging the adequacy of mechanistically
based data include the following:
• mechanistic relevance of the data to
carcinogenicity, , ,
• number of studies of each endpoint,
• consistency of results in different
test systems and different species,
• similar dose response relationships
for tumor and mode of action-related
effects,
• tests conducted in accordance with
generally accepted protocols, and
• degree of consensus and general
acceptance among scientists regarding
interpretation of the significance and,
specificity of the tests.
Although important information can
be gained from in vitro test systems, a
higher level of confidence is generally
given to data that are derived from in
vivo systems, particularly those results
that show a site concordance with the
tumor data.
2.4. Biomarker Information
Various endpoints can serve as
biological markers of events in
biological systems or samples. In some
cases, these molecular or cellular effects
(e.g., DNA or protein adducts, mutation,
chromosomal aberrations, levels of
thyroid stimulating hormone) can be
measured in blood, body fluids, cells
and tissues to serve as biomarkers of
exposure in both animals and humans
(Callemen et al., 1978; Birner et al.,
1990). As such, they can do the
following:
• act as an internal surrogate measure
of chemical dose, representing as •.
appropriate, either recent (e.g., serum
concentration) or accumulated (e.g.,
hemoglobin adducts) exposure,
• help identify doses at which -
elements of the carcinogenic process are
operating,
• aid in interspecies extrapolations
when data are available from both
experimental animal and human cells,
and
• under certain circumstances,
provide insights into the possible shape
of the dose response curve below levels
where tumor incidences are observed
(e.g., Choy, 1993).
Genetic and other findings (like
changes in proto-oncogenes and tumor
suppressor genes in preneoplastic and
neoplastic tissue or possibly measures
of endocrine disruption) can indicate
the potential for disease and as such
serve as biomarkers of effect. They, too,
can be used in different ways:
• The spectrum of genetic changes in
proliferative lesions and tumors
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following chemical administration to
experimental animals can be
determined and compared with those in
spontaneous tumors in control animals,
in animals exposed to other agents of
varying structural and functional
activities, and in persons exposed to the
agent under study.
• They may provide a linkage to
tumor response.
• They may help to identify
subpopulations of individuals who may
be at an elevated risk for cancer, e.g.,
cytochrome P450 2D6/debrisoquine
sensitivity for lung cancer (Caporaso et
al., 1989) or inherited colon cancer
syndromes (Kinzler et al., 1991;
Peltomaki et al., 1993).
• As with biomarkers of exposure, it
may be justified in some cases to use
these endpoints for dose response
assessment or to provide insight into the
potential shape of the dose response
curve at doses below those at which
tumors are induced experimentally.
In applying biomarker data to cancer
assessment (particularly assessments
based on epidemiologic data), one
should consider the following:
• routes of exposure
• exposure to mixtures
• time after exposure
• sensitivity and specificity of
biomarkers
• dose response relationships.
2.5. Mode of Action—Implications for
Hazard Characterization and Dose
Response
The interaction of the biology of the
organism and the chemical properties of
the agent determine whether there is an
adverse effect. Thus, mode of action
analysis is based on physical, chemical,
and biological information that helps to
explain critical events in an agent's
influence on development of tumors.
The entire range of information
developed in the assessment is reviewed
to arrive at a reasoned judgment. An
agent may work by more than one mode
of action both at different sites and at
the same tumor site. It is felt that at least
some information bearing on mode of
action (e.g., SAR, screening tests for
mutagenicity) is present for most agents
undergoing assessment of
carcinogenicity, even though certainty
about exact molecular mechanisms may
be rare.
Inputs to mode of action analysis
include tumor data in humans, animals,
and among structural analogues as well
as the other key data. The more
complete the data package and generic
knowledge about a given mode of-
action, the more confidence one has and
the more one can replace or refine
default science policy positions with
relevant information. Making reasoned
judgments is generally based on a data-
rich source of chemical, chemical class,
and tumor type-specific information.
Many times there will be conflicting
data and gaps in the information base;
one must carefully evaluate these
uncertainties before reaching any
conclusion.
Some of the questions that need to be
addressed include the following:
• Has a body of data been developed
on the agent that fits with a generally
accepted mode of action?
• Has the mode of action been
published and gained general scientific
acceptance through peer-reviewed
research or is it still speculative?
• Is the mode of action consistent
with generally agreed-upon principles
and understanding of carcinogenesis?
• Is the mode of action reasonably
anticipated or assumed, in the absence
of specific data, to operate in humans?
How is this question influenced by
information on comparative uptake,
metabolism, and excretion patterns
across animals and humans?
• Do humans appear to be more or
less sensitive to the mode of action than
are animals?
• Does the agent affect DNA, directly
or indirectly?
• Are there important determinants in
carcinogenicity other than effects on
DNA, such as changes in cell
proliferation, apoptosis, gene
expression, immune surveillance, or
other influences?
In making decisions about potential
modes of action and the relevance of
animal tumor findings to humans
(Ashby et al., 1990), very often the
results of chronic animal studies may
give important clues. Some of the
important factors to review include the
following:
• tumor types, e.g., those responsive
to endocrine influence, those produced
by reactive carcinogens (Ashby and
Tennant, 1991),
• number of tumor sites, sexes,
studies, and species affected or
unaffected (Tennant, 1993),
• influence of route of exposure;
spectrum of tumors; local or systemic
sites,
• target organ or system toxicity, e.g.,
urinary chemical changes associated
with stone formation, effects on immune
surveillance,
• presence of proliferative lesions,
e.g., hepatic foci, hyperplasias,
• progression of lesions from
preneoplastic to benign to malignant
with dose and time,
• ratio of malignant to benign tumors
as a function of dose and time,
• time of appearance of tumors after
commencing exposure,
• tumors invading locally,
metastasizing, producing death,
• tumors at sites in laboratory
animals with high or low spontaneous
historical incidence,
• biomarkers in tumor cells, both
induced and spontaneous, e.g., DNA or
protein adducts, mutation spectra,
chromosome changes, oncogene
activation, and
• shape of the dose response in the
.range of tumor observation, e.g., linear
vs. profound change in slope.
Some of the myriad of ways that
information from chronic animal studies
influences mode of action judgments
include the following. Multisite and
multispecies tumor effects are often
associated with mutagenic agents. •
Tumors restricted to one sex/species
may suggest an influence restricted to
gender/strain, or species. Late onset of
tumors that are primarily benign or are
at sites with a high historical
background incidence or show reversal
of lesions on cessation of exposure may
point to a growth-promoting mode of
action. The possibility that an agent may
act differently in different tissues or
have more than one mode of action in
a single tissue must also be kept in
mind.
Simple knowledge of sites of tumor
increase in rodent studies can give
preliminary clues as to mode of action.
Experience at the National Toxicology
Program (NTP) indicates that substances
that are DNA reactive and produce gene
mutations may be unique in producing
tumors in certain anatomical sites,
while tumors at other sites may arise
from both mutagenic or nonmutagenic
influences (Ashby and Tennant, 1991;
Huffetal., 1991).
Effects on tumor sites in rodents and
other mode of action information has
been explored for certain agents (Alison
et al., 1994; Clayson, 1989; ECETOC,
1991; MacDonald et al., 1994; McClain,
1994; Tischer et al., 1991; ILSI, 1995;
Cohen and Ellwein, 1991; FASEB, 1994;
Havu et al., 1990; U.S. EPA, 1991; Li et
al., 1987; Grasso and Hinton, 1991;
Larson et al., 1994; IARC, 1990; Jack et
al., 1983; Stitzel et al., 1989; Ingram and
Grasso, 1991; Bus and Popp, 1987;
Prahalada et al., 1994; Yamada et al.,
1994; Hill et al., 1989; Burek et al.,
1988).
The selection of a dose response
extrapolation procedure for cancer risk
estimation considers mode of action
information. When information is
extensive and there is considerable
certainty in a given mode of action, a
biologically based or case-specific
model that incorporates data on
processes involved is preferred.
Obviously, use of such a model requires
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tho existence of substantial data on
component parameters of the mode of
action, and judgments on its
applicability must be made on a case-
by-case basis.
In tho absence of information to
dovolop a biologically based or case-
spccific model, understanding of mode
of action should be employed to the
extent possible in deciding upon one of
three science policy defaults: Low-dose
linear extrapolation, nonlinear, and both
procedures. The overall choice of the
dofault(s) depends upon weighing the
various inputs and deciding which best
reflect the mode of action
understanding. A rationale accompanies
whichever default or defaults are
chosen.
A default assumption of linearity is
appropriate when the evidence supports
a mode of action of gene mutation due
to DNA reactivity or supports another
mode of action that is anticipated to be
linear. Other elements of empirical data
may also support an inference of
linearity, e.g., the background of human
exposure to an agent might be such that
added human exposure is on the linear
part of a dose response curve that is
sublincar overall. The default
assumption of linearity is also
appropriate as the ultimate default
when evidence shows no DNA
reactivity or other support for linearity,
but neither is it sufficient evidence of a
nonlinear mode of action to support a
nonlinear procedure.
A default assumption of nonlinearity
is appropriate when there is no
evidence for linearity and sufficient
evidence to support an assumption of
nonlinoarity and a nonlinear procedure.
The mode of action may lead to a dose
response relationship that is nonlinear,
with response falling much more
quickly than linearly \vith dose, or being
most influenced by individual
differences in sensitivity. Alternatively,
tho mode of action may theoretically
have a threshold, e.g., the
corcinogenicity may be a secondary
effect of toxicity that is itself a threshold
phenomenon.
Both linear and nonlinear procedures
may be used in particular cases. If a
mode of action analysis finds
substantial support for differing modes
of action for different tumor sites, an
appropriate procedure is used for each.
Both procedures may also be
appropriate to discuss implications of
complex dose response relationships.
For example, if it is apparent that an
agent is both DNA reactive and is highly
active as a promoter at high doses, and
there are insufficient data for modeling,
both linear and nonlinear default
procedures may be needed to decouple
and consider the contribution of both
phenomena.
2.6. Weight of Evidence Evaluation for
Potential Human Carcinogenicity
A weight of evidence evaluation is a
collective evaluation of all pertinent
information so that the full impact of
biological plausibility and coherence are
adequately considered. Identification
and characterization of human
Carcinogenicity is based on human and
experimental data, the nature,
advantages and limitations of which
have been discussed in the preceding
sections.
The subsequent sections outline: (I)
the basics of weighing individual lines
of evidence and combining the entire
body of evidence to make an informed
judgment, (2) classification descriptors
of cancer hazard, and (3) some case
study examples to illustrate how the
principles of guidance can be applied to
arrive at a classification.
2.6.1. Weight of Evidence Analysis
Judgment about the weight of
evidence involves considerations of the
quality and adequacy of data and
consistency of responses induced by the
agent in question. The weight of
evidence judgment requires combined
input of relevant disciplines. Initial
views of one kind of evidence may
change significantly when other
information is brought-to the
interpretation. For example, a positive
animal Carcinogenicity finding may be
diminished by other key data; a weak
association in epidemiologic studies
may be bolstered by consideration of
other key data and animal findings.
Factors typically considered are
illustrated in figures below. Generally,
no single weighing factor on either side
determines the overall weight. The
factors are not scored mechanically by
adding pluses and minuses; they are
judged in combination.
Human Evidence. Analyzing the
contribution of evidence from a body of
human data requires examining
available studies and weighing them in
the context of well-accepted criteria for
causation (see section 2.2.1). A
judgment is made about how closely
they satisfy these criteria, individually
and jointly, and how far they deviate
from them. Existence of temporal
relationships, consistent results in
independent studies, strong association,
reliable exposure data, presence of dose-
related responses, freedom from biases
and confounding factors, and high level
of statistical significance are among the
factors leading to increased confidence
in a conclusion of causality.
Generally, the weight of human
evidence increases with the number of
adequate studies that show comparable
results on populations exposed to the
same agent under different conditions.
The analysis takes into account all
studies of high quality, whether
showing positive associations or null
results, or even protective effects. In
weighing positive studies against null
studies, possible reasons for
inconsistent results should be sought,
and results of studies that are judged to
be of high quality are given more weight
than those from studies judged to be
methodologically less sound. See figure
2-1.
BILLING CODE 6560-50-P
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Federal Register / Vol. 61, No. 79 / Tuesday, April 23, 1996 / Notices
Human Evidence Factors
Increase Weight
Decrease Weight
Number of independent studies with
consistent results
Most causal criteria satisfied:
Temporal relationship
Strong association
Reliable exposure data
Dose response relationship
Freedom from bias and confounding
Biological plausibility
High statistical significance
Few studies
Equally well designed and conducted
studies with null results
Few causal criteria satisfied
Figure 2-1. Factors for Weighing Human Evidence
Generally, no single factor is
determinative. For example, the strength
of association is one of the causal
criteria. A strong association (i.e., a large
relatively risk) is more likely to indicate
causality than a weak association.
However, finding of a large excess risk
in a single study must be balanced
against the lack of consistency as
reflected by null results from other
equally well designed and well
conducted studies. In this situation, the
positive association of a single study
may either suggest the presence of
chance, bias or confounding, or reflect
different exposure conditions. On the
other hand, evidence of weak but
consistent associations across several
studies suggests either causality or the
same confounder may be operating in
all of these studies.
Animal Evidence. Evidence from
long-term or other carcinogenicity
studies in laboratory animals constitutes
the second major class of information
bearing on carcinogenicity. See figure 2—
2. As discussed in section 2.2.2., each
relevant study must be reviewed and
evaluated as to its adequacy of design
and conduct as well as the statistical
significance and biological relevance of
its findings. Factors that usually
increase confidence in the predictivity
of animal findings are those of (1)
multiplicity of observations in
independent studies; (2) severity of
lesions, latency, and lesion progression;
(3) consistency in observations.
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17983
Animal Evidence Factors
Increase Weight
Number of independent studies with
consistent results
Same'site across species, structural
analogues
Multiple observations
Species
Sites
Sexes
Severity and progression of lesions
Early in life tumors/malignancy
Dose response relationships
Lesion progression
Uncommon tumor
Route of administration like human
exposure
Decrease Weight
Single study
Inconsistent results
Single site/species/sex
Benign tumors only
High background of incidence tumors
Route of administration unlike human
exposure
A
Figure 2-2. Factors for Weighing Animal Evidence
Other Key Evidence. Additional
Information bearing on the qualitative
assessment of carcinogenic potential
may bo gained from comparative
pharmacokinotic and metabolism
studios, genetic toxicity studies, SAR
analysis, and other studies of an agent's
properties. See figure 2—3. Information
from these studies helps to elucidate
potential modes of action and biological potential.
fate and disposition. The knowledge
gained supports interpretation of cancer
studies in humans and animals and
provides a separate source of
information about carcinogenic
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Federal Register / Vol. 61, No. 79 / Tuesday, April 23, 1996 / Notices
Other Key Evidence Factors
Increase Weight
A rich set of other key data are available
Physicochemical data
Data indicate reactivity with
macromolecules
Structure activity relationships support
hazard potential
Comparable metabolism and toxicokinetics
between species
Toxicological and human clinical 'data
support tumor findings
Biomarker data support attribution of
effects to agent
Mode of action data support causal
interpretation of human evidence or
relevance of animal evidence
Decrease Weight
Few or poor data
or
Inadequate data necessitate use of default
assumptions
or
Data show that animal findings are not
relevant to humans
Figure 2-3. Factors for Weighing Other Key Evidence
Totality of Evidence. In reaching a fact, possible weights of evidence span
view of the entire weight of evidence, a broad continuum that cannot be
all data and inferences are merged. capsulized. Most of the time the data in
Figure 2-4 indicates the generalities. In various lines of evidence fall in the
middle of the weights represented in the
four figures in this section.
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17985
Totality of Evidence Factors
Increase Weight
Evidence of human causality
Evidence of animal effects relevant
to humans
Coherent inferences
Comparable metabolism and toxicokinetics
between species
Mode of action comparable across species
Decrease Weight
Data not available or do not show causality
Data not available or not relevant
Conflicting data
Metabolism and toxicokinetics not
comparable
Mode of action not comparable across
species
Figure 2-4. Factors for Weighing Totality of Evidence
Olt-UWa CODE M60-SO-C
The following section and the weight
of evidence narrative discussed in 2.7.2.
provide a way to state a conclusion and
capture this complexity in a consistent
way.
2.6.2. Descriptors for Classifying Weight
of Evidence
Hazard classification uses three
categories of descriptors for human
carcinogenic potential: "known/likely,"
"cannot bo determined," and "not
likely." Each category has associated
subdoscriptors to further define the
conclusion. The descriptors are not
meant to replace an explanation of the
nuances of the biological evidence, but
rather to summarize it. Each category
spans a wide variety of potential data
sots and weights of evidence. There will
always bo gray areas, gradations, and
borderline cases. That is why the
descriptors are presented only in the
context of a weight of evidence narrative
whoso format is given in section 2.7.2.
Using them within a narrative preserves
and presents the complexity that is an
essential part of the hazard
classification. Applying a descriptor is a
matter of judgment and cannot be
reduced to a formula. Risk managers
should consider the entire range of
information included in the narrative
rather than focusing simply on the
descriptor.
A single agent may be categorized in
more than one way if, for instance, the
agent is likely to bo carcinogenic by one
route of exposure but not by another
(section 2.3.3).
The descriptors and subdescriptors
are standardized and are to be used
consistently from case to case. The
discussions below explain descriptors
and subdescriptors which appear in
italics, and along with Appendix A and
section 2.6.3, illustrate their use.
"Known/Likely"
This category of descriptors is
appropriate when the available tumor
effects and other key data are adequate
to convincingly demonstrate
carcinogenic potential for humans; it
includes:
• Agents known to be carcinogenic in
humans based on either epidemiologic
evidence or a combination of
epidemiologic and experimental
evidence, demonstrating causality
between human exposure and cancer,
• Agents that should be treated as if
they were known human carcinogens,
based on a combination of
epidemiologic data showing a plausible
causal association (not demonstrating it
definitively) and strong experimental
evidence.
• Agents that are likely to produce
cancer in humans due to the production
or anticipated production of tumors by
modes of action that are relevant or
assumed to be relevant to human
carcinogenicity.
Modifying descriptors for particularly
high or low ranking in the "known/
likely" group can be applied based on
scientific judgment and experience and
are as follows:
• Agents that are likely to produce
cancer in humans based on data that are
at the high end of the weights of
evidence typical of this group,
• Agents that are likely to produce
cancer in humans based on data that are
at the low end of the weights of
evidence typical of this group.
"Cannot Be Determined"
This category of descriptors is
appropriate when available tumor
effects or other key data are suggestive
or conflicting or limited in quantity and,
thus, are not adequate to convincingly
demonstrate carcinogenic potential for
humans. In general, further agent
specific and generic research and testing
are needed to be able to describe human
carcinogenic potential. The descriptor
cannot be determined is used with a
subdescriptor that captures the
rationale:
• Agents whose carcinogenic
potential cannot be determined, but for
which there is suggestive evidence that
raises concern for carcinogenic effects,
• Agents whose carcinogenic
potential cannot be determined because
the existing evidence is composed of
conflicting data (e.g., some evidence is
suggestive of carcinogenic effects, but
other equally pertinent evidence does
not confirm any concern),
• Agents whose carcinogenic
potential cannot be determined because
there are inadequate data to perform an
assessment,
• Agents whose carcinogenic
potential cannot be determined because
no data are available to perform an
assessment.
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Federal Register / Vol. 61, No. 79 / Tuesday, April 23, 1996 / Notices
"Not Likely"
This is the appropriate descriptor
when experimental evidence is
satisfactory for deciding that there is no
basis for human hazard concern, as
follows (in the absence of human data
suggesting a potential for cancer effects):
• Agents not likely to be carcinogenic
to humans because they have been
evaluated in at least two well conducted
studies in two appropriate animal
species without demonstrating
carcinogenic effects,
• Agents not likely to be carcinogenic
to humans because they have been
appropriately evaluated in animals and
show only carcinogenic effects that have
been shown not to be relevant to
humans (e.g., showing only effects in
the male rat kidney due to accumulation
of alphaau-globulin),
• Agents not likely to be carcinogenic
to humans when carcinogenicity is dose
or route dependent. For instance, not
likely below a certain dose range
(categorized as likely above that range)
or not likely by a certain route of
exposure (may be categorized as likely
by another route of exposure). To
qualify, agents will have been
appropriately evaluated in animal
studies and the only effects show a dose
range or route limitation or a route
limitation is otherwise shown by
empirical data.
• Agents not likely to be carcinogenic
to humans based on extensive human
experience that demonstrates lack of
effect (e.g., phenobarbital).
2.6.3. Case Study Examples
This section provides examples of
substances that fit the three broad
categories described above. These
examples are based on available
information about real substances and
are selected to illustrate the principles
for weight-of-evidence evaluation and
the application of the classification
scheme.
These case studies show the interplay
of differing lines of evidence in making
a conclusion. Some particularly
illustrate the role that "other key data"
can play in conclusions.
Example 1: "Known Human Carcinogen"—
Route-Dependent/Linear Extrapolation
Human Data
Substance 1 is an aluminosilicate mineral
that exists in nature with a fibrous habit.
Several descriptive epidemiologic studies
have demonstrated very high mortality from
malignant mesothelioma, mainly of the
pleura, in three villages in Turkey, where
there was a contamination of this mineral
and where exposure had occurred from birth.
Both sexes were equally affected and at an
unusually young age.
Animal Data
Substance 1 has been studied in a single
long-term inhalation study in rats at one
exposure concentration that showed an
extremely high incidence of pleural
mesothelioma (98% in treated animals versus
0% in concurrent controls). This is a rare
malignant tumor in the rat and the onset of
tumors occurred at a very early age (as early
as 1 year of age). Several studies involving
injection into the body cavities of rats or
mice (i.e., pleural or peritoneal cavities) also
produced high incidences of pleural or
peritoneal mesotheliomas. No information is
available on the carcinogenic potential of
substance 1 in laboratory animals via oral
and dermal exposures.
Other Key Data
Information on the physical and chemical
properties of substance 1 indicates that it is
highly respirable to humans and laboratory
rodents. It is highly insoluble and is not
likely to be readily degraded in biological
fluid.
No information is available on the
deposition, translocation, retention, lung
clearance, and excretion of the substance
after inhalation exposure or ingestion. Lung
burden studies have shown the presence of
elevated levels of the substance in lung tissue
samples of human cases of pleural
mesotheliomas from contaminated villages
compared with control villages.
No data are available on genetic or related
effects in humans. The substance has been
shown to induce unscheduled DNA synthesis
in human cells in vitro and transformation
and unscheduled DNA synthesis in mouse
cells.
The mechanisms by which this substance
causes cancer in humans and animals are not
understood, but appear to be related to its
unique physical, chemical, and surface
properties. Its fiber morphology is similar to
a known group of naturally occurring silicate
minerals that have been known to cause
respiratory cancers (including pleural
mesothelioma) from inhalation exposure and
genetic changes in humans.
Evaluation
Human evidence is judged to establish a
causal link between exposure to substance 1
and human cancer. Even though the human
evidence does not satisfy all criteria for
causality, this judgment is based on a number
of unusual observations: large magnitude of
the association, specificity of the association,
demonstration of environmental exposure,
biological plausibility, and coherence based
on the entire body of knowledge of the
etiology of mesothelioma.
Animal evidence demonstrates a causal
relationship between exposure and cancer in
laboratory animals. Although available data
are not optimal in terms of design (e.g., the
use of single dose, one sex only), the
judgment is based on the unusual findings
from the only inhalation experiment in rats
(i.e., induction of an uncommon tumor, an
extremely high incidence of malignant
neoplasms, and onset of tumors at an early
age). Additional evidence is provided by
consistent results from several injection
studies showing an induction of the same
tumors by different modes of administration
in more than one species.
Other key data, while limited, support the
human and animal evidence of
carcinogenicity. It can be inferred from
human and animal data that this substance
is readily deposited in the respiratory
airways and deep lung and is retained for
extended periods of time since first exposure.
Information on related fibrous substances
indicates that the modes of action are likely
mediated by the physical and chemical
characteristics of the substance (e.g., fiber
shape, high aspect ratio, a high degree of
insolubility in lung tissues).
Insufficient data are available to evaluate
the human carcinogenic potential of
substance 1 by oral exposure. Even though
there is no information on its carcinogenic
potential via dermal uptake, it is not
expected to pose a carcinogenic hazard to
humans by that route because it is very
insoluble and is not likely to penetrate the
skin.
Conclusion
It is concluded that substance 1 is a known
human carcinogen by inhalation exposure.
The weight of evidence of human
carcinogenicity is based on (a) exceptionally.
increased incidence of malignant
mesothelioma in epidemiologic studies of
environmentally exposed human
populations; (b) significantly increased
incidence of malignant mesothelioma in a
single inhalation study in rats and in several
injection studies in rats and mice; and (c)
supporting information on related fibrous
substances that are known to cause cancer
via inhalation and genetic damage in exposed
mammalian and human mesothelial cells.
The human carcinogenic potential of
substance 1 via oral exposure cannot be
determined on the basis of insufficient data.
It is not likely to pose a carcinogenic hazard
to humans via dermal uptake because it is
not anticipated to penetrate the skin.
The mode of action of this substance is not
understood. In addition to this uncertainty,
dose response information is lacking for both
human and animal data. Epidemiologic
studies contain observations of significant
excess cancer risks at relatively low levels of
environmental exposure. The use of linear
extrapolation in a dose response relationship
assessment is appropriate as a default since
mode of action data are not available.
Example 2: "As If Known Human
Carcinogen"—Any Exposure Conditions/
Linear Extrapolation
Human Data
Substance 2 is an alkene oxide. Several
cohort studies of workers using substance 2
as a sterilant have been conducted. In the
largest and most informative study, mortality
from lymphatic and hematopoietic cancer
was marginally elevated, but a significant
trend was found, especially for lymphatic
leukemia and non-Hodgkin's lymphoma, in
relation to estimated cumulative exposure to
the substance. Nonsignificant excesses of
lymphatic and hematopoietic cancer were
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17987
found in threo other smaller studios of
sterilization personnel.
In one cohort study of chemical workers
exposed to substance 2 and other agents,
moclnlity rote from lymphatic and
hcmatopoiclic cancer was elevated, but the
excess was confined to a small subgroup with
only occasional low-level exposure to
substance 2. Six other studios of chemical
workers are considered more limited due to
a smaller number of deaths. Four studies
found an excess of lymphatic and
licmntopolctic cancer (which were significant
in two); no increase in mortality rate was
observed in (lie other two studios.
Animal Data
Substanco 2 was studied in an oral gavage
study in rats. Treatment of substance 2
resulted in a doso-dopondcnt increased
incidence in forcstomach tumors that were
mainly squnmous-coll carcinomas.
Substanco 2 was also studied in two
inhalation studies in mice and two inhalation
studios In rats. In the first mouse study, dose-
dependent increases in combined benign and
malignant tumors at several tissue sites were
induced in mice of both soxos (lung tumors
and tumors of the Hardorian gland in each
sox, and uterine adenocarcinomas, mammary
carcinomas, and malignant lymphomas in
females). In a second study—a screening
study for pulmonary tumors in mice—
Inhalation exposure to substance 2 resulted
in a dose-dependent increase in lung tumors.
In tlio two inhalation studies in rats,
increased incidences of mononuclear-cell
leukemia and brain tumors were induced in
exposed animals of each sex; increased
incidences of peritoneal tumors in the region
of tho tcstls and subcutaneous fibromas were
Induced in exposed male rats.
Substanco 2 induced local sarcomas in
mice following subcutaneous injection. No
tumors wore found in a limited skin painting
study in mice.
Qtlter Key Data
Substance 2 Is a flammable gas at room
temperature. Tho gaseous form is readily
token up in humans and rats, and in aqueous
solution it can penetrate human skin. Studies
in rats indicate that, once absorbed,
substance 2 is uniformly distributed
throughout the body. It is eliminated
melnbollcally by hydrolysis and by
conjugation with glulathiono. Tho ability to
form glutathlono conjugate varies across
animal species, with tho rat being most
active, followed by mice and rabbits.
Substanco 2 is a directly acting alkylating
agent. It has been shown to form adducts
with hemoglobin in both humans and
animals and with DNA in animals. The
increased frequency of hemoglobin adducts,
which havo been used as markers of internal
doso, has been found to correlate with the
lovel and cumulative exposure to substance
2. Significant increases in chromosomal
aberrations and sister chromatid exchanges
in peripheral lymphocytes and induction of
micronuclol in tho bone marrow cells have
been observed in exposed workers.
Substance 2 also induced chromosomal
aberrations and sister chromatid exchanges
In peripheral lymphocytes of monkeys
exposed in vivo. It also induced gene
mutation, specific locus mutation, sister
chromatid exchanges, chromosomal
aberrations, micronuclei, dominant lethal
mutations, and heritable translocation in
rodents exposed in vivo. In human cells in
vitro, it induced sister chromatid exchanges,
chromosomal aberrations, and unscheduled
DNA synthesis. Similar genetic and related
effects were observed in rodent cells in vitro
and in nonmammalian systems.
Evaluation
Available epidemiologic studies, taken
together, suggest that a causal association
between exposure to substance 2 and
elevated risk of cancer is plausible. This
judgment is based on small but consistent
excesses of lymphatic and hematopoietic
cancer in the studies of sterilization workers.
Interpretation of studies of chemical workers
is difficult because of possible confounding
exposures. Nevertheless, findings of elevated
risks of cancer at similar sites in chemical
workers support the findings in studies of
sterilization workers. Additional support is
provided by observations of DNA damage in
the same tissue in which elevated cancer was
seen in exposed workers.
Extensive evidence indicates that
substance 2 is carcinogenic to laboratory
animals. Positive results were consistently
observed in all well-designed and well-
conducted studies. Substance 2 causes dose-
related increased incidences of tumors at
multiple tissue sites in rats and mice of both
sexes by two routes of exposure (oral and
inhalation). The only dermal study that
yielded a nonpositive finding is considered
of limited quality.
Other key data significantly add support to
the potential carcinogenicity of substance 2.
There is strong evidence of heritable
mutations of exposed rodents and
mutagenicity and clastogenicity both in vivo
and in vitro. These findings are reinforced by
observations of similar genetic damage in
exposed workers. Additional support is
based on SAR analysis that indicates that
substance 2 is a highly DNA-reactive agent.
Structurally related chemicals, i.e., low-
molecular-weight epoxides, also exhibit
carcinogenic effects in laboratory animals.
Conclusion
Substance 2 should be considered as if it
were a known human carcinogen by all
routes of exposure. The weight of evidence
of human carcinogenicity is based on (a)
consistent evidence of carcinogenicity in rats
and mice by oral and inhalation exposure; (b)
epidemiologic evidence suggestive of a
causal association between exposure and
elevated risk of lymphatic and hematopoietic
cancer; (c) evidence of genetic damage in
blood lymphocytes and bone marrow cells of
exposed workers; (d) mutagenic effects in
numerous in vivo and in vitro test systems;
(e) membership in a class of DNA-reactive
compounds that have been shown to cause
carcinogenic and mutagenic effects in
animals; and (f) ability to be absorbed by all
routes of exposure, followed by rapid
distribution throughout the body.
Although the exact mechanisms of
carcinogenic action of substance 2 are not
completely understood, available data
strongly indicate a mutagenic mode of action.
Linear extrapolation should be assumed in
dose response assessment.
Example 3: "Likely Human Carcinogen"—
Any Exposure Conditions/Linear
Extrapolation
Human Data
Substance 3 is a brominated alkane. Three
studies have investigated the cancer
mortality of workers exposed to this
substance. No statistically significant
increase in cancer at any site was found in
a study of production workers exposed to.
substance 3 and several other chemicals.
Elevated cancer mortality was reported in a
much smaller study of production workers.
An excess of lymphoma was reported in
grain workers who may have had exposure to
substance 3 and other chemical compounds.
These studies are considered inadequate due
to their small cohort size; lack of, or poorly
characterized, exposure concentrations; or
concurrent exposure of the cohort to other
potential or known carcinogens.
Animal Data
The potential carcinogenicity of substance
3 has been extensively studied in an oral
gavage study in rats and mice of both sexes,
two inhalation studies of rats of different
strains of both sexes, an inhalation study in
mice of both sexes, and a skin painting study
in female mice.
In the oral study, increased incidences of
squamous-cell carcinoma of the forestomach
were found in rats and mice of both sexes.
Additionally, there were increased
incidences of liver carcinomas in female rats,
hemangiosarcomas in male rats, and alveolar/
bronchiolar adenoma of the lung of male and
female mice. Excessive toxicity and mortality
were observed in the rat study, especially in
the high-dose groups, which resulted in early
termination of study, and similar time-
weighted average doses for the high- and
low-treatment groups.
In the first inhalation study in rats and
mice, increased incidences of carcinomas
and adenocarcinomas of the nasal cavity and
hemangiosarcoma of the spleen were found
in exposed animals of each species of both
sexes. Treated female rats also showed
increased incidences of alveolar/bronchiolar
carcinoma of the lung and mammary gland
fibroadenomas. Treated male rats showed an
increased incidence of peritoneal
mesothelioma. In the second inhalation study
in rats (single exposure only), significantly
increased incidences of hemangiosarcoma of
the spleen and adrenal gland tumors were
seen in exposed animals of both sexes.
Additionally, increased incidences of
subcutaneous mesenchymal tumors and
mammary gland tumors were induced in
exposed male and female rats, respectively.
Lifetime dermal application of substance 3
to female mice resulted in significantly
increased incidences of skin papillomas and
lung tumors.
Several chemicals structurally related to
substance 3 are also carcinogenic in rodents.
The spectrum of tumor responses induced by
related substances was similar to those seen
with substance 3 (e.g., forestomach,
mammary gland, lung tumors).
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Other Key Data
Substance 3 exists as a liquid at room
temperature and is readily absorbed by
ingestion, inhalation, and dermal contact. It
is widely distributed in the body and is
eliminated in the urine mainly as metabolites
(e.g., glutathione conjugate).
Substance 3 is not itself DNA-reactive, but
is biotransformed to reactive metabolites as
inferred by findings of its covalent binding to
DNA and induction of DNA strand breaks,
both in vivo and in vitro. Substance 3 has
been shown to induce sister chromatid
exchanges, mutations, and unscheduled DNA
synthesis in human and rodent cells in vitro.
Reverse and forward mutations have been
consistently produced in bacterial assays and
in vitro assays using eukaryotic cells.
Substance 3, however, did not induce
dominant lethal mutations in mice or rats, or
chromosomal aberrations or micronuclei in
bone marrow cells of mice treated in vivo.
Evaluation
Available epidemiologic data are
considered inadequate for an evaluation of a
causal association of exposure to the
substance and excess of cancer mortality due
to major study limitations.
There is extensive evidence that substance
3 is carcinogenic in laboratory animals.
Increased incidences of tumors at multiple
sites have been observed in multiple studies
in two species of both sexes with different
routes of exposure. It induces tumors both at
the site of entry (e.g., nasal tumors via
inhalation, forestomach tumors by ingestion,
skin tumor with dermal exposure) and at
distal sites (e.g., mammary gland tumors).
Additionally, it induced tumors at the same
sites in both species and sexes via different
routes of exposure (e.g., lung tumors). With
the exception of the oral study in which the
employed doses caused excessive toxicity
and mortality, the other studies are
considered adequately designed and well
conducted. Overall, given the magnitude and
extent of animal carcinogenic responses to
substance 3, coupled with similar responses
to structurally related substances, these
animal findings are judged to be highly
relevant and predictive of human responses.
Other key data, while not very extensive,
are judged to be supportive of carcinogenic
potential. Substance 3 has consistently been
shown to be mutagenic in mammalian cells,
including human cells, and nonmammalian
cells; thus, mutation is likely a mode of
action for its carcinogenic activity. However,
the possible involvement of other modes of
action has not been fully investigated.
Furthermore, induction of genetic changes
from in vivo exposure to substance 3 has not
been demonstrated.
Conclusion
Substance 3 is likely to be a human
carcinogen by any route of exposure. In
comparison with other agents designated as
likely human carcinogens, the overall weight
of evidence for substance 3 puts it at the high
end of the grouping.
The weight of evidence of human
carcinogenicity is based on animal evidence
and other key evidence. Human data are
inadequate for an evaluation of human
carcinogenicity. The overall weight of
evidence is based on (a) extensive animal
evidence showing induction of increases of
tumors at multiple sites in both sexes of two
rodent species via three routes of
administration relevant to human exposure;
(b) tumor data of structural analogues
exhibiting similar patterns of tumors in
treated rodents; (c) in vitro evidence for
mutagenic effects in mammalian cells and
nonmammalian systems; and (d) its ability to
be absorbed by all routes of exposure
followed by rapid distribution throughout the
body.
Some uncertainties are associated with the
mechanisms of carcinogenicity of substance
3. Although there is considerable evidence
indicating that mutagenic events could
account for carcinogenic effects, there is still
a lack of adequate information on the
mutagenicity of substance 3 in vivo in
animals or humans. Moreover, alternative
modes of action have not been explored.
Nonetheless, available data indicate a likely
mutagenic mode of action. Linear
extrapolation should be assumed in dose
response assessment.
Example 4: "Likely Human Carcinogen"—All
Routes/Linear and Nonlinear Extrapolation
Human Data
Substance 4 is a chlorinated alkene
solvent. Several cohort studies of dry
cleaning and laundry workers exposed to
substance 4 and other solvents reported
significant excesses of mortality due to
cancers of the lung, cervix, esophagus,
kidney, bladder, lymphatic and
hematopoietic system, colon, or skin. No
significant cancer risks were observed in a
subcohort of one these investigations of dry
cleaning workers exposed mainly to
substance 4. Possible confounding factors,
such as smoking, alcohol consumption, or
low socioeconomic status were not
considered in the analyses of these studies.
A large case-control study of bladder
cancer did not show any clear association
with dry cleaning. Several case-control
studies of liver cancer identified an increased
risk of liver cancer with occupational
exposure to organic solvents. The specific
solvents to which workers were exposed and
exposure levels were not identified.
Animal Data
The potential carcinogenicity of substance
4 has been investigated in two long-term
studies in rats and mice of both sexes by oral
administration and inhalation.
Significant increases in hepatocellujar
carcinomas were induced in mice of both
sexes treated with substance 4 by oral gavage.
No increases in tumor incidence were
observed in treated rats. Limitations in both
experiments included control groups smaller
than treated groups, numerous dose
adjustments during the study, and early
mortality due to treatment-related
nephropathy.
In the inhalation study, there were
significantly increased incidences of
hepatocellular adenoma and carcinoma in
exposed mice of both sexes. In rats of both
sexes, there were marginally significant
increased incidences of mononuclear cell
leukemia (MCL) when compared with
concurrent controls. The incidences of MCL
in control animals, however, were higher
than historical controls from the conducting
laboratory. The tumor finding was also
judged to be biologically significant because
the time to onset of tumor was decreased and
the disease was more severe in treated than
in control animals. Low incidences of renal
tubular cell adenomas or adenocarcinomas
were also observed in exposed male rats. The
tumor incidences were not statistically
significant but there was a significant trend.
Other Key Data
Substance 4 has been shown to be readily
and rapidly absorbed by inhalation and
ingestion in humans and laboratory animals.
Absorption by dermal exposure is slow and
limited. Once absorbed, substance 4 is
primarily distributed to and accumulated in
adipose tissue and the brain, kidney, and
liver. A large percentage of substance 4 is
eliminated unchanged in exhaled air, with
urinary excretion of metabolites comprising a
much smaller percentage. The absorption and
distribution profiles of substance 4 are
similar across species including humans.
Two major metabolites (trichloroacetic acid
(TCA), and trichloroethanol), which are
formed by a P—450-dependent mixed-
function oxidase enzyme system, have been
identified in all studied species, including
humans. There is suggestive evidence for the
formation of an epoxide intermediate based
on the detection of two other metabolites
(oxalic acid and trichloroacetyl amide). In
addition to oxidative metabolism, substance
4 also undergoes conjugation with
glutathione. Further metabolism by renal
beta-lyases could lead to two minor active
metabolites (trichlorovinyl thiol and
dichlorothiokente).
Toxicokinetic studies have shown that the
enzymes responsible for the metabolism of
substance 4 can be saturated at high
exposures. The glutathione pathway was
found to be a minor pathway at low doses,
but more prevalent following saturation of
the cytochrome P-450 pathway. Comparative
in vitro studies indicate that mice have the
greater capacity to metabolize to TCA than
rats and humans. Inhalation studies also
indicate saturation of oxidative metabolism
of substance 4, which occurs at higher dose
levels in mice than in rats and humans.
Based on these findings, it has been
postulated that the species differences in the
carcinogenicity of substance 4 between rats
and mice may be related to the differences in
the metabolism to TCA and glutathione
conjugates.
Substance 4 is a member of the class of
chlorinated organics that often cause liver
and kidney toxicity and carcinogenesis in
rodents. Like many chlorinated organics,
substance 4 itself does not appear to be
mutagenic. Substance 4 was generally
negative in in vitro bacterial systems and in
vivo mammalian systems. However, a minor
metabolite formed in the kidney by the
glutathione conjugation pathway has been
found to be a strong mutagen.
The mechanisms of induced carcinogenic
effects of substance 4 in rats and mice,are not
completely understood. It has been
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17989
postulated that mouse liver carcinogenesis is
related to livor peroxisomal proliferation and
toxicity of tho metabolite TCA. Information
on whether or not TCA induces peroxisomal
proliferation in humans is not definitive. The
induced renal tumors in male rats may be
related either to kidney toxicity or the
activity of a mutogcnic metabolite. The
mechanisms of Increases in MCL in rats are
not known.
Evaluation
Available opidomiologic studios, taken
together, provide suggestive evidence of a
possible causal association between exposure
to substance 4 and cancer incidence in the
laundry and dry cleaning industries. This is
based on consistent findings of elevated
cancer risks in several studies of different
populations of dry cleaning and laundry
workers. However, each individual study is
compromised by a number of study
deficiencies including small numbers of
cancers, confounding exposure to other
solvents, and poor exposure characterization.
Others may interpret these findings
collectively as inconclusive.
Them is considerable evidence that
substance 4 is carcinogenic to laboratory
animals. It induces tumors in mice of both
sexes by oral and inhalation exposure and in
rats of both sexes via inhalation. However,
duo to incomplete understanding of the mode
of mechanism of action, tho predictivity of
animal responses to humans is uncertain.
Animal data of structurally related
compounds showing common target organs
of toxicity and carcinogenic effects (but lack
of mutagenic effects) provide additional
support for the carcinogcnicity of substance
4. Comparative toxicokinctic and metabolism
information indicates that the mouse may be
more susceptible to liver carcinogenesis than
rats and humans. This may indicate
differences of tho degree and extent of
carcinogenic responses, but does not detract
from tho qualitative weight of evidence of
human carcinogonicity. Tho toxicokinctic
information also indicates that oral and
inhalation are tho major routes of human
exposure.
Conclusion
Substance 4 is likely to be carcinogenic to
humans by all routes of exposure. The weight
of evidence of human carcinogcnicity is
based on: (a) Demonstrated evidence of
carcinogonicity in two rodent species of both
sexes via two relevant routes of human
exposure; (b) tho substance's similarity in
structure to other chlorinated organics that
are known to cause liver and kidney toxicity
and cnrcinogenesis in rodents; (c) suggestive
evidence of a possible association between
exposure to the substance in the laundry and
dry cleaning industries and increased cancer
incidence; and (d) human and animal data
indicating that the substance is absorbed by
oil routes of exposure.
In comparison with other agents
designated as likely carcinogens, the overall
weight of evidence places it the lower end of
tho grouping. This is because there is a lack
of good evidence that observed excess cancer
risk in exposed workers is duo solely to
substance 4. Moreover, there is considerable
scientific uncertainty about the human
significance of certain rodent tumors
associated with substance 4 and related
compounds. In this case, the human
relevance of the animal evidence of
carcinogenicity relies on the default
assumption.
Overall, there is not enough evidence to
give high confidence in a conclusion about
any single mode of action; it appears that
more than one is plausible in different rodent
tissues. Nevertheless, the lack of
mutagenicity of substance 4 and its general
growth-promoting effect on high background
tumors as well as its toxicity toward mouse
liver and rat kidney tissue support the view
that the predominant mode is growth-
promoting rather than mutagenic. A
mutagenic contribution to carcinogenicity
due to a metabolite cannot be ruled out. The
dose response assessment should, therefore,
adopt both default approaches, nonlinear and
linear extrapolations. The latter approach is
very conservative since it likely
overestimates risk at low doses in this case,
and is primarily useful for screening
analyses.
Example 5: "Likely/Not Likely Human
Carcinogen"—Range of Dose Limited,
Margin-of-Exposure Extrapolation
Human Data
Substance 5 is a metal-conjugated
phosphonate. No human tumor or toxicity
data exist on this chemical.
Animal Data
Substance 5 caused a statistically
significant increase in the incidence of
urinary bladder tumors in male, but not
female, rats at 30,000 ppm (3%) in the diet
in a'long-term study. Some of these animals
had accompanying urinary tract stones and
toxicity. No bladder tumors or adverse
urinary tract effects were seen in two lower
dose groups (2,000 and 8,000 ppm) in the
same study. A chronic dietary study in mice
at doses comparable to those in the rat study
showed no tumor response or urinary tract
effects. A 2-year study in dogs at doses up
to 40,000 ppm showed no adverse urinary
tract effects.
Other Key Data
Subchronic dosing of rats confirmed that
there was profound development of stones in
the male bladder at doses comparable to
those causing cancer in the chronic study,
but not at lower doses. Sloughing of the
epithelium of the urinary tract accompanied
the stones.
There was a lack of mutagenicity relevant
to carcinogenicity. In addition, there is
nothing about the chemical structure of
substance 5 to indicate DNA-reactivity or
carcinogenicity.
Substance 5 is composed of a metal,
ethanol, and a simple phosphorus-oxygen-
containing component. The metal is not
absorbed from the gut, whereas the other two
components are absorbed. At high doses,
ethanol is metabolized to carbon dioxide,
which makes the urine more acidic; the
phosphorus level in the blood is increased
and calcium in the urine is increased.
Chronic testing of the phosphorus-oxygen-
containing component alone in rats did not
show any tumors or adverse effects on the
urinary tract.
Because substance a is a metal complex, it
is not likely to be readily absorbed from the
skin.
Evaluation
Substance 5 produced cancer of the
bladder and urinary tract toxicity in male,
but not female rats and mice, and dogs failed
to show the toxicity noted in male rats. The
mode of action developed from the other key
data to account for the toxicity and tumors
in the male rats is the production of bladder
stones. At high but not lower subchronic
doses in the male rat, substance 5 leads to
elevated blood phosphorus levels; the body
responds by releasing excess calcium into the
urine. The calcium and phosphorus combine
in the urine and precipitate into multiple
stones in the bladder. The stones are very
irritating to the bladder; the bladder lining is
eroded, and cell proliferation occurs to
compensate for the loss of the lining. Cell
layers pile up, and finally, tumors develop.
Stone formation does not involve the
chemical per se but is secondary to the
effects of its constituents on the blood and,
ultimately, the urine. Bladder stones,
regardless of their cause, commonly produce
bladder tumors in rodents, especially the
male rat.
Conclusion
Substance 5, a metal aliphatic
phosphonate, is likely to be carcinogenic to
humans only under high-exposure conditions
following oral and inhalation exposure that
lead to bladder stone formation, but is not
likely to be carcinogenic under low-exposure
conditions. It is not likely to be a human
carcinogen via the dermal route, given that
the compound is a metal conjugate that is
readily ionized and its dermal absorption is
not anticipated. The weight of evidence is
based on (a) bladder tumors only in male
rats; (b) the absence of tumors at any other
site in rats or mice; (c) the formation of
calcium-phosphorus-containing bladder
stones in male rats at high, but not low,
exposures that erode bladder epithelium and
result in profound increases in cell
proliferation and cancer; and (d) the absence
of structural alerts or mutagenic activity.
There is a strong mode of action basis for
the requirements of (a) high doses of
substance 5, (b) which lead to excess calcium
and increased acidity in the urine, (c) which
result in the precipitation of stones and (d)
the necessity of stones for toxic effects and
tumor hazard potential. Lower doses fail to
perturb urinary constituents, lead to stones,
produce toxicity, or give rise to tumors.
Therefore, dose response assessment should
assume nonlinearity.
A major uncertainty is whether the
profound effects of substance 5 may be
unique to the rat. Even if substance 5
produced stones in humans, there is only
limited evidence that humans with bladder
stones develop cancer. Most often human
bladder stones are either passed in the urine
or lead to symptoms resulting in their
removal. However, since one cannot totally
dismiss the male rat findings, some hazard
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potential may exist in humans following
intense exposures. Only fundamental
research could illuminate this uncertainty.
Example 6: "Cannot Be Determined"—
Suggestive Evidence
Human Data
Substance 6 is an unsaturated aldehyde. In
a cohort study of workers in a chemical plant
exposed to a mixture of chemicals with
substance 6 as a minor component, an
elevated risk of cancer than was expected
was reported. This study is considered-
inadequate because of multiple exposures,
small cohort, and poor exposure
characterization.
Animal Data
Substance 6 was tested for potential
carcinogenicity in a drinking water study in
rats, an inhalation study in hamsters, and a
skin painting study in mice. No significant
increases in tumors were observed in male
rats treated with substance 6 at three dose
levels in drinking water. However, a
significant increase of adrenal cortical
adenomas was found in the only treated
female dose group administered a dose
equivalent to the high dose of males. This
study used a small number of animals (20 per
dose group).
No significant finding was detected in the
inhalation study in hamsters. This study is
inadequate due to the use of too few animals,
short duration of exposure, and inappropriate
dose selection (use of a single exposure that
was excessively toxic as reflected by high
mortality).
No increase in tumors was induced in the
skin painting study in mice. This study is of
inadequate design for carcinogenicity
evaluation because of several deficiencies:
small number of animals, short duration of
exposure, lack of reporting about the sex and
age of animals, and purity of test material.
Substance 6 is structurally related to
lowmolecularweight aldehydes that generally
exhibit carcinogenic effects in the respiratory
tracts of laboratory animals via inhalation
exposure. Three skin painting studies in mice
and two subcutaneous injection studies of
rats and mice were conducted to evaluate the
carcinogenic potential of a possible
metabolite of substance 6 (identified in vitro).
Increased incidences of either benign or
combined benign and malignant skin tumors
were found in the dermal studies. In the
injection studies of rats and mice, increased
incidences of local sarcomas or squamous
cell carcinoma were found at the sites of
injection. All of these studies are limited by
the small number of test animals, the lack of
characterization of test material, and the use
of single doses.
Other Key Data
Substance 6 is a flammable liquid at room
temperature. Limited information on its
toxicokinetics indicates that it can be
absorbed by all routes of exposure. It is
eliminated in the urine mainly as glutathione
conjugates. Substance 6 is metabolized in
vitro by rat liver and lung microsomal
preparations to a dihydroxylated aldehyde.
No data were available on the genetic and
related effects of substance 6 in humans. It
did not induce dominant lethal mutations in
mice. It induced sister chromatid exchanges
in rodent cells in vitro. The mutagenicity of
substance 6 is equivocal in bacteria. It did
not induce DNA damage or mutations in
fungi.
Evaluation
Available human data are judged
inadequate for an evaluation of any causal
relationship between exposure to substance 6
and human cancer.
The carcinogenic potential of substance 6
has not been adequately studied in laboratory
animals due to serious deficiencies in study
design, especially the inhalation and dermal
studies. There is some evidence of
carcinogenicity in the drinking water study
in female rats. However, the significance and
predictivity of that study to human response
are uncertain since the finding is limited to
occurrence of benign tumors, one sex, and at
the high dose only. Additional suggestion for
animal carcinogenicity comes from
observation that a possible metabolite is
carcinogenic'at the site of administration.
This metabolite, however, has not been
studied in vivo. Overall, the animal evidence
is judged to be suggestive for human
carcinogenicity.
Other key data, taken together, do not add
significantly to the overall weight of evidence
of carcinogenicity. SAR analysis indicates
that substance 6 would be DNA-reactive.
However, mutagenicity data are inconclusive.
Limited in vivo data do not support a
mutagenic effect. While there is some
evidence of DNA damage in rodent cells in
vitro, there is either equivocal or no evidence
of mutagenicity in nonmammalian systems.
Conclusion
The human carcinogenicity potential of
substance 6 cannot be determined on the
basis of available information. Both human
and animal data are judged inadequate for an
evaluation. There is evidence suggestive of
potential carcinogenicity on the basis of
limited animal findings and SAR
considerations. Data are not sufficient to
judge whether there is a mode of
carcinogenic action. Additional studies are
needed for a full evaluation of the potential
carcinogenicity of substance 6. Hence, dose
response assessment is not appropriate.
Example 7: "Not Likely Human
Carcinogen"—Appropriately Studied
Chemical in Animals Without Tumor Effects
Human Data
Substance 7, a plant extract, has not been
studied for its toxic or carcinogenic potential
in humans.
Animal Data
Substance 7 has been studied in four
chronic studies in three rodent species. In a
feeding study in rats, males showed a
nonsignificant increase in benign tumors of
the parathyroid gland in the high-dose group,
where the incidence in concurrent controls
greatly exceeded the historical control range.
Females demonstrated a significant increase
in various subcutaneous tumors in the low-
dose group, but findings were not confirmed
in the high-dose group, and there was no
dose response relationship. These effects
were considered as not adding to the
evidence of carcinogenicity. No tumor
increases were noted in a second adequate
feeding study in male and female rats. In a
mouse feeding study, no tumor increases
were noted in dosed animals. There was
some question as to the adequacy of the
dosing; however it was noted that in the •
mouse 90-d subchronic study, a dose'of twice
the high dose in the chronic study led to
significant decrements in body weight. In a
hamster study there were no significant
increases in tumors at any site. No structural
analogues of substance 7 have been tested for
cancer.
Other Key Data
There are no structural alerts that would
suggest that substance 7 is a DNA-reactive
compound. It is negative for gene mutations
in bacteria and yeast, but positive in cultured
mouse cells. Tests for structural chromosome
aberrations in cultured mammalian cells and
in rats are negative; however, the animals
were not tested at sufficiently high doses.
Substance 7 binds to proteins of the cell
division spindle; therefore, there is some
likelihood for producing numerical
chromosome aberrations, an endpoint that is
sometimes noted in cancers. In sum, there is
limited and conflicting information
concerning the mutagenic potential of the
agent.
The compound is absorbed via oral and
inhalation exposure but only poorly via the
skin.
Evaluation
The only indication of a carcinogenic effect
comes from the finding of benign tumors in
male rats in a single study. There is no
confirmation of a carcinogenic potential from
dosed females in that study, in males and
females in a second rat study, or from mouse
and hamster studies.
There is no structural indication that
substance 7 is DNA-reactive, there is
inconsistent evidence of gene mutations, and
chromosome aberration testing is negative.
The agent binds to cell division spindle
proteins and may have the capacity to induce
numerical chromosome anomalies. Further
information on gene mutations and in vivo
structural and numerical chromosome
aberrations may be warranted.
Conclusion
Substance 7 is not likely to be carcinogenic
to humans via all relevant routes of exposure.
This weight of evidence judgment is largely
based on the absence of significant tumor
increases in chronic rodent studies. Adequate
cancer studies in rats, mice, and hamsters fail
to show any carcinogenic effect; a second rat
study showed an increase in benign tumors
at a site in dosed males, but not females.
2.7. Presentation of Results
The results of the hazard assessment
are presented in the form of an overall
technical hazard characterization.
Additionally, a weight of evidence
narrative is used when the conclusion
as to carcinogenic potential needs to be
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17991
presented separately from the overall
characterization.
2.7.1. Technical Hazard
Characterization
The hazard characterization has two
functions. First, it presents results of the
hazard assessment and an explanation
of how the weight of evidence
conclusion was reached. It explains the
potontial for human hazard, anticipated
attributes of its expression, and mode of
action considerations for dose response.
Second, it contains the information
needed for eventual incorporation into a
risk characterization consistent with
EPA guidance on risk characterization
(U.S. EPA, 1995).
The characterization qualitatively
describes the conditions under which
the agent's effects may be expressed in
human beings. These qualitative hazard
conditions are ones that are observable
in tho toxicity data without having done
either quantitative dose response or
exposure assessment. The description
includes how expression is afffected by
route of exposure and dose levels and
durations of exposure.
Tho discussion of limitations of dose
as a qualitative aspect of hazard
addresses the question of whether
reaching a certain dose range appears to
bo a precondition for a hazard to be
expressed; for example, when
carcinogenic effects are secondary to
another toxic effect that appears only
when a certain dose level is reached.
Tho assumption is made that an agent
that causes internal tumors by one route
of exposure will be carcinogenic by
another route, if it is absorbed by the
second route to give an internal dose.
Conversely, if there is a route of
exposure by which the agent is not
absorbed (does not cross an absorption
barrier; e.g., the exchange boundaries of
skin, lung, and digestive tract through
uptake processes) to any significant
degree, nazard is not anticipated by that
route. An exception to the latter
statement would be when the site of
contact is also the target tissue of
carcinogenic!ty. Duration of exposure
may be a precondition for hazard if, for
example, tho mode of action requires
cy totoxicity or a physiologic change, or
is mitogonicity, for which exposure
must be sustained for a period of tune
before effects occur. The
characterization could note that one
would not anticipate a hazard from
isolated, acute exposures. The above
conditions are qualitative ones
regarding preconditions for effects, not
issues of relative absorption or potency
at different dose levels. The latter are
dealt with under dose response
assessment (section 3), and their
implications can only be assessed after
human exposure data are applied in the
characterization of risk.
The characterization describes
conclusions about mode of action
information and its support for
recommending dose response
approaches.
The hazard characterization routinely
includes the following in support of risk
characterization:
• a summary of results of the
assessment,
• identification of the kinds of data
available to support conclusions and
explanation of how the data fit together,
highlighting the quality of the data in
each line of evidence, e.g., tumor effects,
short-term studies, structure-activity
relationships), and highlighting the
coherence of inferences from the
different kinds of data,
• strengths and limitations
(uncertainties) of the data and
assessment, including identification of
default assumptions invoked in the face
of missing or inadequate data,
• identification of alternative
interpretations of data that are
considered equally plausible,
• identification of any
subpopulations believed to be more
susceptible to the hazard than the
general population,
• conclusions about the agent's mode
of action and recommended dose
response approaches,
• significant issues regarding
interpretation of data that arose in the
assessment. Typical ones may include:
—determining causality in human
studies,
•—dosing (MTD), background tumor
rates, relevance of animal tumors to
humans,
—weighing studies with positive and
null results, considering the influence
of other available kinds of evidence,
—drawing conclusions based on mode
of action data versus using a default
assumption about the mode of action.
2.7.2. Weight of Evidence Narrative
The weight of evidence narrative
summarizes the results of hazard
assessment employing the descriptors
defined in section 2.6.1. The narrative
(about two pages in length) explains an
agent's human carcinogenic potential
and the conditions of its expression. If
data do not allow a conclusion as to
carcinogenicity, the narrative explains
the basis of this determination. An
example narrative appears below. More
examples appear in Appendix A.
The items regularly included in a
narrative are:
• name of agent and Chemical
Abstracts Services number, if available,
• conclusions (by route of exposure)
about human carcinogenicity, using a
standard descriptor from section 2.6.1,
• summary of human and animal
tumor data on the agent or its structural
analogues, their relevance, and
biological plausibility,
• other key data (e.g., structure-
activity data, toxicokinetics and
metabolism, short-term studies, other,
relevant toxicity or clinical data),
• discussion of possible mode(s) of
action and appropriate dose response
approach(es),
• conditions of expression of
carcinogenicity, including route,
duration, and magnitude of exposure.
Example Narrative
Aromatic Compound
CASSXXX
CANCER HAZARD SUMMARY
Aromatic compound (AR) is known to be
carcinogenic to humans by all routes of
exposure.
The weight of evidence of human
carcinogenicity is based on (a) consistent
evidence of elevated leukemia incidence in
studies of exposed workers and significant
increases of genetic damage in bone marrow
cells and blood lymphocytes of exposed
workers; (b) significantly increased incidence
of cancer in both sexes of several strains of
rats and mice; (c) genetic damage in bone
marrow cells of exposed rodents and effects
on intracellular signals that control cell
growth.
AR is readily absorbed by all routes of
exposure and rapidly distributed throughout
the body. The mode of action of AR is not
understood. A dose response assessment that
assumes linearity of the relationship is
recommended as a default.
SUPPORTING INFORMATION
Data include numerous human
epidemiologic and biomonitoring studies,
long-term bioassays, and other data on effects
of AR on genetic material and cell growth
processes. The key epidemiologic studies and
animal studies are well conducted and
reliable. The other data are generally of good
quality also.
Human Effects
Numerous epidemiologic and case studies
have reported an increased incidence or a
causal relationship associating exposure to
AR and leukemia. Among the studies are five
for which the design and performance as well
as follow-up are considered adequate to
demonstrate the causal relationship.
Biomonitoring studies of exposed workers
have found dose-related increases in
chromosomal aberrations in bone marrow
cells and blood lymphocytes.
Animal Effects
AR caused increased incidence of tumors
in various tissues in both sexes of several rat
and mouse strains. AR also caused
chromosomal aberrations in rabbits, mice,
and rats—as it does in humans.
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Other Key Data
AR itself is not DNA-reactive and is not
mutagenic in an array of test systems both in
vitro and in vivo. Metabolism of AR yields
several metabolites that have been separately
studied for effects on carcinogenic processes.
Some have mutagenic activity in test systems
and some have other effects on cell growth
controls inside cells.
MODE OF ACTION
No rodent tumor precisely matches human
leukemia in pathology. The closest parallel is
a mouse cancer of blood-forming tissue.
Studies of the effects of AR at the cell level
in this model system are ongoing. As yet, the
mode of action of AR is unclear, but most
likely the carcinogenic activity is associated
with one or a combination of its metabolites.
It is appropriate to apply a linear approach
to the dose response assessment pending a
better understanding because: (a) genetic
damage is a typical effect of AR exposure in
mammals and (b) metabolites of AR produce
mutagenic effects in addition to their other
effects on cell growth controls; AR is a
multitissue carcinogen in mammals
suggesting that it is affecting a common
controlling mechanism of cell growth.
3. Dose Response Assessment
Dose response assessment first
addresses the relationship of dose2 to
the degree of response observed in an
experiment or human study. When
environmental exposures are outside of
the range of observation, extrapolations
are necessary in order to estimate or
characterize the dose relationship (ILSI,
1995). In general, three extrapolations
may be made: from high to low doses,
from animal to human responses, and
from one route of exposure to another.
The dose response assessment
proceeds in two parts. The first is
assessment of the data in the range of
empirical observation. This is followed
by extrapolations either by modeling, if
there are sufficient data to support a
model, or by a default procedure based
as much as possible on information
about the agent's mode of action. The
following discussion covers the
assessment of observed data and
extrapolation procedures, followed by
sections on analysis of response data
and analysis of dose data. The final
section discusses dose response
characterization.
2 For this discussion, "exposure" means contact
of an agent with the outer boundary of an organism.
"Applied dose" means the amount of an agent
presented to an absorption barrier and available for
absorption. "Internal dose" means the amount
crossing an absorption barrier (e.g., the exchange
boundaries of skin, lung, and digestive tract)
through uptake processes. "Delivered dose" for an
organ or cell means the amount available for
interaction with that organ or cell (U.S. EPA,
1992a).
3.1. Dose Response Relationship
In the discussion that follows,
reference to "response" data includes
measures of tumorigenicity as well as
other responses related to
carcinogenicity. The other responses
may include effects such as changes in
DNA, chromosomes, or other key
macromolecules, effects on growth
signal transduction, induction of
physiological or hormonal changes,
effects on cell proliferation, or other
effects that play a role in the process.
Responses other than tumorigenicity
may be considered part of the observed
range in order either to extend the
tumor dose response analysis or to
inform it. The nontumor response or
responses also may be used in lieu of
tumor data if they are considered to be
a more informative representation of the
carcinogenic process for an agent (see
section 3.2).
3.1.1. Analysis in the Range of
Observation
Biologically Based and Case-Specific
Models. A biologically based model is
one whose parameters are calculated
independently of curve-fitting of tumor
data. If data are sufficient to support a
biologically based model specific to the
agent and the purpose of the assessment
is such as to justify investing resources
supporting use, this is the first choice
for both the observed tumor and related
response data and for extrapolation
below the range of observed data in
either animal or human studies.
Examples are the two-stage models of
initiation plus clonal expansion and
progression developed by Moolgavkar
and Knudson (1981) and Chen and
Farland (1991). Such models require
extensive data to build the form of the
model as well as to estimate how well
it conforms with the observed
carcinogenicity data. Theoretical
estimates of process parameters, such as
cell proliferation rates, are not used to
enable application of such a model
(Portier, 1987).
Similarly preferred as a first choice
are dose response models based on
general concepts of mode of action and
data on the agent. For a case-specific
model, model parameters and data are
obtained from studies on the, agent.
In most cases, a biologically based or
case-specific model will not be
practicable, either because the necessary
data do not exist or the decisions that
the assessment are to support do not
justify or permit, the time and resources
required. In these cases, the analysis
proceeds using curve-fitting models
followed by default procedures for
extrapolation, based, to the extent
possible, on mode of action and other
biological information about the agent.
These methods and assumptions are
described below.
Curve-Fitting and Point of Departure
for Extrapolation. Curve-fitting models
are used that are appropriate to the kind
of response data in the observed range.
Any of several models can be used; e.g.,
the models developed for benchmark
dose estimation for noncancer
endpoints may be applied (Barnes et al.,
1995).
For some data sets, particularly those
with extreme curvature, the impact of
model selection can be significant. In
these cases, the choice is rationalized on
biological grounds as possible. In other
cases, the nature of the data or the way
it is reported will suggest other types of
models; for instance, when longitudinal
data on tumor development are
available, time to tumor or survival
models may be necessary and
appropriate to fit the data.
A point of departure for extrapolation
is estimated. This is a point that is
either a data point or an estimated point
that can be considered to be in the range
of observation, without any significant
extrapolation. The LEDio—the lower
95% confidence limit on a dose
associated"with 10% extra risk—is such
a point and is the standard point of
departure, adopted as a matter of
science policy to remain as consistent
and comparable from case to case as
possible.3 It is also a comparison point
for noncancer endpoints (U.S. EPA,
1991f). The central estimate of the EDi0
also may be appropriate for use in
relative hazard and potency ranking.
For some data sets, a choice of point
of departure other than the LEDio may
be appropriate. For example, if the
observed response is below the LEDio,
then a lower point may be a better
choice. Moreover, some forms of data
may not be amenable to curve-fitting
estimation, but to estimation of a
"low-" or "no-observable-adverse-effect
level" (LOAEL, NOAEL) instead, e.g.,
certain continuous data.
The rationale supporting the use of
the LEDio is that a 10% response is at
or just below the limit of sensitivity of
discerning a significant difference in
most long-term rodent studies. The
lower confidence limit on dose is used
to appropriately account for
experimental uncertainty (Barnes et al.,
1995) and for consistency with the
"benchmark dose" approach for
noncancer assessment; it does not
provide information about human
3 It is appropriate to report the central estimate of
the ED 10, the upper and lower 95% confidence
limits, and a graphical representation of model fit.
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17993
variability. In laboratory studies of
cancer or noncancer endpoints, the level
of doso at which increased incidence of
effects can bo detected, as compared to
controls, is a function of the size of the
sample (e.g., number of animals), dose
spacing, and other design aspects. In
noncancer assessment, the dose at
which significant effects are not
observed is traditionally termed the
NOAEL. This is not, in fact, a level of
zero effect. The NOAEL in most study
protocols is about the same as an LEDS
orLEDio—the lower 95% confidence
limit on a dose associated with a 5% or
10% increased effect (Faustman et al.,
1094; Haseman, 1983). Adopting
parallel points of departure for cancer
and noncancor assessment is intended
to mako discussion and comparison of
tho two kinds of assessment more
comparable because of their similar
science and science policy bases and
similar analytic approaches.
Analysis of human studies in the
observed range is designed case by case,
depending on the type of study and how
doso and response are measured in the
study. In some cases the agent may have
discernible interactive effects with
another agent (e.g., asbestos and
smoking), making possible estimation of
contribution of the agent and others as
risk factors. Also, in some cases,
estimation of population risk in
addition, or in lieu of, individual risk
may bo appropriate.
3.1.2. Analysis in the Range of
Extrapolation
Extrapolation to lower doses is
usually necessary, and in the absence of
a biologically based or case-specific
model, is based on one of the three
default procedures described below.
Tho Agency has adopted these three
procedures as a matter of science policy
based on current hypotheses of the
likely shapes of dose response curves
for differing modes of action. The choice
of tho procedure to be used in an
individual case is a judgment based on
tho agent's modes of action.
Linear, A default assumption of
linearity is appropriate when the
evidence supports a mode of action of
gone mutation due to DNA reactivity or
supports another mode of action that is
anticipated to be linear. Other elements
of empirical support may also support
an inference of linearity, e.g., the
background of human exposure to an
agent might be such that added human
exposure is on the linear part of a dose
response curve that is sublinear overall.
Tho default assumption of linearity is
also appropriate as the ultimate science
policy default \vhon evidence shows no
DNA reactivity or other support for
linearity, but neither does it show
sufficient evidence of a nonlinear mode
of action to support a nonlinear
procedure.
For linear extrapolation, a straight
line is drawn from the point of
departure to the origin—zero dose, zero
response (Flamm and Winbush, 1984;
Gaylor and Kodell, 1980; Krewski et al.,
1984). This approach is generally
conservative of public health, in the
absence of information about the extent
of human variability in sensitivity to
effects. When a linear extrapolation
procedure is used, the risk
characterization summary displays the
degree of extrapolation that is being
made from empirical data and discusses
its implications for the interpretation of
the resulting quantitative risk estimates.
Nonlinear. A default assumption of
nonlinearity is appropriate when there
is no evidence for linearity and
sufficient evidence to support an
assumption of nonlinearity. The mode
of action may lead to a dose response
relationship that is nonlinear, with
response falling much more quickly
than linearly with dose, or being most
influenced by individual differences in
sensitivity. Alternatively, the mode of
action may theoretically have a
threshold, e.g., the carcinogenicity may
be a secondary effect of toxicity or of an
induced physiological change (see
example 5, section 2.6.3) that is itself a
threshold phenomenon.
As a matter of science policy under
this analysis, nonlinear probability
functions are not fitted to the response
data to extrapolate quantitative low-
dose risk estimates because different
models can lead to a, very wide range of
results, and there is currently no basis,
generally, to choose among them.
Sufficient information to choose leads to
a biologically based or case-specific
model. In cases of nonlinearity, the risk
is not extrapolated as a probability of an
effect at low doses. A margin of
exposure analysis is used, as described
below, to evaluate concern for levels of
exposure. The margin of exposure is the
LEDio or other point of departure
divided by the environmental exposure
of interest. The EPA does not generally
try to distinguish between modes of
action that might imply a "true
threshold" from others with a nonlinear
dose response relationship. Except in
unusual cases where extensive
information is available, it is not
possible to distinguish between these
empirically.
The environmental exposures of
interest, for which margins of exposure
are estimated, may be actual or
projected future levels. The risk
manager decides whether a given
margin of exposure is acceptable under
applicable management policy criteria.
The risk assessment provides
supporting information to assist the
decisionmaker.
The EPA often conducts margin of
exposure analyses to accompany
estimates of reference doses or
concentrations (RfD, RfC) for noncancer
endpoints.4 The procedure for a margin
of exposure analysis for a response
related to carcinogenicity is
operationally analogous, the difference
being that a threshold of cancer
response is not necessarily presumed. If,
in a particular case, the evidence
indicates a threshold, as in the case of
carcinogenicity being secondary to
another toxicity that has a threshold, the
margin of exposure analysis for the
toxicity is the same as is done for a
noncancer endpoint, and an RfD or RfC
for that toxicity also may be estimated
and considered in cancer assessment.
The analogy between margin of
exposure analysis for noncancer and
cancer responses begins with the
analogy of points of departure; for both
it is an effect level, either LEDio or other
point (presented as a human equivalent
dose or concentration), as data support.
For cancer responses, when animal data
are used, the point of departure is a
human equivalent dose or concentration
arrived at by interspecies dose
adjustment or toxicokinetic analysis. It
is likely that many of the margin of
exposure analyses for cancer will be for
responses other than tumor incidence.
This is because the impetus for
considering a carcinogenic agent to have
a nonlinear dose response will be a
conclusion that there is sufficient
evidence to support that view, and this
evidence will often be information
about a response that is a precursor to
tumors.
To support a risk manager's
consideration of the margin of exposure,
information is provided in a risk
assessment about current understanding
of the phenomena that may be occurring
as dose (exposure) decreases
substantially below the observed data.
The goal is to provide as much
information as possible about the risk
reduction that accompanies lowering of
exposure. To this end, some important
points to address include:
4 An RfD or RfC is an estimate with uncertainty
spanning perhaps an order of magnitude of daily
exposure to the human population (including
sensitive subgroups) that is anticipated to be
without appreciable deleterious effects during a
lifetime. It is arrived at by dividing empirical data
on effects by uncertainty factors that consider inter-
and intraspecies variability, extent of data on all
important chronic exposure toxicity endpoints, and
availability of chronic as opposed to subchronic
data.
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• The slope of the observed dose
response relationship at the point of
departure and its uncertainties and
implications for risk reduction
associated with exposure reduction (a
shallow slope suggests less reduction
than a steep slope),
• The nature of the response used for
the dose response assessment,
• The nature and extent of human
variability in sensitivity to the
phenomena involved,
• Persistence of the agent in the body,
• Human sensitivity to the
phenomena as compared with
experimental animals.
As a default assumption for two of
these points, a factor of no less than 10-
fold each may be employed to account
for human variability and for
interspecies differences in sensitivity
when humans may be more sensitive
than animals. When humans are found
to be less sensitive than animals, a
default factor of no smaller than a 1/10
fraction may be employed to account for
this. If any information about human
variability or interspecies differences is
available, it is used instead of the
default or to modify it as appropriate. In
the case of analysis based on human
studies, obviously, interspecies
differences are not a factor. It should be
noted that the dose response
relationship and inter- or intraspecies
variability in sensitivity are
independent. That is, reduction of dose
reduces risk; it does not change
variability. To support consideration of
acceptability of a margin of exposure by
the risk manager, the assessment
considers all of the hazard and dose
response factors together; hence, the
factors for inter- and intraspecies
differences alone are not to be
considered a default number for an
acceptable margin of exposure. (See
Section 1.3.2.5.)
It is appropriate to provide a graphical
representation of the data and dose
response modeling in the observed
range, also showing exposure levels of
interest to the decisionmaker. (See
figure 3—1.) In order to provide a frame
of reference, by way of comparison, a
straight line extrapolation may be
displayed to show what risk levels
would be associated with decreasing
dose, if the dose response were linear.
If this is done, the clear accompanying
message is that, in this case of
nonlinearity, the response falls
disproportionately with decreasing
dose.
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Linear and Nonlinear. Both linear and
nonlinear procedures may be used in
particular cases. If a mode of action
analysis finds substantial support for
differing modes of action for different
tumor sites, an appropriate procedure is
used for each. Both procedures may also
be appropriate to discuss implications
of complex dose response relationships.
For example, if it is apparent that an
agent is both DNA reactive and is highly
active as a promotor at high doses, and
there are insufficient data for modeling,
both linear and nonlinear default
procedures may be needed to decouple
and consider the contribution of both
phenomena.
3.1.3. Use of Toxicity Equivalence
Factors and Relative Potency Estimates
A toxicity equivalence factor (TEF)
procedure is one used to derive
quantitative dose response estimates for
agents that are members of a category or
class of agents. TEFs are based on
shared characteristics that can be used
to order the class members by
carcinogenic potency when cancer
bioassay data are inadequate for this
purpose (U.S. EPA, 1991c). The ordering
is by reference to the characteristics and
potency of a well-studied member or
members of the class. Other class
members are indexed to the reference
agent(s) by one or more shared
characteristics to generate their TEFs.
The TEFs are usually indexed at
increments of a factor of 10. Very good
data may permit a smaller increment to
be used. Shared characteristics that may
be used are, for example, receptor-
binding characteristics, results of assays
of biological activity related to
carcinogenicity, or structure-activity
relationships.
TEFs are generated and used for the
limited purpose of assessment of agents
or mixtures of agents in environmental
media when better data are not
available. When better data become
available for an agent, its TEF should be
replaced or revised. Criteria for
constructing TEFs are given in U.S. EPA
(1991b). The criteria call for data that
are adequate to support summing doses
of the agents in mixtures. To date,
adequate data to support use of TEF's
has been found in only one class of
compounds (dioxins) (U.S. EPA, 1989a).
Relative potencies can be similarly
derived and used for agents with
carcinogenicity or other supporting
data. These are conceptually similar to
TEFs, but they are less firmly based in
science and do not have the same level
of data to support them. They are used
only when there is no better alternative.
The uncertainties associated with
both TEFs and relative potencies are
explained whenever they are used.
3.2, Response Data
Response data for analysis include
tumor incidence data from human or
animal studies as well as data on other
responses as they relate to an agent's
carcinogenicity, such as effects on
growth control processes or cell
macromolecules or other toxic effects.
Tumor incidence data are ordinarily the
basis of dose response assessment, but
other response data can augment such
assessment or provide separate
assessments of carcinogenicity or other
important effects.
Data on carcinogenic processes
underlying tumor effects may be used to
support biologically based or case-
specific models. Other joptions for such
data exist. If confidence is high in the
linkage of a precursor effect and the
tumor effect, the assessment of tumor
incidence may be extended to lower
dose levels by linking it to the
assessment of the precursor effect
(Swenberg et al., 1987). Even if a
quantitative link is not appropriate, the
assessment for a precursor effect may
provide a view of the likely shape of the
dose response curve for tumor incidence
below the range of tumor observation
(Cohen and Ellwein, 1990; Choy, 1993).
If responses other than tumor incidence
are regarded as better representations of
the carcinogenicity of the agent, they
may be used in lieu of tumor responses.
For example, if it is concluded that the
carcinogenic effect is secondary to
another toxic effect, the dose response
for the other effect will likely be more
pertinent for risk assessment. As
another example, if disruption of
hormone activity is the key mode of
action of an agent, data on hormone
activity may be used in lieu of tumor
incidence data.
If adequate positive human
epidemiologic response data are
available, they provide an advantageous
basis for analysis since concerns about
interspecies extrapolation do not arise.
Adequacy of human exposure data for
quantification is an important
consideration in deciding whether
epidemiologic data are the best basis for
analysis in a particular case. If adequate
exposure data exist in a well-designed
and well-conducted epidemiologic
study that detects no effects, it may be
possible to obtain an upper-bound
estimate of the poteiitial human risk to
provide a check on plausibility of
available estimates based on animal
tumor or other responses, e.g., do
confidence limits on one overlap the
point estimate of the other?
When animal studies are used,
response data from a species that
responds most like humans should be
used if information to this effect exists.
If this is unknown and an agent has
been tested in several experiments
involving different animal species,
strains, and sexes at several doses and
different routes of exposure, all of the
data sets are considered and compared,
and a judgment is made as to the data
to be used to best represent the observed
data and important biological features
such as mode of action. Appropriate
options for presenting results include:
• Use of a single data set,
• Combining data from different
experiments (Stiteler et al., 1993; Vater
et aL, 1993),
• Showing a range of results from
more than one data set,
• Showing results from analysis of
more than one statistically significant
tumor response based on differing
modes of action,
• Representing total response in a~~~
single experiment by combining animals
with statistically significant tumors at
more than one site, or
• A combination of these options.
The approach judged to best represent
the data is presented with the rationale
for the judgment, including the
biological and statistical considerations
involved. The following are some points
to consider:
• Quality of study protocol and
execution,
• Proportion of malignant neoplasms,
• Latency of onset of neoplasia,
• Number of data points to define the
relationship of dose and response,
• Background incidence in test
animal,
• .Differences in range of response
among species, sexes, strains,
• Most sensitive responding species,
and
• Availability of data on related
precursor events to tumor development.
Analyses of carcinogenic effects other
than tumor incidence are similarly
presented and evaluated for their
contribution to a best judgment on how
to represent the biological data for dose
response assessment.
3.3. Dose Data
Whether animal experiments or
epidemiologic studies are the sources of
data, questions need to be addressed in
arriving at an appropriate measure of
dose for the anticipated environmental
exposure. Among these are:
• Whether the dose is expressed as an
environmental concentration, applied
dose, or delivered dose to the target
organ,
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17997
• Whether the dose is expressed in
terms of a parent compound, one or
more metabolites, or both,
• The impact of dose patterns and
timing where significant,
• Conversion from animal to human
doses, whore animal data are used, and
• The conversion metric between
routes of exposure where necessary and
appropriate.
In practice, there may be little or no
information on the concentration or
identity of the active form at a target;
being able to compare the applied and
delivered doses between routes and
species is the ideal, but is rarely
attained. Even so, the objective is to use
available data to obtain as close to a
measure of internal or delivered dose as
possible.
The following discussion assumes
that the analyst will have data of
varying detail in different cases about
toxicokinctics and metabolism.
Discussed below are approaches to basic
data that are most frequently available,
as well as approaches and judgments for
improving the analysis based on
additional data. The estimation of dose
in human studies is tailored to the form
of dose data available.
3.3.1. Intcrspccios Adjustment of Dose
When adequate data are available, the
doses used in animal studies can be
adjusted to equivalent human doses
using toxicokinelic information on the
particular agent. The methods used
should be tailored to the nature of the
data on a caso-by-case basis. In rare
cases, it may also be possible to make
adjustments based on toxicodynamic
considerations. In most cases, however,
there are insufficient data available to
compare dose between species. In these
cases, the estimate of human equivalent
dose is based on science policy default
assumptions. The defaults described
below are modified or replaced
whenever better comparative data on
toxicokinctic or metabolic relationships
ore available. The availability and
discussion of the latter also may permit
reduction or discussion of uncertainty
in the analysis.
For oral exposure, the default
assumption is that delivered doses are
related to applied dose by a power of
body weight. This assumption rests on
the similarities of mammalian anatomy,
physiology, and biochemistry generally
observed across species. This
assumption is more appropriate at low
applied dose concentrations where
sources of nonlinearity, such as
saturation or induction of enzyme
activity, are loss likely to occur. To
derive an equivalent human oral dose
from animal data, the default procedure
is to scale daily applied doses
experienced for a lifetime in proportion
to body weight raised to the 0.75 power
(W°-75). Equating exposure
concentrations in parts per million units
for food or water is an alternative
version of the same default procedure
because daily intakes of these are in
proportion to W°-75. The rationale for
this factor rests on the empirical
observation that rates of physiological
processes consistently tend to maintain
proportionality with W°-7S. A more
extensive discussion of the rationale
and data supporting the Agency's
adoption of this scaling factor is in U.S.
EPA, 1992b. Information such as blood
levels or exposure biomarkers or other
data that are available for interspecies
comparison are used to improve the
analysis when possible.
The default procedure to derive an
human equivalent concentration of
inhaled particles and gases is described
in U.S. EPA (1994) and Jarabek
(1995a,b). The methodology estimates
respiratory deposition of inhaled
particles and gases and provides
methods for estimating internal doses of
gases with different absorption
characteristics. The method is able to
incorporate additional toxicokinetics
and metabolism to improve the analysis
if such data are available.
3.3.2. Toxicokinetic Analyses
Physiologically based mathematical
models are potentially the most
comprehensive way to account for
toxicokinetic processes affecting dose.
Models build on physiological
compartmental modeling and attempt to
incorporate the dynamics of tissue
perfusion and the kinetics of enzymes
involved in metabolism of an
administered compound.
A comprehensive model requires the
availability of empirical data on the
carcinogenic activity contributed by
parent compound and metabolite or
metabolites and data by which to
compare kinetics of metabolism and
elimination between species. A
discussion of issues of confidence
accompanies presentation of model
results (Monro, 1992). This includes
considerations of model validation and
sensitivity analysis that stress the
predictive performance of the model.
When a delivered dose measure is used
in animal to human extrapolation of
dose response data, the assessment
should discuss the confidence in the
assumption that the toxicodynamics of
the target tissue(s) will be the same in
both species, Toxicokinetic data can
improve dose response assessment by
accounting for sources of change in
proportionality of applied to internal or
delivered dose at various levels of
applied dose. Many'of the sources of
potential nonlinearity involve saturation
or induction of enzymatic processes at
high doses. An analysis that accounts
for nonlinearity (for instance, due to
enzyme saturation kinetics) can assist in
avoiding overestimation or
underestimation of low dose response
otherwise resulting from extrapolation
from a sublinear or supralinear part of
the experimental dose response curve
(Gillette, 1983). Toxicokinetic processes
tend to become linear at low doses, an
expectation that is more robust than
low-dose linearity of response (Hattis,
1990). Accounting for toxicokinetic
nonlinearities allows better description
of the shape of the curve at relatively
high levels of dose in the range of
observation, but cannot determine
linearity or nonlinearity of response at
low dose levels (Lutz, 1990a; Swenberg
et al, 1987).
Toxicokinetic modeling results may
be presented as the preferred method of
estimating human equivalent dose or in
parallel discussion with default
assumptions depending on relative
confidence in the modeling.
3.3.3. Route-to-Route Extrapolation
Judgments frequently need to be made
about the carcinogenicity of an agent
through a route of exposure different
than the one in the underlying studies.
For example, exposures of interest may
be through inhalation of an agent tested
primarily through animal feeding
studies or through ingestion of an agent
that showed positive results in human
occupational studies from inhalation
exposure.
Route-to-route extrapolation has both
qualitative and quantitative aspects. For
the qualitative aspect, the assessor
weighs the degree to which positive
results through one route of exposure in
human or animal studies support a
judgment that similar results would
have been observed in appropriate
studies using the route of exposure of
interest. In general, confidence in
making such a judgment is strengthened
when the tumor effects are observed at
a site distant from the portal of entry
and when absorption through the route
of exposure of interest is similar to
absorption via the tested routes. In the
absence of contrary data, the qualitative .
default assumption is that, if the agent
is absorbed by a route to give an internal
dose, it may be carcinogenic by that
route. (See section 2.7.1.)
When a qualitative extrapolation can
be supported, quantitative extrapolation
may still be problematic in the absence
of adequate data. The differences in
biological processes among routes of
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exposure (oral, inhalation, dermal) can
be great because of, for example, first-
pass effects and differing results from
different exposure patterns. There is no
generally applicable method for
accounting for these differences in
uptake processes in quantitative route-
to-route extrapolation of dose response
data in the absence of good data on the
agent of interest. Therefore, route-to-
route extrapolation of dose data relies
on a case-by-case analysis of available
data. When good data on the agent itself
are limited, an extrapolation analysis
can be based on expectations from
physical and chemical properties of the
agent, properties and route-specific data
on structurally analogous compounds,
or in vitro or in vivo uptake data on the
agent. Route-to-route uptake models
may be applied if model parameters are
suitable for the compound of interest.
Such models are currently considered
interim methods; further model
development and validation is awaiting
the development of more extensive data
(see generally, Gerrity and Henry, 1990).
For screening or hazard ranking, route-
to-route extrapolation may be based on
assumed quantitative comparability as a
default, as long as it is reasonable to
assume absorption by compared routes.
When route-to-route extrapolation is
used, the assessor's degree of confidence
in both the qualitative and quantitative
extrapolation should be discussed in the
assessment and highlighted in the dose
response characterization.
3.3.4. Dose Averaging
The cumulative dose received over a
lifetime, expressed as lifetime average
daily dose, is generally considered an
appropriate default measure of exposure
to a carcinogen (Monro, 1992). The
assumption is made that a high dose of
a carcinogen received over a short
period of time is equivalent to a
corresponding low dose spread over a
lifetime. While this is" a reasonable
default assumption based on theoretical
considerations, departures from it are
expected. Another approach is needed
in some cases, such as when dose-rate
effects are noted (e.g., formaldehyde).
Cumulative dose may be replaced, as
appropriate and justified by the data,
with other dose measures. In such cases,
modifications to the default assumption
are made to take account of these
effects; the rationale for the selected
approach is explained.
In cases where a mode of action or
other feature of the biology has been
identified that has special dose
implications for sensitive
subpopulations (e.g., differential effects
by sex or disproportionate impacts of
early-life exposure), these are explained
and are recorded to guide exposure
assessment and risk characterization.
Special problems arise when the human
exposure situation of concern suggests
exposure regimens (e.g., route and
dosing schedule) that are substantially
different from those used in the relevant
animal studies. These issues are
explored and pointed out for attention
in the exposure assessment and risk
characterization.
3.4. Discussion of Uncertainties
The exploration of significant
uncertainties in data for dose and
response and in extrapolation
procedures is part of the assessment.
The presentation distinguishes between
model uncertainty and parameter
uncertainty. Model uncertainty is an
uncertainty about a basic biological
question. For example, a default, linear
dose response extrapolation may have
been made based on tumor and other
key evidence supporting the view that
the model for an agent's mode of action
is a DNA-reactive process. Discussion of
the confidence in the extrapolation is
appropriately done qualitatively or by
showing results for alternatives that are
equally plausible. It is not useful, for
example, to conduct quantitative
uncertainty analysis running multiple
forms of linear models. This would
obviate the function of the policy
default.
Parameter uncertainties deal with
numbers representing statistical or
analytical measures of variance or error
in data or estimates. Uncertainties in
parameters are described quantitatively,
if practicable, through sensitivity
analysis and statistical uncertainty
analysis. With the recent expansion of
readily available computing capacity,
computer methods are being adapted to
create simulated biological data that are
comparable with observed information.
These simulations can be used for
sensitivity analysis, for example, to
analyze how small, plausible variations
in the observed data could affect dose
response estimates. These simulations
can also provide information about
experimental uncertainty in dose
response estimates, including a
distribution of estimates that are
compatible with the observed data.
Because these simulations are based on
the observed data, they cannot assist in
evaluating the extent to which the
observed data as a whole are
idiosyncratic rather than typical of the
true situation. If quantitative analysis is
not possible, significant parameter
uncertainties are described
qualitatively. In either case, the
discussion highlights uncertainties that
are specific to the agent being assessed,
as distinct from those that are generic to
most assessments.
Estimation of the applied dose in a
human study has numerous
uncertainties such as the exposure
fluctuations that humans experience
compared with the controlled exposures
received by animals on test. In a
prospective cohort study, there is
opportunity to monitor exposure and
human activity patterns for a period of
time that supports estimation of applied
dose (U.S. EPA, 1992a). In a
retrospective study, exposure may be
based on monitoring data but is often
based on human activity patterns and
levels reconstructed from historical
data, contemporary data, or a
combination of the two. Such
reconstruction is accompanied by
analysis of uncertainties considered
with sensitivity analysis in the
estimation of dose (Wyzga, 1988; U.S.
EPA, 1986a). These uncertainties can
also be assessed for any confounding
factor for which a quantitative
adjustment of dose response data is
made (U.S. EPA, 1984).
3.5. Technical Dose Response
Characterization
As with hazard characterization, the
dose response characterization serves
the dual purposes of presenting a
technical characterization of the
assessment results and supporting the
risk characterization.
The characterization presents the
results of analyses of dose data, of
response data, and of dose response.
When alternative approaches are
plausible and persuasive in selecting
dose data, response data, or
extrapolation procedures, the
characterization follows the alternative
paths of analysis and presents the
results. The discussion covers the
question of whether any should be
preferred over others because it (or they)
better represents the available data or
corresponds to the view of the
mechanism of action developed in the
hazard assessment. The results for
different tumor types by sex and species
are provided along with the one(s)
preferred. Similarly, results for
responses other than tumor incidence
are shown if appropriate.
Numerical dose response estimates
are presented to one significant figure.
Numbers are qualified as to whether
they represent central tendency or
upper bounds and whether the method
used is inherently more likely to
overestimate or underestimate (Krewski
et al., 1984).
In cases where a mode of action or
other feature of the biology has been
identified that has special implications
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for carly-Hfo exposure, differential
effects by sex, or other concerns for
sensitive subpopulations, these are
explained. Similarly, any expectations
that high dose-rate exposures may alter
the risk picture for some portion of the
population are described. These and
other perspectives are recorded to guide
exposure assessment and risk
characterization. Whether the lifetime
average daily dose or another measure
of doso should be considered for
differing exposure scenarios is
discussed.
Uncertainly analyses, qualitative or
quantitative if possible, are highlighted
in the characterization.
The doso response characterization
routinely includes the following, as
appropriate for the data available:
• Identification of the kinds of data
available for analysis of dose and
response and for dose response
assessment,
• Results of assessment as above,
• Explanation of analyses in terms of
quality of data available,
• Selection of study/response and
dose metric for assessment,
• Discussion of implications of
variability in human susceptibility,
including for susceptible subpopulation,
• Applicability of results to varying
exposure scenarios—issues of route of
exposure, dose rate, frequency, and
duration,
• Discussion of strengths and
limitations (uncertainties) of the data
and analyses that are quantitative as
well as qualitative, and
* Special issues of interpretation of
data, such as:
—Selecting dose data, response data,
and dose response approach(es),
—Uso of meta-analysis,
—Uncertainty and quantitative
uncertainty analysis.
4. Technical Exposure Characterization
Guidelines for exposure assessment of
carcinogenic and other agents are
published (U.S. EPA, 1992a) and are
used in conjunction with these cancer
risk assessment guidelines. Presentation
of exposure descriptors is a subject of
discussion in EPA risk characterization
guidance (U.S. EPA, 1995). The
exposure characterization is a technical
characterization that presents the
assessment results and supports risk
characterization.
The characterization provides a
statement of purpose, scope, level of
detail, and approach used in the
assessment, identifying the exposure
scennrio(s) covered. It estimates the
distribution of exposures among
members of the exposed population as
the data permit. It identifies and
compares the contribution of different
sources and routes and pathways of
exposure. Estimates of the magnitude,
duration, and frequency of exposure are
included as available monitoring or
modeling results or other reasonable
methods permit. The strengths and
limitations (uncertainties) of the data
and methods of estimation are made
clear.
The exposure characterization
routinely includes the following, as
appropriate and possible for the data
available:
• Identification of the kinds of data
available,
• Results of assessment as above,
• Explanation of analyses in terms of
quality of data available,
• Uncertainty analyses as discussed
in Exposure Assessment Guidelines,
distinguishing uncertainty from
variability, and
• Explanation of derivation of
estimators of "high end" or central
tendency of exposure and their
appropriate use.
5. Risk Characterization
5.1. Purpose
The risk characterization process
includes an integrative analysis
followed by a presentation in a Risk
Characterization Summary, of the major
results of the risk assessment. The Risk
Characterization Summary is a
nontechnical discussion that minimizes
the use of technical terms. It is an
appraisal of the science that supports
the risk manager in making public
health decisions, as do other
decisionmaking analyses of economic,
social, or technology issues. It also
serves the needs of other interested
readers. The summary is an information
resource for preparation of risk
communication information, but being
somewhat technical, is not itself .the
usual vehicle for communication with
every audience.
The integrative analysis brings
together the assessments and
characterizations of hazard, dose
response, and exposure to make risk
estimates for the exposure scenarios of
interest. This analysis is generally much
more extensive than the Risk
Characterization Summary. It may be
peer-reviewed or subject to public
comment along with the summary in
preparation for an Agency decision. The
integrative analysis may be titled
differently by different EPA programs
(e.g., "Staff Paper" for criteria air
pollutants), but it typically will identify
exposure scenarios of interest in a
decisionmaldng and present risk
analyses associated with them. Some of
the analyses may concern scenarios in
several media, others may examine, for
example, only drinking water risks. It
also may be the document that contains
quantitative analyses of uncertainty.
The values supported by a risk
characterization throughout the process
are transparency in environmental
decisionmaking, clarity in
communication, consistency in core
assumptions and science policies from
case to case, and reasonableness. While
it is appropriate to err on the side of
protection of health and the
environment in the face of scientific
uncertainty, common sense and
reasonable application of assumptions
and policies are essential to avoid
unrealistic estimates of risk (U.S. EPA,
1995). Both integrative analyses and the
Risk Characterization Summary present
an integrated and balanced picture of
the analysis of the hazard, dose
response, and exposure. The risk analyst
should provide summaries of the
evidence and results and describe the
quality of available data and the degree
of confidence to be placed in the risk
estimates. Important features include
the constraints of available data and the
state of knowledge, significant scientific
issues, and significant science and
science policy choices that were made
when alternative interpretations of data
existed (U.S. EPA, 1995). Choices made
about using default assumptions or data
in the assessment are explicitly
discussed in the course of analysis, and
if a choice is a significant issue, it is
highlighted in the summary.
5.2. Application
Risk characterization is a necessary
part of generating any Agency report on
risk, whether the report is preliminary
to support allocation of resources
toward further study or comprehensive
to support regulatory decisions. In the
former case, the detail and
sophistication of the characterization
are appropriately small in scale; in the
latter case, appropriately extensive.
Even if a document covers only parts of
a risk assessment (hazard and dose
response analyses for instance), the
results of these are characterized.
Risk assessment is an iterative process
that grows in depth and scope in stages
from screening for priority-making, to
preliminary estimation, to fuller
examination in support of complex
regulatory decisionmaking. Default
assumptions are used at every stage
because no database is ever complete,
but they are predominant at screening
stages and are used less as more data are
gathered and incorporated at later
stages. Various provisions in EPA-
administered statutes require decisions
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based on findings that represent all
stages of iteration. There are close to 30
provisions within the major statutes that
require decisions based on risk, hazard,
or exposure assessment. For example,
Agency review of premanufacture
notices under section 5 of the Toxic
Substances Control Act relies on
screening analyses, while requirements
for industry testing under section 4 of
that Act rely on preliminary analyses of
risk or simply of exposure. At the other
extreme, air quality criteria under the
Clean Air Act rest on a rich data
collection required by statute to undergo
reassessment every few years. There are
provisions that require ranking of
hazards of numerous pollutants—by its
nature a screening level of analysis—
and other provisions that require a full
assessment of risk. Given this range in
the scope and depth of analyses, not all
risk characterizations can or should be
equal in coverage or depth. The risk
assessor must carefully decide which
issues in a particular assessment are
important to present, choosing those
that are noteworthy in their impact on
results. For example, health effect
assessments typically rely on animal
data since human data are rarely
available. The objective of
characterization of the use of animal
data is not to recount generic issues
about interpreting and using animal
data. Agency guidance documents cover
these. Instead, the objective is to call out
any significant issues that arose within
the particular assessment being
characterized and triform the reader
about significant uncertainties that
affect conclusions.
5.3. Presentation of Risk
Characterization Summary
The presentation is a nontechnical
discussion of important conclusions,
issues, and uncertainties that uses the
hazard, dose-response, exposure, and
integrative analyses for technical
support. The primary technical supports
within the risk assessment are the
hazard characterization, dose response
characterization, and exposure
characterization described in this
guideline. The risk characterization is
derived from these. The presentation
should fulfill the aims outlined in the
purpose section above.
5.4. Content of Risk Characterization
Summary
Specific guidance on hazard, dose
response, and exposure characterization
appears in previous sections. Overall,
the risk characterization routinely
includes the following, capturing the
important items covered in hazard, dose
response, and exposure
characterization.
• Primary conclusions about hazard,
dose response, and exposure, including
equally plausible alternatives,
• Nature of key supporting
information and analytic methods,
• Risk estimates and their attendant
uncertainties, including key uses of
default assumptions when data are
missing or uncertain,
• Statement of the extent of
extrapolation of risk estimates from
observed data to exposure levels of
interest (i.e., margin of exposure) and its
implications for certainty or uncertainty
in qualifying risk,
• Significant strengths and
limitations of the data and analyses,
including any major peer reviewers'
issues,
• Appropriate comparison with
similar EPA risk analyses or common
risks with which people may be
familiar, and
• Comparison with assessment of the
same problem by another organization.
Appendix A
This appendix contains several general
illustrations of weight of evidence narratives.
In addition, after narrative #5 is an example
of a briefing summary format.
NARRATIVE #1 Chlorinated Alkene
CASffXXX
CANCER HAZARD SUMMARY
Chlorinated alkene (cl-alkene] is likely to
be carcinogenic to humans by all routes of
exposure. The weight of evidence of human
carcinogenicity of cl-alkene is based on (a)
findings of carcinogenicity in rats and mice
of both sexes by oral and inhalation
exposures; (b) its similarity in structure to
other chlorinated organics that are known to
cause liver and kidney damage, and liver and
kidney tumors in rats and mice; (c)
suggestive evidence of a possible association
between cl-alkene exposure of workers in the
laundry and dry cleaning industries and
increased cancer risk in a number of organ
systems; and (d) human and animal data
indicating that cl-alkene is absorbed by all
routes of exposure.
In comparison with other agents
designated as likely carcinogens, the overall
weight of evidence for cl-alkene places it at
the low end of the grouping. This is because
one cannot attribute observed excess cancer
risk in exposed workers solely to cl-alkene.
Moreover, there is considerable scientific
uncertainty about the human significance
and relevance of certain rodent tumors
associated with exposure to cl-alkene and
other chlorinated organics, but insufficient
evidence about mode of action for the animal
tumors. Hence, the human relevance of the
animal evidence of carcinogenicity relies on
a default assumption of relevance.
There is no clear evidence about the mode
of action for each tumor type induced in rats
and mice. Available evidence suggests that
cl-alkene induces cancer mainly by
promoting cell growth rather than via direct
mutagenic action, although a mutagenic
mode of action for rat kidney tumors cannot
be ruled out. The dose response assessment
should, therefore, adopt both default
approaches, nonlinear and linear. It is
recognized that the latter approach likely
overestimates risk at low doses if the mode
of action is primarily growth-promoting. This
approach, however, may be useful for
screening analyses.
SUPPORTING INFORMATION
Human Data
A number of epidemiologic studies of dry
cleaning and laundry workers that have
reported elevated incidences of lung, cervix,
esophagus, kidney, blood and lymphoid
cancers. Many of these studies are
confounded by co-exposure to other
petroleum solvents, making them limited for
determining whether the observed increased
cancer risks are causally related to cl-alkene.
The only investigation of dry cleaning
workers with no known exposure to other
chemicals did not evaluate other
confounding factors such as smoking, alcohol
consumption, and low socioeconomic status
to exclude the possible contribution of these
factors to cancer risks.
Animal Data
The carcinogenic potential of cl-alkene has
been adequately investigated in two chronic
studies in two rodent species, the first study
by gavage and the second study by
inhalation. Cl-alkene is carcinogenic in the
liver in both sexes of mice when tested by
either route of exposure. It causes marginally
increased incidences of mononuclear cell
leukemia (MCL) in both sexes of rats and low
incidences of a rare kidney tumor in male
rats by inhalation. No increases in tumor
incidence were found in rats treated with cl-
alkene by gavage. This rat study was
considered limited because of high mortality
of the animals.
Although cl-alkene causes increased
incidences of tumors at multiple sites in two
rodent species, controversy surrounds each
of the tumor endpoints concerning their
relevance and/or significance to humans (see
discussion under Mode of Action).
Other Key Data
Cl-alkene is a member of a class of
chlorinated organics that often cause liver
and kidney toxicity and carcinogenesis in
rodents. Like many chlorinated
hydrocarbons, cl-alkene itself is tested
negative in a battery of standard genotoxicity
tests using bacterial and mammalian cells
systems including human lymphocytes and
fibroblast cells. There is evidence, however,
that a minor metabolite generated by an
enzyme found in rat kidney tissue is
mutagenic. This kidney metabolite has been
hypothesized to be related to the
development of kidney tumors in the male
rat. This metabolic pathway appears to be
operative in the human kidney.
Human data indicate that cl-alkene is
readily absorbed via inhalation but to a much
lesser extent by skin contact. Animal data
show that cl-alkene is absorbed well by the
oral route.
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18001
MODE OF ACTION
Tho mechanisms of cl-alkone-induced
mouso Itvor tumors are not completely
understood. One mechanism has been
hypothesized to bo mediated by a genotoxic
cpoxldo metabolite generated by enzymes
found in the mouse liver, but there is a lack
of direct evidence in support of this
mechanism. A more plausible mechanism
that still needs to bo further defined is related
to liver poroxisomal proliferation and
toxlcity by TCA (tricnloroacotic acid), a
major metabolite of cl-alkene. However, there
arc no definitive data indicating that TCA
Induces poroxisomal proliferation in
humans.
Tho mechanisms by which cl-alkene
induces kidney tumors in male rats are even
loss well understood. The rat kidney
rcsponsa may bo related to either kidney
toxlcity or the activity of a mutagenic
metabolite of the parent compound.
Tho human relevance of cl-alkene-induced
MCL In rats is unclear. Tho biological
significance of marginally increased
Incidences of MCL, has been questioned by
some, since this tumor occurs spontaneously
in tho tested rat strain at very high
background rates. On tho other hand, it has
boon considered by others to be a true finding
because there was a decreased time to onset
of tho disease and tho disease was more
severe in treated as compared with untreated
control animals. Tho exact mechanism by
which cl-alkono increases incidences of MCL
In rats is not known.
Overall, (hero is not enough evidence to
givo high confidence in a conclusion about
any single mode of action; it would appear
that more than a single mode operates in
different rodent tissues. The apparent lack of
niutngcnlcily of cl-alkene itself and its
general growth-promoting effect on high
background tumors as well as its toxicity
toward mouso liver and rat kidney tissue
support tho view that its predominant mode
of action is coll growth promoting rather than
mutagenic. A mutagenic contribution to the
renal carcinogonicity due to a metabolite
cannot bo entirely ruled out.
NARRATIVES
Unsaliiratcd Aldehyde
CASSXXX
CANCER HAZARD SUMMARY
Tho potential human hazard of unsaturated
aldehyde (UA) cannot bo determined, but
thoro are suggestive data for carcinogenicity.
Tho evidence on carcinogonicity consists
of (a) data from an oral animal study showing
a response only at tho highest dose in female
rots, with no response in males and (b) the
fact that other low-molecular-weight
aldehydes have shown tumorigenicity in the
respiratory tract after inhalation. The one
study of UA effects by tho inhalation route
was not adequately performed. The available
cvidonco is too limited to describe human
carcinogenicity potential or support dose
response assessment.
SUPPORTING INFORMATION
Human Data
An elevated incidence of cancer was
reported in a cohort of workers in a chemical
plant who were exposed to a mixture of
chemicals including UA as a minor
component. The study is considered
inadequate because of the small size of the
cohort studied and the lack of adequate
exposure data.
Animal Data
In a long-term drinking water study in rats,
an increased incidence of adrenal cortical
adenomas was found in the highest-dosed
females. No other significant finding was
made. The oral rat study was well conducted
by a standard protocol. In a 1-year study in
hamsters at one inhalation dose, no tumors
were seen. This study was inadequate due to
high mortality and consequent short
duration. The chemical is very irritating and
is a respiratory toxicant in mammals. The.
animal data are too limited for conclusions
to be drawn.
Structural Analogue Data
UA's structural analogues, formaldehyde
and acetaldehyde, both have carcinogenic
effects on the rat respiratory tract.
Other Key Data
The weight of results of mutagenicity tests
in bacteria, fungi, fruit flies, and mice result
in an overall conclusion of not mutagenic;
UA is lethal to bacteria to a degree that makes.
testing difficult and test results difficult to
interpret. The chemical is readily absorbed
by all routes.
MODE OF ACTION
Data are not sufficient to judge whether
there is a carcinogenic mode of action.
NARRATIVES
Alkene Oxide
CAStfXXX
CANCER HAZARD SUMMARY
Alkene oxide (AO) should be dealt with as
if it were a known human carcinogen by all
routes of exposure. Several studies in
workers, when considered together, suggest
an elevated risk of leukemia and lymphoma
after long-term exposure to AO, even though
no single study conclusively demonstrates
that AO caused the cancer. In addition,
animal cancer and mutagenicity studies as
well as short-term tests of mutagenicity have
strongly consistent results that support a
level of concern equal to having conclusive
human studies.
The weight of evidence of human
carcinogenicity is based on (a) consistent
evidence of carcinogenicity of AO in rats and
mice by both oral and inhalation exposure;
(b) studies in workers that taken together
suggest elevated risk of leukemia and
lymphoma to workers exposed to AO and
show genetic damage in blood lymphocytes
in exposed workers; (c) mutagenic effects in
numerous test systems and heritable gene
mutations in animals; and (d) membership in
a class of DNA-reactive compounds that are
regularly observed to cause cancer in
animals.
Due to its ready absorption by all routes of
exposure and rapid distribution throughout
the body, AO is expected to pose a risk by
any route of exposure. The strong evidence
of a mutagenic mode of action supports dose ,
response assessment that assumes linearity of
the relationship.
SUPPORTING INFORMATION
Human Data
Elevated risks of lymphatic cancer and
cancer of blood-forming tissue have been
reported in exposed workers in several
studies. The interpretation of the studies
separately is complicated by exposures to
other agents in each so there is no single
study that demonstrates that AO caused the
effects; nevertheless, several of the studies
together are considered suggestive of AO
carcinogenicity because they consistently
show cancer elevation in the same tissues.
Biomonitoring studies of exposed workers
find DNA damage in blood lymphocytes and
the degree of DNA damage correlates with
the level and duration of AO exposure.
Finding this damage in the same tissue in
which elevated cancer was seen in workers
adds further weight to the positive suggestion
from the worker cancer studies. The human
data are from well-conducted studies.
Animal Data
AO causes cancer in multiple tissue sites
in rats and mice of both sexes by oral and
inhalation exposure. The database is more
extensive than usual and the studies are
good. The observation of multisite,
multispecies carcinogenic activity by an
agent is considered to be very strong
evidence and is often the case with highly
mutagenic agents. There are also good studies
showing that AO causes heritable germ cell
mutations in mice after inhalation
exposure—a property that is very highly
correlated with carcinogenicity.
Structural Analogue Data
Organic epoxides are commonly found to
have carcinogenic effects in animals,
particularly the low-molecular-weight ones.
Other Key Data
The structure and DNA reactivity of AO
support potential carcinogenicity. Both
properties are highly correlated with
carcinogenicity. Positive mutagenicity tests
in vitro and in vivo add to this support and
are reinforced by observation of similar
genetic damage in exposed workers.
AO is experimentally observed to be
readily absorbed by all routes and rapidly
distributed through the body.
MODE OF ACTION
All of the available data are strongly
supportive of a mutagenic mode of action,
with a particular human target in lymphatic
and blood-forming tissue. The current
scientific consensus is that there is virtually
complete correspondence between ability of
an agent to cause heritable germ cell
mutations, as AO does, and carcinogenicity.
All of this points to a mutagenic mode of
action and supports assuming linearity of the
dose response relationship.
NARRATIVES
Bis-benzenamine
CAS# XXX
CANCER HAZARD SUMMARY
This chemical is likely to be carcinogenic
to humans by all routes of exposure. Its
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carcinogenic potential is indicated by (a)
tumor and toxicity studies on structural
analogues, which demonstrate the ability of
the chemical to produce thyroid follicular
cell tumors in rats and hepatocellular tumors
in mice following ingestion and (b)
metabolism and hormonal information on the
chemical and its analogues, which
contributes to a working mode of action and
associates findings in animals with those in
exposed humans. In comparison with other
agents designated as likely carcinogens, the
overall weight of evidence for this chemical
places it at the lower end of the grouping.
This is because there is a lack of tumor
response data on this agent itself.
Biological information on the compound is
contradictory in terms of how to quantitate
potential cancer risks. The information on
disruption on thyroid-pituitary status argues
for using a margin of exposure evaluation.
However, the chemical is an aromatic amine,
a class of agents that are DNA-reactive and
induce gene mutation and chromosome
aberrations, which argues for low-dose
linearity. Additionally, there is a lack of
mode of action information on the mouse
liver tumors produced by the structural
analogues, also pointing toward a low-dose
linear default approach. In recognition of
these uncertainties, it is recommended to
quantitate tumors using both nonlinear (to
place a lower bound on the risks) and linear
(to place an upper bound on the risks) default
approaches. Given the absence of tumor
response data on the chemical per se, it is
recommended that tumor data on close
analogues be used to possibly develop
toxicity equivalent factors or relative
potencies.
Overall, this chemical is an inferential case
for potential human carcinogenicity. The
uncertainties associated with this assessment
include (1) the lack of carcinogenicity studies
on the chemical, (2) the use of tumor data on
structural analogues, (3) the lack of definitive
information on the relevance of thyroid-
pituitary imbalance for human
carcinogenicity, and (4) the different
potential mechanisms that may influence
tumor development and potential risks.
SUPPORTING INFORMATION
Human Data
Worker exposure has not been well
characterized or quantified, but recent
medical monitoring of workers exposed over
a period of several years has uncovered
alterations in thyroid-pituitary hormones (a
decrease in T3 and T4 and an increase in
TSH) and symptoms of hypothyroidism. A
urinary metabolite of the chemical has been
monitored in workers, with changes in
thyroid and pituitary hormones noted, and
the changes were similar to those seen in an
animal study.
Animal Data
The concentration of the urinary
metabolite in rats receiving the chemical for
28 days was within twofold of that in
exposed workers, a finding associated with
comparable changes in thyroid hormones and
TSH levels. In addition, the dose of the
chemical given to rats in this study was
essentially the same as that of an analogue
that had produced thyroid and pituitary
tumors in rats. The human thyroid responds
in the same way as the rodent thyroid
following short-term, limited exposure.
Although it is not well established that
thyroid-pituitary imbalance leads to cancer
in humans as it does in rodents, information
in animals and in exposed humans suggests
similar mechanisms of disrupting thyroid-
pituitary function and the potential role of
altered TSH levels in leading to thyroid
carcinogenesis.
Structural Analogue Data
This chemical is an aromatic amine, a
member of a class of chemicals that has
regularly produced carcinogenic effects in
rodents and gene and structural chromosome
aberrations in short-term tests. Some
aromatic amines have produced cancer in
humans.
Close structural analogues produce thyroid
follicular cell tumors in rats and
hepatocellular tumors in mice following
ingestion. The thyroid tumors are associated
with known perturbations in thyroid-
pituitary functioning. These compounds
inhibit the use of iodide by the thyroid gland,
apparently due to inhibition of the enzyme
that synthesizes the thyroid hormones (T3,
T4). Accordingly, blood levels of thyroid
hormones decrease, which induce the
pituitary gland to produce more TSH, a
hormone that stimulates the thyroid to
produce more of its hormones. The thyroid
gland becomes larger due to increases in the
size of individual cells and their proliferation
and upon chronic administration, tumors
develop. Thus, thyroid tumor development is
significantly influenced by disruption in the
thyroid-pituitary axis.
Other Key Data
The chemical can be absorbed by the oral,
inhalation, and dermal routes of exposure.
MODE OF ACTION
Data on the chemical and on structural
analogues indicate the potential association
of carcinogenesis with perturbation of
thyroid-pituitary homeostasis. Structural
analogues are genotoxic, thus raising the
possibility of different mechanisms by which
this chemical may influence tumor
development.
NARRATIVE #5
Brominated Alkane (BA)
CAStfXXX
CANCER HAZARD SUMMARY
Brominated alkane (BA) is likely to be a
human carcinogen by all routes of exposure.
The weight of evidence for human
carcinogenicity is at the high end of agents
in the "likely" group. Findings are based on
very extensive and significant experimental
findings that include (a) tumors at multiple
sites in both sexes of two rodent species via
three routes of administration relevant to
human exposure, (b) close structural
analogues that produce a spectrum of tumors
like BA, (c) significant evidence for the
production of reactive BA metabolites that
readily bind to DNA and produce gene
mutations in many systems including
cultured mammalian and human cells, and
(d) two null and one positive epidemiologic
study; in the positive study, there may have
been exposure to BA. These findings support
a decision that B A might produce cancer in
exposed humans. In comparison to other
agents considered likely human carcinogens,
the overall weight of evidence for BA puts it
near the top of the grouping. Given the
agent's mutagenicity, which can influence
the carcinogenic process, a linear dose-
response extrapolation is recommended.
Uncertainties include the lack of adequate
information on the mutagenicity of BA in
mammals or humans in vivo, although such
effects would be expected.
SUPPORTING INFORMATION
Human Data
The information on the carcinogenicity of
BA from human studies is inadequate. Two
studies of production workers have not
shown significant increases in cancer from
exposure to BA and other chemicals. An
increase in lymphatic cancer was reported in
a mortality study of grain elevator workers
who may have been exposed to BA (and
other chemicals).
Animal Data
BA produced tumors in four chronic
rodents studies. Tumor increases were noted
in males and females of rats and mice
following oral dermal and inhalation
exposure (rat—oral and two inhalation,
mouse—oral and dermal). It produces tumors
both at the site of application (e.g., skin with
dermal exposure) and at sites distal to the
portal of entry into the body (e.g., mammary
gland) following exposure from each route.
Tumors at the same site were noted in both
sexes of a species (blood vessel), both species
(forestomach) and via different routes of
administration (lung). Some tumors.
developed after very short latency,
metastasized extensively, and produced
death, an uncommon findings in rodents.
The rodent studies were well designed and
conducted except for the oral studies, in
which the doses employed caused excessive
toxicity and mortality. However, given the
other rodent findings, lower doses would
also be anticipated to be carcinogenic.
Structural Analogue Data
Several chemicals structurally related to
BA are also carcinogenic in rodents. Among
four that are closest in structure, tumors like
those seen for BA were often noted (e.g.,
forestomach, mammary, lung), which helps
to confirm the findings for BA itself. In sum,
all of the tumor findings help to establish
animal carcinogenicity and support potential
human carcinogenicity for BA.
Other Key Data
BA itself is not reactive, but from its
structure it was expected to be metabolized
to reactive forms. Extensive metabolism
studies have confined this presumption and
have demonstrated metabolites that bind to
DNA and cause breaks in the DNA chain.
These lesions are readily converted to gene
mutations in bacteria, fungi, higher plants,
insects and mammalian and human cells in
culture. There are only a limited number of
reports on the induction of chromosome
aberrations in mammals and humans; thus
far they are negative.
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18003
MODE OF ACTION
Human carcinogens often produce cancer
in multiple sites of multiple animal species
and both soxos and arc mutagenic in multiple
test systems. BA satisfies these findings. It
produces cancer in males and females of rats
and mice. It produces gone mutations in cells
across all life forms—plants, bacteria and
animals—including mammals and humans.
Given the mutagonicity of BA exposure and
the multiplicity and short latency of BA
tumor induction, it is reasonable to use a
linear approach for cancer dose-response
extrapolation.
BRIEFING SUMMARY
Route(s)
All .
Class
Likely
Designa-
tion or
rationale
High end
Dose re-
sponse
Default-
linear.
Basis for classification/dose response
1. Human data: Two studies of production
workers show no increase in cancer (one had
a small sample size; the other had mixed
chemical exposures). An increase in
lymphatic cancer is seen among grain
elevator workers who may have been
exposed to other chemicals.
2. Animal data: BA produces tumors at
multiple sites in male and female rats and
mico following oral, dermal, and inhalation
exposure. Tumors are seen at the site of
administration and distally and are often
consistent across sex, species, and route of
administration; some develop early,
motastaslzo, and cause death.
3. Structural analogue data: Close
analogues produce some of the same tumors
as aro seen with BA.
4. Other key data: BA is metabolized to a
reactive chemical that binds DNA and
produces geno mutations in essentially every
tost system including cultured human cells.
5. Mode of action: Like most known human
carcinogens, BA is mutagenic in most test
systems.
6. Hazard classification/uncertainties:
There is a rich database on BA demonstrating
its potential ability to cause tumors in
humans, including (a) multiple animal
tumors, (b) by appropriate routes of exposure,
(c) a mode of action relevant to human
carcinogenicity, and (d) some information in
humans. Together they lead to a designation
near tha high end of the likely human
carcinogen class.
7. Dose response: Given the anticipated
mode of action, a linear default dose
response relationship should be assumed.
Appendix B
This appendix contains responses to
tho National Academy of Sciences
National Research Council report
Science and Judgment in Risk
Assessment (NRG, 1994).
Recommendations oftlie National Academy
of Sciences National Research Council
In 1994, tho National Academy of Sciences
published a report Science and Judgment in
Risk Assessment. Tho 1994 report was
written by a Committee on Risk Assessment
of Hazardous Air Pollutants formed under
the Academy's Board on Environmental
Studies and Toxicology, Commission on Life
Sciences, National Research Council. The
report was called for under Section
112(o)(l)(A,B) of the Clean Air Act
Amendments of 1990, which provided for the
EPA to arrange for the Academy to review:
• risk assessment methodology used by the
EPA to determine the carcinogenic risk
associated with exposure to hazardous air
pollutants from source categories and
subcategories subject to the requirements of
this section and
• improvements in such methodology.
Under Section 112(o)(2)(A,B), the Academy
was to consider the following in its review:
• the techniques used for estimating and
describing the carcinogenic potency to
humans of hazardous air pollutants and
• the techniques used for estimating
exposure to hazardous air pollutants (for
hypothetical and actual maximally exposed
individuals as well as other exposed
individuals).
To the extent practicable, the Academy
was also to review methods of assessing
adverse human health effects other than
cancer for which safe thresholds of exposure
may not exist [Section 112(o)(3)]. The
Congress further provided that the EPA
Administrator should consider, but need not
adopt, the recommendations in the report
and the views of the EPA Science Advisory
Board with respect to the report. Prior to the
promulgation of any standards under Section
112(f), the Administrator is to publish
revised guidelines for carcinogenic risk
assessment or a detailed explanation of the
reasons that any recommendations contained
in the report will not be implemented
[Section 112(o)(6)].
The following discussion addresses the
recommendations of the 1994 report that are
pertinent to the EPA cancer risk assessment
guidelines. Guidelines for assessment of
exposure, of mixtures, and of other health
effects are separate EPA publications. Many
of the recommendations were related to
practices specific to the exposure assessment
of hazardous air pollutants, which are not
covered in cancer assessment guidelines.
Recommendations about these other
guidelines or practices are not addressed
here.
Hazard Classification
The 1994 report contains the following
recommendation about classifying cancer
hazard:
• The EPA should develop a two-part
scheme for classifying evidence on
carcinogenicity that would incorporate both
a simple classification and a narrative
evaluation. At a minimum, both parts should
include the strength (quality) of the evidence,
the relevance of the animal model and results
to humans, and the relevance of the
experimental exposures (route, dose, timing,
and duration) to those likely to be
encountered by humans.
The report also presented a possible matrix
of 24 boxes that would array weights of
evidence against low, medium, or high
relevance, resulting in 24 codes for
expressing the weight and relevance.
These guidelines adopt a set of descriptors
and subdescriptors of weight of evidence in
three categories: "known/likely," "cannot be
determined," and "not likely," and a
narrative for presentation of the weight of
evidence findings. The descriptors are used
within the narrative. There is no matrix of
alphanumerical weight of evidence boxes.
The issue of an animal model that is not
relevant to humans has been dealt with by
not including an irrelevant response in the
weighing of evidence, rather than by creating
a weight of evidence then appending a
discounting factor as the NRG scheme would
do. The issue is more complex than the NRG
matrix makes apparent. Often the question of
relevance of the animal model applies to a
single tumor response, but one encounters
situations in which there are more tumor
responses in animals than the questioned
one. Dealing with this complexity is more
straightforward if it is done during the
weighing of evidence rather than after as in
the NRG scheme. Moreover, the same
experimental data are involved in deciding
on the weight of evidence and the relevance
of a response. It would be awkward to go
over the same data twice.
In recommending that the relevance of
circumstances of human exposure also be
taken into account, the NRG appears to
assume that all of the actual conditions of
human exposure will be known when the
classification is done. This is not the case.
More often than not, the hazard assessment
is applied to assessment of risks associated
with exposure to different media or
environments at different times. In some
cases, there is no priority to obtaining
exposure data until the hazard assessment
has been done. The approach of these
guidelines is to characterize hazards as to
whether their expression is intrinsically
limited by route of exposure or by reaching
a particular dose range based strictly on
toxicological and other biological features of
the agent. Both the use of descriptors and the
narrative specifically capture this
information. Other aspects of appropriate
application of the hazard and dose response
assessment to particular human exposure
scenarios are dealt with in the
characterization of the dose response
assessment, e.g., the applicability of the dose
response assessment to scenarios with
differing frequencies and durations.
The NRG scheme apparently intended that
the evidence would be weighed, then given
a low, medium, or high code for some
combination of relevance of the animal
response, route of exposure, timing, duration,
or frequency. The 24 codes contain none of
this specific information, and in fact, do not
communicate what the conclusion is about.
To make the codes communicate the ,
information apparently intended would
require some multiple of the 24 in the NRG
scheme. As the number of codes increases,
their utility for communication decreases.
Another reason for declining to use codes
is that they tend to become outdated as
research reveals new information that was
not contemplated when they were adopted.
This has been the case with the classification
system under the EPA, 1986 guidelines.
Even though these guidelines do not adopt
a matrix of codes, the method they provide
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of using descriptors and narratives captures
the information the NRC recommended as
the most important, and in the EPA's view,
in a more transparent manner.
Dose Response
• The 1994 report contains the following
recommendations about dose response
issues:
• EPA should continue to explore, and
when scientifically appropriate, incorporate
toxicokinetic models of the link between
exposure and biologically effective dose (i.e.,
dose reaching the target tissue).
• Despite the advantages of developing
consistent risk assessments between agencies
by using common assumptions (e.g.,
replacing surface area with body weight to
the 0.75 power), EPA should indicate other
methods, if any, that would be more accurate.
• EPA should continue to use the
linearized multistage model as a default
option but should develop criteria for
determining when information is sufficient to
use an alternative extrapolation model.
• EPA should continue to use as one of its
risk characterization metrics upper-bound
potency estimates of the probability of
developing cancer due to lifetime exposure.
Whenever possible, this metric should be
supplemented with other descriptions of
cancer potency that might more adequately
reflect the uncertainty associated with the
estimates.
• EPA should adopt a default assumption
for differences in susceptibility among
humans in estimating individual risks.
• In the analysis of animal bioassay data
on the occurrence of multiple tumor types,
the cancer potencies should be estimated for
each relevant tumor type that is related to
exposure and the individual potencies
should be summed for those tumors.
The use of toxicokinetic models is
encouraged in these guidelines with
discussion of appropriate considerations for
their use. When there are questions as to
whether such a model is more accurate in a
particular case than the default method for
estimating the human equivalent dose, both
alternatives may be used. It should be noted
that the default method for inhalation
exposure is a toxicokinetic model.
The rationale for adopting the oral scaling
factor of body weight to the 0.75 power has
been discussed above in the explanation of
major defaults. The empirical basis is further
explored in U.S. EPA, 1992b. The more
accurate approach is to use a toxicokinetic
model when data become available or to
modify the default when data are available as
encouraged under these guidelines. As the
U.S. EPA, 1992b discussion explores in
depth, data on the differences among animals
in response to toxic agents are basically
consistent with using a power of 1.0, 0.75, or
0.66. The Federal agencies chose the power
of 0.75 for the scientific reasons given in the
previous discussion of major defaults; these
were not addressed specifically in the NRC
report. It was also considered appropriate, as
a matter of policy, for the agencies to agree
on one factor. Again, the default for
inhalation exposure is a model that is
constructed to become better as more agent-
specific data become available.
The EPA proposes not to use a computer
model such as the linearized multistage
model as a default for extrapolation below
the observed range. The reason is that the
basis for default extrapolation is a theoretical
projection of the likely shape of the curve
considering mode of action. For this purpose,
a computer model looks more sophisticated
than a straight line extrapolation, but is not.
The extrapolation will be by straight line as
explained in the explanation of major
defaults. This was also recommended by
workshop reviewers of a previous draft of
these guidelines (U.S. EPA, 1994b). In
addition, a margin of exposure analysis is
proposed to be used in cases in which the
curve is thought to be nonlinear, based on
mode of action. In both cases, the observed
range of data will be modeled by curve fitting
in the absence of supporting data for a
biologically based or case-specific model.
The result of using straight line
extrapolation is thought to be an upper
bound on low-dose potency to the human
population in most cases, but as discussed in
the major defaults section, it may not always
be. Exploration and discussion of uncertainty
of parameters in curve-fitting a model of the
observed data or in using a biologically based
or case-specific model is called for in the
dose response assessment and
characterization sections of these guidelines.
The issue of a default assumption for
human differences in susceptibility has been
addressed under the major defaults
discussion in section 1.3 with respect to
margin of exposure analysis. The EPA has
considered but decided not to adopt a
quantitative default factor for human
differences in susceptibility when a linear
extrapolation is used. In general, the EPA
believes that the linear extrapolation is
sufficiently conservative to protect public
health, Linear approaches (both LMS and
straight line extrapolation) from animal data
are consistent with linear extrapolation on
the same agents from human data (Goodman
and Wilson, 1991; Hoel and Portier, 1994). If
actual data on human variability in
sensitivity are available they will, of course,
be used.
In analyzing animal bioassay data on the
occurrence of multiple tumor types, these
guidelines outline a number of biological and
other factors to consider. The objective is to
use these factors to select response data
(including nontumor data as appropriate)
that best represent the biology observed. As
stated in section 3 of the guidelines,
appropriate options include use of a single
data set, combining data from different
experiments, showing a range of results from
more than one data set, showing results from
analysis of more than one tumor response
based on differing modes of action,
representing total response in a single
experiment by combining animals with
tumors, or a combination of these options.
The approach judged to best represent the
data is presented with the rationale for the
judgment, including the biological and
statistical considerations involved. The EPA
has considered the approach of summing
tumor incidences and decided not to adopt
it. While multiple tumors may be
independent, in the sense of not arising from
metastases of a single malignancy, it is not
clear that they can be assumed to represent
different effects of the agent on cancer
processes. In this connection, it is not clear
that summing incidences provides a better
representation of the underlying mode(s) of
action of the agent than combining animals
with tumors or using another of the several
options noted above. Summing incidences
would result in a higher risk estimate, a step
that appears unnecessary without more
reason.
Risk Characterization
• When EPA reports estimates of risk to
decisionmakers and the public, it should
present not only point estimates of risk, but
also the sources and magnitudes of
uncertainty associated with these estimates.
• Risk managers should be given
characterizations of risk that are both
qualitative and quantitative, i.e., both
descriptive and mathematical.
• EPA should consider in its risk
assessments the limits of scientific
knowledge, the remaining uncertainties, and
the desire to identify errors of either
overestimation or underestimation.
In part as a response to these
recommendations, the Administrator of EPA
issued guidelines for risk characterization
and required implementation plans from all
programs in EPA (U.S. EPA, 1995). The
Administrator's guidance is followed in these
cancer guidelines. The assessments of
hazard, dose response, and exposure will all
have accompanying technical
characterizations covering issues of strengths
and limitations of data and current scientific
understanding, identification of defaults
utilized in the face of gaps in the former,
discussions of controversial issues, and
discussions of uncertainties in both their
qualitative, and as practicable, their
quantitative aspects.
Appendix C
Overview of Cancer Processes
The following picture is changing as
research reveals more about carcinogenic
processes. Nevertheless, it is apparent that
several general modes of action are being
elucidated from direct reaction with DNA to
hormonal or other growth-signaling
processes. While the exact mechanism of
action of an agent at the molecular level may
not be clear from existing data, the available
data will often provide support for deducing
the general mode of action. Under these
guidelines, using all of the available data to
arrive at a view of the mode of action
supports both characterization of human
hazard potential and assessment of dose
response relationships.
Cancers are diseases of somatic mutation
affecting cell growth and differentiation. The
genes that control cell growth, programmed
cell death, and cell differentiation are critical
to normal development of tissues from
embryo to adult metazoan organisms. These
genes continue to be critical to maintenance
of form and function of tissues in the adult
(e.g., Meyn, 1993) and changes in them are
essential elements of carcinogenesis (Hsu et
al., 1991; Kakizuka et al., 1991; Bottaro et al.,
1991; Sidransky et al., 1991; Salomon et al.,
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18005
iaaO; Srivastava ot al., 1990). The genes
Involved arc among tho most highly
conserved in evolution as evidenced by the
groat homology of many of them in DNA
sequence and function in organisms as
phylogonetically distant as worms, insects,
and mammals (Auger et al., 1989a, b;
Hollstein ot al., 1991; Horschman, 1991;
Strausfcld ot al., 1991; Forsburg and Nurse,
1991).
Mutations affecting three general categories
of genes have been implicated in
carcinogcncsis. Over 100 oncogenes have
boon found in human and animal tumors that
act as dominant allelos, whereas there are
about 10 known tumor suppressor genes that
ore recessive in action. The normal allelos of
these genes are involved with control of cell
division and differentiation; mutated alleles
lead to a disruption in these functions. The
third class are mutator genes that predispose
the gonomo to enhanced mutagenic events
that contribute further to the carcinogenic
process.
Adult tissues, oven those that are
composed of rapidly replicating cells,
maintain a constant sizo and cell number
(Nunez et al., 1991) by balancing three cell
fates: (1) continued replication, (2)
differentiation to take on specialized
functions, or (3) programmed cell death
(npoptosis) (Raff, 1992; Mailer, 1991; Naeve
ot al., 1991; Schneider et al., 1991; Harris,
1990). Ncoplastlc growth through clonal
expansion can result from somatic mutations
that inactivate control over cell fate
(Kaklzuka et al., 1991; deThe et al., 1991;
Sidransky ot al., 1992; Nowell, 1976).
Cancers may also bo thought of as diseases
of the coll cycle. For example, genetic
diseases that cause failure of cells to repair
DNA damage prior to cell replication
predispose people to cancer. These changes
are also frequently found in tumor cells in
sporadic cancers. These changes appear to be
particularly involved at points in cell
replication called "checkpoints" where DNA
synthesis or mitosis is normally stopped
until DNA damage is repaired or cell death
induced CToboy, 1975). A cell that bypasses
a checkpoint may acquire a heritable growth
advantage. Similar effects on the cell cycle
occur when mitogons such as hormones or
growth factors stimulate cell growth. Rapid
replication in response to tissue injury may
also lead to unrepaired DNA damage that is
a risk factor forcarcinogenosis.
Normally a cell's fato is determined by a
•limed sequence of biochemical signals.
Signal transduction in tho coll involves
chemical signals that bind to receptors,
generating further signals in a pathway
whose target in many cases is control of
transcription of a specific set of genes
(Hunter, 1991; Cantley et al., 1991; Collum
and Alt, 1990). Colls are subject to growth
signals from the same and distant tissues,
e.g., endocrine tissues (Schuller, 1991). In
addition to hormones produced by endocrine
tissues, numerous soluble polypsptide
growth factors have been identified that
control normal growth and differentiation
(Cross and Dexter, 1991; Wellstein et al.,
1990). Tho colls responsive to a particular
growth factor are those that express
transmombrano receptors that specifically
bind tho growth factor.
Solid tumors develop in stages
operationally defined as initiation,
promotion, and progression (see, for
example, Pitot and Dragan, 1991). These
terms, which were coined in the context of
specific experimental designs, are used for
convenience in discussing concepts, but they
refer to complex events that are not
completely understood. During initiation, the
cell acquires a genetic change that confers a
potential growth advantage. During
promotion, clonal expansion of this altered
cell occurs. Later, during progression, a series
of genetic and other biological events both
enhance the growth advantage of the cells
and enlist normal host processes to support
tumor development and cells develop the
ability to invade locally and metastasize
distally, taking on the characteristics of
malignancy. Many endogenous and
exogenous factors are known to participate in
the process as a whole. These include
specific genetic predispositions or variations
in ability to detoxify agents, medical history
(Harris, 1989; Nehreda et al., 1991),
infections, exposure to chemicals or ionizing
radiation, hormones and growth factors, and
immune suppression. Several such risk
factors likely work together to cause
individual human cancers.
A cell that has been transformed, acquiring
the potential to establish a line of cells that
grow to a tumor, will probably realize that
potential only rarely. The process of
tumorigenesis in animals and humans is a
multistep one (Bouk, 1990; Fearon and
Vogelstein, 1990; Hunter, 1991; Kumar et al.,
1990; Sukumar, 1989; Sukumar, 1990) and
normal physiological processes appear to be
arrayed against uncontrolled growth of a
transformed cell (Weinberg, 1989). Powerful
inhibition by signals from contact with
neighboring normal cells is one known
barrier (Zhang et al., 1992). Another is the
immune system (at least for viral infection).
How a cell with tumorigenic potential •
acquires additional properties that are
necessary to enable it to overcome these and
other inhibitory processes is a subject of
ongoing research. For known human
carcinogens studied thus far, there is an often
decades-long latency between exposure to
carcinogenic agents and development of
tumors (Fidler and Radinsky, 1990; Tanaka et
al., 1991; Thompson et al., 1989). This
latency is also typical of tumor development
in individuals with genetic diseases that
make them prone to cancer (Meyn, 1993;
Srivastava et al., 1990).
The importance of genetic mutation in the
carcinogenic process calls for special
attention to assessing agents that cause such
mutations. Heritable genetic defects that
predispose humans to cancer are well known
and the number of identified defects is
growing. Examples include xeroderma
pigmentosum (DNA repair defect) and Li
Fraumeni and retinoblastoma (both are tumor
suppressor gene mutations). Much of the
screening and testing of agents for
carcinogenic potential has been driven by the
idea of identifying this mode of action.
Cognizance of and emphasis on other modes
of action such as ones that act at the level of
growth signalling within or between cells,
through cell receptors, or that indirectly
cause genetic change, comes from more
recent research. There are" not yet
standardized tests for many modes of action,
but pertinent information may be available in
individual cases.
Agents of differing characteristics
influence cancer development: inorganic and
organic, naturally occurring and synthetic, of
inanimate or animate origin, endogenous or
exogenous, dietary and nondietary. The
means by which these agents act to influence
carcinogenesis are variable, and reasoned
hazard assessment requires consideration of
the multiple ways that chemicals influence
cells in experimental systems and in humans.
Agents exert mutagenic effects either by
interacting directly with DNA or by indirect
means through intermediary substances (e.g.,
reactive oxygen species) or processes. Most
DNA-reactive chemicals are electrophilic or
can become electrophilic when metabolically
activated. Electrophilic molecules may bind
covalently to DNA to form adducts, and this
may lead to depurination, depyrimidation, or
produce DNA strand breaks; such lesions can
be converted to mutations with a round of
DNA synthesis and cell division. Other DNA-
interactive chemicals may cause the same
result by intercalation into the DNA helix.
Still other chemicals may methylate DNA,
changing gene expression. Non-DNA-reactive
chemicals produce genotoxic effects by many
different processes. They may affect spindle
formation or chromosome proteins, interfere
with normal growth control mechanisms, or
affect enzymes involved with ensuring the
fidelity of DNA synthesis (e.g.,
topoisomerase), recombination, or repair.
The "classical" chemical carcinogens in
laboratory rodent studies are agents that
consistently produce gene mutations and
structural chromosome aberrations in short-
term tests. A large database reveals that these
mutagenic substances commonly produce
tumors at multiple sites and in multiple
species (Ashby and Tennant, 1991). Most of
the carcinogens identified in human studies,
aside from hormones, are also gene or
structural chromosome mutagens (Tennant
and Ashby, 1991). Most of these compounds
or their metabolites contain electrophilic
moieties that react with DNA.
Numerical chromosome aberrations, gene
amplification, and the loss of gene
heterozygosity are also found in animal and
human tumor cells and may arise from
initiating events or during progression. There
is reason to believe that accumulation of
additional genetic changes is favored by
selection in the evolution of tumor cells
because they confer additional growth
advantages (Hartwell and Kastan, 1994).
Exogenous agents may function at any stage
of carcinogenesis (Barrett, 1993). Some
aberrations may arise as a consequence of
genomic instability arising from tumor
suppressor gene mutation, e.g., p53 (Harris
and Hollstein, 1993). The frequent
observation in tumor cells that both of a pair
of homologous chromosomes have identical
mutation spectra in tumor suppressor genes
suggests an ongoing, endogenous process of
gene conversion. Currently, there is a paucity
of routine test methods to screen for events
such as gene conversion or gene
amplification and knowledge regarding the
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Federal Register / Vol. 61, No. 79 / Tuesday, April 23, 1996 / Notices
ability of particular agents of environmental
interest to induce them is, for the most part,
wanting. Work is under way to characterize,
measure, and evaluate their significance
(Travis et al., 1991).
Several kinds of mechanistic studies aid in
risk assessment. Comparison of DNA lesions
in tumor cells taken from humans with the
lesions that a tumorigenic agent causes in
experimental systems can permit inferences
about the association of exposure to the agent
and an observed human effect (Vahakangas et
al., 1992; Hollstein et al., 1991; Hayward et
al., 1991). An agent that is observed to cause
mutations experimentally may be inferred to
have potential for carcinogenic activity (U.S.
EPA, 1991a). If such an agent is shown to be
carcinogenic in animals, the inference that its
mode of action is through mutagenicity is
strong. A carcinogenic agent that is not
mutagenic in experimental systems but is
mitogenic or affects hormonal levels or
causes toxic injury followed by
compensatory growth may be inferred to
have effects on growth signal transduction or
to have secondary carcinogenic effects. The
strength of these inferences depends in each
case on the nature and extent of all the
available data. ,
Differing modes of action at the molecular
level have different dose response
implications for the activity of agents. The
carcinogenic activity of a direct-acting
mutageh should be a function of the
probability of its reaching and reacting with
DNA. The carcinogenic activity of an agent
that interferes at the level of signal pathways
with many potential receptor targets should
be a function of multiple reactions. The
carcinogenic activity of an agent that acts by
causing cell toxicity followed by
compensatory growth should be a function of
the toxicity.
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