Mechanistic and other relevant data may provide evidence
of carcinogenicity and also help in assessing the relevance
and importance of findings of cancer in animals and in
humans. The nature of the mechanistic and other relevant
data depends on the biological activity of the agent being
considered. The Working Group considers representative
studies to give a concise description of the relevant
data and issues that they consider to be important; thus,
not every available study is cited. Relevant topics may
include toxicokinetics, mechanisms of carcinogenesis,
susceptible individuals, populations and life-stages,
other relevant data and other adverse effects. When data
on biomarkers are informative about the mechanisms of
carcinogenesis, they are included in this section.
These topics are not mutually exclusive; thus, the same
studies may be discussed in more than one subsection.
For example, a mutation in a gene that codes for an enzyme
that metabolizes the agent under study could be discussed
in the subsections on toxicokinetics, mechanisms and individual
susceptibility if it also exists as an inherited polymorphism.
(a) Toxicokinetic data
Toxicokinetics refers to the absorption, distribution,
metabolism and elimination of agents in humans, experimental
animals and, where relevant, cellular systems. Examples
of kinetic factors that may affect dose-response relationships
include uptake, deposition, biopersistence and half-life
in tissues, protein binding, metabolic activation and
detoxification. Studies that indicate the metabolic fate
of the agent in humans and in experimental animals are
summarized briefly, and comparisons of data from humans
and animals are made when possible. Comparative information
on the relationship between exposure and the dose that
reaches the target site may be important for the extrapolation
of hazards between species and in clarifying the role
of in-vitro findings.
(b) Data on mechanisms of carcinogenesis
To provide focus, the Working Group attempts to identify
the possible mechanisms by which the agent may increase
the risk of cancer. For each possible mechanism, a representative
selection of key data from humans and experimental systems
is summarized. Attention is given to gaps in the data
and to data that suggests that more than one mechanism
may be operating. The relevance of the mechanism to humans
is discussed, in particular, when mechanistic data are
derived from experimental model systems. Changes in the
affected organs, tissues or cells can be divided into
three non-exclusive levels as described below.
(i) Changes in physiology
Physiological changes refer to exposure-related modifications
to the physiology and/or response of cells, tissues and
organs. Examples of potentially adverse physiological
changes include mitogenesis, compensatory cell division,
escape from apoptosis and/or senescence, presence of inflammation,
hyperplasia, metaplasia and/or preneoplasia, angiogenesis,
alterations in cellular adhesion, changes in steroidal
hormones and changes in immune surveillance.
(ii) Functional changes at the cellular level
Functional changes refer to exposure-related alterations
in the signalling pathways used by cells to manage critical
processes that are related to increased risk for cancer.
Examples of functional changes include modified activities
of enzymes involved in the metabolism of xenobiotics,
alterations in the expression of key genes that regulate
DNA repair, alterations in cyclin-dependent kinases that
govern cell cycle progression, changes in the patterns
of post-translational modifications of proteins, changes
in regulatory factors that alter apoptotic rates, changes
in the secretion of factors related to the stimulation
of DNA replication and transcription and changes in gap-junction-mediated
intercellular communication.
(iii) Changes at the molecular level
Molecular changes refer to exposure-related changes in
key cellular structures at the molecular level, including,
in particular, genotoxicity. Examples of molecular changes
include formation of DNA adducts and DNA strand breaks,
mutations in genes, chromosomal aberrations, aneuploidy
and changes in DNA methylation patterns. Greater emphasis
is given to irreversible effects.
The use of mechanistic data in the identification of
a carcinogenic hazard is specific to the mechanism being
addressed and is not readily described for every possible
level and mechanism discussed above.
Genotoxicity data are discussed here to illustrate the
key issues involved in the evaluation of mechanistic data.
Tests for genetic and related effects are described in
view of the relevance of gene mutation and chromosomal
aberration/aneuploidy to carcinogenesis (Vainio et
al., 1992; McGregor et al., 1999). The adequacy
of the reporting of sample characterization is considered
and, when necessary, commented upon; with regard to complex
mixtures, such comments are similar to those described
for animal carcinogenicity tests. The available data are
interpreted critically according to the end-points detected,
which may include DNA damage, gene mutation, sister chromatid
exchange, micronucleus formation, chromosomal aberrations
and aneuploidy. The concentrations employed are given,
and mention is made of whether the use of an exogenous
metabolic system in vitro affected the test result.
These data are listed in tabular form by phylogenetic
classification.
Positive results in tests using prokaryotes, lower eukaryotes,
insects, plants and cultured mammalian cells suggest that
genetic and related effects could occur in mammals. Results
from such tests may also give information on the types
of genetic effect produced and on the involvement of metabolic
activation. Some end-points described are clearly genetic
in nature (e.g. gene mutations), while others are associated
with genetic effects (e.g. unscheduled DNA synthesis).
In-vitro tests for tumour promotion, cell transformation
and gap-junction intercellular communication may be sensitive
to changes that are not necessarily the result of genetic
alterations but that may have specific relevance to the
process of carcinogenesis. Critical appraisals of these
tests have been published (Montesano et al., 1986;
McGregor et al., 1999).
Genetic or other activity manifest in humans and experimental
mammals is regarded to be of greater relevance than that
in other organisms. The demonstration that an agent can
induce gene and chromosomal mutations in mammals in vivo
indicates that it may have carcinogenic activity. Negative
results in tests for mutagenicity in selected tissues
from animals treated in vivo provide less weight,
partly because they do not exclude the possibility of
an effect in tissues other than those examined. Moreover,
negative results in short-term tests with genetic end-points
cannot be considered to provide evidence that rules out
the carcinogenicity of agents that act through other mechanisms
(e.g. receptor-mediated effects, cellular toxicity with
regenerative cell division, peroxisome proliferation)
(Vainio et al., 1992). Factors that may give misleading
results in short-term tests have been discussed in detail
elsewhere (Montesano et al., 1986; McGregor et
al., 1999).
When there is evidence that an agent acts by a specific
mechanism that does not involve genotoxicity (e.g. hormonal
dysregulation, immune suppression, and formation of calculi
and other deposits that cause chronic irritation), that
evidence is presented and reviewed critically in the context
of rigorous criteria for the operation of that mechanism
in carcinogenesis (e.g. Capen et al., 1999).
For biological agents such as viruses, bacteria and parasites,
other data relevant to carcinogenicity may include descriptions
of the pathology of infection, integration and expression
of viruses, and genetic alterations seen in human tumours.
Other observations that might comprise cellular and tissue
responses to infection, immune response and the presence
of tumour markers are also considered.
For physical agents that are forms of radiation, other
data relevant to carcinogenicity may include descriptions
of damaging effects at the physiological, cellular and
molecular level, as for chemical agents, and descriptions
of how these effects occur. 'Physical agents' may also
be considered to comprise foreign bodies, such as surgical
implants of various kinds, and poorly soluble fibres,
dusts and particles of various sizes, the pathogenic effects
of which are a result of their physical presence in tissues
or body cavities. Other relevant data for such materials
may include characterization of cellular, tissue and physiological
reactions to these materials and descriptions of pathological
conditions other than neoplasia with which they may be
associated.
(c) Other data relevant to mechanisms
A description is provided of any structure-activity relationships
that may be relevant to an evaluation of the carcinogenicity
of an agent, the toxicological implications of the physical
and chemical properties, and any other data relevant to
the evaluation that are not included elsewhere.
High-output data, such as those derived from gene expression
microarrays, and high-throughput data, such as those that
result from testing hundreds of agents for a single end-point,
pose a unique problem for the use of mechanistic data
in the evaluation of a carcinogenic hazard. In the case
of high-output data, there is the possibility to overinterpret
changes in individual end-points (e.g. changes in expression
in one gene) without considering the consistency of that
finding in the broader context of the other end-points
(e.g. other genes with linked transcriptional control).
High-output data can be used in assessing mechanisms,
but all end-points measured in a single experiment need
to be considered in the proper context. For high-throughput
data, where the number of observations far exceeds the
number of end-points measured, their utility for identifying
common mechanisms across multiple agents is enhanced.
These data can be used to identify mechanisms that not
only seem plausible, but also have a consistent pattern
of carcinogenic response across entire classes of related
compounds.
(d) Susceptibility data
Individuals, populations and life-stages may have greater
or lesser susceptibility to an agent, based on toxicokinetics,
mechanisms of carcinogenesis and other factors. Examples
of host and genetic factors that affect individual susceptibility
include sex, genetic polymorphisms of genes involved in
the metabolism of the agent under evaluation, differences
in metabolic capacity due to life-stage or the presence
of disease, differences in DNA repair capacity, competition
for or alteration of metabolic capacity by medications
or other chemical exposures, pre-existing hormonal imbalance
that is exacerbated by a chemical exposure, a suppressed
immune system, periods of higher-than-usual tissue growth
or regeneration and genetic polymorphisms that lead to
differences in behaviour (e.g. addiction). Such data can
substantially increase the strength of the evidence from
epidemiological data and enhance the linkage of in-vivo
and in-vitro laboratory studies to humans.
(e) Data on other adverse effects
Data on acute, subchronic and chronic adverse effects
relevant to the cancer evaluation are summarized. Adverse
effects that confirm distribution and biological effects
at the sites of tumour development, or alterations in
physiology that could lead to tumour development, are
emphasized. Effects on reproduction, embryonic and fetal
survival and development are summarized briefly. The adequacy
of epidemiological studies of reproductive outcome and
genetic and related effects in humans is judged by the
same criteria as those applied to epidemiological studies
of cancer, but fewer details are given.
Posted 23 January 2006
