Download Carcinogenesis

Introduction to Toxicology
EV 460/660 & BI 460/660
Fall 2014
Carcinogenesis
1.
Historical perspective – linking chemical exposures to cancer
-- 18th century – Hill - nasal cancer in snuff users
-- Pott – scrotal cancer in chimney sweeps
-- 19th century – Rehn – bladder cancer in synthetic dye factory workers
-- early 20th century – coal tar application and dermal cancer in rabbits
-- polycyclic aromatic hydrocarbon (PAH) extracts of coal later found to be active
components-- dimethylbenz(a)anthracene & benzo(a) pyrene
-- 2-naphthylamine found to be active component in synthetic dyes
2.
Cancer – not a single disease, general term referring to multiple types of malignant tumors
3.
Terminology
-- neoplasia – uncontrolled cell growth and proliferation
-- neoplasm – the mass of cells resulting from neoplasia, a tumor
-- benign vs. malignant tumors -- characteristics
-- carcinogenesis – process of cancer (malignant tumor) development
-- carcinogen – chemical, biological, or physical factor associated with development of malignant
tumors
-- carcinomas – cancer of epithelial tissues
-- sarcomas – cancer of connective and muscle tissues
4.
TNM scoring system for staging of tumors – use in diagnosis and treatment
T = tumor development (0-4)
N = involvement of lymph nodes in region of tumor (0-4)
M = degree of metastasis (0-4)
5.
Multistage model of carcinogenesis: Initiation, Promotion, and Progression
A. Initiation – rapid, irreversible change in cell’s genetic material
-- initiating chemicals are electrophiles or are metabolically activated to electrophiles,
covalently bind to DNA to bring about a mutagenic change
-- cell is primed or initiated, transformed for subsequent neoplastic development
-- initiated cell requires a cycle of cell replication to “fix” the genetic change
-- initiated cell may be latent or dormant until acted upon by promoting agents
-- promoting agent allows growth of clones from initiated cell to produce tumor
B. Promotion – promoting agents are not in themselves carcinogens, but when given repetitively
after an initiating agent, can increase the number of tumors or decrease the latency period
-- typically repetitive exposures to the promoter is required, action of promoter may be
reversible at early stages, these suggest that there may be a threshold level of exposure
-- promoting agents are not usually electrophiles and do not bind to DNA
-- promoters may be non chemical factors such as physical injury
-- tumor promoters are often organ specific
C. Progression – tumor growth and development of malignant tumor
6.
7.
Direct vs. Indirect Carcinogens
A.
Direct carcinogens also called primary carcinogens
-- can cause cancer without being first acted upon by body, e.g., do not require
bioactivation
B.
Indirect carcinogens also called secondary carcinogens
-- require bioactivation to become carcinogenic
-- activation occurs through biotransformation reactions, often in liver
C.
Terminology associated with indirect carcinogens
-- Procarcinogen – parent compound
-- chemical that requires metabolism to exert carcinogenic effect
-- Proximate Carcinogen – metabolites of the parent compound
-- intermediates between procarcinogen and ultimate carcinogen
-- Ultimate Carcinogen – metabolic product that actually interacts with cellular components
-- chemical responsible for carcinogenic activity
Genotoxic and Epigenetic Carcinogens -- different mechanisms in carcinogenesis
A.
Genotoxic chemicals – directly damage or modify DNA
-- induce mutations (see later on mutagenesis) in DNA
-- covalently bind to nucleic acids, often interact with purine and
pyrimidine bases of DNA
-- the damaged DNA with bound chemical is a DNA adduct
B.
Epigenetic chemicals – carcinogenic effect through mechanisms other than genotoxic
action
-- indirect mechanisms may include: alteration in gene expression, hormonal
imbalances, immunosuppression, cytotoxicity, promoting effects, and
cocarcinogenic action
8.
Cocarcinogens – agents not themselves carcinogenic but increase tumor incidence when
applied just before or together with carcinogen
-- contrast with promoter model, e.g., chemical applied after initiation
-- however, some chemicals may act as both a promoter and a cocarcinogen
-- cocarcinogenesis may result from non-chemical factors such as physical trauma
-- cocarcinogenesis may be from hormonal, viral, immunologic, and nutritional factors
-- cocarginogens exert their effect by a variety of mechanisms, including: increased
uptake of carcinogen by cells, increased bioactivation of procarcinogens,
inhibition of DNA repair mechanisms, and others
9.
DNA repair mechanisms – variety of repair mechanisms
-- reduce incidence of cancer development following exposure to genotoxic agents
-- one example, excision repair – DNA region containing adduct is removed and
replaced with new area of DNA synthesized using opposite intact strand
-- repair must occur before cell replication “fixes” the damaged sequence
-- agents altering repair processes or rate of cell division will affect tumor frequency
10.
Oncogenes -- activated oncogenes can transform cells from normal to neoplastic
-- are protooncogenes that have been initiated by chemical or biological agent
-- protooncogenes in normal cells function in control of cellular growth, division, and
differentiation; many encode for proteins involved in biochemical signals
regulating mitosis
-- many protooncogenes are involved in physiological growth factors (i.e., epidermal
growth factor)
11.
Tumor Suppressor Genes – code for proteins that are negative regulators of cell growth
-- limit cell proliferation by holding cell in F stage of cell cycle
-- mutations, deletion, or inactivation of suppressor genes removes
negative regulation of cellular growth and division
-- typically, both copies of gene must be affected for loss of
regulation to occur
12.
Screening for chemical carcinogens
-- many bioassays are actually tests of mutagenicity, but this is reasonable since most known
carcinogens are mutagenic
QSAR (Quantitative Structure Activity Relationships) analysis can be used to tentatively
suggest which compounds may have a higher probability of carcinogenic potential
A.
Ames test
-- depends on ability of chemical to cause reverse mutations in mutant Salmonella
typhimurium strains that have defects in the histidine biosynthesis pathway
-- mutants can not grow on histidine-free media, if get colony growth on histidine-free
media then a reverse mutation back to wild type (capable of producing histidine)
occurred
-- typical experimental design includes:
a. positive control with known mutagen
b. negative control (no test compound) to determine background rate of back mutations
c. experimental – multiple concentrations of chemical to be tested
-- an additional manipulation is to run some plates with the S-9 fraction included
-- S-9 fraction is derived from liver of rats exposed to PAHs, includes enzymes capable of
activating indirect carcinogens
-- different S. typhimurium strains can be used to determine type of mutation induced by
chemical agent
-- a variety of similar, related tests are also available (i.e., reverse mutations in E. coli,
forward mutations in S. typhimurium, etc.)
B.
Mammalian cell culture tests
-- a variety of cell lines and test protocols are available, established cells lines (i.e., Chinese
hamster ovary cells or mouse lymphoma cells) are used because the cells clone well and tend
not to become senescent; however, the cells show little metabolic activation of indirect
carcinogens; tests are rapid and relatively inexpensive, but there is concern regarding
generalizations to whole animals; three commonly used tests are described below
I.
mutation at the HGPRT locus (hypoxanthine guanine phosphorioxyl transferase)
-- HGPRT enzyme is responsible for purines from medium into nucleotide bases
-- test depends on mutation involving uptake of purine bases into DNA
-- if cells can grow on media containing toxic purine base, then mutation has
occurred
II.
mutation at the OU locus (ouabain)
-- ouabain kills cells by acting on the membrane Na+/K+-ATPase
-- if cells grow on media containing ouabain, then a mutation has occurred at the
OU locus
III.
mammalian cell transformation test
-- typical utilizes fibroblast cultures from mouse embryos
-- normal cells grown in dishes show contact inhibition of growth, cells grow until
a monolayer of cells fills the plate, then cell growth stops
-- cells that have been transformed by chemical exposure do not show the growth
inhibition and will develop multi-layered clumps of cells or foci
C.
Whole animals tests
-- distinct advantages to using whole animal with interrelated organ systems and
biotransformation mechanisms
-- difficulties associated with added costs and time requirements; complex statistical
analyses, animals rights ethics, and extrapolation of animals data to humans
-- can calculate potency indices such as TD50 value (TD = tumorigenic dose) in
mg/kg/day
-- variety of rodent test systems available to test for induction of tumors of skin, lungs,
breast, liver, etc
13.
Decision-making on potential human carcinogens
-- if a chemical is labeled as a potential human carcinogen then there may be major economic,
industrial, and legal impacts
-- there are various criteria used in decision-making to identify a chemical as a carcinogen,
i.e. -- increased incidence of neoplasms in two or more test species
-- increased incidence of neoplasms in same species, but in separate laboratories
-- one test species, but exceptionally high incidence of neoplasms
-- typically a multi-tiered approach is used in decision-making and final decisions are made
on the basis of the “weight of evidence” of all available data
14.
Some Useful Web Sites
National Cancer Institute
http://www.nci.nih.gov/
International Agency for Research on Cancer
American Cancer Society
http://www.iarc.fr
http://www.cancer.org/