Toxicogenomics

Dr. M. Zdanowicz
 WHY?
Why didnt everyone living the same
distance from Chernobyl develop
cancer?
Why can some people smoke for their
entire lives and never develop lung
cancer?
Why do certain individuals experience
toxicity from a certain level of a drug or
toxin when others dont?
 Definitions
Toxicology:
The study of the nature, effects and detection
of poisons and the treatment of poisoning.

Genomics:
The study of genes and their function.
Genomics aims to understand the structure of
the genome, including the mapping genes
and sequencing the DNA. Genomics examines
the molecular mechanisms and the interplay
of genetic and environmental factors in
disease.
 Proteomics:
The measurement, identification &
comparison of all proteins (and their
properties) in specific tissues and
body fluids during health, toxicity and
disease
 Pharmacogenomics:
The study of the interaction of an
individual's genetic makeup and
response to a drug.

Branch of pharmaceutics which aims

to tailor medicines to an individuals
genetic makeup
 Toxicogenomics
The collection, interpretation, and
storage of information about gene
and protein activity in order to
identify toxic substances in the
environment, and to help treat people
at the greatest risk of diseases
caused by environmental pollutants or
toxicants.
The dose makes the
poison
Paraclesus
 ENVIRONMENTAL

STRESSORS
GENETIC
INFLUENCES
X TIME = DISEASE
 Key Questions
1. How does exposure to
chemicals (toxins, drugs) effect
the expression of human
genes??
2. Do we all respond the same
when exposed to chemicals,
drugs and toxins??
3. Are some of us more resistant
to their detrimental effects??
 There is a growing number of
potentially toxic compounds in the
human environment.
 National Institute of
Environmental Health
Formed the National Center for
Toxicogenomics
http://www.niehs.nih.gov/nct/home.htm

Charged with gathering large

amounts of gene expression data
related to toxicology.
 Comparison
Classic Toxicology Toxicogenomics

How much Identifying

exposure and for agents that are
how long a time associated with
does the toxicity in either
population the average or
need to be more sensitive
exposed to show individual.
toxicity?
Classic Measures New Measures
of Toxicity
Gene
Histopathology Expression:
Clinical DNA Microarrays
Chemistry RT PCR
Northern Blot
Metabolism
Sequencing
Physiology Protein
Enzymology Expression:
Electron Western Blot
Microscopy Electrophoresis
Chips
 Two Key Advances
Human Genome Project:
Sequences the human genome.
Identifies genetic variations between
individuals (polymorphisms).
 DNA Microarrays:
Revolutionary new tool used to
identify mutations in genes.
The chip consists of a small glass
plate encased in plastic.
Each chip contains thousands of
short, synthetic, single-stranded DNA
sequences which together add up to
the normal gene(s) in question
Hand held DNA microarray
 The populations of expressed genes (mRNA)
from different cells or tissues are labeled
with fluorescent tags and hybridized to the
array of DNA fragments.
The array is then scanned with a laser, at
the appropriate wavelength, and
fluorescence measurements are made at
each location on the array to give a
measure of how many gene transcripts are
seen in that sample.
DNA Microarray

Each dot represents

The activity of a single gene.

The brighter the dot, the

Greater the activity!
 By comparing the levels of all these genes
between many samples, we may understand
what molecular changes are occurring at the
transcription level during biological changes.
The advantage over traditional methods
such as Northern blot is that you can
analyze 40,000 genes per sample versus a
dozen or so.
4 million soon!
 Examples
A 24 hour exposure to ethanol in rat
liver cells induces a change in
expression of 86 hepatocyte genes

A brief exposure to a chemical that

causes toxicity through DNA
alkylation alters the expression of
2000 genes in a single cell!
 DNA microarrays may also be used
test a patients blood for the
presence of mutations that might
be associated with certain
diseases.
Ex. BRCA1 & BRCA2 genes and
breast cancer
 Advantages of Measuring
Gene Expression in
Toxicology
Greater sensitivity:
Can detect changes in gene expression
before gross changes such as tissue
damage, tumor formation, etc

Earlier Indication of a Toxic

Response:
Before tissue is altered or tumors form.
 Advantages of Measuring
Gene Expression in
Toxicology
Greater Specificity:
Many different agents can cause a liver
tumor but they may do so through a
number of different mechanisms.
Many toxic agents have the same endpoint
but get there by various pathways we may
now identify.
Also may provide a way to determine the
mechanism of toxicity for a particular
agent.
 Advantages of Measuring
Gene Expression in
Toxicology
Microarrays allow for study of all
toxicological endpoints in a single
assay.

Traditionally, you need to study an agents

carcinogenicity, mutagenicity,
reproductive toxicity,
immunotoxicity, neurotoxicity, and
endocrine toxicity in separate assays.
 Advantages of Measuring
Gene Expression in
Toxicology
The fact that groups of chemical with a
common toxicological mechanism
produce a characteristic pattern
(fingerprint) of gene expression
means it may be possible to discern the
toxic potential of an agent quickly and
cheaply.
A way to cut back on animal
toxicity testing.
 Applications of
Toxicogenomics
 Liver Metabolizing
Enzymes
Most common and well-studied
genetic variations.

Metabolize numerous drugs and

potential environmental toxins.

Can persons with genetically different

enzyme activity be at increased risk
for toxicity and disease?
Nakajima, 2000
 Examples
CYP1A1:
oxidizes polycyclic aromatic
hydrocarbons..increased lung cancer
if reduced.
Glutathione S-Transferases:
Detoxify carcinogenic metabolites such
as hydrocarbon diol-epoxides and
lipoperoxidation productsincreased
skin & lung cancers if reduced.
 Remember, through metabolism
these liver enzymes detoxify
potentially dangerous agents but
they can also form toxic
metabolites from relatively inert
parent compounds!
High rates of activity for certain
enzymes may likewise be a risk!!
 DNA Repair Genes
DNA Damage and DNA adducts
(DNA with covalent chemicals
attached).

Large difference in DNA damage

and adduct formation have been
documented in different individuals
receiving similar exposures.
BP = Benzo[a] pyrene
A polyclycic aromatic
hydrocarbon
 DNA repair genes are polymorphic.
They function at different levels in
different individuals.

Individuals may have significant

differences in their ability to repair
DNA damaged by exposure to toxins,
uv, radiation, chemo drugs, etc.
 What is a known outcome of
DNA damage?

Cancer!
 How can Joe Schmoe smoke three
packs of butts a day for his entire
life and never get lung cancer???
Cigarettes are really safe!
High activity of genes that detoxify
carcinogens?
Low Activity of genes that form
carcinogenic metabolites?
High activity of DNA repair genes?
 Key Point
How might Toxicogenomics and
DNA microarrays be of value
here with respect to variability in
liver enzymes and DNA repair??

It may be used to determine which

individuals would be at risk for long-
term adverse effects following
exposure to certain toxins..
 Mechanism of Drug
Teratogenicity
Valproic Acid:
Avoided in pregnancy due to birth defects.
Mechanism of teratogenicity is unknown.
Kultima (2000) used toxicogenomics and
DNA microarrays to show that the
expression of a number of mouse fetal
genes is increased following valproic acid
exposure.
One is for metallothionein.fetal Zn+
deficiency!! (A key cofactor)
 ToxChip
Chip has been developed that
contains most of the human genes
known to be involved in a
toxicological response.
ToxChip Genes

Adapted from Nuwaysir, 1999

 Microarray profiling of individuals
who might be exposed to toxic
agents may act as a sensitive
biomarker to define more precisely
the nature and level of toxic
exposure the individual is
experiencing.before it is obvious
through tissue injury or disease..
 Summary
Toxicogenomics is an emerging
area of toxicology that
examines how the genetic
make-up of an individual
makes them or more less
susceptible to toxicity and
exposure related diseases.
 Summary
In contrast with classic
toxicology, toxicogenomics
uses changes in gene and
protein expression to detect
subtle molecular-level changes
in organism that may precede
or predict toxic and deleterious
effects of exposure.
 Summary
The development of so called
Tox-Chips may allow for the
rapid and sensitive detection
of wide-ranging molecular
level changes in an organism
exposed to various substances.
 Summary
Toxicogenomics can provide a
powerful means of detecting
molecular level changes in an
organism before obvious
manifestations of toxicity or
disease are present.