DNA damage
mutagenicity and genotoxicity
DNA:
- principal molecule for life of the cell
- structure and function carefully checked
- changes rapidly repaired
- irreversible changes -> cell death (apoptosis)
Mutagenesis - MUTATIONS
- changes in the sequences of deoxynucleotides
- natural mutations (billions of nucleotides/day)
: variability in genoms; reparations
- chemical-induced mutagenesis
DNA damage
DNA repair
Damage of DNA is carefully controlled
constitutively expressed proteins
Changes in DNA
induction of reparation enzymes ("SOS-repair")
= biomarker of DNA damage
Induced mutations
MUTAGENS
- ionizing radiation and UV
- chemicals
Base analogs - inserted into the DNA strand during replication in
place of the substrates.
Agents reacting with DNA - structural changes leading to
miscopying of the template strand
Indirect mutagens - affect cells that synthesize chemicals with
direct mutagenic effect
Point mutations
BASE - EXCHANGE:
ˇSilent mutations:
* code for the same amino acid.
ˇMissense mutations:
* code for a different amino acid.
ˇNonsense mutations:
* which code for a stop
Point mutation
INSERTION
DELETION
Change of the
reading frame
Large scale mutations / chromosomal
Physical factors & DNA damage
Ionizating radiation
- direct interaction with hydrogen atoms in water
(and bases)
-> OH* radicals; H2O2, O2-
oxidation of bases; dimerization ...
UV radiation
- interaction with aromatic cycles (bases)
- base dimerization (T=T)
Ionizing radiation effects on DNA
Chemical induced DNA damage
Bases analogs
- incorporation
into DNA
during replication
(5-Br-Uracil: AT -> GC)
Chemical induced DNA damage
HNO2, HSO3
-,Hydroxylamine, Methoxyamine
deamination of bases
(GC -> AT)
Chemical induced DNA damage
Alkylsulphates, N-nitroso-alkyles, cis-platinum
- alkylation of bases; crosslinks of dsDNA
cisplatin
cyclophosphamide
Chemical induced DNA damage
INTERCALATION & ADDUCT FORMATION
Polycyclic aromatic hydrocarbons (PAHs) & derivatives (Nacetyl-2-aminofluorene
(AAF), benzo[a]pyrene)
Mycotoxins (aflatoxins) aduct formation with DNA (biomarkers)
Psoralen
DNA intercalation
Trosko and Ruch 1998, Frontiers in Bioscience 3:d208
AhR
ER
GJIC
Oxidative stress
IMPORTANT PROCESSES IN CANCEROGENESIS
Does chemically-induced genotoxicity results in in vivo effects
- adducts from mitochondrial DNA ?
- distance between ,,source of radicals" and nuclear DNA ?
- protection mechanisms (mutation -> death/apoptosis)
Rubin (2002) Oncogene 21:7392
Thilly (2003) Nature Genetics 34(3):255
Mutations are not caused by chemicals
Chemicals only allow ,,unveil" previously existing mutations in
nuclear DNA (non-genotoxic events cause cancer !!!)
Redox homeostasis & oxidative stress
Redox homeostasis
- natural levels of oxidants (O2) and antioxidants in each cell
Disruption of redox homeostasis
-> depletion of oxygen: metabolism disruption, acidosis in
tissues, cell necrosis
rare: INSIDE TUMORS
-> overproduction of oxidants:
= oxidative stress
GENERAL MECHANISM OF TOXICITY
Overproduction of oxidants
Oxygen ­ principal molecule in living organisms
Oxygen increase or reactive derivatives -> toxicity
ROS = Reactive Oxygen Species: Sources
- production in mitochondria (byproducts)
- redox-cycling (quinones of xenobiotics)
- Fenton-reaction (metals)
- oxidations mediated via MFO (CYP)
- depletion of antioxidants (reactive molecules)
Reactive Oxygen Species (ROS)
SOD = Superoxide dismutase
Fenton reaction
Reactive Oxygen Species (ROS)
ROS & mitochondria
Examples of chemical-induced
oxidative stress
- Metals:
fenton reaction -> OH*
- Redox-cycling chemicals:
oxy-PAHs
- Depletion of GSH:
reactive molecules,
GST-conjugation,
metals: SH oxidation ...
O2
O2
O
O
e O
-
O
.
. e
-
O O
-
2e -
Antioxidant depletion
GSH (glutathione)
Biomarkers of oxidative damage