Chemical Carcinogenesis

1
Chemical Carcinogenesis

• David.bell_at_nottingham.ac.uk
• http//dmg.nott.ac.uk/teaching/cancer/pharm1.ppt
• http//dmg.nott.ac.uk/teaching/cancer/
• Poland and Knutson Annu. Rev. Pharmacol. Toxicol.
  22 517-554
• Gu YZ, Hogenesch JB, Bradfield CA The PAS
  superfamily Sensors of environmental and
  developmental signals ANNU REV PHARMACOL 40
  519-561 2000

2
Latency
Cancer takes 10-50 years to develop in man
1900- Cigarette smoking an accepted habit
1940- rise in male lung cancer- alarm
1960- smoking is shown to be the cause of lung
cancer
1960-2000. People who started smoking before
1960 die of lung cancer Lung cancer is the most
frequent site of cancer in UK men
3
Causes of cancer
Note that diet, tobacco, occupation, alcohol and
pollution are principally chemically-induced
cancers.
4
Screening for carcinogens

• Animal tests
• 50 animals per group
• Maximal tolerated dose (MTD), and 25
• Cost 250 000
• Time 2 years (rat or mouse)

5
Problems with bioassay

• The US National Toxicology Programme has tested gt
  500 compounds, with selection based solely on
  environmental relevance
• Approx 59 of all chemicals cause cancer
• Food 1000s of chemicals

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Genotoxins vs non-genotoxins

• Mutagenic (genotoxic) carcinogens can be easily
  detected (e.g. Ames test)
• Non-genotoxic carcinogens account for 30-50 of
  rodent bioassay carcinogens
• No clear test for non-genotoxins
• Consequences for development of pharmaceuticals

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The numbering of the beast
4
6
3
5
7
8
2
10
9
1
2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD) One of
the most potent of a variety of related
compounds Long half-life in humans (6 years)
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TCDD is a potent toxin

• LD50 values
• 1mg/kg guinea pig
• 20-50mg/kg rat
• 70mg/kg monkey
• 5000mg/kg hamster
• Highly toxic in chicken, fish
• Lethality involves a wasting syndrome,
  involving many bodily organs, and taking 2-4 weeks

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TCDD and cancer

• Tumorigenic in rat, mouse, guinea pig and fish
  (everything tested)
• Primate study aborted when total dose of 2 mg/kg
  lead to death
• In rat, the lowest carcinogenic dose was 1.4
  ng/kg/day (Plt0.05) for thyroid tumours
• MTD 71ng/kg/day

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A prototypical non-genotoxin

• TCDD is a potent carcinogen
• Very weak/ no mutagenicity in Ames test
• No activity in tests of DNA damage
• Maximum estimate for binding of TCDD to rat liver
  DNA 1 molecule per 1011 bases
• 1 per 107- 105 for known carcinogens
• TCDD is a non-genotoxic carcinogen

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TCDD is a tumour promoter
DEN
Vehicle control- 6 months
0 tumours
DEN
TCDD- 6 months
5/7 tumours
TCDD- 6 months
0 tumours
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History of TCDD

• 1976 Seveso (http//www.roche.com/roche/about/esev
  esa.htm)
• A reaction vessel containing trichlorophenol
  overheated, and exploded. The resulting TCDD
  contamination (gt100g) led to the evacuation of
  people over a wide area.
• 1999 Belgium
• Chick edema disease- contaminated chicken
  foodstuffs

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Pleiotropic response

• Induction of
• cytochromes P450 1A1, 1A2 and 1B1
• CYP1A1 is known as aryl hydrocarbon hydroxylase
  (AHH)
• all three P450s are potent in the metabolism of
  aromatic hydrocarbons
• UPDGT
• GST

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Related chemicals
Cl
Cl
Cl
Cl

• Chlorinated dibenzofurans
• Chlorinated biphenyls
• Chlorinated naphthalenes
• Polycyclic Aromatic Hydrocarbons
• Similar effects
• Less potent
• Same pleiotropic response

O
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Structure-Activity Relationships

• The ED50 for induction of AHH in chick embryo is
  0.3 nmol/kg with TCDD
• 15 different halogenated DbDs showed an SAR for
  induction of AHH
• gt 3 Cl
• Occupy 2,3,7,8 positions
• gt1 ring position unsubstituted
• Correlation between potency for inducing AHH and
  toxicity

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SARs- PAH

• 3-methylcholanthrene (3MC) is a PAH
• Induces AHH (CYP1A1)
• TCDD and 3MC in rat liver
• Produce similar maximal induction response for
  AHH
• Administration of both compounds produces no
  additive effect
• Parallel dose-response curves
• TCDD 30,000 times more potent (ED50 0.8
  nmol/kg)

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A genetic locus in mice

• PAHs induce AHH (ie P450) in the skin (and liver)
  of mice
• But there is genetic variation
• DBA/2 mice do not induce with 3MC
• C57Bl/6J mice do induce with 3MC
• Responsiveness maps as an autosomal dominant
  between C57 and DBA
• the Ah locus (Aryl Hydrocarbon)
• TCDD induces in all mice strains

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Ah locus

• Two possibilities
• (a) 3MC, TCDD have distinct receptors
• (b) the same receptor
• Dose response for TCDD in responsive and
  non-responsive strains
• The dose of TCDD required for induction is
  10-fold greater in non-responsive strains of mice

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Ah Locus

• AHH responsive mice are about 10-fold more
  sensitive than non-responsive mice to
• Thymic involution
• Hepatic porphyria
• Teratogenesis
• The Ah locus controls toxicity
• How does Ah work ?

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Ah encodes a receptor

• Receptor identified in the liver cytosol of
  C57Bl/6 mice
• High affinity 0.2 nMolar
• 8 pMols receptor/ gm liver
• 50 000 molecules/ liver cell
• Lower affinity receptor in DBA/2 mice
• Correlation between ability to bind various
  compounds and potency for induction

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Ah Receptor

• Human AhR is 96 kDa
• Binds dioxin directly
• Family of cellular signalling receptors

1
805
228
416
HLH
PAS1
PAS2
Transactivation
Ligand-Binding Domain 190 amino acids, 25 kDa
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AhR and dioxin signalling
TCDD
Arnt
AhR
Transcription
hsp90
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AhR -/- mice

• Transgenic mice without AhR gene
• Unable to induce cytochrome P450 by TCDD, PAH
• TCDD toxicity vastly reduced (gt100 fold)
• Greatly reduced tumour incidence in skin after
  application of Benzo(a)Pyrene
• Due to AhR
• Or to lack of induction of cytochrome P450 ?