The Aryl Hydrocarbon Receptor as a Molecular Target of Chemopreventive Phytochemicals

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The Aryl Hydrocarbon Receptor asa Molecular
Target ofChemopreventive Phytochemicals
Pawel Sienkiewicz, Ph.D. Sara Bass, M.S.
Henry Ciolino, Ph.D. Chris MacDonald, Ph.D.
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• Cancer is preventable.
• Diets rich in fruits and vegetables are
  associated with a reduced risk of many types of
  cancers.

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Many phytochemicals are believed to affect every
stage of the cancer process
Chemopreventive Cancer Prevented Mechanisms
Phytochemicals Source (animal models)
of Action Alkyl sulfides and
disulfides Allium Esophagus, Colon, Phase II,
GSTSulfide volatiles LungAllyl
cystenes Monoterpenes Citrus Mammary,
Pancreas, Phase II, GST,Lemonene Skin, Lung,
Liver UDP-GT, Phase I Isothiocyanates Crucifers
Liver, Lung, Phase II, GST Mammary Polyphenols
Teas Colon, Lung, Phase II (GST,Epigallocatechin
gallate Skin, Liver QR), Phase
I, AP-1 Curcumin Turmeric Colon, Skin Phase
II, GST, Cox II
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Carcinogenesis
Activation of Procarcinogens by CYPs
Gene Mutation Proto-oncogene Tumor Suppressor
AngiogenesisProgram Cell Death
DNA Damage
Promotion
Progression
Initiation
NormalCells
MutantCells
BenignTumors
MalignantTumors
X
X
X
PhytochemicalsOther Chemopreventive Agents
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TCDD
Benzoapyrene
DMBA
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The Aryl Hydrocarbon Receptor Pathway
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Aryl Hydrocarbon Receptor Battery of Genes

• Phase 1 enzymes Cytochrome P450 1A1
  Cytochrome P450 1A2 Cytochrome P450 1B1
• Phase 2 enzymes Aldehyde dehydrogenase
  Glutathione-S-transferase g
  UDP-glucuronosyltransferase
  NADPH-quinone-oxidoreductase

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MacDonald, Cancer Res. 61 2001 Ciolino, J. Biol.
Chem. 2741999 Ciolino, Cancer Res. 62 2002
Ciolino, Cancer Res. 581998 Ciolino, Biochem. J.
3401999 Ciolino, Br. J. Cancer 791999
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Cellular Defense and Carcinogenesis Section
Kaempferol, Quercetin
Curcumin, DBM
Galangin
Resveratrol
Diosmetin, Diosmin
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Salicylates

• Salicylates are an important class of
  phytochemicals present in human diets.
• Aspirin, a synthetic salicylate, is the most
  studied chemopreventive agent.

• MacDonald, Cancer Res. 64 2004
• Ciolino, Carcinogenesis 27, 2006

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Salicylamide Inhibits Aryl Hydrocarbon-induced
CYP Enzyme Activity in HepG2 Cells
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Aryl Hydrocarbon Receptor Pathway
Drugs
Ligand Binding
Gel Shift
Transcription
mRNA Level
Protein and Enzyme Activity
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Salicylamide

• Salicylamide is a synthetic salicylate used in
  combination with aspirin and other analgesic
  mixtures. Thus it has already been used
  extensively in humans.
• There is no published literature on salicylamide
  other than as an analgesic.
• Its chemopreventive properties are unknown.

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Effect of Salicylamide on TCDD-induced CYP1A1
Activity in HepG2 Cells
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Effect of Salicylamide on TCDD-induced Binding of
the AhR to the XRE of CYP1A1 in HepG2 Cells
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Salicylamide Competes with TCDDfor Binding to
the AhR in HepG2 Cells
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Novel Drugs That Inhibits the Expressionof
Carcinogen-activating Enzymes in Human Hepatoma
HepG2 Cells

• Aspirin-H2S (ACS30)
• Diclofenac-H2S (ACS15) is a NSAID for the
  treatment of inflammation and pain relief.
• Valproate-H2S (ACS2) is a drug for Epilepsy
  treatment in humans. The LD50 orally in rats is
  670 mg/kg (Food Cosmet. Toxicol. 2 327, 1964).

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Measurement of Cell Viability by MTT Assay
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Aryl Hydrocarbon Receptor Pathway
Drugs
Ligand Binding
ChIP
Transcription
mRNA Level
Protein and Enzyme Activity
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ACS15 Inhibits Aryl Hydrocarbon-induced CYP
Enzyme Activity in HepG2 Cells
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ACS15 Inhibits Aryl Hydrocarbon-induced Enzyme
Activities in Recombinant CYPs
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Novel Drugs That Inhibits the Expressionof
Carcinogen-activating Enzymes in Human Hepatoma
HepG2 Cells

• Valproate-H2S (ACS2) is a drug for Epilepsy
  treatment in humans. The LD50 orally in rats is
  670 mg/kg (Food Cosmet. Toxicol. 2 327, 1964).

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Val-H2S Inhibits Aryl Hydrocarbon-induced CYP
Enzyme Activity in HepG2 Cells
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Yeh, J. Biol. Chem. 276 2001.
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Integrated Study of Chemoprevention against Aryl
Hydrocarbons
To establish whether the AhR and the pathway it
mediates is a molecular target of
chemopreventive compounds and S-NSAIDs. David
Wink, NCIPiero Del Soldato, CTG PharmaEd
Sausville, Univ. of MD
To test the hypothesis thatchemopreventive
agents and S-NSAIDs inhibit carcinogen-inducedlu
ng and breast tumorigenesis in rodents by
affecting the AhR battery of genes. Lucy
Anderson, NCIMariam Poirier, NCIPiero Del
Soldato, Pharma
To examine the effect of chemopreventive
agents and S-NSAIDson novel biochemical pathways
involved inthe detoxification of aryl
hydrocarbons. David Wink, NCIDavid Roberts,
NCITerry Moody, NCI