GENE-ENVIRONMENT INTERACTION Herman Autrup Dept Environmental Medicine Institute of Public Health University of Aarhus

Influence gene expression Interacts with gene – adducts coding sites – mutation transcription site – expression Gene product modifies effect of environmental risk factor GENE –ENVIRONMENT INTERACTIONS MECHANISM

Several Mechanistic Possibilities GENE-ENVIRONMENT- INTERACTION Risk Factor Genotype Disease Genotype enhances risk factor expression

Several Mechanistic Possibilities GENE-ENVIRONMENT- INTERACTION Risk Factor Genotype Disease Both genotype and risk factor affect disease

Several Mechanistic Possibilities GENE-ENVIRONMENT- INTERACTION Risk Factor Genotype Disease Risk factor alters gene/disease association

Several Mechanistic Possibilities GENE-ENVIRONMENT- INTERACTION Risk Factor Genotype Disease Genotype alters risk factor/disease association

Influence gene expression

Ah Arnt Cell membrane Cytoplasm ATP ADP + Pi Nuclear membrane XRE CYP1A1 Arnt Ah Nuclei Steroid Hormones vævsspecifikke transcription faktorer Negative regulatoriske elementer Phosphorylation

METAL TOXICITY - TRANSCRIPTOMES Andrew, Env Health Persp 111: , 2003

Interacts with genes - adducts

BULKY CARCINOGEN - DNA ADDUCT DNA: Lymphocytes Procedure: P32 postlabelling Enrichment:P1 nuclease Quantification: phosphor image DNA component 32 P-label resolve nucleotides

CARCINOGEN-DNA ADDUCTS AND EXPOSURE carc. PAH (ng/m 3 ) DNA adducts / 10 8 nucl.) carc. PAH (ng/m 3 ) DNA adducts / 10 8 nucl No correlation with inhaled PAH or other ambient air pollution parameters Thailand police: No correlation between total PAH and bulky DNA adduct levels Medical students, DK: No correlation between PM2.5 and bulky-carcinogen adduct levels Meta-analysis (Peluso et al, 2001): Significant association between PAH (and BP) and bulky-adducts in industrial workers.

CARCINOGEN-DNA ADDUCTS AND CANCER RISK Study design:Type of cancerRiskReference Cohort (smokers only)Lung1.22H. Bak, poster Porvoo Cohort (current smokers)Lung2.98Perera et al, 2002 Case-controlBladder1.9Benhamou et al, 2003 Case-controlBreast (PAH)1.97Rundle et al, 2002 Case-controlBreast (PhIP)4.03Zhu et al, 2003 Conclusion: Carcinogen-DNA adducts appears to be a risk indicator for cancer, especially in smokers

Effect modification

CANDIDATE GENES Low Penetrance: Phase I CYP1A1 PAH’s cigarette smoke CYP1A2 HAA’s cooked meat CYP2E1Nitrosaminesprocessed meat NAT1HAA’scooked meat NAT2HAA’scooked meat

CANDIDATE GENES Low Penetrance: Phase II NAT1, NAT2Arylaminescigarette smoke GSTPAH’s cigarette smoke HAA’s cooked meat

CANDIDATE GENES Some Problems Expressed in target tissue? Inducible? Polymorphisms functional? Dual role (NAT)

GST and GASTRIC CANCER – Meta analysis GSTM1 Deletion ever smoking 2.39 GSTT1 Deletion ever smoking 2.37 GSTM1 and T1 deletions 2.08 Boccia et al, Cancer Letter, 2005

GSTM1 and NEVER SMOKERS Passive smokers Total group Plus 1.00 Null 1.03 Null + null passive h-years Wenzloft et al, Carcinogenesis 2005

GSTM1 and ALCOHOL INTERACTION Esophegeal Gastric Alcohol GSTM1 Del GSTM1 + and low alcohol = 1 GSTM1 Del high and GSTM high and Del

BIOTRANSFORMATION BENZENE NQO

BENZENE TOXICITY Benzene Benzene oxide Phenol Hydroquinone 1,4-Benzoquinone Risk Lack of NQO1 (2.6) High CYP2E1(2.5) Combined(7.8)

BENZENE : S-PHENYLMERCAPTURIC ACID (PMA) AND MUCONIC ACID (TMA) Low activity genotype = 100 Sørensen et al., 2003 Sources of benzene. Ambient air, cigarette smoking, grilling (charcoal) Population: Non-smoking medical students, Copenhagen, DK (ambient air benzene 2.53 ug/m 3 ) Significant difference TMAPMA

BULKY-ADDUCTS – EFFECT OF GSTM1 GSTM1*1/*1 = 100 LymphocytesTissues * Statistically significant PlacentaBreastLung * * * * ** Occupational studies

GENE-GENE INTERACTION Study population:Greek technical school students, non-smokers DNA adducts 0,00 0,25 0,50 0,75 1,00 1,25 1,50 1,752,00 all subjects high ETS exposure GSTM1 null GSTP1 ile/val & val/val mEH exon CYP1A1*2+/+ CYP1A1*2 +/- & -/- non-inducible individuals (76.3%) inducible individuals (23.7%)

INTERACTION OF GSTM1 AND GSTP1 PAH-adducts – WBC OR GSTM1*2/*21.25 GSTP*2/1.44 GSTM1*2/*2 & GSTP*22.20 GSTM1 no effect in current smokers Strongest effect in former smokers Perera et al., Carcinogenesis, 2002

GENOTYPES AFFECTING BULKY-DNA ADDUCT LEVEL Gene CYP1A1 GSTM1 GSTT1 GSTP1 MPO NQO1 EH NAT2 Genotype *2/*2 *1/*1 Tyr113Tyr Slow Effect No effect Combinations

COMBINED EFFECT OF BIOMARKERS ON LUNG CANCER RISK Adduct levelGSTM1OR Lowpresent1.0 Lownull2.3 Highpresent6.9 High null16.2 Tang et al., Carcinogenesis 1998

GEN-AIR - OBJECTIVES  Risk factors in smoking-related cancers in non-smokers ambient air environmental tobacco smoke susceptibility genes  Bulky carcinogen-DNA adducts as risk indicator  Gene-environment interactions

MOLECULAR EPIDEMIOLOGY Definition: Application of molecular markers of exposure and susceptibility in epidemiological studies

GEN-AIR - OBJECTIVES  Risk factors in smoking-related cancers in non-smokers ambient air environmental tobacco smoke susceptibility genes  Bulky carcinogen-DNA adducts as risk indicator  Gene-environment interactions

GEN-AIR STUDY 271 lung cancer cases Risk factors: Passive smoking Ambient air pollution-residence Genetic polymorphism: 26 genes, 39 polymorphisms Bulky carcinogen-DNA adduct: risk indicator

STATISTICAL ANALYSIS Cox´s proportional hazard models Adjusted: Gender, age, smoking habits, time of recruitment, country, school year, energy intake, fruit & vegetable, physical activity, educational level

AMBIENT AIR POLLUTION Residence – road Estimated exposure – Residence and monitoring stations

LUNG CANCER RISK - RESIDENCE AOR No major road1.0 Moderately busy road1.52 Busy road1.22 Slightly higher risk for people living along roads with medium and high traffic

ETS EXPOSURE IN INFANCY Parental smoking HR Daily Ex-smokers1.65 Never smokers2.04 Daily – many hours Ex-smokers2.98 Never smokers3.63 Increased risk for lung cancer in people who has been exposed to ETS

SUSCEPTIBILITY GENES Xenobiotica metabolism: NAT1, NAT2, GSTM1, GSTM3, GSTT1, GSTP1, CYP1A1, CYP1B1, MPO, NQO1 DNA repair: ERCC1, ERCC3, ERCC5, OGT Others: MnSOD, BBRC1, RAD51, TP53 Total: 29 genes, 36 polymorphisms

SUSCEPTIBILITY GENES Metabolism genesOR CYP1A1*2/*23.0 ( ) CYP1B1*2/*21.2 ( ) GSTM1*2/*21.1 ( ) GSTP1*2/*21.8 ( ) NQO1*8.0 ( ) No major role of these genes on lung cancer risk in low exposure situations

RISK GENOTYPES RISK ALLELE : The genotypes associated with an increased risk of cancer in previous studies

GENE-ENVIRONMENT INTERACTION Ambient air Less than 3 polymorphism 3+ polymorphic alleles No major road 1.0 Type A/B AOR

GENE-ENVIRONMENT INTERACTION Environmental tobacco smoke 3+ polymorphic alleles Less than 3 polymorphism NO YES In case of ETS exposure increased risk with increasing number of at risk genotypes (trend P < 0.01) AOR

CARCINOGEN DNA ADDUCTS Involved in initiation of cancer – mutation Related to exposure Indicator of cancer risk – integrating exposure and genetic susceptibility

BULKY ADDUCT AS RISK INDICATOR Detectable vs non-detectable AOR All subjects1.82 Never smokers2.26 Ex-smokers2.24 Persons with detectable adduct level have an increased risk of developing cancer

ADDUCTS AND LUNG CANCER Only people with detectable adducts All subjectbelow1.93 above1.51 Never smokersbelow1.78 above4.04 Ex-smokersbelow3.91 above1.20 Never-smokers with an above median adduct level have an increased risk of lung cancer

N= DNA ADDUCTS AND SUSCEPTIBILITY Adduct level increases with number of risk alleles

GENETIC POLYMORPHISM IN XENOBIOTICA METABOLIZING ENZYMES Cytochrome P450:CYP1A1, CYP1A2, CYP1B1, CYP2A6, CYP2E1, CYP3A4, Glutathione S-Transferase:GSTM1, GSTT1, GSTP1, GSTA1 Others:NQO1, MPO, mEH