Tobacco Carcinogen and Toxicant Biomarkers

1
Tobacco Carcinogen and Toxicant Biomarkers

• Stephen S. Hecht, Ph.D.
• Masonic Cancer Center
• University of Minnesota

2
Outline of Presentation

• Tobacco and cancer
• Background on tobacco carcinogen biomarkers
• Examples of tobacco carcinogen biomarkers
• Total NNAL
• Formaldehyde-DNA adducts

3
Cancers Caused by SmokingIARC Monograph Series

• Volume 38, 1986
• Lung
• Oral cavity
• Pharynx
• Larynx
• Esophagus
• Pancreas
• Bladder

• Volume 83, 2004, added
• Nasal Cavity
• Stomach
• Liver
• Kidney
• Ureter
• Cervix
• Myeloid leukemia

• Volume 100E, 2009, added
• Colorectum
• Ovary (mucinous)

4
Cancer Deaths Due to Smoking

• Worldwide 21 (1,420,000 per year)
• United States 33 (185,000 per year)

IARC World Cancer Report, 2008
5
Tobacco Use Prevalence, 2008

• Adult smokers, U.S. 46,000,000 (20.6)
• Ex-smokers, U.S. 48,100,000 (21.5)
• Smokers, worldwide 1,300,000,000
• Smokeless, worldwide xxx,000,000

CDC, MMWR, November 13, 2009
6
World Smoking Prevalence Males
O. Shafey, M. Ericksen, H. Ross, J. Mackay (2009)
The Tobacco Atlas, 3rd Ed.
7
World Smoking Prevalence Females
O. Shafey, M. Ericksen, H. Ross, J. Mackay (2009)
The Tobacco Atlas, 3rd Ed.
8
Overall Goal

• Elucidate mechanisms of tobacco-induced
  cancer and apply this knowledge to cancer
  prevention.

9
Conceptual Framework for Understanding Tobacco
Carcinogenesis
S.S. Hecht, JNCI, 911194-1210 (1999), Nature
Rev. Cancer 3733-744 (2003) Cancer Principles
and Practice of Oncology, 8th Edition, 147-155
(2008)
10
Significantly Mutated Genes in Lung
Adenocarcinoma Based on Sequencing of 623 Genes
in 188 Tumors
Ding et al, Nature, 4551069-1075, 2008
11
Other Factors Contributing to Tobacco-Induced
Cancer

• Receptor mediated effectsnicotine, nitrosamines
• Direct activation of EGFR and COX-2
• Down-regulation of FHIT
• Hyper-methylation of tumor suppressors
• Tumor promotion and co-carcinogenesis
• Oxidative damage and inflammation
• Cilia-toxicity

H. Takahashi et al, Cancer Cell 17 89 (2010) H.
Schuller, Nature Rev. Cancer 9 195-205 (2009)
K.A. West et al, J. Clin. Invest. 111 81-90
(2003) S.A. Belinsky, Carcinogenesis 26 1481
(2005) DAgostini et al, Cancer Res 66
3936-3941 (2006) Jin et al, Carcinogenesis 29
1614-1622 (2008) Bhutani et al, Cancer Prev.
Res. 1 39-44 (2008)
12
Conceptual Framework for Understanding Tobacco
Carcinogenesis
S.S. Hecht, JNCI, 911194-1210 (1999), Nature
Rev. Cancer 3733-744 (2003) Cancer Principles
and Practice of Oncology, 8th Edition, 147-155
(2008)
13
IARC Carcinogens in Tobacco Smoke
S.S. Hecht, In DeVita et al, Cancer (2010) IARC
Monographs No. 83 (2004) D. Hoffmann and S.S.
Hecht, Handbook Exp. Pharmacol. 9463-102 (1990)
14
Goal

• Develop and validate tobacco carcinogen and
  toxicant biomarkers
• Urinary metabolites
• DNA and protein adducts
• Metabolites in blood, saliva, breath, nails, hair
• Use these biomarkers to identify those smokers
  susceptible to cancer.

15
Outline of Presentation

• Tobacco and cancer
• Background on tobacco carcinogen biomarkers
• Examples of tobacco carcinogen biomarkers
• Total NNAL
• Formaldehyde-DNA adducts

16
Definitions

• Biomarker A distinctive biological or
  biologically derived indicator (as a metabolite)
  of a process, event, or condition
  (Merriam-Websters Collegiate Dictionary)
• Tobacco carcinogen biomarker Any quantifiable
  substance, such as a metabolite, that can be
  specifically related to human exposure to a given
  tobacco carcinogen.

17
Tobacco Carcinogen Biomarkers

• DNA Adducts
• Protein Adducts
• Hemoglobin
• Albumin
• Metabolites
• Breath
• Saliva
• Nails and Hair
• Urine
• Blood

Reviewed in Carcinogenesis 23 907 and 1979
(2002) Nature Rev. Cancer 3 733 (2003)
18
Applications of Tobacco Carcinogen Biomarkers

• Assessing exposure in smokers, smokeless tobacco
  users, and non-smokers exposed to secondhand
  smoke
• Regulation of tobacco products
• Understanding mechanisms of human carcinogenesis
  and identifying susceptible individuals
• Not specifically designed for early detection of
  cancer, but could have applications in screening

19
A Panel of Tobacco Carcinogen and Toxicant
Biomarkers
Range of recent mean values (nmol/24h unless noted otherwise) Range of recent mean values (nmol/24h unless noted otherwise)
Urinary biomarkers Source Smokers Non-smokers
Nicotine equivalents Nicotine 70.4-154 µmol/24 h Not Detected
Total NNAL NNK 1.1 - 2.9 Not Detected
Total NNN NNN 0.049 - 0.24 Not Detected
1-HOP or PheT PAH 0.50 - 1.45 0.18 - 0.50
MHBMA 1,3-Butadiene 15.5 - 322 0.65 - 7.5
SPMA Benzene 3.2 - 32.1 0.17 - 3.14
HPMA Acrolein 5,869 - 11,190 1,131 - 1,847
HBMA Crotonaldehyde 1,965 - 26,000 242 - 3,200
HEMA Ethylene oxide 19.1 - 102 6.51 - 38.8
Cd Cadmium 2.3 - 12.8 1.34 - 8.04
8-epi-PGF2a Oxidative damage 1.48 - 2.80 0.62 - 1.13
PGE-M Inflammation 54 - 60 31.6 - 45.3
Based on 1.3g creatinine per 24h in smokers and
1.5g creatinine per 24h in non-smokers, or 1.5 l
urine per 24h.
S.S. Hecht, J-M Yuan, and D. Hatsukami, Chem.
Res. Toxicol., 2010
20
Structures of the Urinary Biomarkers
21
A Panel of Biomarkers
Recent data (pmol/g globin mean S.D.) Recent data (pmol/g globin mean S.D.)
Hemoglobin adducts Source Smokers Non-smokers
Cyanoethylvaline Acrylonitrile 112 81 6.5 6.4
Carbamoylethylvaline Acrylamide 84.1 41.8 27.8 7.1
Hydroxyethylvaline Ethylene oxide 132 92 21.1 12.7
4-Aminobiphenyl-globin 4-Aminobiphenyl 0.26 0.006a 0.067 0.009a
(fmol/µmol dN mean S.D.) (fmol/µmol dN mean S.D.)
Leukocyte DNA adducts Source Smokers Non-smokers
N6-hydroxymethyl-dAdo Formaldehyde 179 205 15.5 33.8
N2-ethylidene-dGuo Acetaldehyde 1,310 1,720 705 438
Mean concentrations Mean concentrations
Other Source Smokers Non-smokers
Exhaled CO Carbon monoxide 17.4 - 34.4 ppm 2.6 - 6.5 ppm
Carboxyhemoglobin Carbon monoxide 3.4 - 7.1 0.35 - 1.45
22
Biomarker Validation

• Analytical
• Specificity, sensitivity, accuracy, precision
• With respect to tobacco
• Decreases upon cessation
• Dose-response
• With respect to cancer risk

23
Persistence of Biomarkers Study

• Smokers provide baseline 24h urine samples.
• Eight days later, they quit smoking and receive
  nicotine replacement therapy.
• They provide 24h urine samples on days 3, 7, 14,
  21, 28, 42, and 56 after quitting.
• Urine samples are analyzed for mercapturic acids
  (by LC-MS/MS) and other biomarkers.

S.G. Carmella, M. Chen, S. Han, A. Briggs, J.
Jensen, D. K. Hatsukami, and S. S. Hecht Chem.
Res. Toxicol., 22 734-741 (2009)
24
Metabolism of 1,3-Butadiene to Mercapturic Acids
a. GSH, GSTs b. g-glutamyltranspeptidase c.
cysteinylglycine dipeptidase d. cysteine
S-conjugate N-acetyltransferase
C.L. Sprague and A.A. Elfarra, Chem. Res.
Toxicol., 17 819-826 (2004)
25
Mean Urinary MHBMA Reduction Upon Smoking
Cessation, N17
26
Metabolism of Acrolein, Crotonaldehyde, Benzene,
and Ethylene Oxide to Mercapturic Acids
a. GSH, GSTs b. g-glutamyltranspeptidase c.
cysteinylglycine dipeptidase d. cysteine
S-conjugate N-acetyltransferase
27
Mean Urinary HPMA Reduction Upon Smoking
Cessation, N17
28
Mean Urinary HBMA Reduction Upon Smoking
Cessation, N17
29
Mean Urinary SPMA Reduction Upon Smoking
Cessation, N17
30
Mean Urinary HEMA Reduction Upon Smoking
Cessation, N17
31
Structures of Urinary Biomarkers
S.G. Carmella, et al, Chem. Res. Toxicol., 22
734-741 (2009)
32
Mean Urinary 1-HOP Reduction Upon Smoking
Cessation, N15
33
Mean Urinary Total NNAL Reduction Upon Smoking
Cessation, N17
34
Outline of Presentation

• Tobacco and cancer
• Background on tobacco carcinogen biomarkers
• Examples of tobacco carcinogen biomarkers
• Total NNAL
• Formaldehyde-DNA adducts

35
A Panel of Tobacco Carcinogen and Toxicant
Biomarkers
Range of recent mean values (nmol/24h unless noted otherwise) Range of recent mean values (nmol/24h unless noted otherwise)
Urinary biomarkers Source Smokers Non-smokers
Nicotine equivalents Nicotine 70.4-154 µmol/24 h Not Detected
Total NNAL NNK 1.1 - 2.9 Not Detected
Total NNN NNN 0.049 - 0.24 Not Detected
1-HOP or PheT PAH 0.50 - 1.45 0.18 - 0.50
MHBMA 1,3-Butadiene 15.5 - 322 0.65 - 7.5
SPMA Benzene 3.2 - 32.1 0.17 - 3.14
HPMA Acrolein 5,869 - 11,190 1,131 - 1,847
HBMA Crotonaldehyde 1,965 - 26,000 242 - 3,200
HEMA Ethylene oxide 19.1 - 102 6.51 - 38.8
Cd Cadmium 2.3 - 12.8 1.34 - 8.04
8-epi-PGF2a Oxidative damage 1.48 - 2.80 0.62 - 1.13
PGE-M Inflammation 54 - 60 31.6 - 45.3
Based on 1.3g creatinine per 24h in smokers and
1.5g creatinine per 24h in non-smokers, or 1.5 l
urine per 24h.
S.S. Hecht, J-M Yuan, and D. Hatsukami, Chem.
Res. Toxicol., 2010
36
Essential Facts About NNK,A Tobacco-Specific
Lung Carcinogen

• Present in tobacco and tobacco smoke specific to
  tobacco products
• Systemic lung carcinogen in rats, mice, hamsters,
  and ferrets.
• Also induces tumors of the pancreas, nasal
  cavity, and liver in rats
• Considered to be a cause of lung, oral cavity and
  pancreatic cancer in people exposed to tobacco
  products
• NNK and NNN- Carcinogenic to humans Group 1
  (IARC Volume 89, 2007) reaffirmed (Vol 100E,
  2009)

S.S. Hecht, Chem. Res. Toxicol. 11559 (1998)
Nature Rev. Cancer 3733 (2003)
37
Metabolism of NNK by Carbonyl Reduction
38
NNAL Plus NNAL-Glucs (Total NNAL) A Biomarker
of NNK Exposure

• Quantified by GC-TEA or LC-MS/MS
• High analytical specificity and sensitivity
• Specific to tobacco product exposure
• Responsive to dose
• Measures uptake of a lung carcinogen

S.S. Hecht, Carcinogenesis 23 907 (2002)
S.G.Carmella et al, CEBP 4 635 (1995) 12 1257
(2003) D. Hatsukami et al, Nic. Tob. Res. 8
169 (2006)
39
GC-TEA Chromatogram of NNAL in a Smoker's Urine
40
Applications of the Total NNAL Biomarker

• Cessation of smoking or smokeless tobacco
• Reduction of smoking
• Carcinogen uptake from new and old tobacco
  products
• Omni, light and ultra-light cigarettes
• Snus and other smokeless products
• Ultra low nicotine cigarettes
• Evaluation of carcinogen dose in various groups
• Reducers vs. light smokers
• Smokers of differing numbers of cigarettes
• Ethnic groups, gender, and teen-age smokers
• Smokeless vs. smokers
• Duration of smokeless use
• Carcinogen uptake from secondhand cigarette smoke
• Relationship to lung cancer

D. Hatsukami, J. Jensen, A. Joseph, S. E. Murphy,
S.G. Carmella, S.S. Hecht, and co-workers.
Cancer Res., JNCI, CEBP, Nic. Tob. Res.,
1999-2008
41
Tobacco Harm Reduction Continuum of Risk
Conventional cigarettes Modified tobacco
cigarettes Cigarette reduction Cigarette-like
delivery devices Extra-low nicotine
cigarettes Smokeless tobacco products
Nicotine delivery devices Smoking Cessation
Most toxic
Least toxic
D. Hatsukami et al., Nicotine Tob. Res.
9S537-S553 2007
42
Applications of the Total NNAL Biomarker

• Cessation of smoking or smokeless tobacco
• Reduction of smoking
• Carcinogen uptake from new and old tobacco
  products
• Omni, light and ultra-light cigarettes
• Snus and other smokeless products
• Ultra low nicotine cigarettes
• Evaluation of carcinogen dose in various groups
• Reducers vs. light smokers
• Smokers of differing numbers of cigarettes
• Ethnic groups, gender, and teen-age smokers
• Smokeless vs. smokers
• Duration of smokeless use
• Carcinogen uptake from secondhand cigarette smoke
• Relationship to lung cancer

D. Hatsukami, J. Jensen, A. Joseph, S. E. Murphy,
S.G. Carmella, S.S. Hecht, and co-workers.
Cancer Res., JNCI, CEBP, Nic. Tob. Res.,
1999-2008
43
Non-Smokers Exposure to NNK Throughout Life by
Measurement of Urinary Total NNAL
Exposed Group Type of Exposure Total NNAL (fmol/ml urine) of Amount in Smokers' Urinea
Fetus Transplacental 25 29 (amniotic fluid) 1.3
Newborns Transplacental 130 150 6.5
Infants (lt1 year old) Air 83 20 4.2
Elementary School Children Elementary School Children Elementary School Children Elementary School Children
Minneapolis Air 56 76 2.8
Moldova Air 90 77 4.5
Women Living with Smokers Air 50 68 2.5
Hospital Workers Air 59 28 3.0
Casino Patrons Air 18 15 0.9
Restaurant and Bar Workers Air 33 34 1.7
a based on 2 pmol/ml total NNAL in smokers S.S.
Hecht, Carcinogenesis 23907 (2002) S.S. Hecht
et al, CEBP 15988 (2006)
44
Total NNAL measurements in secondhand
smoke-exposed non-smokers have impact

• It can only come from secondhand smoke.
• It represents uptake of a lung carcinogen.
• It is found in the urine of non-smokers.
• It is the only lung carcinogen biomarker
  consistently elevated in exposed non-smokers.
• These studies should spur clean air legislation
  in the remaining countries, states and localities.

45
Median Serum Cotinine Levels in Non-Smokers, by
Age Group 1988-2002
CDC NHANES Study MMWR 55 1130 (2006)
46
Regulation of Indoor Smoking and Tobacco Control

• Regulation of indoor smoking
• Reduces cues for smoking
• Reduces amount smoked
• Can change social norms
• Regulation of indoor smoking, along with
  counter-advertising and taxation, are the most
  effective methods in tobacco control.

47
Applications of the Total NNAL Biomarker

• Cessation of smoking or smokeless tobacco
• Reduction of smoking
• Carcinogen uptake from new and old tobacco
  products
• Omni, light and ultra-light cigarettes
• Snus and other smokeless products
• Ultra low nicotine cigarettes
• Evaluation of carcinogen dose in various groups
• Reducers vs. light smokers
• Smokers of differing numbers of cigarettes
• Ethnic groups, gender, and teen-age smokers
• Smokeless vs. smokers
• Duration of smokeless use
• Carcinogen uptake from secondhand cigarette smoke
• Relationship to lung cancer

D. Hatsukami, J. Jensen, A. Joseph, S. E. Murphy,
S.G. Carmella, S.S. Hecht, and co-workers.
Cancer Res., JNCI, CEBP, Nic. Tob. Res.,
1999-2008
48
Relationship of Urinary NNAL to Lung Cancer in
Two Prospective Cohorts of Cigarette Smokers

• Collaboration with Professors Mimi Yu and
  Jian-Min Yuan
• Two prospective cohorts of Chinese cigarette
  smokers Shanghai and Singapore
• Nested case control study of 246 cases of lung
  cancer and 245 matched controls
• Total NNAL and cotinine quantified in stored
  urine samples collected prior to lung cancer
  diagnosis

49
Joint Effect of Urinary Total NNAL and Cotinine
on Lung Cancer Risk
NNAL in tertile Cotinine in tertile Cotinine in tertile Cotinine in tertile Cotinine in tertile Cotinine in tertile Cotinine in tertile
NNAL in tertile 1st (low) 1st (low) 2nd 2nd 3rd (high) 3rd (high)
NNAL in tertile Ca/Co1 OR (95 CI)2 Ca/Co1 OR (95 CI)2 Ca/Co1 OR (95 CI)2
1st (low) 9/47 1.00 23/25 3.93 (1.54, 10.05) 11/10 5.08 (1.63, 15.89)
2nd 14/24 3.01 (1.11, 8.10) 31/32 4.15 (1.70, 10.12) 22/26 4.48 (1.78, 11.31)
3rd (high) 8/10 3.41 (1.08, 11.25) 30/25 5.58 (2.25, 13.84) 93/46 8.47 (3.69, 19.46)
1 No. of cases/no. of controls 2 Odds ratios (OR)
were adjusted for age, year of interview, year of
sample collection, gender and dialect group,
study location (Shanghai versus Singapore),
number of cigarettes smoked per day, and number
of years of smoking CI, confidence interval.
J. Yuan, M. Yu, S.E. Murphy, S. Carmella, S.S.
Hecht et al., Cancer Res.,69 2990 (2009)
50
Conclusions of the Shanghai and Singapore Study

• Total NNAL significantly associated with risk of
  lung cancer in a dose-dependent manner, after
  adjustment for smoking history and urinary
  cotinine.
• Cotinine was independently associated with lung
  cancer, consistent with previous data.
• Smokers in the highest tertiles of urinary total
  NNAL and cotinine exhibited an 8.5 fold increased
  risk for lung cancer, relative to those with
  comparable smoking history, but in the lowest
  tertiles.

J. Yuan et al, Cancer Res. 69 2990 (2009)
51
Similar Results in the PLCO Study

• The Prostate, Lung, Colon, and Ovarian Cancer
  Screening Trial started 1993
• 77,468 subjects (25,000 smokers) screened over
  1,000 lung cancer cases diagnosed
• Questionnaire data and blood samples collected
  prospectively
• 100 lung cancer cases and 100 controls without
  lung cancer selected all were smokers of gt 10
  CPD
• Pre-diagnostic serum analyzed for total NNAL and
  cotinine

T. Church, K. Anderson, M. Geisser, Y. Zhong, C.
Le, N. Caporaso, S. Carmella, A. Benoit, S. S.
Hecht, CEBP,18 260 (2009)
52
Total NNAL and Lung Cancer

• Total NNAL is a risk biomarker.
• Results are consistent with all experimental and
  previous clinical and epidemiologic data.
• Results further implicate NNK as an independent
  etiologic risk factor in lung cancer.

53
Cigarette Smoke Constituents Targeted for
Regulation by WHO, and Their Biomarkers

• Benzoapyrene
• NNK, NNN
• Acrolein
• Benzene
• 1,3-Butadiene
• Carbon monoxide
• Acetaldehyde
• Formaldehyde
• Nicotine

• 1-HOP or PheT in urine
• NNAL and NNN in urine
• 3-HPMA in urine
• SPMA in urine
• MHBMA in urine
• Exhaled CO
• Leukocyte DNA adducts
• Leukocyte DNA adducts
• Nicotine metabolites

D.M. Burns et al. Tob. Control. 17 132-141
(2008) S.S. Hecht, Carcinogenesis 23 907-922
(2002) L. Chen et al. Chem. Res. Toxicol. 20
108-133 (2007) S.G. Carmella et al. Chem. Res.
Toxicol., 22 734-741 (2009)
54
Outline of Presentation

• Tobacco and cancer
• Background on tobacco carcinogen biomarkers
• Examples of tobacco carcinogen biomarkers
• Total NNAL
• Formaldehyde-DNA adducts

55
Formaldehyde Genetic Toxicology

• Genotoxic
• Mutagenic
• DNA protein cross-links
• DNA strand breaks
• Sister chromatid exchanges
• Chromosomal aberrations
• These changes initiated by reactions with DNA to
  form adducts
• No previous reports of formaldehyde DNA adducts
  in humans
• International Agency for Research on Cancer
  Monographs, Volume 88 (2006)

56
Structures of Formaldehyde-DNA Adducts
R. Shapiro et al. (1980) F. Beland et al. (1984)
57
Conversion of N6-HOMe-dAdo to N6-Me-dAdo
M. Wang et al. Chem. Res. Toxicol. 20 1141-1148
(2007)
58
Outline of Analytical Method for N6-HOMe-dAdo in
DNA
59
LC-ESI-MS/MS-SRM Chromatograms of N6-Me-dAdo in
Smokers' Leukocyte DNA
60
Typical LC-ESI-MS/MS-SRM Chromatograms of
N6-Me-dAdo in Leukocyte DNA
61
Levels of N6-HOMe-dAdo (as N6-Me-dAdo) in
Leukocyte DNA of Smokers and Non-smokers
62
Previous Studies of Leukocyte DNA Adducts in
Smokers vs. Non-smokers

• Most used 32P-postlabelling and immunoassay
  inconsistent results comparing smokers and
  non-smokers
• Mixed results in 8-OH-dG analyses
• Marginally higher levels of acetaldehyde-DNA
  adducts in smokers before stopping
• Significant difference in BPDE-DNA adducts (2
  per 108 vs 1 per 108 nucleotides)
• Our results 5 per 108 vs. 0.5 per 108 nucleotides

IARC Monographs, Vol 83 (2004) Pavanello et al.
Mutat. Res 611 54 (2006) Chen et al Chem. Res.
Toxicol. 20 108 (2007)
63
Sources of Formaldehyde-DNA Adducts

• Formaldehyde itself in cigarette smoke, but blood
  levels were not elevated in volunteers exposed to
  similar amounts
• Smoking effect on endogenous metabolism
• Released as a metabolite from nicotine, NNK, or
  related compounds
• Transfer from formaldehyde-histone adducts
• Secondary metabolite from lipid peroxidation or
  inflammation caused by smoking

64
Conclusions Formaldehyde DNA Adducts

• First study to detect formaldehyde-DNA adducts in
  humans
• Highly significant differences between smokers
  and non-smokers
• Results indicate a previously unrecognized and
  potentially important role for formaldehyde in
  smoking-induced cancer

M. Wang, G. Cheng, S.S. Hecht et al. Cancer Res.,
69 7170 (2009)
65
Overall Goal

• Elucidate mechanisms of tobacco-induced
  cancer and apply this knowledge to cancer
  prevention.

66
We are Making Progress in Tobacco Control

• Smoke-free legislation
• Increased taxation
• Aggressive anti-tobacco advertising

67
Age-Adjusted Total U.S. Mortality Rates for Lung
and Bronchus Cancer
Source SEER data http//seer.cancer.gov/faststats
/
68
(No Transcript)
69
Dorothy Hatsukami
70
Acknowledgements

• Hecht Lab
• Steven Carmella
• Mingyao Wang
• Irina Stepanov
• Pramod Upadhyaya
• Brad Hochalter
• Silvia Balbo
• Shaomei Han
• Menglan Chen
• Guang Cheng
• Lei Meng
• Yan Zhong
• Aleks Knezevich
• John Muzic
• Core Facilities
• Pete Villalta
• Chap Le
• Xianghua Luo

• Dorothy Hatsukami
• Joni Jensen
• Amanda Anderson
• Rachel Feuer
• Tim Church
• Kristin Anderson
• Mindy Geisser
• Jian-Min Yuan
• Mimi Yu

71
Research Support

• National Cancer Institute
• National Institute of Environmental Health
  Sciences
• National Institute on Drug Abuse/NCI/NIAAA
  (TTURC)
• American Cancer Society
• Wallin Chair in Cancer Prevention