Chemical Carcinogenesis: Initiation, Promotion, Progression Ivan Rusyn Laboratory of Environmental Genomics Department of Environmental Sciences & Engineering MHRC ENVR 430

CANCER: “A multicausal, multistage group of diseases the mechanisms of which are still only partially known” (IARC Scientific Publications, 1992) “Cancer is a group of diseases characterized by uncontrolled growth and spread of abnormal cells […] that can result in death” (American Cancer Society, 2006) Age adjusted Cancer Death Rates, by Site, US,

Benign tissue is not cancer. Although the cell growth is moderately increased, the cells do not invade nearby tissue or spread to other parts of the body Malignant tissue is cancer. The cancer cells divide out of control. They can invade and destroy nearby healthy tissue. Also, cancer cells can break away from the tumor they form and enter the bloodstream and lymphatic system Metastasis: the spread of cancer beyond the organ of origin

WHAT MAY CAUSE CANCER ?  Hereditary disorders  Chemicals  Viruses  Chronic inflammation  ??? From

History of Chemical Carcinogenesis Chemical carcinogenesis was first suggested by clinicians 200 years ago –Scrotal cancer in chimney sweeps - Potts –Nasal cancer and snuff dipping - Hill –Today, >50 chemicals are recognized as human carcinogens First experimental studies in animals were done ~80 years ago

Large numbers of chemicals were tested for carcinogenic potential in the s –Maximum Tolerated Doses (MTD) were used. –60% of rodent carcinogens were genotoxic –40% of rodent carcinogens were nongenotoxic –Some chemicals were single site, single species carcinogens –Others were multisite, multispecies carcinogens –Dose-response varies from <1/2 MTD to <1/1000 MTD Most regulations use straight mathematical extrapolation of high dose rodent data to predict risks History of Chemical Carcinogenesis

WORLD HEALTH ORGANIZATION INTERNATIONAL AGENCY FOR RESEARCH ON CANCER IARC Monograph Evaluations LYON, FRANCE Slide courtesy of V. Cogliano (IARC)

IARC: Slide courtesy of V. Cogliano (IARC)

A tour of IARC’s classifications Preamble, Part B, Section 6(d) Slide courtesy of V. Cogliano (IARC)

IARC (2007) - monographs.iarc.fr Carcinogenic to humans (group 1) – 100 agents to date Probably carcinogenic to humans (group 2A) – 68 Possibly carcinogenic to humans (group 2B) – 246 Not classifiable as to its carcinogenicity to humans (group 3) – 516 Probably not carcinogenic to humans (group 4) – 1 U.S. EPA (2003) Carcinogenic to humans Likely to be carcinogenic to humans Suggestive evidence of carcinogenic potential Inadequate information to assess carcinogenic potential Not likely to be carcinogenic to humans U.S. NTP (2002) (see NTP levels of evidence.pdf) Known to be a human carcinogen Reasonably anticipated to be a human carcinogen Cal/EPA (2004) Known to the state to cause cancer

Cancer Cases Attributable to Environmental Carcinogens (Worldwide, 1990) Infections (viruses, parasites, H. pylori) 16% Tobacco (smoked and smokeless) 14% Occupation 4% Alcohol drinking 3% 37% Diet and dietary components including contaminants 25% Pollution 2% Reproductive factors 2% 29%

IARC Group 1 – Carcinogenic to humans Monographs Volumes 1-84 ( ): 89 Agents and Exposures Medical drugs and treatments24 Industrial processes13 Infectious agents or processes10 Physical agents10 Industrial chemicals 7 Inhaled particulates 5 Metals and inorganic salts 5 Lifestyle factors (incl. herbal remedies) 7 Other 8

Exposures to Chemicals in the Workplace Modified from Cullen et al. (1990).

Carcinogenic Risks of Chemical Agents Associated with Medical Therapy and Diagnosis

Carcinogenic Factors Associated with Lifestyle Modified from Pitot (1986) and Vainio et al. (1991)

Chemical Carcinogenesis in the 21 st Century New perceptions of previously known carcinogens: Combined effects of multiple exposures Examples: oAlcohol drinking and aflatoxins oAlcohol drinking and HBV/HBC oAlcohol drinking and tobacco smoking oAlcohol drinking and asbestos/arsenic/radon

Initiating Event Cell Proliferation (clonal expansion) Progression Cell Proliferation Malignancy Second Mutating Event "N" Mutating Event Initiation Promotion Stages of Carcinogenesis

Cellular and Molecular Mechanisms in Multistage Carcinogenesis: INITIATION Initiating event involves cellular genome – MUTATIONS Target genes: - oncogenes/tumor suppressor genes - signal transduction - cell cycle/apoptosis regulators From “Simple” genetic changes

SOURCES OF MUTATIONS ENDOGENOUS DNA DAMAGE EXOGENOUS DNA DAMAGE Depurination DNA REPAIR MUTATION Life Styles Environmental Agents Free Radicals Polymerase Errors CELL REPLICATION

Chemical Exposure (air, water, food, etc.) Internal Exposure Metabolic Activation Macromolecular Binding Detoxication DNA RNA Protein Biologically Effective Dose Efficiency of Mispairing Cell Proliferation X X Initiation (Biomarker)

Williams J.A., Carcinogenesis 22: (2001)

Accumulation of mutations during tumor progression Loeb L.A. Cancer Res. 61: (2001)

Epigenetic alterations – changes induced in cells that alter the expression of the information on transcriptional, translational, or post- translational levels without changes in DNA sequence EPIGENETICS SAMSAH DNMT1 DNMT3a DNMT3b Methylation of DNA Modifications of histones RNA-mediated modifications RNA-directed DNA methylation RNA-mediated chromatin remodeling RNAi, siRNA, miRNA … A Me P U - acetylation - methylation - phosphorylation - ubiquitination P U Me A

GENETIC AND EPIGENETIC MODELS OF THE CANCER INITIATION Epigenetically reprogrammed cells Mutator phenotype cells Endogenous Environmental ALTERATIONS IN CELLULAR EPIGENOME Normal cells Cancer cells Clonal selection and expression of initiated cells Mutator phenotype cells Endogenous Environmental ACQUISITION OF ADDITIONAL RANDOM MUTATIONS Normal cells Cancer cells

Cellular and Molecular Mechanisms in Multistage Carcinogenesis: PROMOTION Reversible enhancement/repression of gene expression: - increased cell proliferation - inhibition of apoptosis No direct structural alteration in DNA by agent or its metabolites From

Role of Increased Cell Proliferation in Carcinogenesis Decreases time available for DNA repair Converts repairable DNA damage intononrepairable mutations (not DNA damage anymore!) Necessary for chromosomal aberrations, insertions, deletions and gene amplification Clonally expands existing cell populations with mutations

No Tumors Tumors No Tumors Tumors X X X X Time X = Application of Initiator = Application of Promoter

N Basophilic Focus AdenomaCarcinoma M 1 M N PromotionRegressionProgression Adapted from: Marsman and Popp. Carcinogenesis 15: (1994) No Tumors Tumors = Application of Promoter

Cellular and Molecular Mechanisms in Multistage Carcinogenesis: PROGRESSION Irreversible enhancement/repression of gene expression Complex genetic alterations (chromosomal translocations, deletions, gene amplifications, recombinations, etc.) Selection of neoplastic cells for optimal growth genotype/ phenotype in response to the cellular environment From “Complex” genetic changes

Immortalization Transformation Loss of contact growth inhibition Autonomy of proliferation Avoidance of apoptosis Aberrant differentiation Induction of angiogenesis Phenotypic characteristics of cancer cells:

Human Tumors and Stages of Carcinogenesis Hussain et al., Oncogene, 2007