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Cancer Etiology

1. Introduction 2. Chemical Factors in Carcinogenesis 3. Physical Factors in Carcinogenesis4. Viral Oncogenesis5. Genetic Predisposition

邵吉民，教授，病理学与病理生理学系shaojimin@zju.edu.cn

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Tumor Benign tumor Malignant tumor

Introduction

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Cancer Incidence and Mortality

Siegel R, Naishadham D, Jemal A. Cancer statistics, 2012.

CA Cancer J Clin. 2012;62(1):10-29. 1,638,910 new cancer cases and 577,190 deaths from cancer are projected to occur in USA in 2012.One in 4 deaths in USA is due to cancer.

2010 年国际抗癌联盟（ UICC ）： 2008 年全世界 1270 万新增癌症患者，死亡人数 760 万。

全国肿瘤登记中心《 2012 中国肿瘤登记年报》 每年新发肿瘤病例约 312 万例，每天约 8550 人 ; 每年因癌死亡 270 万例 , 居民因癌死亡率 13% ，即每 7-8 人中有 1 人因癌死亡。 恶性肿瘤发病 : 第一位肺癌，其次胃癌、结直肠癌、肝癌和食管癌 ;恶性肿瘤死亡：第一位肺癌，其次肝癌、胃癌、食管癌和结直肠癌；中国近 20 年来癌症呈现年轻化及发病率和死亡率“三线”走高的趋势。癌症种类呈现地域化特点。

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History of Cancer Research

Kiberstis P, Marshall E. Cancer crusade at 40. Celebrating an anniversary. Introduction. Science. 2011;331(6024):1539.

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Chemical Carcinogenesis

Multi-stage Theory of Chemical Carcinogenesis Classification of chemical carcinogens Mechanisms of Chemical Carcinogenesis DNA Damage Induced by Ultimate Carcinogens DNA Repair

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Multi-stage Theory of Chemical Carcinogenesis

Initiation -----------Genetic events

Chemical Carcinogens (Direct and Indirect Carcinogens)

Promotion -------Epigenetic events

Tumor promoters

– Murine skin carcinogenesis model:

• A single dose of polycyclic aromatic hydrocarbon (PAH, initiator)

• Repeated doses of croton oil (promoter)

Malignant conversion

Progression ------Genetic and epigenetic events

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Initiation

• Irreversible genetic damage:

A necessary, but insufficient prerequisite for tumor initiation

• Activation of proto-oncogene, inactivation of a tumor suppressor gene, and etc

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Promotion • Promotion: Selective expansion of initiated cells, which

are at risk of further genetic changes and malignant conversion

• Promoters are usually nonmutagenic, not carcinogenic alone, often do not need metabolic activation, can induce tumor in conjuction with a dose of an initiator that is too low to be carcinogenic alone

• Chemicals capable of both initiation and promotion are called complete carcinogens: benzo[a]pyrene and 4-aminobiphenyl

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Malignant conversion • The transformation of a preneoplastic cell into

that expresses the malignant phenotype• Further genetic changes• Reversible• The further genetic changes may result from

infidelity of DNA synthesis• May be mediated through the activation of

proto-oncogene and inactivation of tumor-suppressor gene

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Progression

• The expression of malignant phenotype, the tendency to acquire more aggressive characteristics, Metastasis

• Propensity for genomic instability and uncontrolled growth

• Further genetic changes: the activation of proto-oncogenes and the inactivation of tumor-suppressor genes

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• Activation of proto-oncogenes:– Point mutations: ras gene family, hotspots– Overexpression:

• Amplification

• Translocation

• Loss of function of tumor-suppressor genes: usually a bimodal fashion– Point mutation in one allele– Loss of second allele by deletion, recombinational

event, or chromosomal nondisjunction

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Gene-environmental interactions

• The metabolism of xenobiotics by biologic systems– Individual variation – The competition between activation and detoxication

• The alteration of genes by xenobiotics

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Classification of chemical carcinogens

1. Based on mechanisms(1) Genotoxic carcinogen (DNA-reactive)• Direct-acting: intrinsically reactive

N-methyl-N’-nitro-N-nitrosoguanidine (MNNG),

methyl methanesulfonate (MMS),

N-ethyl-N-nitrosourea (ENU), nitrogen and sulfur mustards • Indirect-acting: require metabolic activation by cellular enzyme to form the DNA-

reactive metabolite (members of the cytochrome P450 family) benzo[]pyrene, 2-acetylaminofluorene, benzidine, Aflatoxin B1, B2.

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(2) Epigenetic carcinogens

• Promotes cancer in ways other than direct DNA damage/ do not change the primary sequence of DNA

• Alter the expression or repression of certain genes and cellular events related to proliferation and differentiation

• Promoters, hormone modifying agents, peroxisome proliferators, cytotoxic agents, and immunosuppressors

• Organochlorine pesticides, [saccharin], estrogen, cyclosporine A, azathioprine

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2. Based on sturcture(1) Nitrosamines (NA)

MNNG, MMS (direct carcinogen)

(2) Polycyclic aromatic hydrocarbons (PAH)

Benzo(a)pyrene (indirect carcinogen)

(3) Aromatic amines (AA)

2-acetylaminofluorene, benzidine (indirect carcinogen)

(4) Aflatoxin (AF)

(5) Inorganic elements and their compounds: arsenic, chromium,

and nickel are also considered genotoxic agents

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Mechanisms of Initiation in Chemical Carcinogenesis

(1) DNA damages:Pro-carcinogen metabolic activation (Phase I and II) Ultimate carcinogen (electrophiles) Interaction with macromolecules (nucleophiles) DNA damage, mutations, chromosomal aberrations, or cell death

(2) Epigenetic changes

(3)Activation of oncogenes; inactivation of tumor suppressor genes

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(1) Alkylating agents are electrophilic compounds with affinity for

nucleophilic centers in organic macromolecules.[Fu D, Calvo JA, Samson LD. Balancing repair and tolerance of DNA damage caused by alkylating agents. Nat Rev Cancer. 2012 Jan 12;12(2):104-20. doi: 10.1038/nrc3185.]

(2) These agents can be either monofunctional or bifunctional.

---Monofunctional alkylating agents have a single reactive group and

thus interact covalently with single nucleophilic centers in DNA (although varied).

such as MNNG

---Bifunctional alkylating agents have two reactive groups, and each molecule is potentially able to react with two sites in DNA.

Interstrand DNA cross-link: the two sites are on opposite polynucleotide strands;

Intrastrand cross-link: on the same polynucleotide chain of a DNA duplex.

such as Nitrogen and sulfur mustard, mitomycin, cis-platinum

Direct Chemical Carcinogens

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Numerous potential reaction sites for alkylation have been identified in all four bases of DNA (not all of them have equal reactivity:

MNNG N-Methyl-N-nitroso-N'-nitroguanidine

---Monofunctional alkylating agents

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---Bifunctional alkylating agents

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Indirect Chemical Carcinogensand Their Phase I Metabolic derivatives

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BPDE binds DNA covalently, resulting in bulky adduct damage

BPDE intercalates into dsDNA non-covalently, leading to conformational abnormalities

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Types of DNA Damage Induced by Ultimate Carcinogens

• DNA Adduct Formation• DNA Break Single Strand Break Double Strand Break • DNA Linkage DNA-DNA linkage DNA-protein Linkage• Intercalation

Bulky aromatic-type adducts, Alkylation (small adducts),

Oxidation, Dimerization, Deamination

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Repair systems• Direct DNA repair/ Direct reversal :

– DNA alkyltransferase (O6-alkylguanine-DNA alkyl transferase)

– One enzyme per lesion

• Base excision repair (BER)– small adducts, – overlap with direct repair – glycosylase to remove the adducted base

DNA Repair

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• Nucleotide excision repair (NER): – involves recognition, preincision, incision, gap-filling,

and ligation, – large distortions – strand specific, the transcribed strand is preferentially

repaired – xeroderma pigmentosum (XP): NER deficiency

• Mismatch repair (MMR) – transition mispairs are more efficiently repaired (G-T

or A-C) than transversion mispairs – microenvironment influences efficiency – similar to NER – involves the excision of large pieces of the DNA

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• Double-strand breaks (DSBs) – homologous recombination – non-homologous end joining (NHEJ): DNA-PK

• Postreplication repair – a damage tolerance mechanism – occurs in response to replication of DNA on a

damaged template – the gap

• either filled through homologous recombination with parental strand

• or insert an A residue at the single nucleotide gap

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Translesion DNA synthesis

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Hormones and the etiology of cancer

• Major carcinogenic consequence of hormone exposure: cell proliferation

• The emergence of a malignant phenotype depends on a series of somatic mutation

• Germline mutations may also occur• How to get exposure: contraceptives, hormone

replacement therapy, or during prevention of miscarriage

• Epidemiological studies

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Hormone-related cancer

• Breast cancer and estrogen

• Endometrial cancer: Estrogen replacement therapy

• Ovarian cancer: follicle stimulating hormone

• Prostate cancer and androgen

• Vaginal adenocarcinoma: in utero diethylstilbestrol (DES) exposure

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Other hormone-related cancers

• Cervical cancer: OC use might increase the risk, still a lot complicating factors

• Thyroid cancer: the pituitary hormone thyroid stimulating hormone (TSH)

• Osteosarcoma: incidence associates with the pattern of childhood skeleton growth; and hormonal activity is a primary stimulus for skeleton growth

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Physical factors in carcinogenesis

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Physical carcinogens

– Corpuscular radiations

– Electromagnetic radiations (EMF)

– Ultraviolet lights (UV)

– Low and high temperatures

– Mechanical traumas

– Solid and gel materials

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Viral Oncogenesis

• RNA Oncovirus (Retrovirus)

• DNA Oncovirus

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RNA Oncovirus

Retroviruses: ssRNA viruses

Reverse transcriptase

Oncogenes

Rous sarcoma in chickens (RSV): in 1911

Human T-cell lymphotropic virus (HTLV-I,II)

Human immunodeficiency virus (HIV)

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Classification of retrovirus

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Structure of RNA Oncovirus

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Genome of RNA Oncovirus and Gene Products

Genome of Human T-cell Leukemia virus (HTLV)

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Life cycle1. Receptor binding and membrane fusion 2. Internalization and uncoating 3. Reverse transcription of the RNA genome to form double-stranded

linear DNA 4. Nuclear entry of the DNA 5. Integration of the linear DNA into host chromosomal DNA to form the

provirus 6. Transcription of the provirus to form viral RNAs

7. Splicing and nuclear export of the RNAs

8. Translation of the RNAs to form precursor proteins

9. Assembly of the virion and packaging of the viral RNA genome

10. Budding and release of the virions

11. Proteolytic processing of the precursors and maturation of the virions

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Replication of RNA Oncovirus

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Mechanisms of Oncogenesis Induced by RNA Oncovirus

• Transducing Retrovirus v-onc

• cis-Activating Retrovirus c-onc

• trans-Activating Retrovirus tax trans-acting x p40tax

rex repressive expression x p27rex, p21rex

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• Oncogene transduction– Acutely transforming in vivo and in vitro– Transform cells by the delivery (transduction) of an

oncogene from the host cell (v-onc) to a target cell– Cause the formation of polyclonal tumors– Most of this group of viruses are replication defective

(the requirement of a helper virus) – Examples: RSV (v-src); Abelson murine leukemia virus (v-Abl)

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•Insertional activation

– Long latent periods, Less efficient– Do not induce transformation of cells in vitro– Usually are replication competent– No oncogenes– Tumors are usually monoclonal– Provirus (LTR) is found within the vincity of a proto-

oncogene (c-myc)– Examples: lymphoid leukosis virus;

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•Grow stimulation and two-step oncogenesis

– The defective spleen focus-forming virus (SFFV) and its helper, the Friend murine leukemia virus (Fr-MuLV)

– Induce a polyclonal erythrocytosis in mice– Require the continued viral replication– A mutant env protein gp55 of SFFV binds and

stimulated the erythropoietin receptor, thus inducing erythroid hyperplasia

– Fr-MuLV or SFFV integration inactivates p53

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• Transactivation

– HTLV-1 and 2– Like cis-activation group: replication competent, carries no

oncogene, induces monoclonal leukemia, and latent– Like transducing group: can immortalize cells in vitro, has no

specific integration site– Unique 3’ genomic structure: the X region; Encodes at least

three proteins: Tax (p40), Rex (p27, p21)– Tax is the focus

– Transactivate the viral LTR, results in a 100- to 200-fold increase in the rate of proviral transcription

– Transactivate cellular enhancers and promoters, including genes for IL-2, granulocyte-macrophage colony-stimulating factor (GM-CSF), c-fos, and others.

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DNA Oncovirus

Papilloma virus

Polyoma virus

Adenovirus

Herpes virus: EB virus

Hepatitis B virus

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Mechanism of Oncogenesis Induced by DNA Oncovirus

Transforming proteins 1. HPV E6 interact with P53 E7 interact with RB

2. Adenovirus E1a interact with RB E1b

3. Polyoma virus SV40 Large T interact with RB Py virus Large and Middle T

Transcription activators 1. EB virus EBNA-2 and LMP 2. HBV p28 X protein

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Gene Map and Function of HPV

ORF Function

E1 Virus proliferationE2 Regulation of transcriptionE5 、 E6 、 E7 Cell transformationL1 、 L2 Encoding capsid proteinE4 Encoding late cytosolic proteinE3 、 E8 Unkown

E5: activates growth factor receptorE6: ubiquitin-mediated degradation of p53E7: binds and inactivates unphosphorylated pRb

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Genome and Products of HBV

Transforming gene: X gene X protein activates gene transcription via XRE

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Genetic Predisposition • Hereditary Cancer• Tumor Genetic Susceptibility

---Tumor susceptibility genes (Cytochrome P450 family, DNA repair genes, Tumor suppressor genes, etc)

• Immunity• Hormones and metabolism• Psychological factors• others