1. Cancer Etiology 1. Introduction 2. Chemical Factors in Carcinogenesis 3. Physical Factors in Carcinogenesis 4. Viral Oncogenesis 5. Genetic Predisposition
邵吉民，教授，病理学与病理生理学系

2. Introduction
Tumor
Benign tumor
Malignant tumor

3. 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。

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

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

6. 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 6

7. 7

8. Initiation Irreversible genetic damage:
A necessary, but insufficient prerequisite for tumor initiation
Activation of proto-oncogene, inactivation of a tumor suppressor gene, and etc 8

9. 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 9

10. 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 10

11. 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 11

12. 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 12

13. Gene-environmental interactions
The metabolism of xenobiotics by biologic systems
Individual variation
The competition between activation and detoxication
The alteration of genes by xenobiotics 13

14. Classification of chemical carcinogens
1. Based on mechanisms
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[a]pyrene, 2-acetylaminofluorene, benzidine, Aflatoxin B1, B2. 14

15. 15

16. (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 16

17. 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 17

18. 18

19. 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 19

20. Direct Chemical Carcinogens
(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 Jan 12;12(2): doi: /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 20

21. ---Monofunctional alkylating agents
MNNG N-Methyl-N-nitroso-N'-nitroguanidine
Numerous potential reaction sites for alkylation have been identified in all four bases of DNA (not all of them have equal reactivity: 21

22. ---Bifunctional alkylating agents22

23. Indirect Chemical Carcinogens and Their Phase I Metabolic derivatives23

24. BPDE binds DNA covalently, resulting in bulky adduct damage
BPDE intercalates into dsDNA non-covalently, leading to conformational abnormalities

25. 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 25

26. DNA Repair 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 26

27. 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
XP:着色性干皮病患者的皮肤部位缺乏核酸内切酶,不能修复被紫外线损伤的皮肤的DNA,因此在日光照射后皮肤容易被紫外线损伤,先是出现皮肤炎症,继而可发生皮肤癌... 27

28. 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 28

29. Translesion DNA synthesis29

30. 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

31. 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

32. 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

33. Physical factors in carcinogenesis

34. Physical carcinogens Corpuscular radiations
Electromagnetic radiations (EMF)
Ultraviolet lights (UV)
Low and high temperatures
Mechanical traumas
Solid and gel materials

36. Viral Oncogenesis
RNA Oncovirus (Retrovirus)
DNA Oncovirus 36

37. RNA Oncovirus Retroviruses: Rous sarcoma in chickens (RSV): in 1911
ssRNA viruses
Reverse transcriptase
Oncogenes
Rous sarcoma in chickens (RSV): in 1911
Human T-cell lymphotropic virus (HTLV-I,II)
Human immunodeficiency virus (HIV)

38. Classification of retrovirus38

39. Structure of RNA Oncovirus39

40. Genome of RNA Oncovirus and Gene Products
Genome of Human T-cell Leukemia virus (HTLV) 40

41. Life cycle Receptor binding and membrane fusion
Internalization and uncoating
Reverse transcription of the RNA genome to form double-stranded linear DNA
Nuclear entry of the DNA
Integration of the linear DNA into host chromosomal DNA to form the provirus
Transcription of the provirus to form viral RNAs
Splicing and nuclear export of the RNAs
Translation of the RNAs to form precursor proteins
Assembly of the virion and packaging of the viral RNA genome
Budding and release of the virions
Proteolytic processing of the precursors and maturation of the virions 41

42. Replication of RNA Oncovirus42

43. 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 43

44. 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) 44

45. 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; 45

46. 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 46

47. 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. 47

48. DNA Oncovirus Papilloma virus Polyoma virus Adenovirus
Herpes virus: EB virus
Hepatitis B virus 48

49. 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
SV 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 49

50. Gene Map and Function of HPV
ORF Function
E Virus proliferation
E Regulation of transcription
E5、E6、E Cell transformation
L1、L Encoding capsid protein
E Encoding late cytosolic protein
E3、E Unkown
E5: activates growth factor receptor
E6: ubiquitin-mediated degradation of p53
E7: binds and inactivates unphosphorylated pRb 50

51. Genome and Products of HBV
李文辉博士,北京生命科学研究所研究员.本实验室的主要研究兴趣为囊膜病毒侵入及相关过程的分子机制。病毒侵入细胞起始于与细胞表面受体的结合，围绕这一过程，我们希望探索：病毒细胞受体分子的识别；病毒受体是如何介导膜融合的；其它胞内因子是如何参与病毒侵入的；病毒蛋白是如何进行免疫逃逸的；病毒跨物种感染与传播；如何有效地抑制病毒的侵入，等。我们将以几种重要的病毒如新型冠状病毒，出血热病毒，乙脑病毒，乙肝病毒等为研究对象，综合运用以生物化学为主，无生物安全隐忧的多学科方法，以回答上述问题。这些研究将有利于深化对于这些病毒基本生物学及病毒与感染宿主相互作用的认识与理解，并且有助于开发有效的抗病毒制剂及新型疫苗。
乙型肝炎病毒（HBV）及其卫星病毒丁型肝炎病毒（HDV）必须通过结合细胞表面受体分子来实现对宿主细胞的感染。因此，发现该受体，以深入理解乙肝感染过程并为乙肝及其相关疾病提供有效的治疗靶点
在这篇文章中，研究人员从树鼩这种动物入手，开始了他们的探索之旅。树鼩是一种与灵长类动物类似的小动物，也是除人类和黑猩猩以外，唯一能被乙肝病毒感染的物种。生物通
李文辉博士的团队首先绘制了一幅高质量的树鼩肝细胞基因表达图谱，为后续的研究奠定了坚实的基础。有了这个数据库，再结合先进的纯化技术和高分辨质谱分析手段，他们发现，肝脏胆酸转运蛋白（NTCP，钠离子-牛磺胆酸钠共转运多肽）会与乙肝病毒表面包膜大蛋白的关键受体结合区发生特异性相互作用。
随后，他们在HBV/HDV易感的肝细胞中进行的一系列基因敲除实验，证明NTCP的确是病毒感染所需的细胞受体。人肝癌细胞株HepG2细胞通常情况下不表达NTCP，也不能被HBV感染, 他们深入研究发现，如果在该细胞株中外源性表达人或树鼩的 NTCP后，则该细胞可以被HBV及HDV感染。他们还鉴定出NTCP上关键的病毒结合区域。比如猴子的NTCP通常不能结合乙肝病毒，但只要突变其NTCP上一段极小的区域，就能使之变成有效的HBV受体。
Transforming gene: X gene
X protein activates gene transcription via XRE 51

52. 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 52