VIRAL ONCOGENESIS

1
VIRAL ONCOGENESIS

• Professor SJ Nayler
• Division of Anatomical Pathology, School of
  Pathology, University of the Witwatersrand and
  the National Health Laboratory Service
• GEMP II, MBBCH, 2006

2
VIRAL ONCOGENESIS

• DNA Viruses
• Human Papilloma Virus (HPV)
• Epstein Barr Virus (EBV)
• Human Herpes Virus 8 (HHV-8)
• Hepatitis B Virus
• RNA Viruses
• Human T-cell leukaemia virus 1 (HTLV-1)

3
DNA Viruses

• Transforming DNA virus
• Form stable associations with host genome
• Integrated virus cant complete replicative
  cycle, essential genes interrupted
• Viral genes that are transcribed early in life
  cycle are expressed in transformed cells

4
Epstein Barr Virus

• Member Herpes virus
• 4 major cancers
• African Burkitts lymphoma
• B-cell Non-Hodgkins lymphoma in HIV
• Hodgkins Lymphoma
• Nasopharyngeal carcinoma

5
EBV

• Infects cell of naopharynx B-lymphocytes
• CD21 molecule
• Genome (linear) ?circularises ? episome in
  nucleus
• Latent infection ? no viral replication and cells
  immortalised
• Viral genes dysregulate N proliferative and
  survival functions
• LMP-1
• prevents apoptosis, by up-regulating bcl-2
• Activates growth promoting pathways (normally
  induced by T-cell)

6
EBV

• Burkitt Lymphoma
• N EBV infection controlled by immune response to
  membrane EBV proteins
• Endemic BL ? Immune factors (chronic malaria) ?
  proliferation of immortalised B-cells
• Actively dividing cells ? mutations (esp.
• t(814) juxtaposes c-myc with immunoglobulin
  loci) ?significant role in oncogenesis
• Multistep progression

7
EBV

• NASOPHARYNGEAL CARCINOMA
• Endemic in SEA, Africa, Arctic Eskimos
• 100 NPC ? clonal integrated EBV
• Plays and important role in tandem with other
  factors
• Genetic
• Enviromental

8
Hepatitis B

• Strong relationship with hepatocellular carcinoma
• 200X ?risk for developing HCC
• Integrated into DNA
• Does not encode oncoproteins
• Probably indirect effect
• Chronic liver damage ? regenerative hyperplasia ?
  mutations
• HBV encodes for HBx protein
• ?activates growth promoting genes (IGF-II) ?
  growth
• Binds p53 interferes with suppressor function

9
HUMAN HERPES VIRUS 8

• Kaposis sarcoma
• Some Non-Hodgkins lymphoma

10
Cervical CarcinomaEPIDEMIOLOGY

• 2nd most common cancer among all women (after
  breast cancer)
• 4 out of every 5 newly diagnosed cases occur in
  developing countries worldwide
• Until recently, the most frequently diagnosed
  malignant neoplasm among black S. A. women

11
Cervical CarcinomaEPIDEMIOLOGY

• Age standardised incidence rate among S.A. women
  22/100 000
• Calculated lifetime risk among black S.A. females
  1 in 34
• Similar incidence to other developing countries
  ranks among the highest in the world

12
Carcinoma of the Cx
13
Squamous cell Ca
14
Cervical CarcinomaEPIDEMIOLOGY

• Evidence indicates a direct causal relationship
  with sexual activity, i.e.
• Early onset of sexual activity
• Multiple sexual partners
• Exposure to high-risk males
• Human papilloma virus (HPV) has been identified
  as the infectious aetiological agent associated
  with cervical Ca

15
Cervical Carcinoma is a Sexually Transmitted
Disease
FOOD FOR THOUGHT
16
Human Papilloma VirusEVIDENCE FOR ITS ROLE IN
CERVICAL CANCER

• Reported worldwide prevalence of detectable HPV
  DNA sequences in cervical cancer specimens is
  99.7
• Demonstration of HPV integration in the nuclei
  (genome) of cervical cancer cells

17
Human Papilloma VirusPHYSICAL STRUCTURE

• Episomal form exists as an icosahedral-shaped
  virion with a diameter of 55nm

18
Human Papilloma VirusPHYSICAL STRUCTURE

• Icosahedral capsid consists of 72 capsomeres
• Major capsid proteins are antigenically
  cross-reactive among all HPV types

19
(No Transcript)
20
Human Papilloma VirusGENOMIC STRUCTURE

• Closed, circular double-stranded DNA of 8000
  base pairs
• Three functional regions
• - Six early genes/ open reading frames
• (E1, E2, E4, E5, E6 E7)
• - Two late genes/ORFs (L1 L2)
• - An upstream regulatory region (URR)

21
L1
L2
URR
E1
E7
E2
E6
E4
Schematic representation of the HPV genome
E5
22
Human Papilloma VirusGENOMIC STRUCTURE

• Genes in the early region
• Proteins encoded are responsible for
  transcription, replication cellular
  transformation
• E6 E7 are the most important ORFs implicated in
  cervical carcinogenesis ? encoded proteins are
  capable of inducing cellular proliferation
  transformation

23
Human Papilloma VirusGENOMIC STRUCTURE

• Genes in the late region (L1 L2)
• Encode for structural proteins essential to viral
  assembly (i.e. minor major capsid proteins)

24
Human Papilloma VirusGENOMIC STRUCTURE

• Upstream regulatory region (URR)
• Regulates expression of all ORFs, including
  promoter elements transcriptional enhancer
  sequences
• Proteins encoded by E2 interact with URR ? ve
  ve effects on transcription

25
Human Papilloma VirusSUBTYPES

• gt100 HPV types described thus far
• HPVs affecting the Cx are grouped according to
  their risk for neoplastic transformation (i.e.
  risk for developing cervical cancer following
  infection)

26
Human Papilloma VirusSUBTYPES INVOLVING THE
CERVIX

• Low-risk types
• Include HPV 6, 11, 42-44, 53-55
• Associated with genital condylomata low-grade
  dysplasia (CIN 1)

27
(No Transcript)
28
(No Transcript)
29
HPV Infection
30
Human Papilloma VirusSUBTYPES INVOLVING THE
CERVIX

• Intermediate risk types
• Include HPV 33, 35, 39, 41, 52, etc.
• Associated with moderate risk of neoplastic
  transformation

31
Human Papilloma VirusSUBTYPES INVOLVING THE
CERVIX

• High-risk types
• Include HPV 16, 18, 31 45
• Strong association with high-grade dysplasia (CIN
  3) invasive carcinoma

32
Human Papilloma VirusPHYSICAL STATE IN HOST
NUCLEI

• HPV exists in 2 forms, i.e. EPISOMAL or
  INTEGRATED
• Low-risk HPV types (e.g. 6 11)
• Remain episomal (not integrated into host genome)
• Result in low-grade dysplasia, which is
  potentially reversible

33
Human Papilloma VirusPHYSICAL STATE IN HOST
NUCLEI

• High-risk HPV types (e.g. 16 18)
• Integrated into the host genome
• Result in progressive dysplasia ? CIN 2 ? CIN 3
  (carcinoma in situ) ? microinvasive Ca ? frankly
  invasive Ca

34
HPV Infection CIN 3
35
CIN 3 detected in a cervical cytology (Pap) smear
36
HPV Cervical CarcinomaMOLECULAR PATHOGENESIS

• Integration of high-risk HPV into host genome

• Disruption of E2 gene

• Loss of repressive effects on E6 E7 gene couple

• Enhanced expression of E6 E7 oncoproteins

• E6 E7 oncoproteins bind to host p53 Rb tumour
  suppressor gene products, respectively

• Dysregulation of N cell cycle

• Uncontrolled neoplastic proliferation

37
STEP 1 Disruption of E2 following viral
integration, with loss of control over E6 E7
L1
L2
URR
E1
E7
?
E2
E6
E4
E5
38
STEP 2 Enhanced expression of E6 E7
oncoproteins
L1
L2
URR
E1
E7
?
E2
E6
E4
E5
39
STEP 3 Binding of E6 E7 to host tumour
suppressor gene products
L1
L2
URR
E1
Rb
E7
?
E2
p53
E6
E4
E5
40
STEP 4 Dysregulation of normal host cell cycle,
with uncontrolled proliferation
L1
L2
URR
E1
Rb
E7
?
E2
p53
E6
E4
E5
Neoplasia
41
HPV Cervical CarcinomaROLE OF CO-FACTORS

• HPV infection alone is not responsible for
  cervical carcinogenesis
• Possible co-factors implicated include
• - Hormones
• - Cigarette smoking
• - Immune status

42
Cervical CancerHISTOLOGICAL SUBTYPES

• Squamous cell Ca (vast majority)
• Adenocarcinoma
• Adenosquamous Ca
• Other rare types, including
• Small cell Ca
• Adenoid cystic Ca
• Adenoid basal Ca
• Carcinosarcoma

43
Squamous cell Ca
44
Adenocarcinoma
45
Adenosquamous Ca
46
Small cell Ca
47
Adenoid cystic Ca Adenoid
basal Ca
48
Carcinosarcoma of the Cx
49
Human Papilloma VirusOVERVIEW OF DETECTION
METHODS

• Traditional methods
• Light microscopy
• Electron microscopy
• Immunohistochemistry
• Molecular methods
• With preservation of tissue morphology
• Without preservation of tissue morphology

50
Human Papilloma VirusMOLECULAR DETECTION
METHODS WITHOUT PRESERVATION OF MORPHOLOGY

• Hybridisation methods, e.g.
• Southern blot hybridisation
• Solution phase hybridisation
• Amplification methods, e.g.
• Polymerase chain reaction (PCR)

51
PCR detection of the HPV L1 gene in invasive Ca
of the Cx
52
Human Papilloma VirusMOLECULAR DETECTION
METHODS WITH PRESERVATION OF MORPHOLOGY

• In situ hybridisation (ISH)
• Isotopic ISH
• Non-isotopic ISH (NISH)
• In situ polymerase chain reaction
• (In situ PCR)

53
Benefits of NISH in HPV Detection in Cervical
Cancer Specimens

• Preservation of tissue morphology
• Identification of specific HPV type(s)
• Provides information concerning the physical
  state of the virus in neoplastic nuclei (episomal
  and/or integrated) via the generation of specific
  signal types
• Ability to be completed in a single day

54
NISH signals for HPV 16 in CIN 3
55
NISH signals for HPV 16 in invasive Ca of the Cx
56
NISH signals for HPV 16 in invasive Ca of the Cx
57
RNA VIRUES

• HTLV-1
• Leukemia endemic Japan / Caribbean
• Tropism for CD4 cells
• Self reading for mechanism

58
(No Transcript)