Molecular Basis of Cancer

1
Lecture 1 Molecular Basis of Cancer
2
What Is Cancer?
Cancer is a group of diseases caused by the
uncontrolled
multiplication of abnormal cells in the body, a
process called
neoplasia.
Abnormal new tissues called neoplasms are
formed.
Neoplasms usually form masses called tumors
that may be
benign (non cancerous) or malignant (cancerous).
Malignant or cancerous tumors grow rapidly,are
invasive
(to surrounding tissue) and metastatic (traveling
via
blood/lymph to invade distant tissues).
Cancers destroy healthy tissues causing loss
of function and
death.
Cancer is the 2nd major killer in populations
of developed
countries the leading cause of death in
children 3-15 (US).
Cancers are genetic disorders caused by
accumulation of
somatic mutations (gene chromosome) in a
persons cells.
Inherited mutations give a predisposition for
certain cancers.
3
Characteristics of Cancer Cells
Cancer cells are genetically altered via gene
or
chromosome mutations so
- lack normal controls over cell division or
apoptosis.
- may express inappropriate genes (e.g. for
telomerase,
enzyme that maintains length of DNA for continued
division)
- are genetically unstable due to loss of DNA
repair
mechanisms (so are more susceptible to radiation
damage
than normal cells).
Divide excessively (proliferate)
indefinitely producing
neoplasms.
Live indefinitely (do not show apoptosis).
Lose the normal attachment to other cells so
become
metastatic (travelling via blood/lymph to invade
distant
sites).
Secrete signals for angiogenesis (growth of
blood vessels
into tumor).
4
Cancer Cells are Undifferentiated
Malignant
Cancer cells are
undifferentiated to
varying degrees
(even anaplastic, like
stem cells) so divide
do not perform the
normal function of
mature cells.
The less
differentiated the
cancer cell the more
malignant the
cancer (the more
rapidly growing is the
tumor).
5
What Causes Cancer?
Inherited mutations in genes that affect cell
cycle,
DNA repair, or apoptosis these mutations give a
genetic predisposition for cancer.
Somatic mutations to these same genes caused by
Exposure to risk factors
- environmental mutagens (carcinogenic chemicals,
radiation)
- hormones
- weakening of immune system (as in AIDS).
Oncogenic (tumor) Virus infections
- Epstein Barr virus (causes Burkitt lymphoma)
- Human Papilloma Virus (causes cervical cancer).
Tumor viruses transform human cells into cancer
cells by
Introducing viral cancer - causing oncogenes
into host cell DNA
Causing Translocation and overexpression of host
protooncogenes.
6

• Normal cell cycle is controlled by signal
  transduction
• Growth factors bind to surface receptors on the
  cell transmembrane proteins relay signals into
  the cell.
• Two types of growth factors
• Growth factors stimulate cell division.
• Growth-inhibiting factors inhibit cell
  division.
• Healthy cells divide only when growth factor and
  growth-inhibiting factor balance favors cell
  division.
• Cancer cells divide without constraint
• (e.g., mutations in growth and growth-inhibiting
  factor genes).

7
Regulation of cell division by signal
transduction.
8
The Cell Cycle
Oncogenes
G2 (cell growth)
M (mitosis)
S (synthesis)
DNA repair genes
G1
G0 (resting)
Tumor suppressor genes
9
Cell cycle
CDK
p53 is known as the guardian of the genome
10
CONTROL of CELL CYCLE
Differentiated
The Cell has 3 major
G0
Cells enter
checkpoints that are
non dividing
G1
sensitive to signals, the
G0 phase-
checkpoint
G1, the G2 and the M
checkpoints.
If a cell does not
G1
pass G1 checkpoint
Control
system
it enters a non-
S
dividing GO Phase.
Most somatic cells
G2
M
are in GO.
Some cells in GO
(e.g. liver cells) can
reenter cell cycle if
M checkpoint
G2 checkpoint
needed.
In cancer cells genes that control
cell cycle have mutated
so cells divide excessively producing
neoplasms.
11

• Cancer and genes
• Three classes of genes are frequently mutated in
  cancer
• Proto-oncogenes (? oncogenes)
• Tumor suppressor genes
• Mutator genes

12

• Proto-oncogenes ? oncogenes
• Proto-oncogenes
• Proto-oncgenes are genes that possess normal gene
  products and stimulate normal cell development.
• Oncogenes
• Oncogenes arise from mutant proto-oncogenes.
• Oncogenes are more active than normal or active
  at inappropriate times and stimulate unregulated
  cell proliferation.
• Some tumor viruses that infect cells possess
  oncogenes
• RNA tumor viruses possess viral oncogenes
  (derived form cellular proto-oncogenes) capable
  of transforming cells to a cancerous state.
• DNA tumor viruses another class of tumor
  viruses do not carry oncogenes, but induce
  cancer by activity of viral gene products on the
  cell (no transformation per se).

13

• Types effects of different types of mutations
• Point mutations occur in protein coding or
  controlling sequences.
• Deletion frameshifts may lead to defective
  proteins.
• Gene amplification random over-replication of
  small segments of DNA results in extra copies
  (up-regulates cell growth).
• Mutator genes
• Mutator gene increases spontaneous mutation rate
  of other genes.
• Mutator gene products are involved in DNA
  replication and repair mutations make the cell
  error prone.

14
Proteins Control Cell Division
The cell cycle is controlled by
proteins
from inside outside the cell.
Intracellular Cyclins and Cyclin Dependent
Kinases
(CDKs) control the checkpoints.
Hormones or extracellular proteins from other
cells
(called Growth Factors) signal target cell to
divide.
- Hormones (e.g. Growth Hormone) or Growth Factors
bind to receptor proteins of target cell membrane.
- This triggers a molecular signaling pathway.
- A series of linked proteins activate Cyclin-
CDKs which
Allows Cells to Pass Cell Cycle Checkpoints
divide.
15
How Growth Factors Trigger Cell
Division
Growth Factor
or Hormone
Plasma
membrane
Relay
G1 checkpoint prevents
Proteins open
damaged DNA from replicating
checkpoints
Checkpoint controlled by
Receptor
Cyclin - CDK
protein
Signal
Transduction
Control
Pathway
G1
S
system
G2
M
16
Tumor Suppressor Proteins Inhibit
Cell
Division Prevent Cancer
Tumor suppressor proteins are
proteins that
bind to checkpoint proteins to stop
the cell
cycle prevent cell division if
DNA is damaged.
Tumor suppressor proteins stop division
of
mutated cells until mistakes in DNA
are
repaired by enzymes.
TS proteins keep most mutations from
being
passed on to daughter cells developing
into
cancer.
If the genes for TS proteins mutate
the brake
on cell division is removed cancers may
result.
Two important TS proteins are the
p53 protein
the RB protein.
17
The p53 Tumor Suppressor Protein
The p53 tumor suppressor protein is
activated
when DNA is damaged. The p53 gene is
called the
guardian angel of the genome
P53 protein
activates
Internal
signalling
genes for
pathway
proteins that
Prevent cell
entering S
phase
Repair DNA
Cause
DNA repair
Apoptosis
apoptosis (if
DNA is
Cell cannot enter S phase
irreparable)
18
Gene Mutations That Cause Cancer
Mutations in 4 types of genes cause Cancer
Proto - oncogenes genes that code for normal
proteins
used in cell division
Growth factors
Membrane Receptors for Growth Factors
Signaling Proteins (e.g. ras proto- oncogene
mutates in 30 of cancers).
Tumor Suppressor genes gene that code for
proteins that
help prevent uncontrolled cell division by
blocking key steps
(e.g. DNA replication).
- Retinoblastoma susceptibilty (RB) gene
- p53 gene mutates in gt50 of cancers.
DNA Repair genes
Genes for Apoptosis
19
How
Carcinogens
Cause
Inactivation
Cancer
of DNA
Repair Genes
Inactivation
Inactivation of
Activation of
of Genes for
Tumor
Oncogenes
Apoptosis
Suppressor
Genes
20
Oncogenes Are Mutated Proto-oncogenes
A cell can acquire a cancer - causing
oncogene
from
A virus
A mutation in a proto-oncogene.
Oncogenes still code for the proteins
needed for
cell division but they cause cancer by
producing
Too much of the protein
An abnormally active protein, e.g.
protein that
activates division by itself
Protein that is made when it is not
needed
Protein that should be made by a
different (i.e.
dividing) cell.
21
Cancer causing Mutations
Proto-oncogenes form active oncogenes by
- being misplaced (e.g. by translocation) to a
site where
the gene is continually expressed resulting in
overproduction of a protein that stimulates cell
division
(e.g. in Chronic Myeloid Leukemia)
- By mutating to a form that is over expressed.
Cancer causing Mutations in Tumor Suppressor
genes
inactivate the genes so normal protein product is
not formed.
Mutated
oncogene
Tumor
Suppressor
gene
neoplasm
22
Over
Growth
Stimulation
Membrane
factor
Hyperactive
Receptor
of Cell
relay protein
Division by
(product of
Normal product
Oncogene
of ras gene
ras oncogene)
Relay
issues signals
proteins
on its own.
Transcription factor
(activated)
DNA
Transcription
Protein that
Translation
stimulates
cell division
23
Tumor-suppressor gene
Mutated tumor-suppressor gene
Cancer
Normal
from
Protein
Mutation
Defective,
prevents
of Tumor
nonfunctioning
cell
protein
Supressor
division
Gene
if DNA is
Protein
damaged
absent
(cell division
Cell division
not blocked)
allowed if DNA
repaired
Mutations accumulate
in cancer cells
24
Multiple Genetic Changes Cause Cancer
Cancers result from a series of genetic changes
in a cell lineage
Inherited (germline) cancers begin with an
inherited cancer
susceptibility mutation in every cell that is
passed on to
offspring.
Inherited cancers may follow a dominant
pattern, e.g. Inherited
Retinoblastoma caused by a mutation in the Rb
tumor
supressor gene increases cancer risk 10,000 x.
However, Inherited cancers need at least one
more somatic
mutation for cancer to develop (2 hit hypothesis
for cancer
causation) .
Sporadic cancers are caused solely by somatic
mutations
occurring in certain body cells so are not passed
on to
offspring.
Accumulation of somatic mutations in a cell
over time
eventually leads to uncontrolled cell division
and cancer.
Therefore sporadic cancers tend to appear much
later in life than
inherited cancers.
25
Accumulation of Mutations Cause
Cancer
inheritance of a germ cell mutation
acts as a risk
factor for cancers by reducing the
number of
somatic mutations required to cause
cancer.
Early mutations show up in all
subsequent stages
of a cancer.
1
Normal
4
3
2
Chromosomes mutation
mutations
mutations
mutations
Normal cell
Malignant cell
26
Oncogenes
Normal genes (regulate cell growth)
1st mutation (leads to accelerated cell division)
1 mutation sufficient for role in cancer
development
27
Tumor Suppressor Genes
Normal genes (prevent cancer)
1st mutation (susceptible carrier)
2nd mutation or loss (leads to cancer)
28
The Two-Hit Hypothesis
First hit
First hit in germline of child
Second hit (tumor)
29
Pathogenesis of Colon Cancer
Colon
Colon Cancer is usually
Sporadic develops in a series
of steps caused by a series of
somatic cell mutations
Loss of
Removal of polyps
tumor-
prevents cancer
suppressor
Colon wall gene (e.g. APC)
Small benign
Normal colon
growth (polyp)
epithelial cells
adenoma
APC adenomatous polypodsis coli
30
Pathogenesis of Colon Cancer -2
Loss of
Activation of
Campbell, Reece
tumor-
ras oncogene
Biology fig. 19.13
suppressor
gene p53
Loss of
Additional
tumor-
mutations
Malignant tumor
suppressor
Larger benign
(adenocarcinoma)
Small benign gene
growth (adenoma)
growth (polyp)
Environmental Risk factors for Colon
Cancer
Low fiber diet
Smoked meats (contain heterocyclic
aromatic amines that are
converted to mutagens in the liver)
Low intake of fruits vegetables
(antioxidants)
Low intake of cruciferous vegetables (a
chemical in brocolli,
brussels sprouts, cabbage activates
enzymes that block
formation of mutagens)
31
Genetic Abnormalities Associated With Hematologic
Malignancies

• A- Point Mutaion
• Mutaions within the RAS oncogenes or P53 tumor-
  suppressor gene are common in many haempoietic
  malignancies. The point mutation may involve
  several base pairs. In 35 of cases of AML the
  nucleophosmin gene shows an insertion of 4 base
  pairs.
• B- Translocation
• Includes two main mechanisms
• 1- Fusion of parts of two genes to generate a
  chimeric fusion gene that codes a novel fusion
  protein. Ex BCR- ABL in t(9 22) in chronic
  myeloid leukaemia.
• 2- Overexpression of a normal cellular gene. Ex
  overexpression of BCL-2 in the t(14 18)
  translocation of follicular lymphoma or MYC gene
  in Burkitt,s lymphoma.

32

• C- Deletions
• May involve a small part of a chromosome,
  the short or long arm or the entire chromosome.
  Losses most commonly affect chromosomes 5, 6, 7,
  11, 20 and Y. The critical event is probably loss
  of a tumor suppressor gene.
• D- Duplication or amplification
• Gains are common in chromosomes 8, 12, 19,
  21 and Y. It is not a common feature in
  haematologic malignancy but has been described
  involving the MLL gene.
• E- Epigenetic alterations
• Means alterations in the mechanism by
  which genes are transcribed and are stably
  inherited with each cell division so they are
  passed on as the malignant cell divides. The most
  important mechanisms are methylation of cytosine
  residues in DNA and enzymatic alterations such as
  acetylation or methylation of the histone protein
  that package DNA.

33
Prostate Cancer
Prostate Cancer is the most common cancer
among men (esp.
gt65 yrs, African - Americans) 2nd in cancer
deaths in men.
Risk factors include increasing age, race,
family history, fat diet,
male hormones over many years.
Adenocarcinoma occurs in periphery of prostate
gland.
Metastasises to lungs, bones (bone pain often
first symptom as
early stage of primary tumor may be asymptomatic).
Tumors are graded from 1 (well differentiated
cells) to 5 (least
differentiated cells, high malignancy).
Manifestations (some similar to Benign
Prostatic Hyperplasia)
- changes in voiding pattern, dysuria, hematuria,
from metastasis low back pain from bone, wt
loss, anemia,
shortness of breath.
Screening (important for early asymptomatic
cancers) includes
- digital rectal exam (palpation of prostate by
DRE detects
nodular lump)
- transrectal ultrasonography (measures prostate
vol more
sensitive than DRE)
- PSA blood test
34
PSA Test for Prostate Cancer
Tumor cells express abnormal
genes so form abnormal
proteins (antigens) so can act as tumor
markers.
Prostate Specific Antigen (PSA)
a glycoprotein
released by prostate gland into the blood
identified as a
marker of prostate cancer in 1980.
PSA is highly specific to prostate
gland but not specific to
prostate cancer. Elevated blood PSA
can also occur with
non cancerous conditions (i.e. false
positives occur with
prostatitis or with benign prostatic
hyperplasia - BPH)
The need to treat stage 1 cancers
detected by PSA test is
controversial stage 1 tumors are
asymptomatic, not detected
by digital exam present in 80 of men
over 80 yrs.
Treatment of Prostate Cancer
includes surgery,
radiation hormonal manipulation (e.g.
androgen inhibitors) if
expected survival gt10yrs, otherwise
watchful waiting is
preferred in elderly patients.
PSA test is used to assess treatment
(correlates with
prostate size cancer stage).
35
Etiology of Breast Cancer
Breast cancer is most common cancer in women
2nd most
common in cancer deaths in women (after lung
cancer).
Risk Factors for Breast Cancer
Prolonged exposures to estrogens (early
menarche late
menopause). Breast cancers that are estrogen
receptor ve are
treated with drugs (e.g. tamoxifen) that bind to
these receptors.
Late Childbearing (having first child after
age 30)
Breasts with a high proportion of lobular
(milk producing)
and ductal tissue density.
Not breast feeding babies increases post
menopausal BC.
Exposure to radiation.
High alcohol consumption.
Family History of BC Genetic Predisposition
in 5-20 of
cases (inheriting mutated breast cancer
susceptibility
genes, BRCA-1 or BRCA-2).
36
Genetics of Breast Cancer
5 - 20 of breast cancers are Familial .
Most involve mutations in 2 Tumor Suppressor
genes
involved in DNA repair so are used as genetic
markers.
Both genes also increase the risk of ovarian
cancer.
- Breast Cancer Susceptibility Gene 1 (BRCA1) on
chromosome 17
- Breast Cancer Susceptibility Gene 2 (BRCA2) on
chromosome 13.
Mutated HER-2/neu Gene (Human Epidermal Growth
Factor Receptor 2 gene is an Oncogene for a
protein that
stimulates cell division occurs in 25- 30 of
Breast
Cancers.
Her-2/neu breast cancers strike early in
adulthood spread
quickly.
Herceptin is a monoclonal antibody based drug
that binds
to Her-2/neu receptors blocks cell division in
tumors.
37
Pathogenesis of Breast Cancer
Tumor (usually
adenocarcinoma
in milk ducts)
Campbell, Reece,
Biology, fig. 12.19
Glandular
tissue
Note lumpectomy a possible treatment
in early stages
A tumor grows from a
Cancer cells invade
single cancer cell.
neighboring tissue.
38
Pathogenesis of Breast Cancer -2
Lymph
vessel
Blood
vessel
Cancer cell
Metastatic tumor
Cancer cells spread via
Small percentage
lymph blood to other
of metastisised cancer
parts of the body.
cells may survive and
Sentinel lymph node
establish a new tumor in
a
biopsy determines if
another part of the body.
cancer has spread if
further lymph node
removal required.
39
Diagnostic methods used to study malignant cells

• 1- Karyotype analysis It is a direct
  morphological analysis of chromosomes from tumor
  cells under the microscope.
• 2- Fluorescent in situ hybridization analysis
• FISH analysis involves the use of
  fluorescent- labelled genetic probes which
  hybridize to specific parts of the genome. This
  can detect extra copies of genetic material or
  reveal chromosomal translocation.
• 3- Southern blot analysis It involves extraction
  of DNA from leukaemic cells followed by
  restriction enzyme digestion, gel electrophoresis
  and transfer by blotting to a suitable membrane.
  The DNA is then hybridized to a probe
  complementary to the gene of interest.
• 4- Polymerase chain reaction Can be performed
  on blood or bone marrow for a number of specific
  translocations such as t(9 22) and t(15 17). It
  is very sensitive and can detect one abnormal
  cell in one million normal cells. It is of great
  value to diagnose minimal residual disease.

40

• 5- DNA microarrayAllows rapid and comprehensive
  analysis of cellular transcription by hybridizing
  labelled cellular mRNA to DNA probes which are
  immobilized on a solid support. This approach can
  rapidly determine mRNA expression from a large
  number of genes and may be used to determine the
  mRNA expression pattern of different leukaemia or
  lymphoma subtypes.
• 6- Flow cytometry Normal cells each have a
  characteristic profile but malignant cells often
  express an aberrant phenotype that can be useful
  in allowing their detection.
• 7- Immunohistochemistry Antibodies can be used
  to stain tissue sections with fluorescent
  markers.
• Value of using these methods
• a- Initial diagnosis.
• b- Establishing treatment protocol.
• c- Monitoring response to therapy.

41
Cytogenetics
Cytogenetics is the original cancer
genetic test
used to identify abnormal mutated
chromosomes by karyotype analysis.
Cytogenetics identified the
Philadelphia
chromosome resulting from a translocation
error in chromosome 22 forming an
oncogene
for chronic myelogenous leukemia
(CML) in
1960.
Karyotype Analysis is done by
culturing tumor
cells, arresting them in metaphase
spreading
chromosomes via use of hypotonic solutions.
Chromosomes are stained and
interpreted by a
cytogeneticist.
Process may take weeks (as many tumors
do not
grow in vitro).
42

• Thank You

Thank You