Download Neoplasia 6

Neoplasia 6
Dr Heym Awad
FRCPath
Cyclins and cyclin dependent kinases
Normal cell cycle
• All the stimuli mentioned in the previous lecture aim for quiescent
cells to enter the cell cycle
• Replication of cells is stimulated by growth factors or signaling from
ECM components through integrins
• Each phase in the cell cycle depends on successful completion of the
previous one
• Cycle stops when essential gene function is lost
• G1 to S transition is critical because if this checkpoint is passed the
cells are committed for DNA replication
• G1 to S is called the restriction point
• Progression through cell cycle, especially G1-S is regulated by proteins
called cyclins
• Cyclins activate kinases CDK (cyclin dependent kinases)
• Cyclin and CDK form complexes that phosphorylate target proteins
that drive the cell through the cell cycle.
• Cyclins : D, E, A , B (they appear in this sequence
The DEAB sequence of cyclins !
• SO: cyclin/CDK complexes cause proliferation.
• Theses are inhibited by cyclin dependent kinase inhibitors (CDKI)
• CDKI important for enforcing the checkpoints and delaying cell cycle.
• Checkpoints important to discover DNA damage and prevent cells
with DNA damage from continuing cell cycle
• G1-S checkpoint monitors integrity of chromosomes before
replicating
• G2-M checks DNA integrity after replication to decide if cells can
safely go into mitosis
• When there is DNA damage the checkpoints are activated.. They
delay the cell cycle and trigger DNA repair
• If damage is severe: apoptosis or senescence are stimulated.
CDKI
• Some inhibit CDK broadly p21, p27, p57
• Others are specific p15,p16,p18,p19
• (you don’t need to remember which is which!!)
• Mutations causing increased cyclins or CDK cause self sufficiency in
growth signals.
• Mutations inhibiting CDKI will cause increased growth.
• Examples: cyclin D is activated in several tumors mainly lymphomas.
Second hallmark of cancer
• 2. insensitivity to growth inhibitors
• Growth inhibition is achieved by tumor suppressor genes
• Loss or decreased functions of tumor suppressor genes is essential to
cause cancer
• RB gene= retinoblastoma gene = governor of cell cycle
retinoblastoma
• Rare childhood tumor
• 60 % of cases are sporadic, 40% familial
• Predisposition to develop the tumor is inherited as autosomal
dominant trait
• To develop retinoblastoma: two hit hypothesis
retinoblastoma
Two hit hypothesis
• Two mutations (hits) required to develop retinoblastoma
• The 2 mutations involve the RB gene on chromosome 13 (13q14)locus
• Both copies of RB gene need to be deactivated to develop
retinoblastoma
• In familial cases, one hit is inherited (germ line mutation)
the other is acquired
-In sporadic cases, both mutations are acquired
• In familial cases , one single somatic mutation is needed .. So
dominant pattern of inheritance.
• NOTE: retinoblastoma disease follows autosomal dominant
inheritance. Because the susceptibility of the disease is what is
inherited. BUT, RB gene is a recessive gene.
Two hit hypothesis
Two hit hypothesis
• People with inherited RB have increased risk of other cancers.. Mainly
osteosarcomas and soft tissue sarcomas
• Homozygous loss (both copies lost) of RB gene can be seen in many
cancers like breast, bladder…
• A cell heterozygous in RB locus is not neoplastic ( one normal and
one abnormal allele)
• The two hits are essential for neoplastic transformation
RB gene
• RB gene product is a DNA binding protein expressed in all cells
• This protein has two forms: active hypophsphorylated state and
inactive hyperphosphorylated
• Bb regulates G1/S checkpoint
• S phase requires cyclin E/CDK2 complex
• Expression of cyclin E needs E2F family of transcription factors
• Early in G1 , RB is in its active hypophosphorylated form, it binds and
inhibits E2F .. So no cyclin E2
• This is done by two ways: hyposphorylated RB sequesters E2f so it
doesn’t interact with transcription factors also RB recruits chromatin
remodeling proteins that bind to promoters of E2F, so DNA becomes
insensitive to transcription factors
• Mitogenic signals ..lead to cyclin d expression and activate cyclin
D/CDK 4/6 complexes.. These complexes phosphorylate RB..
• Phosphorylated RB is inactivate … this leads to release of E2F
• E2F now free and can cause transcription of cyclin E..
• Cyclin E stimulates DNA replication
• So cells enter S phase
• Once in s phase cells are committed to division.. They don’t need
additional growth signals
• In M phase phosphate removed from RB ,so it goes back to its
inactive state .
How RB works
• E2F is not the only transcription factor targeted by RB
• Rb stimulates myocyte, adipocyte and other cell specific transcription
fatcors… so important for G0-G1 with differentiation
• Rb is important for tumerogenesis but it is not mutated in all cancers
• If it is not mutated, other gene mutations mimicking RB mutation
must play a role
• Mutations in genes affecting RB phosphorylation: Mutatiomnal
activation of cdk4 or overexpression of cyclin D favour cell
proliferation by inactivating RB
NOTE
• Loss of nomal cell cycle control is found in all tumors through
mutations of RB, cyclin D, CDK4 or CDKN2A (which is a CDKI)
• Some virus like HPV have protein E7 which binds to the
hypophsphorylated RB so prevents it from inhibiting E2F
• So RB is functionally deleted
TP53 gene: the guardian of the genome
• Tp53 is one of the most commonly mutated genes in cancer
• It encodes p53 protein
P53 causes growth inhibition by three mechanisms
1. Temporary cell cycle arrest: quiescence
2. permenant cell cycle arrest: senescence
3. triggers apoptosis
Check this pic in the book…
• P53 monitors internal stress whereas RB senses external signals
• P53 is triggered by several stresses: anoxia, inappropriate oncogene
activity (MYC or RAS) or DNA damage.
• In non-stressed healthy cell, p53 is short lived: 20 minutes because it
binds MDM2 which is a protein that targets it for destruction
• When cells are stressed ..sensors that include protein kinases are
activated (ATM is one of these kinases)
• These activated kinases catalyze post translational modifications of
p53 and release it from MDM2
• Now p53 has longer life span and can drive transcription of certain
genes.. hundreds of them
Genes transcribed by p53
• Suppress growth by three mechanisms
• 1. mediating cell cycle arrest. This occurs late in G1. caused by p53
dependent transcription of CDK1 gene= p21= CDKNIA
• P21 protein inhibit cyclin/CDK complexes and prevent
phosphorylation of RB
• So cell is arrested in G1
• Pause to repair any DNA damage
• P53 also induces expression of DNA damage repair genes
• If DNA is repaired successfully ,p53 upregulates transciption of
MDM2.. Destruction of p53.. Removal of the block on cell cycle.
• If DNA not repaired p53 makes cells enter apoptosis or senescence
Senescence by p53
• Senescence needs activation f p53 and or RB and expression of their
mediators like CDKI
• Mechanisms of senescence unclear but seem to involve global
chromatin change, which permanent change gene expression
P53 induced apoptosis
• Induced by pro-apoptotic genes including BAX and PUMA
• P53 also represses proliferative and anti-apoptotic genes (bcl2)
• ? P53 is a transcriptional activator so how could it repress certain
gene expression
• Answer by miRNAs
• More than 70% of human cancers have mutated TP53
• Both copies of the gene need to be lost for cancer to develop
• Mostly somatic
• Rare li Fraumini syndrome: inherit defect in one allele.. More
predisposition to cancer (Sarcoma, breast carcinoma , leukemia and
brain tumor)
• P53 can become nonfunctional by some DNA viruses
• HPV, Hep B, EBV.. Proteins can bind to p53 and deactivate it
• Note p53 activated by phosphorylation