cell cycle checkpoint and cancer

Cell Cycle Checkpoint and Cell Cycle Checkpoint and CancerCancer

R4 R4 李日翔李日翔 / VS / VS 林璟宏林璟宏

Introduction and Cell CycleIntroduction and Cell Cycle DNA damage and CheckpointDNA damage and Checkpoint Checkpoint and CarcinogenesisCheckpoint and Carcinogenesis Checkpoint and Drug TherapyCheckpoint and Drug Therapy

Introduction and Cell CycleIntroduction and Cell Cycle

Nature Medicine 2004;10:789-799

Oncogene

Tumor Suppressor

Apoptosis

Cancer predisposing gene

Control of cell cycle by cdk-cyclin complexesControl of cell cycle by cdk-cyclin complexes

Molecular cell biology

S / M phase kinase :

cdk + cyclin

Regulation of Cell CycleRegulation of Cell Cycle

Cell 2004;116:221-34

INK4

(RB)

Cell 2004;116:235-46

DNA damage and DNA damage and checkpointcheckpointTheory of checkpointTheory of checkpointATM proteinATM protein

CheckpointCheckpoint

A control loop that makes the initiation of one A control loop that makes the initiation of one event in the cell cycle dependent on the event in the cell cycle dependent on the successful completion of an earlier eventsuccessful completion of an earlier event A cell must replicate every sequence of DNA A cell must replicate every sequence of DNA

once and only onceonce and only once The mass of the cell must The mass of the cell must doubledouble

Gene VIII p845-6

CheckpointCheckpoint Initiation of late events is dependent on Initiation of late events is dependent on

the completion of early eventsthe completion of early events Substrate-product orderSubstrate-product order

If… then…If… then… Complement cascade, apoptosis cascadeComplement cascade, apoptosis cascade Give enough substrate, and yield productGive enough substrate, and yield product

Control mechanismControl mechanism ……unless…unless… Found some chemical / mutant relieve a Found some chemical / mutant relieve a

dependent relationshipdependent relationship Relief of dependenceRelief of dependence

Hartwell and Weinerg, Science 1989;246:629-633

Checkpoint

Molecular biology of the cell 2002

p53 and checkpointp53 and checkpoint


p53 mutant let DNA damaged cell enter S phase but not impact G2 checkpoint

RAD9 system in yeastRAD9 system in yeast

cdc9 (DNA ligase) mutation cdc9 (DNA ligase) mutation DNA replicat DNA replicated but unligated ed but unligated arrest at S phase arrest at S phase

cdc9-rad9 double mutant:cdc9-rad9 double mutant: Enter into mitosisEnter into mitosis Cell die much more rapidlyCell die much more rapidly Loss chromosome spontaneously 21X than wilLoss chromosome spontaneously 21X than wil

d-typed-type

Science 1989;246:629-633


Damage of checkpoint

Stopped by next

checkpoint

ATM ATM (Ataxia Telangiectasia mutated)(Ataxia Telangiectasia mutated)

Heterozygous for AT mildly manifest cancer Heterozygous for AT mildly manifest cancer predisposition and radiation sensitivitypredisposition and radiation sensitivity

Cell of AT patients have a reduced life-span,Cell of AT patients have a reduced life-span, higher requirements for serum growth facto higher requirements for serum growth factor, defect in cytoskeletonr, defect in cytoskeleton

Inhibition of semiconservative DNA synthesiInhibition of semiconservative DNA synthesis and cell cycle progression by ionizing irrads and cell cycle progression by ionizing irradiation is reduced in AT celliation is reduced in AT cell

Science 1995;26:1749-53

ATMATM

AT cell failed to induced p53 in response AT cell failed to induced p53 in response to irradiationto irradiation ATM act upstream to p53ATM act upstream to p53

Identify ATM gene from patient with ATIdentify ATM gene from patient with AT Similarity between ATM and rad3Similarity between ATM and rad3

rad3 protein is required for the G2-M rad3 protein is required for the G2-M checkpoint, it monitor the completion of DNA checkpoint, it monitor the completion of DNA damage repairdamage repair

Similarity between ATM and PI3-kinaseSimilarity between ATM and PI3-kinaseCell 1992;71:587-95

Science 1995;26:1749-53

TIBS 1995;20:426-30

mTOR, p110…

TIBS 1995;20:426-30

p53p53

Nature 2004;432:316-23

Trends in Cell Biology 2002;12:509-16

Cancer cell 2003;3:421-429

ATM

Checkpoint in S. cerevisiaeCheckpoint in S. cerevisiae

Science 2002;297:552-7

DNA damage

Hus1/Rad1/Rad9

Chk1 BRCA1

DNA repairCell cyclearrest

Others

ATM

ATRRad17/RFC

Apoptosis

or

Human DNA damage response pathway

Replication block

p53

Sensors

Effectors

Signals

Transducers

Common intermediate

Nature 2000;408:433-9

Checkpoint and Checkpoint and CarcinogenesisCarcinogenesis

Checkpoint and carcinogenesisCheckpoint and carcinogenesis

Nature 2004;432:316-23

Adenoma-Carcinoma Sequence

Why the patient so unlucky?Why the patient so unlucky? Dose each oncogene mutate randomly anDose each oncogene mutate randomly an

d independently?d independently? Dose tumor progress steadily?Dose tumor progress steadily?

Science 2002;297:565-9

Oncogene induced a ‘oncogenic stress’ (cyclin E), which activate DNA repair response (ATM, pChk2)

Nature 2005;434:864-70

U-2-OS cell

Cyclin E expressed in a Tetracyclin-repressible manner

Mcm7: loading control

Nature 2005;434:864-70

Absent

Low level

Medium level

High level

pT-Chk2 level in bladder cancer

Checkpoint damage in cancerCheckpoint damage in cancer(bladder cancer)(bladder cancer)

Nature 2005;434:864-70

Genome instability during cancer progressionGenome instability during cancer progression

Nature 2005;434:907-13

Nature 2005;434:907-13

TUNEL stain: hallmark of apoptosis

SummarySummary

Oncogene mutationOncogene mutation hyperplasia hyperplasia DNA replication stress DNA replication stress Double strand brea Double strand brea

kk Activate checkpoint Activate checkpoint cell cycle arrest, DNA r cell cycle arrest, DNA r

epair, or apoptosis epair, or apoptosis no tumor progress no tumor progress Defect of checkpoint (ATM-Chk2-p53)Defect of checkpoint (ATM-Chk2-p53) Genomic instability and cancerGenomic instability and cancer

Nature 2005;434:864-70

Nature 2005;434:907-13

A model for carcinogenesisA model for carcinogenesis

Nature 2005;434:907-13

Science 2008;319:1352-5

Science 2008;319:1352-5

P.I.: proliferation index

DDR.I.: DNA damage reponse index

A./S.I.: apoptosis or senescence index

Checkpoint and Drug Checkpoint and Drug TherapyTherapy

Chemotherapy with intact checkpointChemotherapy with intact checkpointChemotherapy with defective checkpointChemotherapy with defective checkpointTargeting checkpointTargeting checkpoint

Mechanism of alkylating agentMechanism of alkylating agent

Cancer Principles and proctice of Oncology, 7ed p339

Therapy with intact checkpointTherapy with intact checkpoint

DNA damage activate DNA repair and DNA damage activate DNA repair and checkpoint pathwaycheckpoint pathway

ATM->Chk1/2->p53->p21ATM->Chk1/2->p53->p21 Cell cycle arrest or death?Cell cycle arrest or death? Therapy shift arrest to deathTherapy shift arrest to death

Arrest after SN-38Arrest after SN-38

(CPT-11)

JCO 2005;23:4499-4508

FlavopiridolFlavopiridol

CDK inhibitor Inhibit bcl-2, XIAP, p21,

mcl-1, cyclin D1,30 and phospho-survivin

Inhibit p21-induced cell cInhibit p21-induced cell cycle arrest at G2 checkpycle arrest at G2 checkpointoint

JCO 2005;23:4499-4508

JCO 2005;23:4499-4508

Therapy with defective checkpointTherapy with defective checkpoint Response to Flavopiridol according to pResponse to Flavopiridol according to p

53 status53 status Resistance to anti-CD33 in AML cell unResistance to anti-CD33 in AML cell un

able to activate Chk1/Chk2 able to activate Chk1/Chk2

JCO 2005;23:4499-4508

Blood 2003;101:4589-97

Targeting checkpointTargeting checkpoint

Loss of ATM sensitized p53-deficient cell to Loss of ATM sensitized p53-deficient cell to topoisomerase poisons and anti-metabolitetopoisomerase poisons and anti-metabolitess

SN-38 followed by UCN-01 (inhibit Chk1 /CSN-38 followed by UCN-01 (inhibit Chk1 /Chk2) enhance apoptosis in cell with mutated hk2) enhance apoptosis in cell with mutated p53 but not intact p53 p53 but not intact p53

Under development: CHIR124, inhibitor of Under development: CHIR124, inhibitor of Chk1, and 17-AAG: suppresses Chk1expreChk1, and 17-AAG: suppresses Chk1expressionssion JCO 2005;23:4499-4508

Annals of Oncology 2003;14:938-45

Flavopiridol inhibit cell cycle arrestFlavopiridol inhibit cell cycle arrestApoptosis

Flavopiridol


Chk1/p21

UCN-01

A model for targeting checkpointA model for targeting checkpoint

JCO 2005;23:4499-4508

So Simple?

Down stream p53 inhibit G1/S checkpoint

Up stream p53 inhibit G2 checkpoint

To be definedTo be defined

What determines DNA repair, What determines DNA repair, cell cycle arrest, or cell cycle arrest, or apoptosis?apoptosis?

Some drug induced cell cycle Some drug induced cell cycle arrest, some apoptosis, and arrest, some apoptosis, and some necrosis?some necrosis?

Different cell line had different Different cell line had different response?response?

Undefined pathway? Undefined pathway? ∆∆p53?p53?

Cancer cell 2003;3:421-429

?Cell 2005;122:21-32

Thanks for your attentionThanks for your attention

Mitotic checkpointMitotic checkpoint

Nature review cancer 2005;5:773-85

Mitotic checkpoint defect in cancerMitotic checkpoint defect in cancer

Nature review cancer 2005;5:773-85

∆∆p53 in checkpointp53 in checkpoint

Cell 2005;122:21-32

cell cycle checkpoint and cancer

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