Harliansyah .Ph.D Kuliah Blok Neoplasia 2013

7/27/2019 Harliansyah .Ph.D Kuliah Blok Neoplasia 2013

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Oncogene andTumor Supressor Gene

Harliansyah, Ph.D

YARSI UniversityDepartment of Biochemistry

harliansyah.hanif @ yarsi.ac.id

March, 2013


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Cancer Pathogenesis

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Cellular Basis of Cancer

Cancer is a collection of diseases

characterized by abnormal anduncontrolled growth

Cancer arises from a loss of normalgrowth control

In normal tissues, the rates of newcell growth and old cell death arekept in balance

In cancer, this balance is disrupted

This disruption can result from

1) uncontrolled cell growth or

2) loss of a cell's ability to undergoapoptosis


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Cancer Cell Do Not Grow Faster Than

Normal Cells

Rather, Their Growth is Just

Uncontrolled


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1 fertilized egg

50x1012

Proliferation Differentiation Death

1016 cell divisions/lifetime


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Proliferation Differentiation Death

Transit

Proliferating

Exiting

Renewing

Cellular equilibrium


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10Proliferation Differentiation Death

Cancer: disruption of

cellular equilibrium


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Post mitoticStem cell

Differentiated Normalsenescent

differentiated

cell

Benign

tumor

Grade 2malignancy

Grade 3 or 4

malignancy

Stem cells as the target of carcinogens


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Invasion and Metastasis

Abnormal cells proliferateand spread (metastasize) toother parts of the body

Invasion - directmigration andpenetration intoneighboring tissues

Metastasis - cancer cellspenetrate into lymphatic

system and blood vessels


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Benign tumors

generally do not

spread by

invasion or

metastasis Malignant

tumors are

capable of

spreading by

invasion and

metastasis

Malignant versus Benign Tumors


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What causes Cancer? Cancer is caused by

alterations or mutations in

the genetic code

Can be induced in somatic

cells by: Carcinogenic

chemicals

Radiation Some viruses

Heredity - 5%


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Hanahan and Weinberg, Cell 100: 57, 2000

Apoptosis

Oncogenes

Tumor Suppressor

Inv. and MetsAngiogenesis

Cell cycle


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What is the molecular basis of cancer?

Cancer is a genetic disease.

Mutations in genes result in alteredproteins

During cell division

External agentsRandom event

Most cancers result from mutations insomatic cells

Some cancers are caused by mutations ingermline cells


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Theories o f cancer genesis

Standard Dogma

Proto-oncogenes (Ras melanoma)

Tumor suppressor genes (p53 variouscancers)

Modified Dogma

Mutation in a DNA repair gene leads to theaccumulation of unrepaired mutations(xeroderma pigmentosum)

Early-Instability Theory

Master genes required for adequate cellreproduction are disabled, resulting inaneuploidy (Philadelphia chromosome)


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CANCER AND GENETICS

Cancer: genome disease

Causes of genomic changes

Effects of genomic changes

Revolution in cancer treatment: Smart Bullets Period


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CANCER: GENOME DISEASE

Loss of DNA

Gain of DNA

Changes in nucleotides

Epigenetic effects


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Signs for Genom ic Changes in

Cancer

Changes in chromosome numbers

- Aneuploidy

Chromosomal changes- Increase in DNA copy number-15 different

region

- Loss in chromosomal -200.000 regions

Micro changes- Microsatellite changes Mikrosatellite - 100.000

- Nucleotide changes


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Chromosomal changes in the genome of cancer

cells: tip of the iceberg

Terminal

Deletion

http://www.tokyo-med.ac.jp/genet/cai-e.htm

Ring

Chromosome

Robertsonian

Translocation

DeletionReciprocal

translocation

IsochromosomesInsertion Inversion

Duplication


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Nucleotide changes in the genome of cancer

cells: unseen site of the iceberg

NucleotideDeletions

NucleotideInsertions

NucleotideSubstitutions

http://www.tokyo-med.ac.jp/genet/cai-e.htm


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DNA Loss in cancer cells


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Early Brain Tumor

(Astrocytoma Stage II)

Advance Brain Tumor

Glioblastoma Multiform (Stage IV)

DNA Loss in cancer cells: beyond coincidence ...


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p53

locus

Chromosomal loss:

Mostly, it is a sign

for the loss of atumor suppressor

gene

PTEN

locus

CDKN2

locus

RB1

locus

???

locus


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Cancer: Genome Disease

Epigenetic effects


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Genetic and Epigenetic Silencing of Tumor Suppressor Genes

Plass - 2002
http://hmg.oupjournals.org/content/vol11/issue20/images/large/ddf21901.jpeg


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Carcinogenic

chemicals

UV

Replication Errors

Radiation

Viruses

Rearrangements

(translocation, deletions,

amplifications)

Point mutations

Alters DNA of genes controlling cell proliferation.

(Proliferation becomes abnormal)

Cancer cell

Normal cellDamaged DNA

THE CAUSES OF GENOMIC CHANGES IN CANCER


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Hasar

Etken Tr

Hasar

EtkeniKanser Riski areti

FizikselMortesi Inlar Deri Ka., Melanoma P53 (CC-TT)

Radyasyon Tiroid Ka., Lsemi Translokasyon

Kimyasal

Benzopren Akcier Ka. p53 (G-T)

Aflatoksin Karacier Ka. p53 (249 G-T)

Oksidatif Stres Yallk Kanserleri P53 (C-T)

Biyolojik HBV Karacier Ka.Virus DNA

ntegrasyonu

THE CAUSES OF GENOMIC CHANGES IN CANCER:

Somatic Changes


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Genes Disease Function Inheretance Cancer Risk

FA Genes F-A DNA Damage respose ? OR Lsemi

XP GenesX-P

NER Type

DNA RepairOR Skin Ca.

BLM Bloom DNA Helicase ? OR Various cancers

WRN Werner DNA Helicase ? OR Sarcoma

RECQ4 Rothmund-Thomson

DNA Helicase OR Sarcoma

MLH1, MSH2,

PMS1, PMS2

MMR

DNA Repair

OD Colon,

Endometrium Ca.

OR Lsemi, NF1

BRCA1, BRCA2 DNA Repair

OD Breast, Ovary,

Prostate, Pancreas

Ca

ATMA-T

DNA Damage sense ?

OR Lymphoma,

Leukemia

OD Breast Ca. ?

p53 Li-Fraumeni DNA Damage sense OD Various cancers

THE CAUSES OF GENOMIC CHANGES IN CANCER:

Hereditary Predisposition


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Approximately 90-95% of all cancers

are sporadic.

5-10% are inherited.

CANCER AND

GENETICS


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Oncogenes Tumor suppressor genes

DNA repair genes

GENES PLAYING ROLE IN

CANCER DEVELOPMENT


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What are the genes responsible for tumorigenic

cel l growth?

Normal

Cancer

Proto-oncogenes Cell growth

and

proliferationTumor suppressor genes

+

-

Mutated or activated

oncogenes Malignant

transformationLoss or mutation of

Tumor suppressor genes

++


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ONCOGENES

Oncogenes are mutated forms of

cellular proto-oncogenes.

Proto-oncogenes code for cellular

proteins which regulate normal cell

growth and differentiation.


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Class I: Growth Factors

Class II: Receptors for Growth Factors and Hormones

Class III: Intracellular Signal Transducers

Class IV: Nuclear Transcription Factors

Class V: Cell-Cycle Control Proteins

Five types of proteins encoded by proto-oncogenes participate in control of cell growth:

F ti f C ll l P t O


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4. Nuclear

Proteins:

Transcription

Factors

5. Cell Growth

Genes

3. CytoplasmicSignal Transduction

Proteins

1. Secreted Growth Factors

2. Growth Factor Receptors

Functions of Cellular Proto-Oncogenes


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A generic signalling

pathway


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Oncogenes

proto-oncogene = ras

Oncogene = mutated ras

Always activated

Always stimulating

proliferation


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amino acid posi t ion

Ras gene 12 59 61 Tumor

c-ras (H, K, N) Gly Ala Gln normal cells

H-ras Gly Ala Leu lung carcinoma

Val Ala Gln bladder carcinoma

K-ras Cys Ala Gln lung carcinoma

Arg Ala Gln lung carcinoma

Val Ala Gln colon carcinoma

N-ras Gly Ala Lys neuroblastoma

Gly Ala Arg lung carcinoma

Murine sarcoma virus

H-ras Arg Thr Gln Harvey strain

K-ras Ser Thr Gln Kirsten strain

Am ino acid subst i tu t ions in Ras fam i ly p roteins

(inac tivates GTPase)


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Act ivat ion mechanisms of proto-oncogenes

proto-oncogene --> oncogene


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CHROMOSOMAL REARRANGEMENTS OR TRANSLOCATIONS

Neoplasm Translocation Proto-oncogene

Burkitt lymphoma t(8;14) 80% of cases c-myc1

t(8;22) 15% of cases

t(2;8) 5% of cases

Chronic myelogenous t(9;22) 90-95% of cases bcr-abl2

leukemia

Acute lymphocytic t(9;22) 10-15% of cases bcr-abl2

Leukemia

1c-myc is translocated to the IgG locus, which results in its activated expression

2bcr-abl fusion protein is produced, which results in a constitutively active abl kinase


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GENE AMPL IFICATION

Oncogene Amplification Source of tumor

c-myc ~20-fold leukemia and lung carcinoma

N-myc 5-1,000-fold neuroblastoma

retinoblastoma

L-myc 10-20-fold small-cell lung cancer

c-abl ~5-fold chronic myoloid leukemia

c-myb 5-10-fold acute myeloid leukemia

colon carcinoma

c-erbB ~30-fold epidermoid carcinoma

K-ras 4-20-fold colon carcinoma

30-60-fold adrenocortical carcinoma


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Oncogenes are usually dominant

(gain of function)

cellular proto-oncogenes that have been mutated

(and activated)

cellular proto-oncogenes that have been captured byretroviruses and have been mutated in the process

(and activated)

virus-specific genes that behave like cellular proto-

oncogenes that have been mutated to oncogenes (i.e.,

activated)


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The resu lt:

Overproduction of growth factors

Flooding of the cell with replication

signals Uncontrolled stimulation in the

intermediary pathways

Cell growth by elevated levels of

transcription factors


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Tumor suppressor genes

Normal function - inhibit cell proliferation

Absence/inactivation of inhibitor --> cancer

Both gene copies must be defective

KNUDSON TWO HIT HYPOTHESIS IN FAMIL IAL CASES


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KNUDSON TWO HIT HYPOTHESIS IN FAMIL IAL CASES

RB rb

rb rbRB

Familial RB (%30)

Tumor cells Normal cells

Normal cells

Inactivation of a tumor suppressor

gene requires two mutations, inherited

mutation and somatic mutation.

RBLOH

KNUDSON TWO HIT HYPOTHESIS IN SPORADI C CASES


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RB RB

RB

LOH

RB

Mutation

NormalCells

Tumor cells

KNUDSON TWO HIT HYPOTHESIS IN SPORADI C CASES

RB RB

Inactivation of a tumorsuppressor gene

requires two somatic

mutations.


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TUMOR SUPPRESSOR GENES

Disorders in w hich gene is af fected

Gene (locu s) Func t ion Famil ial Sporad ic

DCC (18q) cell surface unknown colorectal

interactions cancer

WT1 (11p) transcription Wilms tumor lung cancer

Rb1 (13q) transcription retinoblastoma small-cell lung

carcinoma

p53 (17p) transcription Li-Fraumeni breast, colon,syndrome & lung cancer

BRCA1(17q) transcriptional breast cancer breast/ovarian

tumors

BRCA2 (13q) regulator/DNA repair


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CELL CYCLE

Daugther cell

Mitosis

DNA replication

Control Point

Gateway

GrowthFactors

Cell cycleinhibitors

CELL CYCLES

Rb gene


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Rb gene Rb protein controls cell cycle moving past G1 checkpoint

Rb protein binds regulatory transcription factor E2F

E2F required for synthesis of replication enzymes

E2F - Rb bound = no transcription/replication

Growth factor --> Ras pathway

--> G1Cdk-cyclin synthesized

Active G1 Cdk-cyclin kinase phosphorylates Rb

Phosphorylated Rb cannot bind E2F --> S phase Disruption/deletion ofRb gene

Inactivation of Rb protein

--> uncontrolled cell proliferation --> cancer

53


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p53 Phosphyorylated p53 activates transcription ofp21 gene

p21 Cdk inhibitor (binds Cdk-cyclin complex --> inhibits kinase

activity) Cell cycle arrested to allow

DNA to be repaired

If damage cannot be repaired

--> cell death (apoptosis)

Disruption/deletion ofp53 gene

Inactivation of p53 protein

--> uncorrected DNA damage

--> uncontrolled cell proliferation --> cancer


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These are genes that ensure each strand of geneticinformation is accurately copied during cell division of the

cell cycle.

Mutations in DNA repair genes lead to an increase in thefrequency of mutations in other genes, such as proto-

oncogenes and tumor suppressor genes.

i.e. Breast cancer susceptibility genes (BRCA1 and BRCA2)

Hereditary non-polyposis colon cancer susceptibility genes(MSH2, MLH1, PMS1, PMS2) have DNA repair functions.

Their mutation will cause tumorigenesis.

DNA REPAIR GENES


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Molecular

mechanisms of

DNA double

strand break

repair

BRCA1/2

IMPORTANCE OF DNA REPAIR


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IMPORTANCE OF DNA REPAIR


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Multiple mutations lead to colon cancer

Genetic changes --> tumor changes

Cellular

Tumor Progress ion


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Revolution in cancer treatment:

Smart Bullets Period

Summary of 30 years of research (1971-2001)


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Hanahan & Weinberg 2000

y y ( )


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Bilimsel Aratrmalarn

Kanserle Savaa KatksRCEPTN

HERCEPTIN

STI-571


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Translocation and Bcr-Abl fusion in CML


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STI-571 against Bcr-Abl


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Smart bul let STI -571 lockes itself to the target molecule

STI-571


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Thousands of Targets

HERCEPTIN

STI-571

?

??

??

?

?

?

??

?

?

?

?

?

??

?

MOLECULAR BIOLOGY & INFORMATICS


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MOLECULARBIOLOGY & INFORMATICS

Biyoinformatik

~30.000 genes

~300.000 protein

~3.000.000 interaction

1 human cell

~3.000.000.000 bp

DNA


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Harliansyah .Ph.D Kuliah Blok Neoplasia 2013

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