2.1 molecular biology cancer

8/8/2019 2.1 Molecular Biology Cancer

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Genetic Basis of Cancer: the Origins

Circa 1860: microscopist Mueller discovers thattumor is made of cells

1908: Ellerman and Bang demonstrate that afilterable agent induces leukemia in chicken

1911: Rous demonstrates that a filterable agentinduces sarcomas in chicken

1936: Bittner that mammary cancer can betransmitted through milk in a specific mouse strain

1951: Gross discovered the first murine leukemiavirus


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The SV40 oncogenic

polyomavirus


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Retrovirus vital cycle


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The largest part

of retroviral

oncogenes aretyrosin kinase

proteins (src, fps,

yes, fgr, ros, abl,

erbB, kit, sea)

Other oncogenes

functions as

transcription

factors (myc,

myb, fos, jun,

erbA)


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HRAS was the first proto-oncogene found activated by point mutations in

human tumor cells: about 20% of all human tumors carry a mutated RAS

Its oncogenic function was demonstrated by the finding that the mutant RAS

can transform NIH3T3

Activated RAS cooperates with other oncogenes in tranforming primary

human cells.

Its activity is essential to maintain the transformed phenotype

Point Mutation Activated-RAS

Oncoprotein is Required to Maintain the

Malignant Phenotype


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Il cromosoma Ph nella

Leucemia Mieloide Cronica:


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La scoperta del cromosoma Ph nella Leucemia

Mieloide Cronica: pi del 95% dei casi presenta

questa aberrazione cromosomica

Peter Nowell, Universit di Pennsylvania, Philadelphia


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Il cromosoma Ph : una traslocazione

reciproca tra il cromosoma 9 ed il 22

Janet Rowley, Universit di Chicago


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Scoperta del difetto genetico:

la traslocazione t(9;22) causa la fusione dei

geni BCR-ABLEli Canaani, Israel

Carlo M. Croce, Italia-USA

Il proto-oncogene ABL, una tirosin-chinasi,

diviene attivata dalla fusione con BCR: la sua

attivita` chinasica diviene costitutivamente

attiva a livelli elevati


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Translocation

t(8;14)(q24;q32) in

Burkitts lymphoma


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MYC is activated by juxtaposition to the Ig locus


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Nonrandom chromosomal aberrations

in myeloid malignancies


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Multiple translocation partners for

MLL and TEL


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Nonrandom chromosomal aberrations

in lymphoid malignancies


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DNA amplification leads to

oncogene over-expression


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Proto-oncogenes can be activated by a

variety of mechanisms

Point mutations, leading to aberrant biochemical function

Fusion with other genes, leading to aberrant biochemicalfunction

Abnormally elevated level due to promoter/enhancer activation

or to gene amplification


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Il fenotipo tumorigenico

recessivo a livello cellulare

FUSIONE

Cellula

Tumorale

Cellula

Non Tumorale

Ibrido non tumorigenico


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TUMOR SUPPRESSOR GENES AND CANCER

X

X

X XMutation 1

Mutation 2

Heterozygous Loss of

Heterozygosity

Normal

Other: Gene Methylation

Expression Levels


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Knudsons Two-Hits Hypothesis


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Mechanisms for RB1 tumor suppressor

gene inactivation


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DNA polymorphisms and loss of

heterozygosity in cancer


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In mammalian cells, cytosines present inthe dinucleotide CpG are generally

methylated

CpG islands are regions 500-2000

nucleotides long enriched in CpG

dinucleotides that are found in about 60%of gene promoters.

CpG islands are usually not methylated

DNA Methylation in

Mammalian CellsCpG methylation is the prototype of a mammalian epigenetic mark


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DNA methylation of CpG island

promoters and gene regulation


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DNA methylation in development


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DNA methylation at replication

maintains a stable epigenotype

DNA

replication

DNA

Methylation

(DNMT1)

Inheritance of cytosine methylation:

1. the symmetrical nature of the

modified sequences (CpG)

2. The preference of the

maintenance DNMT1 for

hemimethylated DNA


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Methylation

ofCpG Islands

in Gene PromotersRepresses

Gene Expression


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Tumor suppressor gene inactivation

by promoter methylation


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LOH Mutation Methylation

Homozygous

deletion

Mutation

+ LOH

Methylation

+ LOH

Mutation +

Methylation

Biallelic

Methylation

RB1 Retinoblastoma yes yes yes rare yes yes yes ?

VHL Renal Ca yes yes yes rare yes yes yes ?

BRCA1 Breast/Ovarian Ca yes yes yes no yes ? ? yes

CDKN2A Melanoma yes yes yes yes yes yes yes yes

MLH1 Colorectal Ca yes yes yes ? yes ? yes yes

APC Colorectal Ca yes yes yes ? yes ? ? ?

MGMT Lung ca yes no yes no no ? no yes

DAP Lung ca yes no yes no no ? no yes

Double Hit MechanismsMechanisms of Inactivation

Examples of tumor suppressor genes

inactivated by promoter methylation


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Tumor suppressor genes are

inactivated by a variety of mechanisms

Point mutations, leading to aberrant biochemical function

Deletion leading to loss of gene

Aberrant DNA methylation leading to loss of transcriptional

activity


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Moss (2002) Current Biology 12, R138

microRNAs are expressed as longer precursor RNAs


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RISCRepressione TraduzioneRepressione Traduzione

Alcuni miRNAs

Pre-miRNAPre-miRNA

DroshaDrosha

miRNA genemiRNA gene

Pri-miRNAPri-miRNA

DicerDicer

DuplexmiRNAmiRNA

Exportin-5Exportin-5

Helicase

Regione

codificante

RISC RISC

3 non codificante

Taglio mRNATaglio mRNA

RISC

RNA bicatenarioRNA bicatenario

siRNAsiRNA

siRNA e miRNA, per la loro

struttura, biogenesi e meccanismo

di funzionamento potrebbero

rappresentare variazioni dellostesso processo evolutivo

Da trascrizione elementi ripetitivi e trasposoniDa trascrizione elementi ripetitivi e trasposoni,


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mir-15 and mir-16 are within the minimal

region of 13q deletion in human CLL

k


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BM

PerBlLeu

k

CD5+

Kidney

Prostate

Liver

Pancreas

Skmuscle

Testicle

miR16

miR15

70bp

20bp

20bp

70bp

CLL

13qLOH

1

+/+

2

ND

3

+/-

4

+/-

5

+/-

6

ND

7

+/-

8

NI

9

+/+

10

+/+

11

NI

12

+/-

13

NI

14

NI

15

+/-

16

+/-

17

+/-

18

NI

miR16

miR15

70bp

20bp

20bp

70bp

CD5+

Expression of

mir-15 and mir-16in normal tissues

and human CLLs


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BCL2 is target of mir-15/16


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BCL2 is target of mir-15/16

Saggi luciferasici per la conferma di


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Saggi luciferasici per la conferma di

bersagli genici di miRNA deregolati in

cancro

Luciferasi

3UTR gene bersaglio

microRNA

EFFETTO: RIDUZIONE ATTIVITA LUCIFERASICA

Effetto di mir-15a / mir-16-1 su BCL2


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Let-7 down-regulates human RAS

Johnson et al. Cell 120: 635, 2005


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Let-7 down-regulate human RAS



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Let-7 is down-regulated in human cancer,

resulting in up-regulation of RAS protein



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Let-7 is down-regulated in human cancer,

resultin in up-regulation of RAS protein


Main mechanism of miRNA involvement in cancer is aberrant expression


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miRprom

ote

r

miRgene

miRtranscr

ipt

Specific effects

Apoptosis

Proliferation

Invasion

Angiogenesis

Target mRNA

overexpressionoverexpression

Target mRNA

downregulation

Delet

ion+Pr

omote

rHo

mozy

gous

deletio

n

miR

15/16,m

iR-26a?

Deleti

on+Mu

tation

Translo

cation

miR

Amplification(miR155)

miR 142

hyper

methy

lation

miR1

5/16?

Othergene

p

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