ARF IN CANCER

1
ARF IN CANCER
Cloned in 1995 550 papers 55,000 p53 papers
2
IN THE BEGINNING
Virologists using viruses to cause cancer
1st place Varmus Bishop Nobel Prize 2nd
place Sherr
src fms CSF-1R
macrophage proliferation
3
DISCOVERING THE INK4A/ARF LOCUS LUCK
CSF-1R (sherr)
PATENT
MYRIAD PHARMACEUTICALS
CYCLIN D (sherr)
p16INK4a (beach sherr)
CYCLIN D-CDK4 COMPLEXES (sherr)
re-activate
INACTIVATE RB (sherr weinberg)
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NORTHERNS REVEAL TWO TRANSCRIPTS
PATENT
MYRIAD PHARMACEUTICALS
PATENT
ST. JUDE
1a
2
3
Quelle 5 10 kb RACE
1b
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ARF IS A TUMOR SUPPRESSOR IN MICE!
mice die from many tumor types
THE COOL EXPERIMENT!!!
cells are immortal
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THE BIG MISTAKE
BECAUSE THEY USED MOUSE ARF!!!
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(No Transcript)
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FOUR GROUPS GET THERE FIRSTTHREE HAVE IT RIGHT
WRONG!!!
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ARF IS A SENSOR OF HYPERPROLIFERATION
SELECTION AGAINST ARF or p53
10
PICK YOUR POISON
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CONTROVERSY AGAIN!!!!!!
NUCLEAR BODIES?
NUCLEOLAR SEQUESTRATION
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THREE MODELS IN 1999
Nucleolar sequestration
Nuclear bodies
Nuclear binding
Sherr Levine Vousden Bar-Sagi
Xiong Zhang
Peters Quelle
13
IDENTIFYING THE NUCLEOLAR SIGNALS
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Re-visited in 2000
Nucleolar sequestration
Nuclear bodies
Nuclear binding
Sherr Levine Vousden Bar-Sagi
Xiong Zhang
Peters Quelle
ARTIFACT OF ADENOVIRUS
ONLY IN SAOS2 CELLS
WHEN IN DOUBT, GO WITH THE PHYSIOLOGY!
15
P53-INDEPENDENT FUNCTION FOR ARF DISCOVERED
16
BELIEVE THEM OR NOT???
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HERE WE GO AGAIN!
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NPM IS ESSENTIAL FOR PROLIFERATION
b
a
TKO MEFs
NPM RNAi
uninfected
NPM RNAi
DAPI
100
TKO MEFs
Hours post-transduction
80
24
48
72
96

-
-
-
-




BrdU Incorporation
NPM RNAi
60
a-NPM
a-NPM
40
20
a-BrdU
a-tubulin
0
TKO MEFs (96 hrs post-transduction)
19
ARF PREVENTS NPM FROM SHUTTLING
a
b
c
d
His-NPM
Myc-NPC-M9 GFP-ARF
His-NPM GFP-ARF
His-NPM GFP-ARF?1-62
m
h
h
m
DIC
h
m
h
h
m
DAPI
a-His or a-Myc
GFP
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NPM SHUTTLING IS REQUIRED FOR PROLIFERATION
a
His-NPM ?NES Myc-NPC-M9
c
vector
TKO MEFs
DIC
DAPI
a-His
a-Myc
NPM
?NES
His-NPM ?NES
100
?121-139
DAPI
h
80
m
60
BrdU Incorporation
a-His

b
40
His-NPM Flag-NPM ?NES
20
DIC
DAPI
a-His
a-Flag
a-BrdU
m
0
TKO MEFs (96 hrs post-transduction)
h
RIDING THE RIBOSOME MODEL
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THREE NEW MODELS FOR 2004
RIDE THE RIBOSOME
DEGRADE NPM/B23
PREVENT RIBOSOME SYNTHESIS
WEBER
ZHANG
SHERR
WHO WILL BE RIGHT THIS TIME?
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THE LOGIC OF OVERLAPPING TUMOR SUPPRESSORS
EGFR
p16INK4A
AKT
ARF
PTEN
p53
NPM
b-CATENIN
GSK3-b
CDK4/CYCLIN D
RB
MDM2
CDK2/CYCLIN E
E2F
p21CIP
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NPM IN HUMAN CANCERS
E2Fs
MYC
NPM
LIFE
ARF
DEATH
p53
MDM2
- Overexpressed in many cancers - Fusion partners
in lymphoma leukemia
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EVERYONES FAVORITE MODEL
Cells of origin
PDGF, FGF overexpression
Mdm2 overexpression
50
p53 mutation
EGFR amplification
65
50
Low grade glioma
CDK4 amplification RB or INK4a/ARF loss
INK4A /ARF loss
65
65
Anaplastic glioma
PTEN loss
PTEN loss
30
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PRIMARY
SECONDARY
Adapted from Holland, Nat. Rev. Genet. 2 120
(2001)
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TWO WAYS TO GET RID OF ARF
9p21 locus
hypermethylation
1a
2
3
CpG
1b

• Delete the entire 50 kb locus
• Methylate the promoter

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MASS CONFUSION
Methylation
Breast carcinomas 24-50 Colon
carcinomas 20-33 Bladder carcinomas 16-56 P
ancreas (PEN) 40-80 Salivary Gland 28 NSCLC
16-55 Glioblastomas 50-76 HNSCC 13-43 HC
C 10-16
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BUT WHAT WE DO KNOW IS
ARF IS FREQUENTLY TARGETED IN HUMAN
CANCERS USUALLY MORE-SO THAN INK4A INACTIVATION
IS USUALLY SINGULAR LOSING BOTH INK4A ARF IS
BAD LOSS OF ARF DOES NOT INVERSELY CORRELATE
WITH p53 MUTATION