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THE CHORDOMA GENOME
PETER CAMPBELL,
WELLCOME TRUST SANGER INSTITUTE
Chordoma
• 1% of primary malignant bone
tumours.
32%
• 0.5-1/1000,000/year.
• 0 - 90 yrs: 55, any age,
33%
• Male to female ratio 2:1
29%
•37% metastasize
•5% dedifferentiation
•Overall median survival ~7 years
Chordoma
H&E
BRACHYURY
Coding point mutations – 23 chordomas
1. Whole exome
sequencing
Sample
ascertainment
Sample QC
Whole exome library
preparation
2. Variant
discovery
NGS data QC &
filtering
Whole exome
mapping
Normal –Tumour
sequence
comparison
Raw variant
discovery
Massively parallel
sequencing
(Illumina)
Variant annotation
Variant selection
3. Validation and
confirmation
Variant resequencing
Roche 454
Somatic variant
confirmation
Define driver
mutations
Targeted
resequencing studies
Calculate prevalence
PD4929a
PD3814a
PD3804a
PD6370a
PD4188a
PD4926a
PD7185a
PD3808a
PD7187a
PD7186a
PD4927a
PD3820a
PD3812a
PD4187a
PD3821a
PD3815a
PD4928a
40
PD3819a
50
PD3807a
60
PD4183a
70
PD3806a
PD7184a
PD7183a
Frequency
Number of coding mutations per patient
complex indel
insertion
deletion
microRNA
essential splice
nonsense
30
missense
20
10
0
Mutations in known cancer genes
PIK3CA mutations
E545K
M1043I
C
T
C
A
G
T
G
G
C
G
G
C
G
C
C
C
A
C
G
T
G
C
G
T
C
G
A
missense
C essential splice
T
missense
G
missense
A
missense
ag
deletion
C
missense
A
missense
C
missense
T
missense
tc
deletion
T
insertion
ac
deletion
ataac
deletion
A
missense
T
missense
A
missense
T
missense
A
missense
G
missense
G
missense
T
missense
T
missense
g
deletion
A
missense
C
missense
C
missense
T
missense
A
missense
A
missense
A
missense
T
missense
c.
3780
3780
4344
4344
13458
13458
12186
12186
2736
2736
3504
3504
11406
11406
3207
3207
1885
1885
6948
6948
10383
10383
6018
6018
3534
3534
2547
2547
10191
10191
10650
10650
p.
gene cds (bp)
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
consequence
gene freq (excl
silent)
AUTS2
AUTS2
DCC
DCC
DNAH17
DNAH17
FLG
FLG
GPR110
GPR110
ITGA10
ITGA10
LYST
LYST
PIK3CA
PIK3CA
PRICKLE3
PRICKLE3
PTPRZ1
PTPRZ1
RELN
RELN
SCN2A
SCN2A
SLC9A10
SLC9A10
SLCO5A1
SLCO5A1
VCAN
VCAN
XIRP2
XIRP2
mut allele
gene
PD3814a
PD7186a
PD3820a
PD7186a
PD3819a
PD7185a
PD4929a
PD4929a
PD4929a
PD7184a
PD4188a
PD4188a
PD4187a
PD4927a
PD3814a
PD7187a
PD3814a
PD7185a
PD3804a
PD6370a
PD4187a
PD6370a
PD3808a
PD4929a
PD4928a
PD4929a
PD3807a
PD4926a
PD4928a
PD7185a
PD3815a
PD3821a
wt allele
sample
Recurrently mutated genes
p.T1047N
p.?
p.A455V
p.T1110A
p.R1515C
D_ag
p.H2505R
p.S3149F
p.N437K
p.G253R
D_tc
I_T
D_ac
D_ataac
p.E545K
p.M1043I
p.V130L
p.A301D
p.T957N
p.A2267G
p.E2910Q
p.S594T
p.S1974L
D_g
p.R721C
p.I911V
p.P341A
p.R596Q
p.V132I
p.D1366E
p.N2480K
p.Q1680H
c.3140C>A
c.660+2T>C
c.1364C>T
c.3328A>G
c.4543C>T
2
c.7514A>G
c.9446C>T
c.1311C>G
c.757G>A
2
1
2
5
c.1633G>A
c.3129G>T
c.388G>T
c.902C>A
c.2870C>A
c.6800C>G
c.8728G>C
c.1780T>A
c.5921C>T
1
c.2161C>T
c.2731A>G
c.1021C>G
c.1787G>A
c.394G>A
c.4098T>A
c.7440C>A
c.5040G>T
Summary of exome data
• 10-30 coding mutations per patient
• No frequently mutated novel genes
• Recurrent mutations in genes regulating histone
modification
• Recurrent mutations leading to activation of PI3K
signalling
Finding genomic rearrangements
500bp
Chr 1
Chr 4
Chr 1
Chr 4
Chordoma
Other chordomas
Hallmarks of chromothripsis
• Massive genomic rearrangement in localised chromosomal
regions
• Whole chromosomes, chromosome arms or chromosome bands
• Alternating copy number states
• 2, 3 or occasionally 4 discrete states with many switches
• Retention of heterozygosity in higher copy number state
• Clustering of breakpoints
• Ends essentially randomly joined in random orientation
• Approx equal numbers of rearrangements with ‘deletion-type’,
‘tandem-duplication-type’ and ‘inverted’ orientation
Catastrophe model
Catastrophic chromosome breakage
Non-homologous end joining
Rearrangement screens in sarcoma
Histology
Number
Number with
chromothripsis
Osteosarcoma
10
3
Chordoma
14
2
Myxofibrosarcoma
5
3
Pleomorphic sarcoma
2
2
MPNST
2
2
Liposarcoma
2
0
Leiomyosarcoma
2
2
Chondrosarcoma
2
0
Solitary fibrous tumour
1
0
Total
40
14 (35%)
Triple jeopardy – chordoma
Role of inherited genetic factors
SNP upstream of T (brachyury)
Level of association – rs2305089
CASES
A allele freq
CONTROLS
A allele freq
P value
OR
95% CI
Discovery
0.88
0.53
4.4x10-9
6.1
3.1-12.1
Replication
0.83
0.53
2.8x10-4
4.1
1.8-9.5
4.6x10-12
5.3
3.1-8.9
Combined
SNP associated with T expression
Conclusions
• Chordomas show 10-30 coding mutations per patient
• Frequent incidence of chromothripsis
• Germline predisposition with brachyury SNP
Acknowledgements
University College London
Cancer Genome Project
• Jose Tubio & Susie Cooke
• Patrick Tarpey
•
•
•
•
Adrienne Flanagan
Roberto Tibrabosco
Fernanda Amary
Nischalan Pillay
• David McBride
• John Marshall & Keiran Raine
Chordoma Foundation
• Adam Butler & Jon Teague
•
• Lucy Stebbings & Catherine Leroy
• Sarah O’Meara, Laura Mudie
• Mike Stratton & Andy Futreal
Josh Sommer