Supplemental Figure 1 - Cancer Discovery · PDF file Supplemental Figure 6 0 1 Haplotype ratio...
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TCGA-06-0214
TCGA-06-5415
TCGA-14-2554
TCGA-26-5132
EGFR vIII var. 1
EGFR vIII var. 2
EGFR vIII var. 3
EGFR C-ter deletion
EGFR vIII var. 1
EGFR vIII var. 2
EGFR vIII var. 1
Δ13-15
EGFR vIII var. 2
EGFR vIII var. 3
EGFR vIII var. 1
EGFR vIII var. 2
EGFR vIII var. 3
A
B
C
D
Supplemental Figure 1
Number of read pairs (million) 10
0 10
1 10
2 0
0.5
1
Different blood normal libraries
Bulk primary tumor libraries
Down-sampled bulk tumor libraries
Selected single-cell WGA libraries
Poor WGA libraries
0.5
1
10 0
10 1
10 2
Number of read pairs (million)
K S
m et
ric (
~ 60
K b
bi ns
) K
S m
et ric
( ~
57 K
b bi
ns )
0 0.5 1 0
0.5
1
KS metric
% o
f p ile
up r
ea ds
r em
ov ed
Down-sampled bulk tumor libraries Single-cell WGA libraries
A
C
BBT340
BT325
0
Supplemental Figure 2
7: 54,560,250--55,510,789
p11.1
A
p12.1 p11.2
BT325
EGFR
p12.1 p11.1
7: 54,861,307--55,396,737
p11.2
B
C
D
EGFR CEP7
EGFR 12 13 14 15 16 Iso A (1210aa)
BT340
EGFR
Supplemental Figure 3
Chr. 7 Chr. 7
p12.1 p11.1p11.2
Two distinct deletions spanning ex. 14-15
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 1617 18
19 20
21 22
X
T /N
c op
y ra
tio
0.5
1
2
Subclonal
Homozygous deletion
Hemizygous deletion
Trisomic
Tetrasomic
Disomic
Clonal 54
1
2
0.5
1.5
0
T /N
c op
y ra
tio
a
a. Subclonal deletions in chrs. 1, 4, 6
EGFR
CDKN2A/2B
BA Bulk copy number of BT325 Clonality analysis of SCNA in BT325
Supplementary Figure 4
A Germline heterozygous site
Reference base Alternate base
Genotype
Possibe haplotypes
~1Kb
Cluster 1 Cluster 2
Clustering based on pairwise distance
Haplotype inference
Retained allelotypeDeleted allelotype
LOH populationHeterozygous population
RD
R R D R D
. . R R . . R
. . . . .. . . . .
1
2
3
4
5
6
Single-cell allelotyping
D D RD D R
. . . R . . R
R R . . R ..
D
0
0.5
1
Allelotype ratio (deleted vs. retained) within SNS
Allelotype distribution within two clusters
Cluster 1
Cluster 2
R D
R D
T /N
m ix
di pl
oi d
F Cell
B
A
Loss -of-heterozygosity
Haplotypes
Heterozygous
Genotypes from incomplete single-nucleus sequencing
Homozygous loss
A
B
Genotype from complete single-nucleus sequencing
1
2
3
4
5
6
Pairwise distance
2/3
C
cell 3 (LOH) vs.
cell 1(Het)
cell 3 (LOH) vs.
cell 4 (LOH) 0
Pairwise comparison
E
G
0
0.01
0.02
0.03
0.04
0.05 % of bases containing at least 1 error read avg: 0.019; std. dev: 0.005 % of reads containing at least 1 error base avg: 0.012; std. dev: 0.003
Tumor cells with chr. 10 deletion
H
Supplemental Figure 5
1
0.5
0
Haplotype ratio in Chr. 10 (Deleted vs. Retained)
Chromosome 7 copy ratio
Cells with less even coverage Cells with more even coverage
0
1
2
3
Normal (diploid)
Tumor cells with complete 6q deletion Tumor cells retaining multiple 6q segments
A
B
C
Chromosome 10 deletion
Chromosome 7 amplification
Chromosome 6 haplotyping Chr. 9
haplotyping
D E
Haplotype compositions in regions with subclonal deletions
Retained haplotype Deleted haplotype
0 0.5 1 0 1000
Haplotype ratio
D:R Total # of SNPs
53.6--57 Mb 63.3--66.1 Mb
27.1-30.9 Mb 31.8-32.0 Mb
28--33 Mb
Clonal Events
Subclonal deletions in 6q and 9p Subclonal Chr. 16 loss
Subclonal Events
Supplemental Figure 6
0 1
Haplotype ratio D:R
C hr
. 1 6
LO H
C hr
. 1 6
no n-
LO H
CDKN2A hom. loss Chr.10 hem. loss
Deletions in Chrs. 6 and 9
Chr. 16 loss
A
Chr9: 9--35Mb
Retained allelotype Deleted allelotype
B
0 0.5 1
Haplotype ratio (Deleted vs. Retained)
0 0.5 1 1.5 2
Chr. 7 copy ratio
C9p deletion Chr. 10 deletion Trisomy 7 Haplotyping
Clonal Events
Chr7: 54560159-54560356 Chr7: 55510459-55510566
Tumor/normal mixture 1
Tumor/normal mixture 2
E
Amplicon 7: 54,560,250--55,510,789
F
Tumor/normal mixture 1
Tumor/normal mixture 2
Chr3: 22615037-22615438Chr9: 21029084-21029485
Presence of EGFR amplification in T/N mixtures
Presence of Chr. 9-Chr. 3 translocation in T/N mixtures
D
1B1A
8B8A
9B9A
Rearranged segments by the reference assembly
Rearranged segments in derivative chromosomes
Complex rearrangement disrupting CDKN2A
Bulk 2 1 0
C N
Representative SNS library
Pool of SNS libraries
Chr. 1
Chr. 8
Chr. 9
Chr. 3 fragments
20,900--22,150Kb
CDKN2A/CDKN2B
9A 8B
1A 9B
8A 1B
der (1,9)
der (8,1)
der (9,8) × 2
Supplementary Figure 7
Chr. 2
Chr. 8
2:187389362
2:187389633
8:32908298
8:32909007
2:1892626238:2261740
2:1894072648:2261928 A
8p-tel.
2q-tel.
8p-tel.
8:32908298
B -a5 • -a6 • -a1
a7 • -a3 • -a4 • a9 • a8 • -a2 • -a10
Chr. 12
a b c de f g 12a • 12e • 12b • 12f • -12d • 12c • 12f
C Chr. 14
a
b
c d
e
f g nlkjih
m 14a • -14l • 14g • -14f • 14d • 14c • 12f
? • 14i • -14j • 14k • 14h • ?
D
Chr. 9Chr. 6 9B6A
Simple deletion
Chr.6 Chr.9 CEP6
Chr. 9
20 25 30 35 40
E
Bulk CN 50 60 7055 65
FISH probes Chr. 6 Bulk CN
Chr.6 Chr.9 CEP9
extrachromosomal
amplicons
FISH probes Chr. 9 chr6:71,726,042-71,726,935 chr9:27,138,441-27,140,176
+
connects to the Chr. 6 amplicon
connects to the Chr. 6 segment
connects to another Chr. 6 segment
connects to another Chr. 6 amplicon
Non-amplified Chr. 6 segment Non-amplified Chr. 9 segment
Chr. 6 amplicon Chr. 9 amplicon
Inferred derivative chromosome in cluster 1
9B6A
9B
Inferred derivative chromosome in cluster 2
6A 9B
6A
Chromothripsis
6.5Mb
9c
9b
6d
6c
6b 6a
dm 6:9
Supplemental Figure 8
Chromosome 1 (Mb) Chromosome 4 (Mb) Chromosome 6 (Mb)
20 22 24 26 183 185 187 110 120 130 140 150 160
0.5
M in
or a
lle le
fre
qu en
cy
0
1
Bulk CN
0.50 1 1.5
All tumor nuclei
Haplotype ratio (Deleted vs. Retained)
Nuclei retaining heterozygosity
Nuclei having lost heterozygosity
Two nucleus mixture
A
Retained allelotype Deleted allelotype
0. 5
0 1
1. 5
Nuclei retaining heterozygosity
Nuclei having lost heterozygosity in chr. 1, 4, 6
1:1 mixture
0 0.
5 1
Chr. 10
Deletions in chrs 1, 4, 6
Tumor nuclei Non-tumor nuclei
CDKN2A homozygous deletion Chr. 10 hemizygous deletion Heterozygous Chr. 10
Diploid normal 1:1 mixture T/N
H ap
lo ty
pe r
at io
D :R
H ap
lo ty
pe r
at io
D :R
Chromosome 1 (Mb) Chromosome 4 (Mb) Chromosome 6 (Mb) 20 22 24 26 183 185 187 110 120 130 140 150 160
0.5
M in
or a
lle le
fre
qu en
cy
0
1
Bulk CN
LOH tumor nuclei only
1a 1b
1A 4a 4b
4A4A4A
6a 6b 6c 6d 6e 6f 6h6g 6i
6D 6E 6F6G
Rearranged segments by the reference assembly
