Supplemental Figure 1 - Cancer Discovery · PDF file Supplemental Figure 6 0 1 Haplotype ratio...

Click here to load reader

  • date post

    10-Jul-2020
  • Category

    Documents

  • view

    0
  • download

    0

Embed Size (px)

Transcript of Supplemental Figure 1 - Cancer Discovery · PDF file Supplemental Figure 6 0 1 Haplotype ratio...

  • 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