99476 2 supp 1770090 mjgxq7 - cancer...
TRANSCRIPT
Supplementary Figure S1. RNA-Seq reads are concordant over multiple runs. Scatter plots show log-transformed, entire transcriptome RNA-Seq reads for prostate cancer cell line DU145 (A) across 4 technical replicates (Lanes 1 - 4) and for breast cancer cell line MCF-7 (B) across 3 biological replicates during early estrogen time-course experiment (0 - 6 hours). Dashed black lines are linear regressions. R2, correlation coefficients.
DU145
log (Absolute expression [RPKM]),Lane 1
log
(Abs
exp
ress
ion
[RPK
M])
-4 -2 0 2 4-6
-4
-2
0
2
4R2 =R2 =R2 =
0.96720.96750.9666
Lane 2Lane 3Lane 4
MCF-7
log (Abs expression [RPKM]), 0h
log
(Abs
exp
ress
ion
[RPK
M])
-4 -2 0 2 4 6-4
-2
0
2
4
63h6h
R2 =R2 =
0.92240.9119
B
A
AKT2
RNA-Seq(log[RPKM])
qPC
R(lo
g [R
Q])
0 1 2 3-0.5
0.0
0.5
1.0
1.5
2.0R2 = 0.7049
AXL
RNA-Seq(log[RPKM])
-2 -1 0 1 2 3-2
0
2
4
60.9073
EGFR
RNA-Seq(log[RPKM])
-3 -2 -1 0 1 2 30
2
4
60.9048
ERBB2
RNA-Seq(log[RPKM])
qPC
R(lo
g [R
Q])
0 1 2 3 40
1
2
3
40.8731
FGFR4
RNA-Seq(log[RPKM])
-2 -1 0 1 2 3-1
0
1
2
3
40.7018
PLK2
RNA-Seq(log[RPKM])
qPC
R(lo
g [R
Q])
0 1 2 30
1
2
3
40.7771
RET
RNA-Seq(log[RPKM])
-3 -2 -1 0 1 2-1
0
1
2
3
4
50.9251
RPS6KB1
RNA-Seq(log[RPKM])
0.0 0.5 1.0 1.5 2.0 2.5-0.5
0.0
0.5
1.0
1.50.7097
Supplementary Figure S2. RNA-Seq reads correlate with gene expression. Log-transformed RNA-Seq expression for 8 different kinases, measured as RPKM, are plotted against log-transformed gene expression, measured as relative quantity (RQ) by qPCR. Each point represents a unique sample. Dashed black lines are linear regressions. R2, correlation coefficients.
qPC
R(lo
g [R
Q])
qPC
R(lo
g [R
Q])
qPC
R(lo
g [R
Q])
qPC
R(lo
g [R
Q])
qPC
R(lo
g [R
Q])
R2 = R2 =
R2 = R2 = R2 =
R2 = R2 =
AKT2
RNA-
Seq
(RPK
M) qPC
R(R
Q)
PAN
C-1
SU.8
6.86
PL45
HC
C22
18
SK-M
EL-5
HBL
-100
MM
576
MD
A-M
B-41
5
HC
C14
28
MD
A-M
B-23
1
hTER
T-H
ME1
Hs-
700T
HC
C13
95
Panc
-08.
13
MM
96L
Panc
-02.
13
MD
A-M
B-15
7
Panc
-05.
04
AsPC
-1
Hs-
766T
0
100
200
300
400
500
0
20
40
60
80RNA-SeqqPCR
FGFR4
RNA-
Seq
(RPK
M) qPC
R(R
Q)
HC
C22
18
MD
A-M
B-3
30
UA
CC
-812
_T
MD
A-M
B-4
15
UA
CC
-893
_T
Pan
c-10
.05
BT-
20
Pan
c-02
.03
PA
NC
-1
BT-
549
HC
C13
95_B
L
Pan
c-08
.13
Pan
c-02
.13
HC
C14
28_B
L
MC
F10A
Pan
c-05
.04
MD
A-M
B-2
31
BxP
C-3
hTE
RT-
HM
E1
HP
NE
010
0100
20
70
120
170
200
2200
4200
6200PLK2
RNA-
Seq
(RPK
M) qPC
R(R
Q)
HC
C22
18
Panc
-05.
04
MIA
-PaC
a-2
PL45
MM
96L
BT-5
49
Panc
-10.
05
Panc
-03.
27
Cap
an-2
hTER
T-H
ME1
Hs-
700T
BxPC
-3
L3.3
SK-M
EL-5
UAC
C-8
12_T
HC
C14
28
MC
F7
MM
576
MD
A-M
B-3
30
HC
C14
28_B
L
0
100
200
300
400
0
200
400
600
800
RPS6KB1
RNA-
Seq
(RPK
M) qPC
R(R
Q)
MC
F7
BT-4
74
HC
C22
18
Panc
-05.
04
MIA
-PaC
a-2
DU
-145
Panc
-10.
05
HC
C19
37
HPD
E
HC
C13
95_B
L
AsPC
-1
BxPC
-3
MD
A-M
B-15
7
Hs-
700T
HC
C14
28_B
L
HC
C11
87
BT-2
0
BT-5
49
HPN
E
HC
C13
95
0
20
40
60
80
100
0
5
10
15
20
25
Supplementary Figure S3. Validation of outlier expression predicted by RNA-Seq. RNA-Seq reads (blue) and qPCR gene expression (purple) of 8 kinases are plotted for 20 different samples. Samples with predicted outlier expression of the respective kinase by RNA-Seq are highlighted by the red bars. Samples with predicted non-outlier expression are highlighted by the green bars.
RNA-SeqqPCR
RNA-SeqqPCR
RNA-SeqqPCR
AXL
RNA-
Seq
(RPK
M) qPC
R(R
Q)
MD
A-M
B-23
1
Panc
-03.
27
HPN
E
CFP
AC-1
HBL
-100
DU
-145
Panc
-10.
05
HC
C14
28
HC
C13
95_B
L
HC
C14
28_B
L
ZR-7
5-30
HC
C22
18
UAC
C-8
12_T
MD
A-M
B-33
0
BT-2
0
MC
F7
SK-M
EL-5
HC
C11
87
MM
576
UAC
C-8
93_T
050
020100100100200100300100400100
100200300400500
ERBB2
RNA-
Seq
(RPK
M) qPC
R(R
Q)
ZR-7
5-30
UAC
C-8
93_T
UAC
C-8
12_T
HC
C22
18
MD
A-M
B-33
0
BT-4
74
SU
.86.
86
Hs-
700T
BT-5
49
MD
A-M
B-23
1
Hs-
766T
PAN
C-1
hTE
RT-
HM
E1
HC
C13
95
MD
A-M
B-15
7
MM
576
MM
96L
HC
C13
95_B
L
SK-
ME
L-5
HC
C14
28_B
L
050
0250500
2000
3500
5000
100
1100
2100
3100
RET
RNA-
Seq
(RPK
M) qPC
R(R
Q)
UAC
C-8
93_T
UAC
C-8
12_T
HC
C14
28
MC
F7
MD
A-M
B-33
0
BxP
C-3
Cap
an-1
Cap
an-2
HC
C13
95_B
L
HC
C14
28_B
L
HP
AC
MC
F10A
MD
A-M
B-23
1
MM
576
MM
96L
Pan
c-02
.03
Pan
c-03
.27
Pan
c-05
.04
Pan
c-08
.13
Pan
c-10
.05
0.00.51.0
010202020402060208020
20406080 RNA-Seq
qPCR
RNA-SeqqPCR
RNA-SeqqPCR
EGFR
BT-
20
AsP
C-1
HP
DE
L3.3
MD
A-M
B-2
31
BxP
C-3
Pan
c-02
.03
HC
C22
18
HC
C14
28
SK
-ME
L-5
BT-
474
MC
F7
UA
CC
-812
_T
MD
A-M
B-4
15
ZR-7
5-30
MD
A-M
B-3
30
MM
96L
MM
576
HC
C14
28_B
L
HC
C13
95_B
L
010
02500500025000450006500085000
2070
120170220 RNA-Seq
qPCR
RNA-
Seq
(RPK
M) qPC
R(R
Q)
Supplementary Figure S4. Gene expression is concordant for matched cell lines and xenografts. (A, left) Log-transformed RNA-Seq expression of all 468 kinases for primary pancreatic adenocarcinoma tissue xenograft DS-08-947 are plotted against log-transformed RNA-Seq expression for its derived cell line. Kinases highlighted in purple have outlier expression in both xenograft tissue and matched cell line. (A, right) Log-transformed expression levels of random genes for DS-08-947 are plotted against log-transformed expression levels for its derived cell line by qPCR. (B) Log-transformed expression levels of random genes for commercial pancreatic adenocarcinoma cell lines, BxPC-3 and PANC-1, are plotted against log-transformed expression levels for the corresponding xenografts by qPCR. Genes with outlier expression nominated by RNA-Seq are highlighted in purple. Dashed black lines are linear regressions. R2, correlation coefficients.
B
A
BxPC-3
Cell line qPCR(log[RQ])
Xeno
graf
t qPC
R(lo
g[R
Q])
0 2 4 6 80
2
4
6MET
EGFRR2 = 0.8060
PANC-1
Cell line qPCR(log[RQ])
Xeno
graf
t qPC
R(lo
g[R
Q])
0 2 4 6 80
2
4
6
8
AKT2
MET
R2 = 0.7765
DS-08-947: qPCR
Xenograft cell line qPCR(log[RQ])
Xeno
graf
t tis
sue
qPC
R(lo
g[R
Q])
0 2 4 60
2
4
6
METR2 = 0.7940
DS-08-947: RNA-Seq
Xenograft cell line RNA-Seq(log[RPKM])
Xeno
graf
t tis
sue
RN
A-Se
q(lo
g[R
PKM
])
-3 -2 -1 0 1 2 3-3
-2
-1
0
1
2
3EPHA2PLK2
R2 = 0.8330
TNIK
MET