Download Ma-1/HGF

HCC827ER
HGF(-)
140
120
120
100
100
% cell viability
% cell viability
140
HCC827ER
HGF(+)
80
60
40
20
Erlotinib
17-DMAG
17-DMAG+Erlotinib1μM
80
60
40
20
0
0
0.01
0.03
0.1
0.3
Drug concentration (μM)
1
0
0
0.01
0.03
0.1
0.3
1
Drug concentration (μM)
Supplementary Figure 1. The effect of 17-DMAG on the growth of lung cancer
cells with Met amplification
Tumor cells were continuously treated with increasing concentrations of EGFR-TKI,
erlotinib, or 17-DMAG, with or without HGF (20 ng/ml), and cell growth was determined
after 72 hours by MTT assay. Data shown are the representative of 3 independent
experiments. Error bars indicate SD of triplicate cultures.
PC-9 HGF(-)
140
120
100
% cell viability
80
60
40
100
EGFR-TKI
80
17-DMAG
60
17-DMAG+EGFR-TKI 1μM
40
20
20
0
0
0
0.01
0.1
Drug concentration (μM)
0
1
PC-9 HGF(+)
140
0.01
0.1
Drug concentration (μM)
1
H1975 HGF(+)
120
100
120
% cell viability
100
80
60
40
20
100
80
60
40
0
0.01
0.1
Drug concentration (μM)
1
80
60
40
20
20
0
Ma-1/HGF
120
140
% cell viability
% cell viability
120
% cell viability
H1975 HGF(-)
140
0
0
0
0.01
0.1
Drug concentration (μM)
1
0
0.01
0.1
Drug concentration (μM)
1
Supplementary Figure 2. The effect of combined therapy with 17-DMAG and EGFRTKI on the growth of lung cancer cells with mutated EGFR
Tumor cells were continuously treated with increasing concentrations of EGFR-TKI,
erlotinib (PC-9 and Ma-1/HGF), CL-387,785 (H1975), or 17-DMAG, with or without HGF (20
ng/ml), and cell growth was determined after 72 hours by MTT assay. Data shown are the
representative of 3 independent experiments. Error bars indicate SD of triplicate cultures.
Supplementary Figure 3.
30
25
20
15
5
< 0.1
10
< 0.1
HGF (ng/ 2×105cells)
35
0
Ma-1
Ma-1/Vec
Ma-1/HGF
Ma-1/Vec
Ma-1/HGF
PI
Ma-1
Control
Erlotinib
17-DMAG
HGF
HGF+Erlotinib
HGF+17-DMAG
2.90
31.52
35.49
3.05
5.44
24.89
1.64
14.01
12.11
0.69
2.85
15.64
2.64
4.04
17.81
AnnexinV
Supplementary Figure 4. 17-DMAG induces apoptosis even in the presence of HGF.
Ma-1, Ma-1/Vec, and Ma-1/HGF cells were incubated with HGF (20 ng/mL) and erlotinib
(0.3 μmol/L) or 17-DMAG (0.3 μmol/L) for 48 hour and washed twice with PBS. The
apoptotic cells were determined by Annexin V assays according to the manufactor’s
protocol. Values shown are percentage of apoptotic cells. FL1-H and FL2-H, heights of
fluorescence intensity.
Tumor Volume (mm3)
A
450
400
350
300
250
200
150
100
50
0
Erlotinib
Tumor Volume (mm3)
1
5
2
6
3
7
4
8
17-DMAG
Combination
0
B
C
Control
450
400
350
300
250
200
150
100
50
0
7
10
14
Days after inoculation
17
21
Control
Erlotinib
17-DMAG
Combination
0
7
10
14
Days after inoculation
17
21
1: Ma-1 Control
2: Ma-1 Erlotinib
3: Ma-1 17-DMAG
4: Ma-1 Combination
5: Ma-1/HGF Control
6: Ma-1/HGF Erlotinib
7: Ma-1/HGF 17-DMAG
8: Ma-1/HGF Combination
Supplementary Figure 5. The effect of combination treatment with 17-DMAG plus erlotinib to
HGF-induced erlotinib resistance in vivo.
Ma-1 /Vec (A) or Ma-1/HGF (B) (5 × 106) cells were inoculated subcutaneously into SCID mice on
day 0. Mice received oral erlotinib (20 mg/kg/d) and/or intraperitoneal 17-DMAG (10 mg/kg/d),
starting on day 7. Tumor size was measured twice a week and tumor volumes were calculated as
described in Materials and Methods. Error bars indicate standard errors of 6tumors. C, macroscopic
appearances of representative tumors harvested on day 21.
Supplementary Figure 6.
Ma-1/HGF
Ma-1/Vec
Control
Erlotinib
17-DMAG
Supplementary Figure 7.
Ma-1/HGF
Ma-1/Vec
Control
Erlotinib
17-DMAG