Actin

PEA-15

2774 Hey

KOC-7c

OVCA420

OVCA432

OVTOKO

RMG-1

SKOV3.ip

1

Supplementary Fig. S1. Expression level of endogenous PEA-15 in ovarian cancer cell lines by Western blotting analysis. A total of 15 g of proteins was used for SDS-PAGE. The expression levels of PEA-15 protein for each cell line were evaluated using LI-COR software.

Supplementary Figure S1.Lee et al.

B

MM

P-7

Vec

tor

PE

A-1

5-A

AP

EA

-15-

DD

A

Supplementary Fig. S2. PEA-15-AA-expressing cells showed downregulation of -catenin target genes. To test the effect of PEA-15-AA-induced--catenin inhibition on -catenin target genes, we performed quantitative RT-PCR and immunohistochemical staining. (A) Quantitative RT-PCR analysis of c-myc and c-met. (B) Immunohistochemical staining of MMP-7 using xenograft tumor samples. Scale bars: 50 m.

Supplementary Figure S2.Lee et al.

Tumor tissue Cancer-adjacent normal tissue

pP

EA

-15

-S1

04

pP

EA

-15

-S1

16

-c

ate

nin

Supplementary Fig. S3. PEA-15 phosphorylated at Ser104 (pPEA-15-S104) and PEA-15 phosphorylated at Ser116 (pPEA-15-S116) were highly overexpressed in breast tumor tissues. To investigate the expression status of pPEA-15-S104, pPEA-15-S116, and b-catenin, we performed immunohistochemical staining using a human breast tumor microarray (US Biomax Inc). Original magnification, ×200 (insets, ×40).

Supplementary Figure S3.Lee et al.

Supplementary Figure S4.Lee et al.

Tu

mo

r w

eig

ht

(mg

)

Vecto

r

PEA-15-

WT

PEA-15-

DD-3

PEA-15-

AA-10

200

400

600

800

1000

1200

1400

1600

P<0.0001

P=0.0039

P<0.0001

Supplementary Fig. S4. Effect of PEA-15s in an ovarian cancer xenograft model. (A). Upper table showed incidence of tumor formation of vector, PEA-15-WT, PEA-15-WT, PEA-15-DD-3, and PEA-15-AA-1 clone. Lower panel, tumor weights were significantly smaller with PEA-15-AA-1 clone than with vector (P<0.0001), PEA-15-WT (P=0.0039), and PEA-15-DD-3 (P<0.0001) clone. (B). Representative images showed tumor formation of SKOV3.ip1-vector, SKOV3.ip1-PEA-15-WT, SKOV3.ip1-PEA-15-DD-3 and SKOV3.ip1-PEA-15-AA-1 cells in mouse peritoneal area. White arrows indicate tumors.

Vector

PEA-15-W

TPEA-15-DD-3

PEA-15-AA-1

A BTotal number

of miceIncidence of tumor

formation (%)

Vector 10 10/10 (100%)

PEA-15-WT 10 10/10 (100%)

PEA-15-DD-3 10 10/10 (100%)

PEA-15-AA-1 8 8/10 (80%)

Supplementary Table S1. Incidence of high and low -catenin expression levels in PEA-15-AA, PEA-15-DA, and PEA-15-DD tumor tissues.

  PEA-15 phosphorylation status

-catenin expression PEA-15-AA PEA-15-DA PEA-15-DD

High (intensity=2 or 3) 2 (25%) 7 (41.2%) 11 (33.3%)

Low (intensity=0 or 1) 6 (75%) 10 (58.8%) 22 (66.7%)

Total sample number 8 17 33

Supplementary Table S1.Lee et al.