A83-01 inhibits TGF-β-induced upregulation of Wnt3 and epithelial to mesenchymal transition in HER2-overexpressing breast cancer cells.
OBJECTIVE
The aim of this study is to investigate the mechanisms of interactions between TGF-β and Wnt/β-catenin pathways that induce and regulate EMT and promote breast cancer cells to become resistant to treatment.


METHODS
The effect of TGF-β on Wnt/β-catenin signaling pathway was examined by using a human Wnt/β-catenin-regulated cDNA plate array and western blot analysis. The interaction of Twist at promoter of Wnt3 was examined by chromatin immunoprecipitation (ChIP) assay. Secreted Wnt3 level was determined by ELISA assay.


RESULTS
HER2-overexpressing breast cancer cells treated with TGF-β have a reduced response to trastuzumab and exhibited EMT-like phenotype. The TGF-β-induced EMT in HER2-cells was concordant with upregulation of Wnt3 and β-catenin pathways. The TGF-β-induced induction of Wnt3 during EMT was found to be Smad3-dependent. ChIP analysis identified occupancy of Twist at promoter region of Wnt3. Knock-down of Twist by shRNA confirmed the significance of Twist in response to TGF-β regulating Wnt3 during EMT. Subsequently, TGF-β-induced matrix metalloproteinases, MMP1, MMP7, MMP9, MMP26, Vascular endothelial growth factors (VEGF), and activation of Wnt/β-catenin signaling were repressed by the shRNA treatment. TGF-βR1 ALK5 kinase inhibitor, A83-01 can effectively prevent the TGF-β-induced Twist and Wnt3. Co-treating A83-01 and trastuzumab inhibited TGF-β-induced cell invasion significantly in both trastuzumab responsive and resistant cells.


CONCLUSIONS
Our data demonstrated an important interdependence between TGF-β and Wnt/β-catenin pathways inducing EMT in HER2-overexpressing breast cancer cells. Twist served as a linkage between the two pathways during TGF-β-induced EMT. A83-01 could inhibit the TGF-β-initiated pathway interactions and enhance HER2-cells response to trastuzumab treatment.
Division of Cancer Research and Training, Department of Medicine, Charles R. Drew University of Medicine and Science, 1731 East 120th Street, Los Angeles, CA, 90059, USA. yanyuanwu@cdrewu.edu.|David Geffen School of Medicine, Jonsson Comprehensive Cancer Center, University of California at Los Angeles, Los Angeles, CA, USA. yanyuanwu@cdrewu.edu.
Yanyuan
Y
Wu
Division of Cancer Research and Training, Department of Medicine, Charles R. Drew University of Medicine and Science, 1731 East 120th Street, Los Angeles, CA, 90059, USA.
Trinh
T
Tran
Division of Cancer Research and Training, Department of Medicine, Charles R. Drew University of Medicine and Science, 1731 East 120th Street, Los Angeles, CA, 90059, USA.
Sami
S
Dwabe
Division of Cancer Research and Training, Department of Medicine, Charles R. Drew University of Medicine and Science, 1731 East 120th Street, Los Angeles, CA, 90059, USA.
Marianna
M
Sarkissyan
Division of Cancer Research and Training, Department of Medicine, Charles R. Drew University of Medicine and Science, 1731 East 120th Street, Los Angeles, CA, 90059, USA.
Juri
J
Kim
Division of Cancer Research and Training, Department of Medicine, Charles R. Drew University of Medicine and Science, 1731 East 120th Street, Los Angeles, CA, 90059, USA.
Miguel
M
Nava
Division of Cancer Research and Training, Department of Medicine, Charles R. Drew University of Medicine and Science, 1731 East 120th Street, Los Angeles, CA, 90059, USA.
Sheilah
S
Clayton
Division of Cancer Research and Training, Department of Medicine, Charles R. Drew University of Medicine and Science, 1731 East 120th Street, Los Angeles, CA, 90059, USA.|David Geffen School of Medicine, Jonsson Comprehensive Cancer Center, University of California at Los Angeles, Los Angeles, CA, USA.
Richard
R
Pietras
Division of Cancer Research and Training, Department of Medicine, Charles R. Drew University of Medicine and Science, 1731 East 120th Street, Los Angeles, CA, 90059, USA.|David Geffen School of Medicine, Jonsson Comprehensive Cancer Center, University of California at Los Angeles, Los Angeles, CA, USA.
Robin
R
Farias-Eisner
Division of Cancer Research and Training, Department of Medicine, Charles R. Drew University of Medicine and Science, 1731 East 120th Street, Los Angeles, CA, 90059, USA.|David Geffen School of Medicine, Jonsson Comprehensive Cancer Center, University of California at Los Angeles, Los Angeles, CA, USA.
Jaydutt V
JV
Vadgama
D001943:Breast Neoplasms / Q000188:drug therapy* / Q000235:genetics / Q000473:pathology; D045744:Cell Line, Tumor; D019008:Drug Resistance, Neoplasm / Q000235:genetics; D058750:Epithelial-Mesenchymal Transition / Q000235:genetics; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000235:genetics; D011720:Pyrazoles / Q000008:administration & dosage; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000235:genetics*; D000077293:Receptor, Transforming Growth Factor-beta Type I; D018125:Receptors, Transforming Growth Factor beta / Q000037:antagonists & inhibitors / Q000235:genetics; D013882:Thiosemicarbazones / Q000008:administration & dosage; D016212:Transforming Growth Factor beta / Q000235:genetics*; D000068878:Trastuzumab / Q000008:administration & dosage; D051879:Twist-Related Protein 1 / Q000235:genetics; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics; D060449:Wnt Signaling Pathway / Q000187:drug effects; D060508:Wnt3 Protein / Q000235:genetics*; D051176:beta Catenin / Q000235:genetics
D017346:Protein Serine-Threonine Kinases; D011720:Pyrazoles; D018719:Erb-b2 Receptor Tyrosine Kinases; D000077293:Receptor, Transforming Growth Factor-beta Type I; D018125:Receptors, Transforming Growth Factor beta; D013882:Thiosemicarbazones; D016212:Transforming Growth Factor beta; D000068878:Trastuzumab; D051879:Twist-Related Protein 1; D042461:Vascular Endothelial Growth Factor A; D060508:Wnt3 Protein; D051176:beta Catenin; C507011:A-83-01; C495270:CTNNB1 protein, human; C467484:VEGFA protein, human; C555507:WNT3 protein, human; C508053:ERBB2 protein, human; C000625841:TGFBR1 protein, human
EMT; Smad3; TGF-β; Twist; Wnt3