Related Experiment Video
Updated: Jun 5, 2026
Experimental Approaches to Study Mitochondrial Localization and Function of a Nuclear Cell Cycle Kinase, Cdk1
Published on: February 25, 2016
2D-DIGE analysis of phospho-enriched fractions from dasatinib-treated melanoma cell lines
Alex J Eustace1, Paul Dowling, Michael Henry
1National Institute for Cellular Biotechnology, Dublin City University, Dublin 9, Ireland. Alex.Eustace@dcu.ie
Abstract:
Current therapeutic regimes for metastatic melanoma have failed to provide robust clinical responses. Dasatinib has shown anti-proliferative and anti-invasive effects in vitro; however, not all melanoma cells tested were sensitive to dasatinib. We used 2D-DIGE analysis of phospho-enriched fractions to identify phosphoproteins involved in regulating response to dasatinib in an isogenic pair of melanoma cell lines, one sensitive to dasatinib (WM-115) and the other resistant (WM-266-4). In WM-115 cells treated with dasatinib, 18 unique protein species with altered phosphorylation levels were detected. Dasatinib treatment of WM-266-4 cells resulted in phosphoprotein alterations to four unique protein species. Four phosphorylated forms of Annexin-A2 (ANXA2) were increased in WM-115 cells treated with dasatinib, whilst dasatinib treatment did not alter ANXA2 phosphoprotein levels in WM-266-4 cells. Immunoblotting confirmed that phosphorylation of ANXA2, on tyrosine residues, was increased in WM-115 cells treated with dasatinib. Subsequent knockdown of ANXA2 by siRNA significantly inhibited proliferation of WM-115 cells but did not significantly reduce proliferation of WM-266-4 cells. Therefore, ANXA2 plays a role in regulating proliferation in dasatinib-sensitive WM-115 cells and could potentially play a role in sensitivity to dasatinib in melanoma cells.
Insights
Dasatinib shows promise for metastatic melanoma, but resistance is an issue. Researchers identified Annexin-A2 (ANXA2) phosphorylation as a key factor in dasatinib sensitivity in melanoma cells.
Area of Science:
- Oncology
- Molecular Biology
- Biochemistry
Background:
- Metastatic melanoma treatments often lack efficacy.
- Dasatinib exhibits anti-cancer properties but faces variable sensitivity in melanoma cells.
Purpose of the Study:
- To identify phosphoproteins regulating dasatinib response in melanoma.
- To investigate the role of Annexin-A2 (ANXA2) in dasatinib sensitivity.
Main Methods:
- Utilized 2D-DIGE analysis of phospho-enriched fractions.
- Compared dasatinib-treated sensitive (WM-115) and resistant (WM-266-4) melanoma cell lines.
- Confirmed ANXA2 phosphorylation via immunoblotting and assessed proliferation after ANXA2 knockdown using siRNA.
Main Results:
- Identified 18 and 4 unique phosphoprotein alterations in sensitive and resistant cells, respectively.
- Observed increased phosphorylation of Annexin-A2 (ANXA2) in dasatinib-sensitive WM-115 cells.
- ANXA2 knockdown significantly inhibited WM-115 cell proliferation but not WM-266-4 cells.
Conclusions:
- ANXA2 phosphorylation is elevated in dasatinib-sensitive melanoma cells.
- ANXA2 plays a role in regulating proliferation in dasatinib-sensitive melanoma.
- ANXA2 may be a potential therapeutic target for improving dasatinib sensitivity in melanoma.
