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Proteomic identification of E6AP as a molecular target of tamoxifen in MCF7 cells
Savita Lochab1, Pooja Pal, Jitendra K Kanaujiya
1DTDD Division, CSIR-Central Drug Research Institute, Lucknow, Uttar Pradesh, India.
Abstract:
Tamoxifen (Tam) is most widely used selective estrogen receptor modulator (SERM) for treatment of hormone-responsive breast cancer. Despite being regularly used in clinical therapy for breast cancer since 1971, the mechanism of Tam action remains largely unclear. In order to gain insights into Tam-mediated antibreast cancer actions, we applied 2DE and MS based proteomics approach to identify target proteins of Tam. We identified E6-associated protein, i.e. E6AP (UBE3A) among others to be regulated by Tam that otherwise is upregulated in breast tumors. We confirmed our 2DE finding by immunoblotting and further show that Tam leads to inhibition of E6AP expression presumably by promoting its autoubiquitination, which is coupled with nuclear export and subsequent proteasome-mediated degradation. Furthermore, we show that Tam- and siE6AP-mediated inhibition of E6AP leads to enhanced G0-G1 growth arrest and apoptosis, which is also evident from significant upregulation of cytochrome-c, Bax, p21, and PARP cleavage. Taken together, our data suggest that, Tam-targeted E6AP inhibition is in fact required for Tam-mediated antibreast cancer actions. Thus, E6AP may be a therapeutic target in breast cancer.
Insights
Tamoxifen, a breast cancer drug, works by reducing E6-associated protein (E6AP) levels. This inhibition triggers cancer cell death, suggesting E6AP as a potential therapeutic target for breast cancer treatment.
Area of Science:
- Oncology
- Molecular Biology
- Biochemistry
Background:
- Tamoxifen (Tam) is a widely used selective estrogen receptor modulator (SERM) for hormone-responsive breast cancer.
- The precise mechanism of Tam action in breast cancer therapy remains largely undefined.
- E6-associated protein (E6AP), also known as UBE3A, is upregulated in breast tumors.
Purpose of the Study:
- To elucidate the molecular targets and mechanisms underlying Tamoxifen's anti-breast cancer effects.
- To identify proteins regulated by Tamoxifen using a proteomics approach.
- To investigate the role of E6AP in Tamoxifen's therapeutic action.
Main Methods:
- Proteomics approach utilizing 2D electrophoresis (2DE) and mass spectrometry (MS) to identify Tamoxifen targets.
- Immunoblotting to confirm protein expression changes.
- Analysis of cell cycle arrest, apoptosis markers, and protein degradation pathways.
Main Results:
- Tamoxifen treatment was found to regulate E6AP expression.
- Tamoxifen induces E6AP degradation through autoubiquitination, nuclear export, and proteasomal breakdown.
- Inhibition of E6AP, either by Tamoxifen or siRNA, enhances G0-G1 growth arrest and apoptosis, evidenced by increased cytochrome-c, Bax, p21, and PARP cleavage.
Conclusions:
- Tamoxifen-mediated inhibition of E6AP is crucial for its anti-breast cancer activity.
- E6AP represents a potential therapeutic target for breast cancer treatment.
- Understanding Tamoxifen's interaction with E6AP provides insights into breast cancer therapy mechanisms.
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