Immunohistochemical validation of multiple phospho-specific epitopes for estrogen receptor α (ERα) in tissue microarrays of ERα positive human breast carcinomas

Skliris, Georgios; Rowan, Brian G.; Al-Dhaheri, Mariam; Williams, Christopher C.; Troup, Sandy; Begic, Sanela; Parisien, Michelle; Watson, Peter H.; Murphy, Leigh C.

doi:10.1007/s10549-008-0267-z

Cited by 33 publications

(48 citation statements)

References 23 publications

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“…One concern in the detection of phospho proteins regards tissue collection routines. Validation of several ER phospho antibodies did not show a significant decreased detection of phospho ERs in relation to increased time of collection (15). The specificity of the Pak1 antibody was previously confirmed by Western blot and immunohistochemistry (16 test was applied.…”

Section: Translational Relevancementioning

confidence: 77%

Estrogen Receptor-α Phosphorylation at Serine 305, Nuclear p21-Activated Kinase 1 Expression, and Response to Tamoxifen in Postmenopausal Breast Cancer

Bostner

Skoog

Fornander

et al. 2010

Clinical Cancer Research

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Purpose: In vitro, p21-activated kinase 1 (Pak1) phosphorylates the serine 305 residue of the estrogen receptor α (ERα) and influences the response of breast cancer cells to tamoxifen. We investigated the influence of Pak1 and pERα ser305 on breast cancer prognosis and results of tamoxifen therapy. Experimental Design: We examined Pak1 and pERα ser305 protein by immunohistochemistry in a series of 912 tumors from node-negative breast cancer patients randomized to tamoxifen or no adjuvant endocrine treatment.Results: Cytoplasmic Pak1 correlated to large tumors and ER negativity, whereas nuclear Pak1 and pERα ser305 correlated to small tumors and ER positivity. Nuclear expression of Pak1 and pERα ser305 predicted reduced response to tamoxifen in patients with ERα-positive tumors (tamoxifen versus no tamoxifen: hazard ratio (HR), 1.33; 95% confidence interval (95% CI), 0.42-4.2; P = 0.63), whereas patients lacking this combination benefitted significantly from tamoxifen (HR, 0.43; 95% CI, 0.30-0.62; P < 0.0001). Similar nonsignificant trends were detected in analyses of the proteins separately. Pak1 in the cytoplasm was an independent prognostic marker, indicating increased recurrence rate (HR, 1.79; 95% CI, 1.17-2.74; P = 0.0068) and breast cancer mortality (HR, 1.98; 95% CI, 1.14-3.46; P = 0.016) for patients randomized to no adjuvant treatment. Conclusion: Our results suggest that patients with tumors expressing Pak1 and pERα ser305 in combination are a group in which tamoxifen treatment is insufficient. In addition, the pathway may be of interest as a drug target in breast cancer. Furthermore, the findings support previous studies showing that Pak1 has differential roles in the cytoplasm and the nucleus. Clin Cancer Res; 16(5); 1624-33. ©2010 AACR.

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Section: Translational Relevancementioning

confidence: 77%

Estrogen Receptor-α Phosphorylation at Serine 305, Nuclear p21-Activated Kinase 1 Expression, and Response to Tamoxifen in Postmenopausal Breast Cancer

Bostner

Skoog

Fornander

et al. 2010

Clinical Cancer Research

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“…Other phospho-ERα specific antibodies used for IHC were validated previously [8]. HER-2 (mouse monoclonal, clone CB11) and ERα (6F11, mouse monoclonal, Abcam, Cambridge, MA) were used as previously described [9].…”

Section: Antibodiesmentioning

confidence: 99%

Estrogen Receptor Alpha Phosphorylated at Tyrosine 537 is Associated with Poor Clinical Outcome in Breast Cancer Patients Treated with Tamoxifen

et al. 2010

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A phospho-tyrosine 537-ERα (p-Y537-ERα) antibody was validated for immunohistochemistry of formalin-fixed paraffin-embedded human breast cancer biopsies and used to interrogate multiple ER positive breast cancer cases present on tissue microarrays already constructed by the Manitoba Breast Tumor Bank. Nuclear p-Y537-ERα protein expression was positively associated with positive nodes (Spearman r = 0.20, P = 0.0002) and large tumor size (r = 0.13, P = 0.02). On univariate analysis, high levels of p-Y537 were associated with poor overall survival (HR = 1.65, 95% CI 1.08-2.52, P = 0.02) but not relapse free survival from breast cancer. The association with overall survival was not significant on multivariate analysis. These data support the relevance of phosphorylation at p-Y537-ERα in human breast cancers and add further support to the presence of a phosphorylation code for ERα in human breast cancer in vivo.

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“…Indeed, several causes and contributing factors for inducing TAM resistance have been described in breast cancer patients or cell models, including upregulation of growth factor receptors (like EGFR (epidermal growth factor receptor), 8,9 IGFR (insulin-like growth factor receptor) and HER2 (human epidermal growth factor receptor 2) 10,11 ), activation of kinases (such as AKT, 12 MAPK, 13,14 protein kinase A (PKA) in combination with PAK1 6,15,16 ), and the resulting phosphorylation status of ERa. [17][18][19][20] The effects of PKA-induced phosphorylation of ERa at Serine residue 305 (ERaS305-P) in the region between the ligand-binding domain and the DNA-binding domain are understood in molecular detail. Phosphorylation at Ser305 results in a conformational arrest when exposed to TAM, 17 which affects recruitment of coregulators.…”

Section: Introductionmentioning

confidence: 99%

PKA phosphorylation redirects ERα to promoters of a unique gene set to induce tamoxifen resistance

et al. 2012

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Protein kinase A (PKA)-induced estrogen receptor alpha (ERa) phosphorylation at serine residue 305 (ERaS305-P) can induce tamoxifen (TAM) resistance in breast cancer. How this phospho-modification affects ERa specificity and translates into TAM resistance is unclear. Here, we show that S305-P modification of ERa reprograms the receptor, redirecting it to new transcriptional start sites, thus modulating the transcriptome. By altering the chromatin-binding pattern, Ser305 phosphorylation of ERa translates into a 26-gene expression classifier that identifies breast cancer patients with a poor disease outcome after TAM treatment. MYC-target genes and networks were significantly enriched in this gene classifier that includes a number of selective targets for ERaS305-P. The enhanced expression of MYC increased cell proliferation in the presence of TAM. We demonstrate that activation of the PKA signaling pathway alters the transcriptome by redirecting ERa to new transcriptional start sites, resulting in altered transcription and TAM resistance.

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Immunohistochemical validation of multiple phospho-specific epitopes for estrogen receptor α (ERα) in tissue microarrays of ERα positive human breast carcinomas

Cited by 33 publications

References 23 publications

Estrogen Receptor-α Phosphorylation at Serine 305, Nuclear p21-Activated Kinase 1 Expression, and Response to Tamoxifen in Postmenopausal Breast Cancer

Estrogen Receptor-α Phosphorylation at Serine 305, Nuclear p21-Activated Kinase 1 Expression, and Response to Tamoxifen in Postmenopausal Breast Cancer

Estrogen Receptor Alpha Phosphorylated at Tyrosine 537 is Associated with Poor Clinical Outcome in Breast Cancer Patients Treated with Tamoxifen

PKA phosphorylation redirects ERα to promoters of a unique gene set to induce tamoxifen resistance

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