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Epidermal growth factor (EGF) receptor-ligand based molecular staging predicts prognosis in head and neck squamous cell carcinoma partly due to deregulated EGF- induced amphiregulin expression

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ABSTRACT

Background: Increased expression of epidermal growth factor receptor (EGFR) and its ligands is associated with poor prognosis and chemoresistance in many carcinoma types, but its role in head and neck squamous cell carcinoma (HNSCC) is unclear. Our aim was to clarify whether mRNA expression of EGFR-ligands was linked to prognosis and cisplatin resistance, and if so, which ligand was most important and how was the expression regulated.

Methods: To examine the prognostic effect of EGFR-ligand expression, we analyzed tumorous mRNA expression in 399 HNSCC patients. The intracellular signaling pathways controlling epidermal growth factor (EGF)-induced amphiregulin (AREG) expression were examined in three oral squamous cell carcinoma (OSCC) cell lines. Effect of AREG on cisplatin resistance was examined by viability assays in four-, and by association in 11 OSCC cell lines.

Results: The patients were divided into five groups according to the median mRNA expression levels of four EGFR ligands, i.e. AREG, EGF, heparin-binding EGF-like growth factor (HBEGF) and beta-cellulin (BTC). The number of increased-expressed EGFR-ligands were progressively correlated to five-year survival, even in advanced TNM-stage IV patients, where five-year mortality increased from 26 % if tumor expressed none to one EGFR-ligand, to 45 % in three to four ligand expressing tumors. Thus, staging the tumor according to these EGFR-ligand mRNA expression pattern completely out performed TNM staging in predicting prognosis. Multivariate analysis identified AREG as the dominating predictor, and AREG was overexpressed in OSCC compared to tumors from other sites. Both EGF and HBEGF stimulation induced strong AREG increase in OSCC cell lines, which was partially mediated by the extracellular signal-regulated kinase 1/2 pathway, and negatively regulated by p38, c-Jun N-terminal kinase, and phosphoinositide-3 kinase. Although increased AREG mRNA expression predicted unfavorable prognosis in platinum treated HNSCC patients, AREG did not mediate cisplatin resistance in the OSCC cell lines.

Conclusions: Increased tumorous mRNA expression of four EGFR ligands was progressively associated with poor prognosis in HNSCC. Thus, EGFR-ligands mRNA expression pattern may be a new prognostic biomarker. The tightly regulated EGF-induced AREG mRNA expression was partly lost in the OSCC cell lines and restoring its regulation may be a new target in cancer treatment.

Trial registration: Not applicable as the clinical data of the 498 HNSCC patients and their mRNA expression profiles were collected from the open TCGA database: http://cancergenome.nih.gov/cancersselected/headandneck.

No MeSH data available.


Related in: MedlinePlus

Cisplatin resistance was independent of AREG expression in 11 OSCC cell lines. a Neither exogenous EGF nor AREG affected cisplatin resistance in OSCC cell lines. Cells were grown in the presence (+) or absence (−) of human recombinant EGF (50 ng/ml), human recombinant AREG (100 ng/ml) for 24 h and then treated with (+) or without (−) 10 μM cisplatin (NS: not significant. p > 0.05, student’s t-test). b Cisplatin IC50 value did not correlate to AREG mRNA expression (left) or protein production (right) in 11 OSCC cell lines. The cisplatin IC50 value in sensitive (n = 4) or resistant (n = 7) OSCC cell lines were plotted against AREG mRNA expression (left) or 24 h AREG production (right). The cell lines were clustered into cisplatin sensitive (IC50 < 3.5 μM) or resistant (IC50 > 8.5 μM). Median indicated with horizontal lines (Wilcoxon rank-sum test)
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Fig6: Cisplatin resistance was independent of AREG expression in 11 OSCC cell lines. a Neither exogenous EGF nor AREG affected cisplatin resistance in OSCC cell lines. Cells were grown in the presence (+) or absence (−) of human recombinant EGF (50 ng/ml), human recombinant AREG (100 ng/ml) for 24 h and then treated with (+) or without (−) 10 μM cisplatin (NS: not significant. p > 0.05, student’s t-test). b Cisplatin IC50 value did not correlate to AREG mRNA expression (left) or protein production (right) in 11 OSCC cell lines. The cisplatin IC50 value in sensitive (n = 4) or resistant (n = 7) OSCC cell lines were plotted against AREG mRNA expression (left) or 24 h AREG production (right). The cell lines were clustered into cisplatin sensitive (IC50 < 3.5 μM) or resistant (IC50 > 8.5 μM). Median indicated with horizontal lines (Wilcoxon rank-sum test)

Mentions: Patients with high AREG expression had poor clinical response to cisplatin treatment (Fig. 3a), suggesting that AREG increased cisplatin resistance in the HNSCC, as revealed for several other carcinoma types [17, 18]. However, neither EGF nor AREG increased cisplatin resistance in any of the four cisplatin sensitive cell lines (Fig. 6a). Moreover, neither AREG mRNA nor protein expression was associated with cisplatin resistance in the 11 OSCC cell lines (Fig. 6b), and cisplatin treatment did not change the AREG mRNA expression or secretion (data not shown).Fig. 6


Epidermal growth factor (EGF) receptor-ligand based molecular staging predicts prognosis in head and neck squamous cell carcinoma partly due to deregulated EGF- induced amphiregulin expression
Cisplatin resistance was independent of AREG expression in 11 OSCC cell lines. a Neither exogenous EGF nor AREG affected cisplatin resistance in OSCC cell lines. Cells were grown in the presence (+) or absence (−) of human recombinant EGF (50 ng/ml), human recombinant AREG (100 ng/ml) for 24 h and then treated with (+) or without (−) 10 μM cisplatin (NS: not significant. p > 0.05, student’s t-test). b Cisplatin IC50 value did not correlate to AREG mRNA expression (left) or protein production (right) in 11 OSCC cell lines. The cisplatin IC50 value in sensitive (n = 4) or resistant (n = 7) OSCC cell lines were plotted against AREG mRNA expression (left) or 24 h AREG production (right). The cell lines were clustered into cisplatin sensitive (IC50 < 3.5 μM) or resistant (IC50 > 8.5 μM). Median indicated with horizontal lines (Wilcoxon rank-sum test)
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Fig6: Cisplatin resistance was independent of AREG expression in 11 OSCC cell lines. a Neither exogenous EGF nor AREG affected cisplatin resistance in OSCC cell lines. Cells were grown in the presence (+) or absence (−) of human recombinant EGF (50 ng/ml), human recombinant AREG (100 ng/ml) for 24 h and then treated with (+) or without (−) 10 μM cisplatin (NS: not significant. p > 0.05, student’s t-test). b Cisplatin IC50 value did not correlate to AREG mRNA expression (left) or protein production (right) in 11 OSCC cell lines. The cisplatin IC50 value in sensitive (n = 4) or resistant (n = 7) OSCC cell lines were plotted against AREG mRNA expression (left) or 24 h AREG production (right). The cell lines were clustered into cisplatin sensitive (IC50 < 3.5 μM) or resistant (IC50 > 8.5 μM). Median indicated with horizontal lines (Wilcoxon rank-sum test)
Mentions: Patients with high AREG expression had poor clinical response to cisplatin treatment (Fig. 3a), suggesting that AREG increased cisplatin resistance in the HNSCC, as revealed for several other carcinoma types [17, 18]. However, neither EGF nor AREG increased cisplatin resistance in any of the four cisplatin sensitive cell lines (Fig. 6a). Moreover, neither AREG mRNA nor protein expression was associated with cisplatin resistance in the 11 OSCC cell lines (Fig. 6b), and cisplatin treatment did not change the AREG mRNA expression or secretion (data not shown).Fig. 6

View Article: PubMed Central - PubMed

ABSTRACT

Background: Increased expression of epidermal growth factor receptor (EGFR) and its ligands is associated with poor prognosis and chemoresistance in many carcinoma types, but its role in head and neck squamous cell carcinoma (HNSCC) is unclear. Our aim was to clarify whether mRNA expression of EGFR-ligands was linked to prognosis&nbsp;and cisplatin resistance, and if so, which ligand was most important and how was the expression regulated.

Methods: To examine the prognostic effect of EGFR-ligand expression, we analyzed tumorous mRNA expression in 399 HNSCC patients. The intracellular signaling pathways controlling epidermal growth factor (EGF)-induced amphiregulin (AREG) expression were examined in three oral squamous cell carcinoma (OSCC) cell lines. Effect of AREG on cisplatin resistance was examined by viability assays in four-, and by association in 11 OSCC cell lines.

Results: The patients were divided into five groups according to the median mRNA expression levels of four EGFR ligands, i.e. AREG, EGF, heparin-binding EGF-like growth factor (HBEGF) and beta-cellulin (BTC). The number of increased-expressed EGFR-ligands were progressively correlated to five-year survival, even in advanced TNM-stage IV patients, where five-year mortality increased from 26&nbsp;% if tumor expressed none to one EGFR-ligand, to 45&nbsp;% in three to four ligand expressing tumors. Thus, staging the tumor according to these EGFR-ligand mRNA expression pattern completely out performed TNM staging in predicting prognosis. Multivariate analysis identified AREG as the dominating predictor, and AREG was overexpressed in OSCC compared to tumors from other sites. Both EGF and HBEGF stimulation induced strong AREG increase in OSCC cell lines, which was partially mediated by the extracellular signal-regulated kinase 1/2 pathway, and negatively regulated by p38, c-Jun N-terminal kinase, and phosphoinositide-3 kinase. Although increased AREG mRNA expression predicted unfavorable prognosis in platinum treated HNSCC patients, AREG did not mediate cisplatin resistance in the OSCC cell lines.

Conclusions: Increased tumorous mRNA expression of four EGFR ligands was progressively associated with poor prognosis in HNSCC. Thus, EGFR-ligands mRNA expression pattern may be a new prognostic biomarker. The tightly regulated EGF-induced AREG mRNA expression was partly lost in the OSCC cell lines and restoring its regulation may be a new target in cancer treatment.

Trial registration: Not applicable as the clinical data of the 498 HNSCC patients and their mRNA expression profiles were collected from the open TCGA database: http://cancergenome.nih.gov/cancersselected/headandneck.

No MeSH data available.


Related in: MedlinePlus