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Anterior gradient protein 2 expression in high grade head and neck squamous cell carcinoma correlated with cancer stem cell and epithelial mesenchymal transition.

Ma SR, Wang WM, Huang CF, Zhang WF, Sun ZJ - Oncotarget (2015)

Bottom Line: Anterior gradient protein 2 (AGR2) is a novel biomarker with potential oncogenic role.We found that AGR2 protein levels were higher in HNSCC than in normal oral mucosa.These results suggest that AGR2 is involved in EMT and self-renewal of CSC and may present a potential therapeutic target (oncotarget) for HNSCC.

View Article: PubMed Central - PubMed

Affiliation: The State Key Laboratory Breeding Base of Basic Science of Stomatology & Key Laboratory of Oral Biomedicine Ministry of Education, Wuhan University, Wuhan, China.

ABSTRACT
Anterior gradient protein 2 (AGR2) is a novel biomarker with potential oncogenic role. We sought to investigate the diagnostic and prognostic role of AGR2 on head and neck squamous cell carcinoma (HNSCC) with an emphasis on its correlation of cancer stemloid cells (CSC) and epithelial mesenchymal transition (EMT). We found that AGR2 protein levels were higher in HNSCC than in normal oral mucosa. High levels of AGR2 were associated with the T category, pathological grade and lymph node metastasis of HNSCC. Expression of AGR2 increased in recurring HNSCC after radiotherapy and in post cisplatin-based chemotherapeutic tissues. In HNSCC cell lines, knock-down of AGR2 induced apoptosis, reduced sphere formation, and down-regulated Survivin, Cyclin D1, Bcl2, Bcl2l1, Slug, Snail, Nanog and Oct4. In addition, over-expressed AGR2 in transgenic mice with spontaneous HNSCC was associated with lost function of Tgfbr1 and/ or lost function of Pten. In vitro knockdown TGFBR1 in HNSCC cell lines increased AGR2 expression. These results suggest that AGR2 is involved in EMT and self-renewal of CSC and may present a potential therapeutic target (oncotarget) for HNSCC.

No MeSH data available.


Related in: MedlinePlus

AGR2 expression in human head neck cancer(A) AGR2 DNA copy number from Peng's dataset and (B) mRNA expression from Ginos's dataset (as log2 median –centered ratio) for head neck cancer versus normal counterpart shown as raw data. (C) AGR2 DNA copy number of TCGA head neck cancer as shown as raw data. (D) Representative immunohistochemistry staining of AGR2 in oral mucosa as well as in HNSCC tissue with quantification in (E, One way ANOVA, P <0.001). Dys, dysplasia; SCC, squamous cell carcinoma; ***, P <0.001.
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Figure 1: AGR2 expression in human head neck cancer(A) AGR2 DNA copy number from Peng's dataset and (B) mRNA expression from Ginos's dataset (as log2 median –centered ratio) for head neck cancer versus normal counterpart shown as raw data. (C) AGR2 DNA copy number of TCGA head neck cancer as shown as raw data. (D) Representative immunohistochemistry staining of AGR2 in oral mucosa as well as in HNSCC tissue with quantification in (E, One way ANOVA, P <0.001). Dys, dysplasia; SCC, squamous cell carcinoma; ***, P <0.001.

Mentions: To determine whether AGR2 expression was associated with HNSCC in humans, we interrogated the Tissue Cancer Genome Atlas dataset [23] and Oncomine database [24]. In a meta-analysis of gene expression profiling, increased AGR2 DNA copy number of mRNA expression was significantly increased in many cancers (e.g., esophagus, thyroid, ovarian, pancreatic, breast, prostate, lung, and HNSCC) compared with the normal counterpart (P < 0.001, Supplementary Fig. 1), thereby suggesting that AGR2 may act as an oncogene in human cancer cells. In the Peng et al. dataset [25], which independently performed DNA copy number analysis on oral squamous cell carcinoma, 38 out of 112 OSCCs showed amplification of AGR2 copy number (Fig. 1A). Through analyze the raw data of Ginos et al. dataset [26] using Oncomine, we found the significantly enhanced mRNA expression of AGR2 in 21 out of 54 HNSCCs compared with normal oral mucosa (Fig. 1B). TCGA data sheet indicated an increase in the DNA copy number of HNSCC (n = 290) compared with the normal counterpart (blood, n = 338, Fig. 1C). Furthermore, to assess the protein expression of AGR2 in human HNSCC tissues, we performed immunohistochemistry in human HNSCC tissue microarray (Fig. 1D). AGR2 exhibited high expression in the cytoplasm and membrane of the cancer cells (Fig. 1D). This analysis showed significantly increased immunoreactivity of AGR2 in HNSCC (n = 59) compared with dysplasia (n = 13, P < 0.001) and normal oral mucosa (n = 39, P < 0.001, Fig. 1D with quantification in Fig. 1E). High-grade (Grades II and III) HNSCC presented intense AGR2 immunoreactivity compared with low-grade (Grade I, n=20) HNSCC (P < 0.001, Fig. 2A with quantities in Fig. 2B). The expression of AGR2 was also associated with tumor size and with increased AGR2 expression in T2 (n = 37) and T3 category (n = 13) compared with the T1 category (n = 9, Fig. 2C). We also noted a remarkable increase in AGR2 immunoreactivity in the original tumor of the pathological lymph node-positive patient (pN1, n = 20, P < 0.01, Fig. 2D) compared with the pathological lymph node-negative patient (pN0, n = 39). The results above indicated that AGR2 protein expression correlated with high-grade HNSCC. To further explore the prognosis value of AGR2 in HNSCC, Kaplan–Meier method was used. As shown in Supplementary Fig. S2, high AGR2 expression may indicate a rather poor prognosis of HNSCC patient, whereas log-Rank analysis indicated that cumulative rate of the patients with high AGR2 (P = 0.1161, n = 29) expression did not reach statistical significance.


Anterior gradient protein 2 expression in high grade head and neck squamous cell carcinoma correlated with cancer stem cell and epithelial mesenchymal transition.

Ma SR, Wang WM, Huang CF, Zhang WF, Sun ZJ - Oncotarget (2015)

AGR2 expression in human head neck cancer(A) AGR2 DNA copy number from Peng's dataset and (B) mRNA expression from Ginos's dataset (as log2 median –centered ratio) for head neck cancer versus normal counterpart shown as raw data. (C) AGR2 DNA copy number of TCGA head neck cancer as shown as raw data. (D) Representative immunohistochemistry staining of AGR2 in oral mucosa as well as in HNSCC tissue with quantification in (E, One way ANOVA, P <0.001). Dys, dysplasia; SCC, squamous cell carcinoma; ***, P <0.001.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4496185&req=5

Figure 1: AGR2 expression in human head neck cancer(A) AGR2 DNA copy number from Peng's dataset and (B) mRNA expression from Ginos's dataset (as log2 median –centered ratio) for head neck cancer versus normal counterpart shown as raw data. (C) AGR2 DNA copy number of TCGA head neck cancer as shown as raw data. (D) Representative immunohistochemistry staining of AGR2 in oral mucosa as well as in HNSCC tissue with quantification in (E, One way ANOVA, P <0.001). Dys, dysplasia; SCC, squamous cell carcinoma; ***, P <0.001.
Mentions: To determine whether AGR2 expression was associated with HNSCC in humans, we interrogated the Tissue Cancer Genome Atlas dataset [23] and Oncomine database [24]. In a meta-analysis of gene expression profiling, increased AGR2 DNA copy number of mRNA expression was significantly increased in many cancers (e.g., esophagus, thyroid, ovarian, pancreatic, breast, prostate, lung, and HNSCC) compared with the normal counterpart (P < 0.001, Supplementary Fig. 1), thereby suggesting that AGR2 may act as an oncogene in human cancer cells. In the Peng et al. dataset [25], which independently performed DNA copy number analysis on oral squamous cell carcinoma, 38 out of 112 OSCCs showed amplification of AGR2 copy number (Fig. 1A). Through analyze the raw data of Ginos et al. dataset [26] using Oncomine, we found the significantly enhanced mRNA expression of AGR2 in 21 out of 54 HNSCCs compared with normal oral mucosa (Fig. 1B). TCGA data sheet indicated an increase in the DNA copy number of HNSCC (n = 290) compared with the normal counterpart (blood, n = 338, Fig. 1C). Furthermore, to assess the protein expression of AGR2 in human HNSCC tissues, we performed immunohistochemistry in human HNSCC tissue microarray (Fig. 1D). AGR2 exhibited high expression in the cytoplasm and membrane of the cancer cells (Fig. 1D). This analysis showed significantly increased immunoreactivity of AGR2 in HNSCC (n = 59) compared with dysplasia (n = 13, P < 0.001) and normal oral mucosa (n = 39, P < 0.001, Fig. 1D with quantification in Fig. 1E). High-grade (Grades II and III) HNSCC presented intense AGR2 immunoreactivity compared with low-grade (Grade I, n=20) HNSCC (P < 0.001, Fig. 2A with quantities in Fig. 2B). The expression of AGR2 was also associated with tumor size and with increased AGR2 expression in T2 (n = 37) and T3 category (n = 13) compared with the T1 category (n = 9, Fig. 2C). We also noted a remarkable increase in AGR2 immunoreactivity in the original tumor of the pathological lymph node-positive patient (pN1, n = 20, P < 0.01, Fig. 2D) compared with the pathological lymph node-negative patient (pN0, n = 39). The results above indicated that AGR2 protein expression correlated with high-grade HNSCC. To further explore the prognosis value of AGR2 in HNSCC, Kaplan–Meier method was used. As shown in Supplementary Fig. S2, high AGR2 expression may indicate a rather poor prognosis of HNSCC patient, whereas log-Rank analysis indicated that cumulative rate of the patients with high AGR2 (P = 0.1161, n = 29) expression did not reach statistical significance.

Bottom Line: Anterior gradient protein 2 (AGR2) is a novel biomarker with potential oncogenic role.We found that AGR2 protein levels were higher in HNSCC than in normal oral mucosa.These results suggest that AGR2 is involved in EMT and self-renewal of CSC and may present a potential therapeutic target (oncotarget) for HNSCC.

View Article: PubMed Central - PubMed

Affiliation: The State Key Laboratory Breeding Base of Basic Science of Stomatology & Key Laboratory of Oral Biomedicine Ministry of Education, Wuhan University, Wuhan, China.

ABSTRACT
Anterior gradient protein 2 (AGR2) is a novel biomarker with potential oncogenic role. We sought to investigate the diagnostic and prognostic role of AGR2 on head and neck squamous cell carcinoma (HNSCC) with an emphasis on its correlation of cancer stemloid cells (CSC) and epithelial mesenchymal transition (EMT). We found that AGR2 protein levels were higher in HNSCC than in normal oral mucosa. High levels of AGR2 were associated with the T category, pathological grade and lymph node metastasis of HNSCC. Expression of AGR2 increased in recurring HNSCC after radiotherapy and in post cisplatin-based chemotherapeutic tissues. In HNSCC cell lines, knock-down of AGR2 induced apoptosis, reduced sphere formation, and down-regulated Survivin, Cyclin D1, Bcl2, Bcl2l1, Slug, Snail, Nanog and Oct4. In addition, over-expressed AGR2 in transgenic mice with spontaneous HNSCC was associated with lost function of Tgfbr1 and/ or lost function of Pten. In vitro knockdown TGFBR1 in HNSCC cell lines increased AGR2 expression. These results suggest that AGR2 is involved in EMT and self-renewal of CSC and may present a potential therapeutic target (oncotarget) for HNSCC.

No MeSH data available.


Related in: MedlinePlus