IJC International Journal of Cancer
Osteopontin is a useful predictor of bone metastasis and survival in patients with locally advanced nasopharyngeal carcinoma Xue Hou1,2,3*, Xuan Wu4*, Peiyu Huang2,3,5, Jianhua Zhan2,3, Ting Zhou1,2,3, Yuxiang Ma2,3,6, Tao Qin1,2,3, Rongzhen Luo2,3,7, Yanfen Feng2,3,7, Ying Xu8, Likun Chen1,2,3 and Li Zhang1,2,3 1
Department of Medical Oncology, Sun Yat-sen University Cancer Center, Guangzhou City, Guangdong Province, People’s Republic of China State Key Laboratory of Oncology in South China, Guangzhou City, Guangdong Province, People’s Republic of China 3 Collaborative Innovation Center for Cancer Medicine, Guangzhou City, Guangdong Province, People’s Republic of China 4 Department of Oncology, Peking University Shenzhen Hospital, Shenzhen City, Guangdong Province, People’s Republic of China 5 Department of Nasopharyngeal Carcinoma, Sun Yat-sen University Cancer Center, Guangzhou City, Guangdong Province, People’s Republic of China 6 Department of Clinical Research, Sun Yat-sen University Cancer Center, Guangzhou City, Guangdong Province, People’s Republic of China 7 Department of Pathology, Sun Yat-sen University Cancer Center, Guangzhou City, Guangdong Province, People’s Republic of China 8 Institute of Medical Statistics and Epidemiology, Sun Yat-sen University, Guangzhou City, Guangdong Province, People’s Republic of China
Early Detection and Diagnosis
2
Bone is the most common metastatic site in nasopharyngeal carcinoma (NPC). Osteopontin (OPN) and bone sialoprotein (BSP) are demonstrated to be involved in multiple steps of distant metastasis and correlate with bone metastasis (BM) in cancers. We aim to explore the impacts of OPN and BSP on the prognosis of the patients with locally advanced NPC. A tissue microarray including 162 locally advanced NPC specimens was generated for immunohistochemical evaluation. All of the patients received curative treatment. Twenty-two patients developed BM during follow-up. The OPN expression level was higher in patients with BM than in those without BM (p 5 0.005), whereas no significant difference of the BSP expression level was noted (p 5 0.634). Univariate analysis demonstrated that a higher level of OPN expression associated with a poorer 8-year metastasis-free survival (MFS) rate (p < 0.001), 8-year bone metastasis-free survival (BMFS) rate (93.6 vs. 87.5 vs. 64.5% for immunoreactivity score 1, 2 and 3, respectively; p 5 0.001) and median overall survival (OS) time (p < 0.001). Multivariate Cox analysis confirmed that high level of OPN expression was independent factor associated with decreased BMFS (p 5 0.02), MFS (p < 0.001) and OS (p < 0.001). Our findings indicate that OPN is a prognostic biomarker for BM and survival in patients with locally advanced NPC, and therefore it is useful in identifying the patients with an increased risk of cancer progression and BM to guide tailored therapy.
Nasopharyngeal carcinoma (NPC) is highly prevalent in South China.1–3 The development of radiotherapeutic techniques has provided significant survival benefits for patients with early-stage NPC, with a 5-year overall survival (OS) rate above 90%.4,5 However, for locally advanced disease, which constitutes approximately 70–80% of patients with NPC at the first diagnosis, many of them will develop distant
metastasis even after curative treatment,2,6,7 whereas the bone is the most common metastatic site.8,9 Therefore, it is important to select the patients with a high risk of bone metastasis (BM) to provide them with tailored therapy to improve long-term survival. Tumor–node–metastasis (TNM) staging is commonly used in the assessment of clinical outcomes in patients with
Key words: nasopharyngeal neoplasms, osteopontin, bone sialoprotein, bone metastasis, prognosis Abbreviations: AJCC: American Joint Committee on Cancer; BM: bone metastasis; BMFS: bone metastasis-free survival; BSP: bone sialoprotein; CCRT: concurrent chemoradiotherapy; CT: computed tomography; IHC: immunohistochemical; IRS: immunoreactivity score; MFS: metastasis-free survival; MRI: magnetic resonance imaging; NPC: nasopharyngeal carcinoma; OPN: osteopontin; OS: overall survival; SIBLING: small integrin-binding ligand N-linked glycoprotein; SYSUCC: Sun Yat-sen University Cancer Center; TNM: tumor– node–metastasis; UICC: Union Internationale Against Cancer *X.H. and X.W. contributed equally to this work and share the first authorship Grant sponsor: Guangdong Provincial Science and Technology Planning Foundation; Grant number: 2012B031800280; Grant sponsor: National High Technology Research and Development Program of China; Grant number: 2012AA02A502; Grant sponsor: Excellent Young Talent Project of Sun Yat-sen University Cancer Center; Grant number: 04140601. DOI: 10.1002/ijc.29540 History: Received 15 Jan 2015; Accepted 23 Mar 2015; Online 30 Mar 2015 Correspondence to: Li Zhang, Department of Medical Oncology, Sun Yat-sen University Cancer Center, 651 East Dongfeng Road, Guangzhou City, Guangdong Province 510060, People’s Republic of China, Tel.: +86-20-87343458, Fax: +86-20-87342480, E-mail: [email protected]
C 2015 UICC Int. J. Cancer: 137, 1672–1678 (2015) V
1673
Hou et al.
What’s new? Nasalpharyngeal carcinoma frequently metastasizes to the bones. These authors sought a way to identify which patients are most likely to get bone metastases? They found that patients with bone metastasis also had high levels of osteopontin, an integrin-binding glycophosphoprotein which has been shown in previous work to help cancer cells infiltrate the bone. Patients with lots of osteopontin also had shorter metastasis-free survival, suggesting that testing for it could help clinicians target and treat those patients most likely to suffer metastases.
Material and Methods Patients and tissue specimens
Our study was approved by the Ethics Committee of Sun Yat-sen University Cancer Center (SYSUCC), and informed consent was obtained from each participant. The data were obtained from an NPC database at SYSUCC between January 2001 and April 2005. All of the patients included in the analysis fit the following criteria: (i) histologically diagnosed as NPC; (ii) their disease was at Stage III–IVa at diagnosis; (iii) adequate tumor specimens were available for tissue microarray construction; (iv) they were administered curative therapies (induction chemotherapy and curative radiotherapy) and (v) they were under follow-up for at least 5 years. Clinical stage was determined according to the staging system of the American Joint Committee on Cancer (AJCC) and Union Internationale Against Cancer (UICC), seventh edition (AJCC/UICC-7). We excluded patients with a history of concurrent malignant disease or other previous primary cancers. As a result, 162 cases fit the inclusion criteria and were included in our study. Table 1 summarizes the detailed information of the included cases. All of the patients were administered two to three cycles of cisplatin plus 5-Fu induction chemotherapy and then underwent a uniform conventional C 2015 UICC Int. J. Cancer: 137, 1672–1678 (2015) V
two-dimensional radiotherapy protocol in accordance with the treatment policy for NPC at SYSUCC. Paraffin-embedded primary tumor tissues of the 162 patients were obtained to construct the NPC tissue microarray. Hematoxylin- and eosin-stained sections were made from each block to define representative tumor regions. For each patient’s sample, two tissue cylinders were taken from different representative tumor regions. Tissue cylinders with a diameter of 1 mm were punched from selected areas of each “donor” block using the Manual Tissue Arrayer (Beecher Instruments, Silver Spring, MD), and arrayed into a recipient paraffin block. Serial 5-lm sections were mounted on silane-coated slides for hematoxylin and eosin staining and immunohistochemical (IHC) staining. OPN and BSP IHC and scoring
IHC staining was performed to assess the protein expression of OPN and BSP. The detailed procedure was similar to that of our previous study.15 The monoclonal antibody concentrations were 1:100 for OPN (#sc-21742, Santa Cruz Biotechnology, Santa Cruz, CA) and 1:2,000 for BSP (#MAB 1061, Chemicon, Temecula, CA). Two independent observers (Luo and Feng) blinded to the clinical information determined the immunoreactivity score (IRS) for IHC staining. Semi-quantitative scoring, based on the percentage of positive tumor cells, was used to assess the IRS. For protein OPN, IRS 1 denotes no positive tumor cell, IRS 2 denotes the percentage of positive tumor cells
Osteopontin is a useful predictor of bone metastasis and survival in patients with locally advanced nasopharyngeal carcinoma Xue Hou1,2,3*, Xuan Wu4*, Peiyu Huang2,3,5, Jianhua Zhan2,3, Ting Zhou1,2,3, Yuxiang Ma2,3,6, Tao Qin1,2,3, Rongzhen Luo2,3,7, Yanfen Feng2,3,7, Ying Xu8, Likun Chen1,2,3 and Li Zhang1,2,3 1
Department of Medical Oncology, Sun Yat-sen University Cancer Center, Guangzhou City, Guangdong Province, People’s Republic of China State Key Laboratory of Oncology in South China, Guangzhou City, Guangdong Province, People’s Republic of China 3 Collaborative Innovation Center for Cancer Medicine, Guangzhou City, Guangdong Province, People’s Republic of China 4 Department of Oncology, Peking University Shenzhen Hospital, Shenzhen City, Guangdong Province, People’s Republic of China 5 Department of Nasopharyngeal Carcinoma, Sun Yat-sen University Cancer Center, Guangzhou City, Guangdong Province, People’s Republic of China 6 Department of Clinical Research, Sun Yat-sen University Cancer Center, Guangzhou City, Guangdong Province, People’s Republic of China 7 Department of Pathology, Sun Yat-sen University Cancer Center, Guangzhou City, Guangdong Province, People’s Republic of China 8 Institute of Medical Statistics and Epidemiology, Sun Yat-sen University, Guangzhou City, Guangdong Province, People’s Republic of China
Early Detection and Diagnosis
2
Bone is the most common metastatic site in nasopharyngeal carcinoma (NPC). Osteopontin (OPN) and bone sialoprotein (BSP) are demonstrated to be involved in multiple steps of distant metastasis and correlate with bone metastasis (BM) in cancers. We aim to explore the impacts of OPN and BSP on the prognosis of the patients with locally advanced NPC. A tissue microarray including 162 locally advanced NPC specimens was generated for immunohistochemical evaluation. All of the patients received curative treatment. Twenty-two patients developed BM during follow-up. The OPN expression level was higher in patients with BM than in those without BM (p 5 0.005), whereas no significant difference of the BSP expression level was noted (p 5 0.634). Univariate analysis demonstrated that a higher level of OPN expression associated with a poorer 8-year metastasis-free survival (MFS) rate (p < 0.001), 8-year bone metastasis-free survival (BMFS) rate (93.6 vs. 87.5 vs. 64.5% for immunoreactivity score 1, 2 and 3, respectively; p 5 0.001) and median overall survival (OS) time (p < 0.001). Multivariate Cox analysis confirmed that high level of OPN expression was independent factor associated with decreased BMFS (p 5 0.02), MFS (p < 0.001) and OS (p < 0.001). Our findings indicate that OPN is a prognostic biomarker for BM and survival in patients with locally advanced NPC, and therefore it is useful in identifying the patients with an increased risk of cancer progression and BM to guide tailored therapy.
Nasopharyngeal carcinoma (NPC) is highly prevalent in South China.1–3 The development of radiotherapeutic techniques has provided significant survival benefits for patients with early-stage NPC, with a 5-year overall survival (OS) rate above 90%.4,5 However, for locally advanced disease, which constitutes approximately 70–80% of patients with NPC at the first diagnosis, many of them will develop distant
metastasis even after curative treatment,2,6,7 whereas the bone is the most common metastatic site.8,9 Therefore, it is important to select the patients with a high risk of bone metastasis (BM) to provide them with tailored therapy to improve long-term survival. Tumor–node–metastasis (TNM) staging is commonly used in the assessment of clinical outcomes in patients with
Key words: nasopharyngeal neoplasms, osteopontin, bone sialoprotein, bone metastasis, prognosis Abbreviations: AJCC: American Joint Committee on Cancer; BM: bone metastasis; BMFS: bone metastasis-free survival; BSP: bone sialoprotein; CCRT: concurrent chemoradiotherapy; CT: computed tomography; IHC: immunohistochemical; IRS: immunoreactivity score; MFS: metastasis-free survival; MRI: magnetic resonance imaging; NPC: nasopharyngeal carcinoma; OPN: osteopontin; OS: overall survival; SIBLING: small integrin-binding ligand N-linked glycoprotein; SYSUCC: Sun Yat-sen University Cancer Center; TNM: tumor– node–metastasis; UICC: Union Internationale Against Cancer *X.H. and X.W. contributed equally to this work and share the first authorship Grant sponsor: Guangdong Provincial Science and Technology Planning Foundation; Grant number: 2012B031800280; Grant sponsor: National High Technology Research and Development Program of China; Grant number: 2012AA02A502; Grant sponsor: Excellent Young Talent Project of Sun Yat-sen University Cancer Center; Grant number: 04140601. DOI: 10.1002/ijc.29540 History: Received 15 Jan 2015; Accepted 23 Mar 2015; Online 30 Mar 2015 Correspondence to: Li Zhang, Department of Medical Oncology, Sun Yat-sen University Cancer Center, 651 East Dongfeng Road, Guangzhou City, Guangdong Province 510060, People’s Republic of China, Tel.: +86-20-87343458, Fax: +86-20-87342480, E-mail: [email protected]
C 2015 UICC Int. J. Cancer: 137, 1672–1678 (2015) V
1673
Hou et al.
What’s new? Nasalpharyngeal carcinoma frequently metastasizes to the bones. These authors sought a way to identify which patients are most likely to get bone metastases? They found that patients with bone metastasis also had high levels of osteopontin, an integrin-binding glycophosphoprotein which has been shown in previous work to help cancer cells infiltrate the bone. Patients with lots of osteopontin also had shorter metastasis-free survival, suggesting that testing for it could help clinicians target and treat those patients most likely to suffer metastases.
Material and Methods Patients and tissue specimens
Our study was approved by the Ethics Committee of Sun Yat-sen University Cancer Center (SYSUCC), and informed consent was obtained from each participant. The data were obtained from an NPC database at SYSUCC between January 2001 and April 2005. All of the patients included in the analysis fit the following criteria: (i) histologically diagnosed as NPC; (ii) their disease was at Stage III–IVa at diagnosis; (iii) adequate tumor specimens were available for tissue microarray construction; (iv) they were administered curative therapies (induction chemotherapy and curative radiotherapy) and (v) they were under follow-up for at least 5 years. Clinical stage was determined according to the staging system of the American Joint Committee on Cancer (AJCC) and Union Internationale Against Cancer (UICC), seventh edition (AJCC/UICC-7). We excluded patients with a history of concurrent malignant disease or other previous primary cancers. As a result, 162 cases fit the inclusion criteria and were included in our study. Table 1 summarizes the detailed information of the included cases. All of the patients were administered two to three cycles of cisplatin plus 5-Fu induction chemotherapy and then underwent a uniform conventional C 2015 UICC Int. J. Cancer: 137, 1672–1678 (2015) V
two-dimensional radiotherapy protocol in accordance with the treatment policy for NPC at SYSUCC. Paraffin-embedded primary tumor tissues of the 162 patients were obtained to construct the NPC tissue microarray. Hematoxylin- and eosin-stained sections were made from each block to define representative tumor regions. For each patient’s sample, two tissue cylinders were taken from different representative tumor regions. Tissue cylinders with a diameter of 1 mm were punched from selected areas of each “donor” block using the Manual Tissue Arrayer (Beecher Instruments, Silver Spring, MD), and arrayed into a recipient paraffin block. Serial 5-lm sections were mounted on silane-coated slides for hematoxylin and eosin staining and immunohistochemical (IHC) staining. OPN and BSP IHC and scoring
IHC staining was performed to assess the protein expression of OPN and BSP. The detailed procedure was similar to that of our previous study.15 The monoclonal antibody concentrations were 1:100 for OPN (#sc-21742, Santa Cruz Biotechnology, Santa Cruz, CA) and 1:2,000 for BSP (#MAB 1061, Chemicon, Temecula, CA). Two independent observers (Luo and Feng) blinded to the clinical information determined the immunoreactivity score (IRS) for IHC staining. Semi-quantitative scoring, based on the percentage of positive tumor cells, was used to assess the IRS. For protein OPN, IRS 1 denotes no positive tumor cell, IRS 2 denotes the percentage of positive tumor cells
