Cancer Letters

Cancer Letters

Volume 253, Issue 1, 8 August 2007, Pages 34-42
Cancer Letters

Recombination of CXCR4, VEGF, and MMP-9 predicting lymph node metastasis in human breast cancer

https://doi.org/10.1016/j.canlet.2007.01.005Get rights and content

Abstract

We investigated the expression of CXC chemokine receptor 4 (CXCR4), vascular endothelial growth factor (VEGF) and matrix metalloproteinase (MMP)-9 to elucidate whether these markers could predict lymph node metastasis in human breast cancer. Higher rates of CXCR4 (61%), VEGF (68%), and MMP-9 (63%) expression were found in breast cancer tissues than in normal and atypical hyperplasia tissues. The expression of these markers was significantly associated with primary tumor progression, histological grade, and lymph node status. We found there were significant correlations between the expressions of any two of the three markers (P < 0.001). Furthermore, our studies indicated that concomitant expression of CXCR4/VEGF (P = 0.007), CXCR4/MMP-9 (P < 0.001) or VEGF/MMP-9 (P = 0.003) had stronger correlation with lymph node metastasis than did each alone and that combined expression of all three makers strongly correlated with lymph node metastasis (P < 0.001). Thus, simultaneously examining the expression of CXCR4, VEGF, and MMP-9 in cancer tissues of breast cancer will provide valuable prognostic diagnosis of lymph node metastasis.

Introduction

Malignant metastasis in patients with breast cancer usually involves in regional lymph nodes, bone marrow and liver, resulting in significant morbidity and mortality. Tumor metastasis consists of complex processes including degradation of extracellular matrix (EM), intravasation into lymphatic or blood vessels, clonogenic growth in secondary sites and neoangiogenesis [1]. A number of molecules such as cytokines, growth factors and matrix metalloproteinases (MMPs) have been implicated in the lymph node metastasis of cancer [2]. However, whether these metastasis-related molecules can be used as prognostic markers for such metastasis of breast cancer has yet to be evaluated.

Chemokine is a superfamily of small cytokines with the ability to chemoattract cells to the target tissues. CXC chemokine receptor 4 (CXCR4)–CXCL12 interaction plays an important role in the regulation of hematopoiesis, migration of hematopoietic cells and angiogenesis [3], [4]. Accumulating evidence indicates that chemokines and their receptors regulate homing and proliferation of cancer cells at specific secondary sites. It has been shown that human malignant breast tumors and metastases expressed increased levels of chemokine receptor CXCR4. By comparison, the CXCR4 ligand CXCL12 is abundantly expressed in the local cancer site and draining lymph node [5]. Ex vivo studies further demonstrated that the metastasis of breast cancer cells with CXCR4 gene deficiency was impaired [6]. Moreover, it was shown that CXCR4 was highly expressed in a number of human tumors such as oral squamous cell carcinoma, colorectal cancer and breast cancer and was an important prognostic factor in patients. [7], [8], [9].

Angiogenesis has been known to play an important role in the development of tumor growth and metastasis [10]. Vascular endothelial growth factor (VEGF) potently increases vascular permeability and promotes the formation of new blood vessels in tumor and thus is regarded as the main growth stimulatory factor in the tumor-related angiogenesis [11]. The prognostic value of high expression of VEGF has been demonstrated in various types of human tumors [12], [13]. Most recent studies demonstrated that stimulation of CXCR4 enhanced the secretion VEGF, leading to accelerated angiogenesis in tumors and the subsequent tumor metastasis [14], [15], [16]. These observations suggest that the expression level of CXCR4 and VEGF may be an appropriate biomarker for predicting the metastatic ability of tumor cells.

Tumor metastasis also involves the degradation of EM [17]. EM degradation is largely mediated by several types of proteolysis, but MMPs appear to be the most potent players in this process. In addition, MMPs such as MMP-9, MMP-2, and MMP-3 have been implicated in regulating angiogenesis and tumor progression [18], [19]. Notably, MMP-9 also plays an important role in tumor angiogenesis via signal pathways related to the VEGF/VEGF receptor system [20]. All these data suggest that combined evaluation of the expression of CXCR4, VEGF, and MMP-9 in tumor will be useful for predicting the probability of malignant metastasis.

In this study, we demonstrated that high expression of CXCR4, VEGF, and MMP-9 had a significant correlation with clinicopathologic factors and lymph node status in a series of breast cancer patients undergoing surgery. Furthermore, we found that concomitant expression of any two of these molecules had a stronger correlation with lymph node metastasis than did each of the three markers alone and that combined expression of all three markers strongly correlated with lymph node metastasis. Thus, recombination of CXCR4, VEGF, and MMP-9 will prove to be valuable prognostic markers for predicting the metastatic ability of breast cancer cells.

Section snippets

Patients and specimens

Sections of paraffin-embedded tissue samples were provided by the Department of Pathology, Pudong New Area People’s Hospital, Shanghai, China. They were obtained from patients with breast cancer (n = 84) and included 76 breast cancer tissue samples and eight normal tissue samples adjacent to mammary tumor. There were also six atypical hyperplasia samples. All patients underwent curative-intent surgery at the Department of Surgery from 2002 to 2005. The clinical, epidemiologic, and histopathologic

CXCR4, VEGF, and MMP-9 immunostaining in different mammary lesions

CXCR4 exhibited cytoplasmic and nuclear staining. Normal tissue adjacent to tumor cells occasionally showed weak CXCR4 staining that was mostly cytoplasmic. All six atypical hyperplasia cases showed positive but weak CXCR4 staining. Compared with benign lesions, 46 cancer cases (61%) showed high CXCR4 expression (Fig. 1A and B). VEGF immunoreactivity, localized to the cytoplasm, was weak in eight normal and six atypical hyperplasia samples, but the staining was much stronger in 52 cancer cases

Discussion

In this study, we found that the high expression of CXCR4, VEGF and MMP-9, which had been demonstrated to be associated with increased metastatic potential in cancer in ex vivo and animal studies, were valuable predictive biomarkers of lymph node metastases in human breast cancer. Furthermore, concomitant expression of three markers was significantly associated with an increased risk of lymph node metastasis.

Malignant tumor metastasis is the result of a series of complex processes that depend

Acknowledgments

We express our gratitude to Drs. Qiang Liu (Department of Pathology, Renji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China) and Dengshan Wang (Department of Pathology, Pudong New Area People’s Hospital, Shanghai, China) for their help in the immunohistochemical assay and Dr. Hongmei Li (Soochow University, Suzhou, China) in statistical analysis. We highly appreciate Dr. Sheri M. Skinner (Baylor College of Medicine, Houston, USA) for his critical review of the

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