Chemokine receptor CXCR7 is an independent prognostic biomarker in glioblastoma

1. Introduction

Glioblastoma (GBM), also known as Grade IV astrocytoma, is the most fatal primary brain cancer in adults with five-year survival rate of only 5% [1]. In most cases, GBM manifest very rapidly without any recognizable precursor lesions. Standard treatment includes complete surgical resection, followed by radiotherapy and adjuvant chemotherapy with temozolomide; however in most cases GBM relapses rapidly. Available GBM management at relapse are usually ineffective. As a result, the median survival of GBM patients with standard-of care treatment is only 15 months currently [2]. Although considerable progress has been made in the last several decades, the exact pathological mechanisms are still unclear and GBM remains an incurable cancer. Therefore, it is of great clinical value to identify the independent and significant prognostic biomarkers, not only improving poor prognosis but also providing promising therapeutic targets.

Chemokines are a group of homologous proteins that interact with G protein-coupled receptors [3]. Chemokines could bind to their receptors on target cells and trigger a series of signal transduction involving the release of intracellular calcium, the generation of inositol triphosphate and activation of protein kinase C [4]. The chemokine CXCL12, also known as SDF-1, regulates many important physiological functions like neuronal development, pro-angiogenic factor and stem cell motility [5]. CXCL12 and its receptor CXCR4 are highly expressed in various types of cancers [6], and play important roles in angiogenesis, proliferation and invasion [7, 8, 9]. In glioblastoma, CXCL12 and CXCR4 are also over-expressed and associated to the regions of angiogenesis and necrosis [10]. CXCR7, another receptor for CXCL12, is expression in many types of cancers such as breast, prostate and lung cancer [11, 12]. It is believed that the CXCR7/CXCR4 heterodimeric complex could enhance CXCL12 signaling and contribute to adhesion to endothelial cells and the inhibition of apoptosis [12]. Although CXCR4-dependent signaling pathway in tumorigenesis and metastasis is actively investigated; the expression of CXCR7 in brain tumor, especially in malignant GBM, has been poorly studied. Previous evidences have demonstrated that CXCR7 was expressed in glioma surgical specimens and various cell lines [13, 14]. In cell lines, CXCR7 was highly expressed and mediated anti-apoptotic effects [13], and targeting CXCR7 could inhibit glioma cell proliferation and mobility [15]. In mice and rats, it was also demonstrated that inhibition of CXCR7 extended survival of brain tumor [16]. Unfortunately, in human glioma specimens, there was no difference for CXCR7 mRNA expression between glioblastomas and non-tumor controls or low-grade gliomas [14]. Here, by examining the expression of CXCR7, our aim was to determine whether CXCR7 was an independent prognostic biomarker of overall survival (OS) in patients with GBM

2. Materials and methods

3. Results

3.2 CXCR7 protein expression and overall survival

Using immunohistochemical techniques, CXCR7 expression was observed in the cytoplasm and the cell membrane of neoplastic cells (Fig. 1). Of all 146 GBM tissues examined, 69 were classified as low-expression, the rest 77 samples were high-expression. As shown in Table 1, there is no statistical differences between these two subgroups in term of age, gender, family history of cancer, extent of surgery, chemotherapy, radiotherapy, KPS, MGMT methylation status and diameter of tumor. Interestingly, compared with CXCR7 positive group, more IDH-1 Mutated patients were included in CXCR7 negative group ( 0.05). In addition, the median survival time for CXCR7 negative group (12.4 months) was significantly longer than the median survival time for CXCR7 positive patients (6.1 months).

Figure 2 showed the Kaplan-Meier survival curves of all recruited GBM subjects. The overall survivals of patients with high levels of CXCR7 were worse than those with low levels of CXCR7 (Fig. 2). Moreover, the multivariate Cox proportional hazard analysis were carried out to assess the potential prognostic factors (Table 2). These included age, gender, extent of surgery, KPS, diameter of tumor, chemotherapy, radiotherapy, MGMT methylation status, IDH-1 mutation status and CXCR7 expression; among which only age, KPS, chemotherapy MGMT methylation status, IDH-1 mutation status and CXCR7 were independent factors.

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Figure 1.

Representative samples with CXCR7 staining in human glioblastoma tissues ( × 400). (A) High expression; (B) Low expression.

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Figure 2.

Kaplan-Meier overall survival curves for CXCR7 expression in 146 glioblastoma samples.

Understanding the pathological mechanisms underlying GBM is important for developing target-specific medicines. CXCL12/CXCR4 axis has been implicated in the etiology of numerous types of cancers [7]. However, little attention has been devoted to the role of CXCR7, the second receptor for CXCL12, in the clinical significance of GBM patients. Here our data demonstrated that high level of CXCR7 protein was an independent biomarker of worse prognosis in GBM patients. It also might suggest CXCR7 is a potential therapeutic target in GBM treatment.

The complex nature of GBM has not been completely elucidated, therefore only very few therapies could provide significant improvement in overall survivals currently. It has long been known that the heterogeneities in GBM patients are prominent, and early diagnosis and treatment do not always improve the clinical outcomes. However, with the increasing knowledge, GBM can be classified into different subgroups based on the molecular signature, and clinician are better understanding this lethal disease. Many predictive and prognostic biomarkers have been discovered that could contribute to the GBM management in clinic [19]. For example, IDH 1 and 2 mutation, MGMT promoter methylation and loss of heterozygosity 1p/19q have been proved to be important in GBM treatment and prognosis [20]. Other known characteristics that could affect OS include age, tumor histological grade, its location and KPS [21]. Even so, evaluation the clinical outcome of GBM was still very difficult. In this study, we mainly focus on the significance between overall survival and CXCR7 in GBM patients. Furthermore, the other OS prognostic characteristics, including age, gender, extent of surgery, KPS, diameter of tumor, chemotherapy MGMT methylation status, IDH-1 mutation status and radiotherapy were subjected to multivariate analysis. The results revealed that age, KPS, chemotherapy, MGMT methylation status, IDH-1 mutation status and CXCR7 were independent prognostic factors.

CXCL12 is a chemokine that is expressed in numerous types of tumor cells and involves in invasion and metastasis [9, 22]. For many years, CXCR4 is believed to be the unique receptor for CXCL12 and CXCL12 the only ligand for CXCR4 [23]. Moreover, CXCR4 has been discovered to be most widely expressed in various types of cancers and contributes to cancer cell proliferation, invasion, as well as in metastasis and survival [7, 8, 9]. Although CXCR7 showed ten-fold higher affinity to CXCL12 compare with CXCR4; little knowledge is available on the biological role of this newly discovered receptor. CXCR7 is absent or expressed at very low levels in most normal tissues, but presents in several different malignant cell types [24, 25]. Typical chemokine responses, such as modulation of adenylyl cyclase activity or intracellular calcium mobilization, are not generated after CXCL12/CXCR7 binding. It is originally believed that the major role of CXCR7 is to modulate the biological function of CXCR4 [26]. This opinion was now challenged by the discoveries that CXCR7 could elicit MAPK, Akt and JAK/STAT3 activation through a β -arrestin-dependent pathway [27]. Moreover, CXCR7 played an essential role in proliferation, adhesion, growth and trans-endothelial migration of different types of cancer cells [24, 25]. In neural system, CXCR7/CXCL12 signaling pathway could control the differentiation and growth of Schwann cell, astrocytes as well as human glioma cells [22, 28]. Accordingly, there is no surprise that CXCR7 is associated with disease-free and disease-specific survival in tumors, like cervical cancer, non-small cell lung cancer and renal cancer [29, 30, 31]. In the present study, we examined whether high levels of CXCR7 was independently correlated with poor clinical survivals. Our data showed that in GBM, the OS was worse in high CXCR7 expression subjects than in patients with low levels of CXCR7, even after adjust for other prognostic factors. Previous studies had shown that antagonist of CXCR7 could suppress the tumor growth in animal models [32]. Although blocking CXCR7 in human had not been conducted currently, it might be a better therapeutic strategy to inhibit both CXCR7 and CXCR4 than just one receptor in the future.

In conclusion, the CXCR7 expression in GBM tissue is an important prognostic tool in predicting overall survival independent of the traditionally used clinicopathological characteristics. CXCR7 expression may be a valuable prognostic biomarker and worthy of further studies in other patient cohorts to validate our results.