Purpose: We herein examined whether the single nucleotide polymorphism (SNP) at −2518 of the MCP-1 gene promoter region influences clinical outcomes among nasopharyngeal carcinoma (NPC) patients.

Experimental Design: The study population consisted of 411 NPC patients without metastasis at diagnosis. All patients were treated at the Chang Gung Memorial Hospital from March 1994 to November 2004. The MCP-1 SNP−2518 genotype of each patient was determined by TaqMan genotyping kit. Statistical analyses were conducted to compare disease-specific survival (DSS), progression-free survival (PFS), local recurrence-free survival (LRFS), and distant metastasis-free survival (DMFS) of patients according to genotype. MCP-1 expression in tumor biopsies was examined by immunohistochemistry.

Results: Among 411 NPC patients, carriers of AA and AG genotypes were prone to distant metastasis than that of GG genotype (hazard ratio, 2.21; P = 0.017, and hazard ratio, 2.23; P = 0.005, for AA and AG genotype, respectively) after initial radiotherapy. No genotype-specific significant difference was found in DSS, PFS, and LRFS. Furthermore, immunohistochemistry revealed that MCP-1 expression level was higher in NPC tumor cells from GG carriers compared with those from AA and AG carriers.

Conclusions:MCP-1 SNP−2518 may be a valuable genetic marker for assessing the risk of developing distant metastasis after the radiotherapy in NPC patients. Carriers of A allele may require more aggressive chemotherapy implicating a potential marker for personalized medicine. We speculate that a regulatory SNP may be associated with the distant metastasis of NPC. Validation studies are warranted.

Nasopharyngeal carcinoma (NPC) is a unique tumor with a remarkably geographic and ethnic distribution. NPC is more prominent in southeastern Asia, including southern China, Hong Kong, and Taiwan, where the annual incidence rate is ∼25-fold higher than that in Western world (1). NPC is closely associated with EBV infection (2), but EBV infection is ubiquitous in human, and the incidence of NPC remains high among Chinese people who have migrated to North America. Moreover, the survival of NPC patients varied among different ethnic groups. Together, these reports suggested an underlying genetic component to NPC risk, progression, and outcomes (35). Consistent with this notion, host genetic factors are likely to influence cancer susceptibility and outcomes (6). In the context of NPC, HLA haplotype (7, 8) and polymorphisms in DNA repair enzymes such as XRCC1 and hOGG1 (9) have been associated with NPC susceptibility, but no genetic predictor has been reported.

Monocyte chemoattractant protein-1 [MCP-1 or tumor-derived chemotactic factor (TDCF)], belongs to CC chemokine family, a member of the small inducible gene, and is encoded by the CCL2 gene, which was mapped to chromosome 17q11. Induction of MCP-1 by inflammatory cytokine interleukin-1 (IL-1)can be modulated through a functional single nucleotide polymorphism (SNP−2518G/A) in MCP-1 distal regulatory region (10), and cells with AG or GG genotype produce ∼2-fold more MCP1 compared with cells with AA genotype. MCP-1 is known as a potent chemoattractant for monocytes and T lymphocytes (11); it is thought to be involved in several diseases characterized by intense macrophage infiltration, including atherosclerosis (12) and cancers. The functional SNP−2518 has been reported to be associated with increased susceptibility or severity of diseases such as asthma (13), atherosclerosis (14), and rheumatoid arthritis (15). Previous studies have shown that MCP-1 is overexpressed in many tumors such as glioma (16), ovarian (17), breast (18), and esophagus cancers (19). Furthermore, expression of MCP-1 in tumor cells correlates with macrophage infiltration and tumor vascularity (19, 20). Notably, the patient's serum with high MCP-1 level is correlated with better prognosis (2123), but the molecular mechanism remains to be investigated. To date, no studies have been carried out to examine the role of this SNP in tumor progression.

In NPC, the overexpression of MCP-1 has been detected in the infiltrated macrophages, leading to intensive leukocyte infiltration (24). However, it is unclear whether (a) MCP-1 is overexpressed in NPC tumor cells; (b) MCP-1 expression level can be modulated by the functional SNP; and (c) MCP-1 SNP−2518 genotype is associated with NPC patients' prognosis.

In this study, we hypothesized that MCP-1 SNP−2518 may be associated with the clinical outcome of NPC patients under current therapeutic protocols. We retrospectively analyzed MCP-1 SNP−2518 genotypes and their association with clinical outcomes in 411 NPC patients receiving complete radiotherapy. This is the first report suggesting a host genetic polymorphism that may serve as a prognostic predictor for NPC.

Patients, clinical staging protocol, oncological treatment, and clinical outcome assessment. This retrospective cohort comprises 411 NPC patients who had been admitted to Chang Gung Memorial Hospital (CGMH), Lin-Kou, from March 1994 to November 2004. The tumor-node-metastasis (TNM) stage was defined according to the 2002 cancer staging system revised by the American Joint Committee on Cancer (AJCC; ref. 25), and histologic typing was done according to the WHO classification criteria (26). This study was reviewed and approved by the institutional review board and ethics committee of CGMH. Informed consent was obtained from all patients.

All enrolled patients had been treated with definitive radiotherapy (cumulative dose of external beam radiotherapy ≧64.8 Gy). Among them, 109 patients received additional chemotherapy in the Department of Radiation Oncology at CGMH. Patients who were diagnosed with distant metastatic disease at presentation (M1 stage) and/or who had undergone previous treatment at another institute were excluded from the present study. For all enrolled patients, pathology records were retrieved from pathologic databases and medical records and reviewed for confirmation of the NPC diagnosis. Information on stage, treatment, follow-up, and limited information on family history were collected from hospital tumor registries and medical files.

Patients were followed-up at 2- to 3-month intervals during the first 3 years after therapy and at 6-month intervals thereafter. The minimal follow-up period was 28 months. The primary end point was disease-specific survival (DSS), which was calculated from the date of diagnosis to the date of death or the last follow-up. Progression-free survival (PFS), distant metastasis-free survival (DMFS), and local recurrence-free survival (LRFS) were also assessed. The time to local recurrence or distant metastasis was calculated using the date on which local recurrence or distant metastasis status was detected as the end point. Patients who died without occurrence of local recurrence or distant metastasis were censored in the analyses of LRFS and DMFS.

DNA extraction and genotyping. Blood samples were collected at the time of enrollment, and genomic DNA was obtained using a DNA isolation kit (Qiagen). The target MCP-1 SNP−2518 (CCL2 gene −2518G/A, rs1024611) was detected using a commercially available TaqMan genotyping assay kit and a GeneAmp PCR System 9700 (both from Applied Biosystems Inc.) according to the manufacturer's instructions. Two template-free controls and six DNA samples of known genotype were included in each plate as negative and positive controls, respectively. The Sequence Detection Software provided by ABI was used for genotyping analysis.

Immunohistochemical staining analysis. Immunohistochemical analyses were done using an automatic immunohistochemistry (IHC) staining device according to the manufacturer's procedures (Bond, Vision Biosystems). Anti–MCP-1 antibody (R&D Systems) and anti-CD68 antibody (DAKO, Dakopatts) were used. The detailed procedure of immunohistochemical staining was described in Supplementary Data. The MCP-1 intensity was scored as negative or 0; mild or 1; moderate or 2; or intense or 3 at 100× magnification according on the scoring method described in Supplementary Data. The amount of CD68+ macrophages infiltrated was measured using ImageScope (Aperio Technologies) software at 200× magnification. A total of 3 to 10 fields per slide were selected, counted, and averaged.

In situ hybridization.In situ hybridization (ISH) for detecting EBV-encoded RNA transcripts (EBER) was done using the EBV Probe ISH kit (Novocastra) according to the manufacturer's instructions.

MCP-1 reporter construct and luciferase activity assay. The distal regulatory region, which contains either G or A at the MCP-1 SNP−2518 was amplified from NPC patient genomic DNA by PCR. IL-1β–induced promoter activity was examined in 293T cells as described in Supplementary Data.

Statistical analysis. Patients were separated into subgroups based on their MCP-1 SNP−2518 (CCL2 gene −2518G/A, rs1024611) genotypes (AA, AG, or GG). Box plots were used to show the MCP-1 IHC score in subjects with different genotype and number of macrophages infiltrated in NPC biopsies with different MCP-1 expression levels. The boundary of the box closet to the zero indicates the 25th percentile; and the boundary farthest from zero indicates the 75th percentile; a line in the box marks the median. Whiskers above and below the box indicate the 90th and 10th percentiles. ANOVA was used to evaluate the MCP-1 IHC score by genotype and amount of infiltrated macrophages by MCP-1 expression levels. The age at diagnosis, distribution of gender, overall stage, N stage, T stage, and use of chemotherapy were compared between patient subgroups using the χ2 test. Kaplan-Meier plots of DSS, PFS, DMFS, and LRFS were established, and statistical significance was measured by the log-rank test. The Cox proportional regression model was used to evaluate the effect of MCP-1 SNP−2518 genotype and other potential prognostic factors on survival tests. All statistical tests were two sided, and a P value of ≦0.05 was considered statistically significant. All statistical analyses were done using the SPSS software version 13.0 (SPSS Inc.).

In vivo correlation of mCP-1 expression in tumor cells with genotypes. To test whether MCP-1 expression can be detected in NPC tumors, 37 NPC biopsies were examined by IHC. The distribution of NPC tumor cells was assayed by the presence of EBV noncoding transcripts (EBERs) as previously described (27). Among 37 NPC biopsies, all of them had EBV infection, and 30 samples (81%) had MCP-1 overexpression. MCP-1 was observed in both tumor and stromal cells but mainly in the cytoplasm of tumor cells (Fig. 1A). However, little or comparatively low expression of MCP-1 was detected in the adjacent cells. To correlate the MCP-1 expression level with the MCP-1 SNP−2518 genotype, genomic DNA collected from the same patients was genotyped by TaqMan genotyping kit. The expression levels of MCP-1 in biopsies was scored and compared among patients with three genotypes. As shown in Fig. 1A and B and Supplementary Table S1, the biopsies from the GG-genotype patients had the highest MCP-1 expression level, whereas AA genotype had the lowest (P < 0.001). Together, these results suggested that MCP-1 is overexpressed by NPC tumor cells, and the expression level is associated with the MCP-1 SNP−2518 genotype.

Fig. 1.

Correlation between MCP-1 expression level and MCP-1 SNP−2518 genotype in NPC biopsies. A, MCP-1 expression (brown signals) from GG and AA patients was detected by IHC. The EBERs signal (dark blue) indicated the distribution of NPC tumor cells, and the CD68 signal (brown) marked the macrophages in NPC biopsies. Images in the box (left, 100×) were enlarged and shown in the right (400×). B, box plots showed a significant correlation of MCP-1 IHC score in NPC biopsies among three genotypes (χ2 test, P < 0.001). C, box plots showed increased infiltrated macrophages (CD68+) in NPC biopsies correlated with higher MCP-1 expression level.

Fig. 1.

Correlation between MCP-1 expression level and MCP-1 SNP−2518 genotype in NPC biopsies. A, MCP-1 expression (brown signals) from GG and AA patients was detected by IHC. The EBERs signal (dark blue) indicated the distribution of NPC tumor cells, and the CD68 signal (brown) marked the macrophages in NPC biopsies. Images in the box (left, 100×) were enlarged and shown in the right (400×). B, box plots showed a significant correlation of MCP-1 IHC score in NPC biopsies among three genotypes (χ2 test, P < 0.001). C, box plots showed increased infiltrated macrophages (CD68+) in NPC biopsies correlated with higher MCP-1 expression level.

Close modal

Previous reports suggested that the extent of macrophage accumulation in tumors is correlated with the MCP-1 expression level (19, 28). Therefore, we counted the number of infiltrated macrophage in NPC biopsies by IHC using the anti-CD68 antibody. As shown in Fig. 1A and C, the increased amount of infiltrated macrophages is correlated with the high MCP-1 expression levels detected in tumor cells (P = 0.012).

In vitro MCP-1 SNP−2518 A or G allele promoter assay. To further confirm if MCP-1 SNP−2518 genotype influences its expression in response to a given stimulus, we did an in vitro promoter assay in the presence of IL-1β, a potent stimulator of MCP-1 protein expression (29) and can be detected in NPC biopsies (30). A luciferase reporter gene was fused with MCP-1 promoter region (−2906 to +215, with A or G allele, respectively) isolated from NPC patients in this cohort. Compared with the A-allele–containing promoter construct (4.9-fold activation), G-allele construct had higher promoter activity (7.2-fold activation) under IL-1β stimulation (Supplementary Fig. S1; P = 0.012). Thus, results indicated that the G allele isolated from NPC biopsy showed higher enhancer activity than A allele after treated with an inflammatory cytokine.

Patient characteristics and genotypic frequencies. To evaluate the association of MCP-1 SNP−2518 with clinical outcome under current therapeutic protocols, a retrospective cohort of 411 NPC patients were subjected to clinical outcome assessment study. The patient characteristics and clinical features are summarized in Table 1. The median age at diagnosis was 47.6 years (range, 13-87), with a male-to-female ratio of 3:1. The frequencies of the MCP-1 SNP−2518 AA, AG, and GG genotypes among the studied NPC patients were 20% (82/411), 47% (194/411), and 33% (135/411), respectively. The clinicopathologic features were comparable among patient subgroups classified according to the MCP-1 −2518 genotype.

Table 1.

Clinicopathologic features of NPC patients

CharacteristicNumber of patientsMCP-1 SNP−2518 genotype
AAAGGGP
 411 82 (20%) 194 (47%) 135 (33%)  
Age at diagnosis (y)      
    Mean 48.7 49.31 48.56 48.52 0.875* 
    Median 47.63 48.37 48.09 46.95  
Gender      
    Male 308 63 (21%) 139 (45%) 106 (34%) 0.334 
    Female 103 19 (18%) 55 (54%) 29 (28%)  
Tumor stage      
    T1 143 22 (15%) 73 (51%) 48 (34%) 0.462 
    T2 115 22 (19%) 54 (47%) 39 (34%)  
    T3 71 20 (28%) 28 (40%) 23 (32%)  
    T4 82 18 (22%) 39 (48%) 25 (30%)  
Node stage      
    N0 152 31 (20%) 69 (46%) 52 (34%) 0.798 
    N1 138 27 (20%) 66 (48%) 45 (32%)  
    N2 84 20 (24%) 39 (46%) 25 (30%)  
    N3 37 4 (11%) 20 (54%) 13 (35%)  
AJCC stage      
    I 62 9 (15%) 33 (53%) 20 (32%) 0.391 
    II 133 22 (16%) 66 (50%) 45 (34%)  
    III 110 30 (27%) 45 (41%) 35 (32%)  
    IV 106 21 (20%) 50 (47%) 35 (33%)  
Use of chemotherapy      
    Yes 109 24 (22%) 46 (42%) 39 (36%) 0.474 
CharacteristicNumber of patientsMCP-1 SNP−2518 genotype
AAAGGGP
 411 82 (20%) 194 (47%) 135 (33%)  
Age at diagnosis (y)      
    Mean 48.7 49.31 48.56 48.52 0.875* 
    Median 47.63 48.37 48.09 46.95  
Gender      
    Male 308 63 (21%) 139 (45%) 106 (34%) 0.334 
    Female 103 19 (18%) 55 (54%) 29 (28%)  
Tumor stage      
    T1 143 22 (15%) 73 (51%) 48 (34%) 0.462 
    T2 115 22 (19%) 54 (47%) 39 (34%)  
    T3 71 20 (28%) 28 (40%) 23 (32%)  
    T4 82 18 (22%) 39 (48%) 25 (30%)  
Node stage      
    N0 152 31 (20%) 69 (46%) 52 (34%) 0.798 
    N1 138 27 (20%) 66 (48%) 45 (32%)  
    N2 84 20 (24%) 39 (46%) 25 (30%)  
    N3 37 4 (11%) 20 (54%) 13 (35%)  
AJCC stage      
    I 62 9 (15%) 33 (53%) 20 (32%) 0.391 
    II 133 22 (16%) 66 (50%) 45 (34%)  
    III 110 30 (27%) 45 (41%) 35 (32%)  
    IV 106 21 (20%) 50 (47%) 35 (33%)  
Use of chemotherapy      
    Yes 109 24 (22%) 46 (42%) 39 (36%) 0.474 
*

P value was calculated using ANOVA.

P value was computed using the χ2 test.

MCP-1 genotype and NPC DSS. The 5-year DSS rate across all studied patients was 75.3%. As shown by Kaplan-Meier plot, the 5-year DSS rates for patient subgroups defined by MCP-1 SNP−2518 genotype AA, AG, and GG were 71.3%, 74.5% and 78.7%, respectively (Fig. 2A). When DSS was compared among MCP-1 SNP−2518 genotypes using a log-rank test, no significant difference was observed between different subgroups of patients. We conducted a multivariate analysis of the genotype effect on DSS using the Cox proportional regression models adjusted by other prognostic factors, such as tumor stage, node stage, sex, use of chemotherapy, and age at onset. Our results indicated that DSS was significantly dependent on age at onset, primary tumor stage, and node stage, which is similar to those previously reported (31), whereas MCP-1 SNP−2518 was not an independent predictor for DSS in NPC patients (Supplementary Table S2).

Fig. 2.

Kaplan-Meier survival curves in NPC patients according to MCP-1 SNP−2518 genotype. A, DSS. B, PFS. C, LRFS. D, DMFS.

Fig. 2.

Kaplan-Meier survival curves in NPC patients according to MCP-1 SNP−2518 genotype. A, DSS. B, PFS. C, LRFS. D, DMFS.

Close modal

MCP-1 genotype and PFS. All analyses were repeated using the date of disease progression as the end point. Kaplan-Meier plots showed that no significant difference was observed between patients of different MCP-1 SNP−2518 genotype (Fig. 2B). The Cox proportional regression model indicated that only tumor stage and node stage were independent predictors for PFS time, and MCP-1 SNP−2518 was not an independent predictor for PFS in NPC patients (Supplementary Table S2).

MCP-1 genotype and LRFS. Among the 411 NPC patients, 93 patients (22.6%) developed local recurrence after the completion of the radiotherapy. Genotypic frequencies of MCP-1 SNP−2518 AA, AG, and GG carriers were 14% (13/93), 47.3% (44/93), and 38.7% (36/93), respectively. All analyses were repeated using the date of local recurrence diagnosed as the end point. The survival curves revealed no significant difference in LRFS among different subgroups (Fig. 2C). However, in a Cox proportional regression model (Table 2) that adjusted the effect of MCP-1 SNP−2518 by other potential prognostic factors, patients of AA genotype showed a lower risk for developing local recurrence when compared with AG- and GG-genotype patients (hazard ratio, 0.53; P = 0.05).

Table 2.

Cox multivariate regression analysis for LRFS in NPC patient

FactorsPHazard ratio (95% confidence interval)
Sex (female versus male) 0.521  
Age (≥48 y versus <48 y) 0.263  
Tumor stage   
    T2 versus T1 0.127  
    T3 versus T1 0.005* 2.54 (1.33-4.84) 
    T4 versus T1 <0.001* 4.084 (2.21-7.54) 
Node stage   
    N1 versus N0 0.717  
    N2 versus N0 0.67  
    N3 versus N0 0.631  
Use of chemotherapy 0.311  
MCP1 SNP−2518 genotype   
    AA versus GG 0.05* 0.53 (0.28-1.0) 
    AG versus GG 0.678  
FactorsPHazard ratio (95% confidence interval)
Sex (female versus male) 0.521  
Age (≥48 y versus <48 y) 0.263  
Tumor stage   
    T2 versus T1 0.127  
    T3 versus T1 0.005* 2.54 (1.33-4.84) 
    T4 versus T1 <0.001* 4.084 (2.21-7.54) 
Node stage   
    N1 versus N0 0.717  
    N2 versus N0 0.67  
    N3 versus N0 0.631  
Use of chemotherapy 0.311  
MCP1 SNP−2518 genotype   
    AA versus GG 0.05* 0.53 (0.28-1.0) 
    AG versus GG 0.678  
*

With statistical significance.

With marginal significance.

MCP-1 genotype and DMFS. A total of 86 out of 411 (20.9%) NPC patients developed distant metastasis after the initial radiotherapy. Among the 86 patients, 24.4% (21/86), 55.8% (48/86), and 19.8% (17/86) belonged to MCP-1 SNP−2518 AA, AG, and GG genotypes, respectively. All analyses were repeated using the date of distant metastasis diagnosed as the end point. As shown in Fig. 2D, the 5-year DMFS rate of AA, AG, and GG genotype patients was 74.1%, 73.1%, and 86.4%, respectively. The presence of one or two copies of the A allele at the MCP-1 SNP−2518 strongly predicted inferior DMFS (AA versus GG, P = 0.011; and AG versus GG, P = 0.005) in NPC patients.

When a Cox proportional regression model was used to assess the prognostic significance of MCP-1 SNP−2518 for an NPC patient to develop distant metastasis, the results (Table 3) revealed that, in addition to tumor stage and node stage, NPC patients with the AA and AG genotype had a higher risk for developing metastasis compared with patients with GG genotype (AA versus GG: hazard ratio, 2.21; P = 0.017; AG versus GG: hazard ratio, 2.23; P = 0.005). These results suggest that in addition to the tumor stage and node stage, the MCP-1 SNP−2518 A allele was an independent predictor for higher risk of developing distant metastasis in NPC patients after treatment.

Table 3.

Cox multivariate regression analysis for DMFS in NPC patients

FactorsPHazard ratio (95% confidence interval)
Sex (female versus male) 0.521  
Age (≥48 y versus <48 y) 0.607  
Tumor stage   
    T2 versus T1 0.173  
    T3 versus T1 0.061* 1.89 (0.97-3.69) 
    T4 versus T1 0.013 2.24 (1.19-4.24) 
Node stage   
    N1 versus N0 0.002 2.52 (1.41-4.52) 
    N2 versus N0 0.004 2.57 (1.35-4.92) 
    N3 versus N0 <0.001 4.56 (2.15-9.68) 
Use of chemotherapy 0.484  
MCP1 SNP−2518 genotype   
    AA versus GG 0.017 2.21 (1.16-4.23) 
    AG versus GG 0.005 2.23 (1.28-3.90) 
FactorsPHazard ratio (95% confidence interval)
Sex (female versus male) 0.521  
Age (≥48 y versus <48 y) 0.607  
Tumor stage   
    T2 versus T1 0.173  
    T3 versus T1 0.061* 1.89 (0.97-3.69) 
    T4 versus T1 0.013 2.24 (1.19-4.24) 
Node stage   
    N1 versus N0 0.002 2.52 (1.41-4.52) 
    N2 versus N0 0.004 2.57 (1.35-4.92) 
    N3 versus N0 <0.001 4.56 (2.15-9.68) 
Use of chemotherapy 0.484  
MCP1 SNP−2518 genotype   
    AA versus GG 0.017 2.21 (1.16-4.23) 
    AG versus GG 0.005 2.23 (1.28-3.90) 
*

With marginal significance.

With statistical significance.

Our data indicate that a single nucleotide polymorphism, −2518G/A, in the regulatory region of MCP-1 gene, is an independent genetic risk predictor for developing distant metastasis after complete treatment in NPC patients. It is based on the correlation studies of a retrospective cohort of 411 NPC patients, collected over 10 years, with MCP-1 frequencies and disease outcomes. Patients with AA or AG genotype that have relatively lower MCP-1 expression are more prone to distant metastasis in the first 3 years after treatment than those with the GG genotype. Previous studies and our data confirm that the MCP-1 expression level is indeed associated with genotype. The MCP-1 expression level is likely to be regulated through the binding of transcription factors. A nuclear factor complex containing IRF-1 has shown greater affinity to the A allele than the G allele (32). Although the effect of IRF-1 binding is unclear, AG or GG genotype patients showed slightly higher MCP-1 level. Consistent with the above results, our in vitro MCP-1 promoter-reporter assay showed that the construct with the G allele has ∼1.5-fold higher activity than its counterpart A allele under IL-1β stimulation. Hence, genetic background may be involved in the predetermination of the expression level of MCP-1.

No significant difference was found among three genotypes in DSS and PFS. However, our statistical analysis suggested that among these NPC patients that had recurrence, patients with AA genotype tend to develop distant metastasis, whereas patients with GG genotype tend to develop local recurrence. Therefore, MCP-1 SNP−2518 genotype cannot foretell the risk of NPC relapse, but it may predict, if relapse does occur, which genotype may associate with a particular cancer phenotype. Because the present work is focused on the study of a candidate gene, future search of new and more SNPs related to NPC risk by whole genome association study may be needed. In addition, this study is limited by its retrospective nature and the patients collected in a single institute; validation studies by others are awaited.

A recent report suggested that the tumor microenvironment plays a critical role in the control of local tumor recurrence and distant metastasis (33). Antitumor immune response can act as an environmental stress, which may select the specific cancer cell clones with metastatic potential (34). MCP-1 is a key player in innate and adaptive immunity, which mediates macrophage recruitment to inflammation sites and modulates T helper cell polarization (35). As shown in Fig. 1C, the number of CD68+ macrophages infiltrated in the NPC tumor mass is correlated with the MCP-1 expression level, which can be modulated by the MCP-1 genotype. As a result, the NPC tumor microenvironment may be altered due to the different MCP-1 levels, which is governed by genetic background. However, how MCP-1 may lead to different cancer phenotypes such as distant metastasis and local recurrence is still obscured.

Host genetic factors are likely to influence cancer susceptibility and outcome. The role of germ line polymorphisms in modulating the outcome or metastatic potential of tumor cells has been proposed (36). For example, in human breast cancer, the −313 G/A polymorphism located in the regulatory region of the SIPA1 gene has been strongly associated with axillary node involvement at the time of diagnosis (37). Here, we report that MCP-1 SNP−2518 genotype seems to be a novel candidate for NPC distant metastasis, although the detailed mechanism is still unclear. Because overexpression of MCP-1 has been reported in a variety of cancers, it is worth testing if the MCP-1 SNP−2518 genotypes can predict the metastasis potential of patients with other cancers.

Despite its radiosensitive nature, NPC is characterized by its high metastatic potential; 20% to 25% of the patients develop distant metastasis after initial treatment in our hospital. Current clinical prognostic indicators for NPC patients include TNM stage (31), histopathologic classification (38), age of onset (39), cumulative dose of radiotherapy (40), and pretherapy circulating EBV DNA load (4143). Elevated plasma EBV DNA in combination with the Unio Internationale Contra Cancrum staging data improves risk discrimination in early-stage disease (44). The practical value of the genetic predictor, such as MCP-1 SNP−2518, should be validated by other cohorts and also judged in the light of plasma EBV DNA level.

MCP-1 induction through NF-κB activation has been reported in HHV8-infected endothelial cells (45) and in EBV-infected monocytes (46). EBV-encoded latent membrane protein 1 (LMP1), which expressed in NPC tumor cells, can activate NF-κB signaling (47). It would be interesting to investigate if EBV infection or LMP1 expression in NPC tumor cells can play a role in MCP-1 induction.

With the increasing emphasis on personalized medicine and the life quality of cancer patients, the assistance of genetic metastasis predictor may provide a simple and reproducible assay for clinical outcome prediction, which can facilitate an appropriate allocation of adjuvant therapy.

Grant support: Ministry of Education (to Chang Gung University), National Science Council (NSC 94-2314-B-182A-188, 94-3112-B-182-005 and 95-2320-B-182-001) and Chang Gung Memorial Hospital (CMRPD150961 and CMRPG360221), Taiwan.

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Note: Supplementary data for this article are available at Clinical Cancer Research Online (http://clincancerres.aacrjournals.org/).

1
Parkin DM, Whelan SL, Ferlay J, Raymond L, Young J, editors. Cancer incidence in five continents. Volume VII.
IARC Sci Publ
1997
;
143
:
814
–5.
2
zur Hausen H, Schulte-Holthausen H, Klein G, et al. EBV DNA in biopsies of Burkitt tumours and anaplastic carcinomas of the nasopharynx.
Nature
1970
;
228
:
1056
–8.
3
Marks JE, Phillips JL, Menck HR. The National Cancer Data Base report on the relationship of race and national origin to the histology of nasopharyngeal carcinoma.
Cancer
1998
;
83
:
582
–8.
4
Su CK, Wang CC. Prognostic value of Chinese race in nasopharyngeal cancer.
Int J Radiat Oncol Biol Phys
2002
;
54
:
752
–8.
5
Wei WI, Sham JS. Nasopharyngeal carcinoma.
Lancet
2005
;
365
:
2041
–54.
6
Erichsen HC, Chanock SJ. SNPs in cancer research and treatment.
Br J Cancer
2004
;
90
:
747
–51.
7
Grulich AE, McCredie M, Coates M. Cancer incidence in Asian migrants to New South Wales, Australia.
Br J Cancer
1995
;
71
:
400
–8.
8
Feng BJ, Huang W, Shugart YY, et al. Genome-wide scan for familial nasopharyngeal carcinoma reveals evidence of linkage to chromosome 4.
Nat Genet
2002
;
31
:
395
–9.
9
Cho EY, Hildesheim A, Chen CJ, et al. Nasopharyngeal carcinoma and genetic polymorphisms of DNA repair enzymes XRCC1 and hOGG1.
Cancer Epidemiol Biomarkers Prev
2003
;
12
:
1100
–4.
10
Rovin BH, Lu L, Saxena R. A novel polymorphism in the MCP-1 gene regulatory region that influences MCP-1 expression.
Biochem Biophys Res Commun
1999
;
259
:
344
–8.
11
Lu B, Rutledge BJ, Gu L, et al. Abnormalities in monocyte recruitment and cytokine expression in monocyte chemoattractant protein 1–deficient mice.
J Exp Med
1998
;
187
:
601
–8.
12
Gu L, Tseng SC, Rollins BJ. Monocyte chemoattractant protein-1.
Chem Immunol
1999
;
72
:
7
–29.
13
Szalai C, Kozma GT, Nagy A, et al. Polymorphism in the gene regulatory region of MCP-1 is associated with asthma susceptibility and severity.
J Allergy Clin Immunol
2001
;
108
:
375
–81.
14
Szalai C, Duba J, Prohaszka Z, et al. Involvement of polymorphisms in the chemokine system in the susceptibility for coronary artery disease (CAD). Coincidence of elevated Lp(a) and MCP-1 −2518 G/G genotype in CAD patients.
Atherosclerosis
2001
;
158
:
233
–9.
15
Gonzalez-Escribano MF, Torres B, Aguilar F, et al. MCP-1 promoter polymorphism in Spanish patients with rheumatoid arthritis.
Hum Immunol
2003
;
64
:
741
–4.
16
Yoshimura T, Robinson EA, Tanaka S, Appella E, Kuratsu J, Leonard EJ. Purification and amino acid analysis of two human glioma-derived monocyte chemoattractants.
J Exp Med
1989
;
169
:
1449
–59.
17
Negus RP, Stamp GW, Relf MG, et al. The detection and localization of monocyte chemoattractant protein-1 (MCP-1) in human ovarian cancer.
J Clin Invest
1995
;
95
:
2391
–6.
18
Valkovic T, Lucin K, Krstulja M, Dobi-Babic R, Jonjic N. Expression of monocyte chemotactic protein-1 in human invasive ductal breast cancer.
Pathol Res Pract
1998
;
194
:
335
–40.
19
Ohta M, Kitadai Y, Tanaka S, et al. Monocyte chemoattractant protein-1 expression correlates with macrophage infiltration and tumor vascularity in human esophageal squamous cell carcinomas.
Int J Cancer
2002
;
102
:
220
–4.
20
Kuroda T, Kitadai Y, Tanaka S, et al. Monocyte chemoattractant protein-1 transfection induces angiogenesis and tumorigenesis of gastric carcinoma in nude mice via macrophage recruitment.
Clin Cancer Res
2005
;
11
:
7629
–36.
21
Ueno T, Toi M, Saji H, et al. Significance of macrophage chemoattractant protein-1 in macrophage recruitment, angiogenesis, and survival in human breast cancer.
Clin Cancer Res
2000
;
6
:
3282
–9.
22
Monti P, Leone BE, Marchesi F, et al. The CC chemokine MCP-1/CCL2 in pancreatic cancer progression: regulation of expression and potential mechanisms of antimalignant activity.
Cancer Res
2003
;
63
:
7451
–61.
23
Dehqanzada ZA, Storrer CE, Hueman MT, et al. Correlations between serum monocyte chemotactic protein-1 levels, clinical prognostic factors, and HER-2/neu vaccine-related immunity in breast cancer patients.
Clin Cancer Res
2006
;
12
:
478
–86.
24
Tang KF, Tan SY, Chan SH, et al. A distinct expression of CC chemokines by macrophages in nasopharyngeal carcinoma: implication for the intense tumor infiltration by T lymphocytes and macrophages.
Hum Pathol
2001
;
32
:
42
–9.
25
Sobin LH, Wittekind C. International Union against Cancer. TNM: classification of malignant tumours. 6th ed. New York: Wiley-Liss; 2002.
26
Shanmugaratnam K. Histological typing of nasopharyngeal carcinoma. IARC Sci Publ 1978. p. 3–12.
27
Niedobitek G, Young LS, Sam CK, Brooks L, Prasad U, Rickinson AB. Expression of Epstein-Barr virus genes and of lymphocyte activation molecules in undifferentiated nasopharyngeal carcinomas.
Am J Pathol
1992
;
140
:
879
–87.
28
Varney ML, Johansson SL, Singh RK. Tumour-associated macrophage infiltration, neovascularization and aggressiveness in malignant melanoma: role of monocyte chemotactic protein-1 and vascular endothelial growth factor-A.
Melanoma Res
2005
;
15
:
417
–25.
29
Martin T, Cardarelli PM, Parry GC, Felts KA, Cobb RR. Cytokine induction of monocyte chemoattractant protein-1 gene expression in human endothelial cells depends on the cooperative action of NF-κB and AP-1.
Eur J Immunol
1997
;
27
:
1091
–7.
30
Huang YT, Sheen TS, Chen CL, et al. Profile of cytokine expression in nasopharyngeal carcinomas: a distinct expression of interleukin 1 in tumor and CD4+ T cells.
Cancer Res
1999
;
59
:
1599
–605.
31
Sham JS, Choy D. Prognostic factors of nasopharyngeal carcinoma: a review of 759 patients.
Br J Radiol
1990
;
63
:
51
–8.
32
Gonzalez E, Rovin BH, Sen L, et al. HIV-1 infection and AIDS dementia are influenced by a mutant MCP-1 allele linked to increased monocyte infiltration of tissues and MCP-1 levels.
Proc Natl Acad Sci U S A
2002
;
99
:
13795
–800.
33
Kaur P, Mulvaney M, Carlson JA. Basal cell carcinoma progression correlates with host immune response and stromal alterations: a histologic analysis.
Am J Dermatopathol
2006
;
28
:
293
–307.
34
Seymour K, Pettit S, O'Flaherty E, Charnley RM, Kirby JA. Selection of metastatic tumour phenotypes by host immune systems.
Lancet
1999
;
354
:
1989
–91.
35
Gu L, Tseng S, Horner RM, Tam C, Loda M, Rollins BJ. Control of TH2 polarization by the chemokine monocyte chemoattractant protein-1.
Nature
2000
;
404
:
407
–11.
36
Hunter K. Host genetics influence tumour metastasis.
Nat Rev
2006
;
6
:
141
–6.
37
Crawford NP, Ziogas A, Peel DJ, Hess J, Anton-Culver H, Hunter KW. Germline polymorphisms in SIPA1 are associated with metastasis and other indicators of poor prognosis in breast cancer.
Breast Cancer Res
2006
;
8
:
R16
.
38
Kurniawan AN, Susworo R, Sumanto. Nasopharyngeal carcinoma: correlation of histopathology with radiation response.
Southeast Asian J Trop Med Public Health
1985
;
16
:
613
–8.
39
Tan BC, Khor TH, Chia KB. Radiotherapy in treatment of nasopharyngeal carcinoma.
Ann Acad Med Singapore
1980
;
9
:
347
–9.
40
Perez CA, Devineni VR, Marcial-Vega V, Marks JE, Simpson JR, Kucik N. Carcinoma of the nasopharynx: factors affecting prognosis.
Int J Radiat Oncol Biol Phys
1992
;
23
:
271
–80.
41
Lo YM, Chan LY, Chan AT, et al. Quantitative and temporal correlation between circulating cell-free Epstein-Barr virus DNA and tumor recurrence in nasopharyngeal carcinoma.
Cancer Res
1999
;
59
:
5452
–5.
42
Chan AT, Lo YM, Zee B, et al. Plasma Epstein-Barr virus DNA and residual disease after radiotherapy for undifferentiated nasopharyngeal carcinoma.
J Natl Cancer Inst
2002
;
94
:
1614
–9.
43
Lin J, Wang W, Chen K, et al. Quantification of plasma Epstein-Barr virus DNA in patients with advanced nasopharyngeal carcinoma.
N Engl J Med
2004
;
350
:
2461
–70.
44
Leung SF, Zee B, Ma BB, et al. Plasma Epstein-Barr viral deoxyribonucleic acid quantitation complements tumor-node-metastasis staging prognostication in nasopharyngeal carcinoma.
J Clin Oncol
2006
;
24
:
5414
–8.
45
Caselli E, Fiorentini S, Amici C, Di Luca D, Caruso A, Santoro MG. Human herpesvirus 8 acute infection of endothelial cells induces monocyte chemoattractant protein 1–dependent capillary-like structure formation: role of the IKK/NF-κB pathway.
Blood
2007
;
109
:
2718
–26.
46
Gaudreault E, Fiola S, Olivier M, Gosselin J. Epstein-Barr virus induces Mcp-1 secretion by human monocytes via Tlr2.
J Virol
2007
;
81
:
8016
–24.
47
Li HP, Chang YS. Epstein-Barr virus latent membrane protein 1: structure and functions.
J Biomed Sci
2003
;
10
:
490
–504.