Liang-Ru Ke

CCL2-CCR2 axis promotes metastasis of nasopharyngeal carcinoma by activating ERK1/2-MMP2/9 pathway

Distant metastasis remains the major failure of nasopharyngeal carcinoma (NPC). In this study, the roles of chemokine C-C motif ligand 2 (CCL2), and its receptor chemokine C-C motif receptor type 2 (CCR2) on NPC metastasis were investigated. Serum CCL2 and CCL2/CCR2 expression level were remarkably increased in NPC patients compared to non-tumor patients by ELISA and IHC analyses. High expressions of CCL2/CCR2 were significantly associated with NPC metastasis and poor overall survival (OS). High expression of CCR2 is an independent adverse prognostic factor of OS and distant metastasis free survival (DMFS). Overexpressions of CCL2 and CCR2 were detected in high-metastatic NPC cell lines. Upregulating CCL2 and CCR2 respectively in low-metastatic NPC cell lines could promote cell migration and invasion, and exogenous CCL2 enhanced the motility in CCR2-overexpressing cells. On the other hand, downregulating CCL2 and CCR2 respectively in high-metastatic NPC cell lines by shRNA could decrease cell migration and invasion. However, exogenous CCL2 could not rescue the weaken ability of motility of CCR2-silencing cells. In nude mouse model, distant metastasis was significantly facilitated in either CCL2-overexpressing or CCR2-overexpressing groups, which was more obvious in CCR2-overexpressing group. Also, distant metastasis was considerably inhibited in either CCL2-silencing or CCR2-silencing groups. Dual overexpression of CCL2/CCR2 could activate extracellular signal-regulated kinase (ERK1/2) signaling pathway, which sequentially induced matrix metalloproteinase (MMP) 2 and 9 upregulations in the downstream. In conclusion, CCL2-CCR2 axis could promote NPC metastasis by activating ERK1/2-MMP2/9 pathway. This study helps to develop novel therapeutic targets for distant metastasis in NPC.

The Impact of Baseline Serum C-Reactive Protein and C-Reactive Protein Kinetics on the Prognosis of Metastatic Nasopharyngeal Carcinoma Patients Treated with Palliative Chemotherapy

Background The aim of this study was to determine whether baseline C-reactive protein (CRP) levels and CRP kinetics predict the overall survival in metastatic nasopharyngeal carcinoma (mNPC) patients. Methods A total of 116 mNPC patients from January 2006 to July 2011 were retrospectively reviewed. Serum CRP level was measured at baseline and thereafter at the start of each palliative chemotherapy cycle for all patients. Results Patients with higher values of baseline CRP (≥ 3.4 mg/L) had significantly worse survival than those with lower baseline CRP values (< 3.4 mg/L). Patients were divided into four groups according to baseline CRP and CRP kinetics: (1) patients whose CRP < 3.4 mg/L and never elevated during treatment; (2) patients whose CRP < 3.4 mg/L and elevated at least one time during treatment; (3) patients whose CRP ≥ 3.4 mg/L and normalized at least one time during treatment; and (4) patients whose CRP ≥ 3.4 mg/L and never normalized during treatment. The patients were further assigned to non-elevated, elevated, normalized, and non-normalized CRP groups. Overall survival rates were significantly different among the four groups, with three-year survival rates of 68%, 41%, 33%, and 0.03% for non-elevated, elevated, normalized, and non-normalized CRP groups respectively. When compared with the non-elevated group, hazard ratios of death were 1.69, 2.57, and 10.34 in the normalized, elevated, and non-normalized groups (P < 0.001). Conclusions Baseline CRP and CRP kinetics may be useful to predict the prognosis of metastatic NPC patients treated with palliative chemotherapy and facilitate individualized treatment. A prospective study to validate this prognostic model is still needed however.

c-Src activation promotes nasopharyngeal carcinoma metastasis by inducing the epithelial-mesenchymal transition via PI3K/Akt signaling pathway: A new and promising target for NPC

Aberrant activation of cellular Src (c-Src), a non-receptor tyrosine kinase, could promote cancer progression through activating its downstream signaling pathways. However, the roles of c-Src and phosphorylated-Src (p-Src) in nasopharyngeal carcinoma (NPC) progression are rarely investigated. Herein, we have identified high c-Src concentrations in the serum of NPC patients with distant metastasis using high-throughput protein microarrays. Levels of c-Src in serum and p-Src in human primary NPC samples were unfavorable independent prognostic factors for cancer-specific survival, disease-free survival, and distant metastasis-free survival. Depletion or inactivation of c-Src in NPC cells using sgRNA with CRISPR/Cas9 system or PP2 decreased cell viability, colony formation, migration and invasion in vitro and metastasis in vivo. In contrast, these malignancies could be up-regulated by overexpressed c-Src in a NPC cell line with low-metastasis potential. Furthermore, p-Src was involved in promoting NPC cell metastasis by inducing the epithelial-mesenchymal transition (EMT) process via activating the PI3K/Akt pathway and cytoskeleton remodeling. The p-Src-induced EMT process could be retarded by PP2, which mediated by down-regulating the PI3K/Akt pathway. In conclusion, elevated levels of c-Src in serum and p-Src in primary NPC tissue correlated with poor outcomes of NPC patients. And aberrant activation of c-Src facilitated NPC cells with malignant potential, especially metastasis ability, which mediated by the PI3K/Akt pathway activation and sequentially induced the EMT process. These findings unveiled a promising approach for targeted therapy of advanced NPC.

A prognostic model predicts the risk of distant metastasis and death for patients with nasopharyngeal carcinoma based on pre-treatment interleukin 6 and clinical stage ☆

Because inflammation plays a critical role in nasopharyngeal carcinoma (NPC), this study aims to investigate the correlation between the pro-inflammation cytokine interleukin-6 (IL6) and the prognosis of NPC and develop a new prognostic model. IL6 levels were measured in the serum of 290 NPC patients by ELISA and the correlation between IL6 and prognosis of NPC was evaluated by Kaplan-Meier analysis and multivariate analysis. The results showed that elevated IL6 levels were positively correlated with poorer 9-year overall survival (OS), disease-free survival (DFS), distant metastasis-free survival (DMFS) and lung metastasis-free survival (lung-MFS). IL6 level was an independent prognostic factor for OS, DFS, DMFS and lung-MFS. The CI-model based on TNM stage and IL6 level could better predict the OS, DFS, DMFS and lung-MFS of NPC patients. Here, the newly developed prognostic CI-model for predicting distant metastasis and death of NPC patients could facilitate patients consulting and individualized immunotherapy.

Comparison of the short-term efficacy between docetaxel plus carboplatin and 5-fluorouracil plus carboplatin in locoregionally advanced nasopharyngeal carcinoma

Objective Platinum-based chemotherapy in combination with radiotherapy is a standard treatment strategy for locoregionally advanced nasopharyngeal carcinoma (NPC). This study aimed to investigate the long-term efficacy and tolerability of inductive chemotherapy with docetaxel plus carboplatin (TC) or 5-fluorouracil plus carboplatin (FC) followed by concurrent radiation therapy in patients with NPC. Methods Patients (N=88) were randomized to receive TC or FC as inductive therapy followed by concurrent radiotherapy (60–70 Gy) with two cycles of carboplatin (area under the curve =5 mg·h/L). Patients were followed up for 8 years. Primary end point was progression-free survival (PFS). Secondary end points included overall survival (OS), toxicity, tumor response, distant metastasis-free survival, and local recurrence-free survival. Results At the end of the follow-up period, 31 patients died, 32 had disease progression, eleven had cancer recurrence, and 25 had distant metastasis. Overall, there was no difference between treatment groups with regard to response or survival. We found that following induction and concurrent chemoradiotherapy, the majority of patients showed a complete response (~96%–98% for induction therapy and 82%–84% for comprehensive therapy) to both therapies. PFS and OS were also similar between groups. The rate of PFS was 63.6% for both FC and TC and that of OS was 65.9% and 63.5%, respectively. The overall incidence of grade 3–4 adverse events in the TC group (20.5%) was higher than in the FC group (10.7%). Neutropenia and leukopenia were the most common grade 3–4 adverse events in the TC group, and mucositis was the most common in the FC group. Conclusion These data indicate that TC and FC therapies have similar efficacy in treating locally advanced NPC and both are well tolerated.

The plasma Epstein–Barr virus DNA level guides precision treatment for nasopharyngeal carcinoma in the intensity-modulated radiotherapy era: a large population-based cohort study from an endemic area

Background: In the intensity-modulated radiotherapy (IMRT) era, the survival benefit of concurrent chemotherapy for locoregionally advanced nasopharyngeal carcinoma (LA-NPC) remains undetermined. This study aimed to evaluate the benefits of IMRT with concurrent chemotherapy compared with IMRT alone for LA-NPC patients with different plasma Epstein–Barr virus (EBV) DNA levels. Methods: Patients were identified from a prospectively maintained database in an endemic area between November 2002 and December 2013. Cox proportional hazards models, propensity score matching, and inverse probability weighting models were established for survival analysis. Stratification analysis was performed based on interaction effects analysis. Finally, sensitivity analysis was performed considering unmeasured confounders. Results: A total of 1357 eligible patients were enrolled (median follow up 62.4 months; range 3.5–155.8 months). No significant survival differences were observed between groups in the entire cohort. Notably, a significant interaction effect was observed between treatment regimens and EBV DNA levels. In patients with high EBV DNA levels (>4000 copies/ml), all three models showed that IMRT with concurrent chemotherapy significantly improved overall survival [hazard ratio (HR) 2.521, 95% confidence interval (CI) 1.218–5.216], disease-free survival (HR 2.168, 95% CI 1.349–3.483), and distant metastasis-free survival (HR 2.331, 95% CI 1.194–4.551) compared with IMRT alone. No differences were found in patients with low EBV DNA levels. Sensitivity analysis confirmed the robustness of the results. Conclusion: In the IMRT era, concurrent chemotherapy treatment of LA-NPC patients with high EBV DNA levels is reasonable. However, the optimal regimen for LA-NPC patients with low EBV DNA levels needs further validation in randomized clinical trials.

Concurrent Chemoradiotherapy versus Intensity-modulated Radiotherapy Alone for Elderly Nasopharyngeal Carcinoma Patients with Pre-treatment Epstein-Barr Virus DNA: A Cohort Study in an Endemic Area with Long-term Follow-up.

Purpose: To date, no guidelines exist for elderly nasopharyngeal carcinoma (NPC) patients (60 years of age or older) due to a lack of prospective clinical trials. This study evaluated the efficacy of concurrent chemotherapy (CCRT) for NPC in elderly patients treated with intensity-modulated radiotherapy (IMRT). Methods: Patients were identified from a prospectively maintained database. A total of 198 consecutive cases of elderly patients with NPC receiving IMRT, including 103 patients treated with IMRT plus CCRT and 95 patients treated with IMRT alone, were analysed from January 2002 to December 2013. Multivariate analysis (MVA) using the Cox proportional hazards model and propensity score analysis (PSA) were performed for overall survival (OS) and disease-free survival (DFS). Finally, sensitivity analysis was performed. Results: The median follow-up time was 55.3 months (range, 3-135.6 months). In the entire cohort, both MVA and PSA models showed that compared with IMRT alone, IMRT plus CCRT significantly improved survival (hazard ratio [HR] 2.143, 95% confidence interval [95% CI] 1.180-3.890; HR 1.961, 95% CI, 1.117-3.443, for OS and DFS, respectively). Similar results were found in the subgroups with high levels of Epstein-Barr virus (EBV) DNA, except in the low-EBV-DNA cohort. The total rates of severe acute toxicity, including leukopenia, neutropenia, stomatitis, and emesis, were significantly higher in the IMRT+CCRT group than in the IMRT-alone group (P < 0.001) but were similar to the rates of severe late toxicity (P = 0.818). Sensitivity analysis confirmed the robustness of our analysis. Conclusions: In the era of IMRT, CCRT retained survival benefits at high EBV DNA levels but not at low EBV DNA levels for elderly NPC patients. Randomized clinical trials are needed to confirm our findings.

The prognostic significance of carcinoma-associated fibroblasts and tumor-associated macrophages in nasopharyngeal carcinoma

Purpose Tumor stroma cells play an important role in the carcinogenesis and progression of cancer. The aim of the present investigation was to explore the predictive role of carcinoma-associated fibroblasts (CAFs) and tumor-associated macrophages (TAMs) in nasopharyngeal carcinoma (NPC). Patients and methods The densities of CAFs and TAMs were measured by immunohistochemistry staining for α-smooth muscle actin (α-SMA), CD68, and CD163 in two sets of tissue microarrays including 260 pretreatment NPC tissues, that is, a training test comprising of 152 patients and a validation set comprising of 108 patients. Chi-square tests were performed for comparisons among the groups. Survival rates were estimated by using the Kaplan–Meier method and compared with log-rank tests. Cox proportional hazards models were used to identify significant independent variables. Results Patients older than 50 years showed a lower expression of CD68, and there was a positive relationship between the densities of CAFs and CD163+ TAMs (p=0.001). In the multivariate analysis of the training test, both α-SMA and CD163 were independent prognostic factors for overall survival and progression-free survival (all p<0.05). Based on the expression levels of α-SMA and CD163, patients were categorized into three groups: high-risk, intermediate-risk, and low-risk groups according to both high, either high, and both low, respectively. Survival analysis and Cox multivariate analysis showed that the risk groups based on α-SMA and CD163 expression were independent predictors for the survival of patients with NPC in the training test, which was also confirmed by the validation test. Conclusion A patient’s risk group based on the level of CD163+ TAMs and CAFs was an independent predictor of survival, which may facilitate patient counseling and individualized treatment.

Effect of induction chemotherapy with cisplatin, fluorouracil, with or without taxane on locoregionally advanced nasopharyngeal carcinoma: a retrospective, propensity score-matched analysis

BackgroundAvailable data in the literature comparing different induction chemotherapy (IC) regimens on locoregionally advanced nasopharyngeal carcinoma (NPC) are scarce. The purpose of the present study was to evaluate the outcomes of locoregionally advanced NPC patients who were treated with taxane, cisplatin and 5-fluorouracil (TPF) or cisplatin and 5-fluorouracil (PF) as IC followed by concurrent chemoradiotherapy (CCRT).MethodsIn total, 1879 patients with locoregionally advanced NPC treated with IC and CCRT from a prospectively maintained database were included in the present observational study. We compared overall survival (OS), disease-specific survival (DSS), distant metastasis-free survival (DMFS), and locoregional relapse-free survival, using the propensity score method.ResultsIn total, 1256 patients received TPF or PF as IC backbone. The TPF group showed significantly better OS (hazard ratio [HR], 0.660; 95% confidence interval [CI] 0.442–0.986; P = 0.042), DSS (HR, 0.624; 95% CI 0.411–0.947; P = 0.027) and DMFS (HR, 0.589; 95% CI 0.406–0.855; P = 0.005) compared with the PF group in multivariable analyses. Propensity score matching identified 294 patients in each cohort and confirmed that TPF was associated with significantly improved 5-year OS (88.1% vs. 80.7%; P = 0.042), DSS (88.5% vs. 80.7%; P = 0.021) and DMFS (87.9% vs. 78.6%; P = 0.012) rates compared with the PF group. There were no significant differences in locoregional relapse-free survival before or after matching.ConclusionsIn our study, IC with the TPF regimen combined with CCRT showed improved long-term survival for the patients with locoregionally advanced NPC compared with the PF regimen. However, a prospective randomized clinical trial to validate these findings is necessary.

A phase II trial of induction NAB-paclitaxel and cisplatin followed by concurrent chemoradiotherapy in patients with locally advanced nasopharyngeal carcinoma

OBJECTIVES Nanoparticle albumin-bound paclitaxel (NAB-paclitaxel) was designed to avoid solvent-related toxicities, and improve anti-tumor efficacy via increasing paclitaxels intratumoral concentration and its uptake by tumor cells. This trial aimed to determine the safety and efficacy of induction NAB-paclitaxel combined with cisplatin followed by concurrent chemoradiotherapy (CCRT) in patients with locally advanced nasopharyngeal carcinoma (LA-NPC). PATIENTS AND METHODS Patients with stage III-IVb NPC received NAB-paclitaxel (260mg/m2) combined with cisplatin (80mg/m2) intravenously on days 1 and 22, followed by cisplatin (80mg/m2) on days 43 and 64, concomitant with intensity-modulated radiation therapy. This trial is registered with the Chinese Clinical Trials Registry, number ChiCTR-ONC-12002615. RESULTS From July 2010 to November 2013, 36 eligible patients with nonmetastatic stage III-IVb NPC were enrolled. The objective response rates were 97.2% (eight complete responses [CRs] and 27 partial responses [PRs]) and 100% (30 CRs and six PRs) after two cycles of induction chemotherapy (ICT) and CCRT, respectively. With a median follow-up time of 45months, the estimated 3-year progression-free survival and cancer-specific survival were 86.1% (95% confidence interval [CI], 69.8-99.8%) and 91.7% (95% CI, 68.9-100.0%), respectively. The most frequent grade 3-4 toxicities were neutropenia (8.6%) and nausea (8.6%) after ICT and thrombocytopenia (34.3%) and leukopenia (28.6%) after CCRT. CONCLUSION NAB-paclitaxel combined with cisplatin as an ICT regimen showed encouraging anti-tumor effects and manageable toxicities in LA-NPC. Further randomized controlled trials in phase III of NAB-paclitaxel in patients with LA-NPC are warranted.