Clinical Management Of Nasopharyngeal Carcinoma Research Articles

Inhibition of MNK pathway sensitizes nasopharyngeal carcinoma to radiotherapy.

Improving the clinical management of nasopharyngeal carcinoma (NPC) is an unmet need owing to the high incidence of treatment failure caused by radioresistance. In our study, we observed increased phosphorylation of translation initiation factor 4E (eIF4E), regulated by MAP kinase-interacting kinase (MNK), in NPC cells following irradiation treatment. Using siRNA to deplete MNK, we found that radiation-induced eIF4E phosphorylation was eliminated, NPC cell sensitivity to radiation was enhanced, and radioresistant NPC cell viability was reduced. Furthermore, we tested three pharmacological MNK inhibitors (eFT508, CGP57380, and cercosporamide) and found that they were effective against radioresistant NPC cells and synergized with irradiation. In-vivo experiments confirmed that eFT508, at a tolerable dose, inhibited the growth of radioresistant NPC and synergized with radiation in a radiosensitive NPC xenograft model. Our research highlights the activation of MNK-mediated survival mechanisms in NPC in response to radiotherapy and the potential of combining radiation with MNK inhibitors as a sensitizing strategy. Notably, eFT508 is currently being investigated in clinical trials for cancer treatment, and our findings may prompt the initiation of clinical trials using eFT508 in radioresistant NPC patients.

Depletion of SOD2 enhances nasopharyngeal carcinoma cell radiosensitivity via ferroptosis induction modulated by DHODH inhibition

BackgroundRecurrence due to the development of radioresistance remains a major challenge in the clinical management of nasopharyngeal carcinoma. The objective of this study was to increase the sensitivity of nasopharyngeal carcinoma cells to ionizing radiation by enhancing oxidative stress and ferroptosis caused by disrupting the mitochondrial anti-oxidant enzyme system.MethodsOxidative stress cell model was constructed by SOD2 knockdown using shRNA. The expression and activity of DHODH was suppressed by siRNA and brequinar in SOD2 depleted cells. Protein levels were determined by western blotting and ferroptosis was assessed by C11 BODIPY and malondialdehyde assay. Cell viability was evaluated using CCK-8 assay while radiotoxicity was assessed by colony formation assay. Cellular ATP level was determined by ATP assay kits, ROS was determined by DCFD and DHE, while mitochondrial oxygen consumption was determined by seahorse assay. Data were analyzed by two-tailed independent t-test.ResultsRadiation upregulated SOD2 expression and SOD2 depletion increased cellular O2.−, malondialdehyde, and the fluorescence intensity of oxidized C11 BODIPY. It also resulted in mitochondrial damage. Its depletion decreased colony formation both under ionizing and non-ionizing radiation conditions. The ferroptosis inhibitor, deferoxamine, rescued cell viability and colony formation in SOD2 depleted cells. Cellular level of malondialdehyde, fluorescence intensity of oxidized C11 BODIPY, O2.− level, ATP, and mitochondrial oxygen consumption decreased following DHODH inhibition in SOD2 depleted cells. Cell viability and colony formation was rescued by DHODH inhibition in SOD2 depleted cells.ConclusionInducing oxidative stress by SOD2 inhibition sensitized nasopharyngeal carcinoma cells to ionizing radiation via ferroptosis induction. This was found to be dependent on DHODH activity. This suggests that DHODH inhibitors should be used with caution during radiotherapy in nasopharyngeal carcinoma patients.

Cell-free EBV DNA as a biomarker during clinical management of nasopharyngeal carcinoma in a nonendemic region.

Nasopharyngeal carcinoma (NPC) is the most common malignant tumor of the nasopharynx. Although NPC is not endemic in India,higher incidences were observed in its North-Eastern regions particularly Sikkim, Nagaland, Manipur, and Mizoram. Early detection of NPC is difficult because the nasopharynx is not readily amenable to clinical examination and symptoms of NPC are nonspecific.The development of suitable biomarkers for early diagnosis of NPC as well as accurate monitoring of treatment response is needed urgently. In this exploratory pilot study, we have investigated the clinical significance of assessing plasma Epstein-Barr virus (EBV) DNA load at diagnosis and during treatment. We found that EBV DNA is detectable at diagnosis in the majority of patients with nonendemic NPC and the absolute copy number of circulating EBV DNA per milliliter increases progressively with the stage of the disease. The viral load declined significantly with induction chemotherapy and definitive chemoradiation but showed a sharp rise at relapse. Patients with EBV DNA levels ≥1500 copies/ml had a higher risk of disease progression or relapse when compared with patients who had EBV DNA <1500 copies/ml at baseline. Estimation of plasma EBV DNA may serve as an excellent noninvasive tool to monitor disease extent, response to therapy, and for better prediction of future relapse or progression-free survival in a nonendemic NPC patient population.

LncRNA linc00312 suppresses radiotherapy resistance by targeting DNA-PKcs and impairing DNA damage repair in nasopharyngeal carcinoma

Radioresistance is the main obstacle in the clinical management of nasopharyngeal carcinoma (NPC). linc00312 is deregulated in a number of human cancers, including NPC. However, the detailed functions and underlying mechanisms of linc00312 in regulating radiosensitivity of NPC remains unknown. In this study, cox regression analysis was used to assess the association between linc00312 and NPC patients’ survival after radiotherapy. Our results reveal that linc00312 is significantly down-regulated in NPC tissues and patients with higher expression of linc00312 are significantly associated with longer overall survival and better short-term radiotherapy efficacy. Overexpression of linc00312 could increase the sensitivity of NPC cells to ionizing radiation, as indicated by clonogenic survival assay, comet assay, and flow cytometry. Mechanistically, RNA pull down and RNA immunoprecipitation were performed to investigate the binding proteins of linc00312. linc00312 directly binds to DNA-PKcs, hinders the recruitment of DNA-PKcs to Ku80, and inhibits phosphorylation of AKT–DNA–PKcs axis, therefore inhibiting the DNA damage signal sensation and transduction in the NHEJ repair pathway. In addition, linc00312 impairs DNA repair and cell cycle control by suppressing MRN–ATM–CHK2 signal and ATR–CHK1 signal. In summary, we identified DNA-PKcs as the binding protein of linc00312 and revealed a novel mechanism of linc00312 in the DNA damage response, providing evidence for a potential therapeutic strategy in NPC.

Effects of baohuoside-I on epithelial-mesenchymal transition and metastasis in nasopharyngeal carcinoma.

To investigate the effect of baohuoside-I against nasopharyngeal carcinoma (NPC) and its underlying mechanism, baohuoside-I was employed to treat NPC cell lines CNE1 and CNE2 in vitro, followed by attachment and detachment assays to evalute the epithelial-mesenchymal transition (EMT) phenotype markers. Baohuoside-I was also administered to experimental mice to assess its effect on xenograft tumor growth and NPC cell metastasis. A microRNA (miRNA, miR) microarray was performed to screen for miRNA altered by baohuoside-I in NPC cells. Bioinformatic tools and luciferase activity assay was conducted to identify the downstream molecules mediating the anti-tumor property of baohuoside-I. Baohuoside-I inhibited EMT and metastasis and upregulated miR-370-3p in NPC cells, which was shown to directly recognize and inhibit expression of Hedgehog pathway component Smoothened (SMO). Baohuoside-I suppresses metastasis as well as EMT of NPC cells through targeting the Hedgehog pathway component SMO, and may serve as a potent anti-tumor agent in the clinical management of NPC.

Abstract 2254: CCL2/CCR2 axis signaling promotes metastasis of nasopharyngeal carcinoma by activating ERK1/2 pathway via upregulating MMP2 and MMP9

Abstract Despite tremendous progress in diagnosis and treatments of nasopharyngeal carcinoma (NPC) has been established nowadays, distant metastasis remains the major cause of death.The underlying molecular mechanisms are largely unknown. In the present study,we investigated the roles in NPCs for an axis of candidate metastasis-related genes located at 17q11.2-q12, chemokine C-C motif ligand 2 (CCL2), and located at 3p21.31, chemokine C-C motif receptor 2 (CCR2). ELISA analyses of human serum from 50 cases with NPC and 50 cases without NPC revealed the mean serum CCL2 concentration of cases with NPC was significantly higher than cases without NPC (P&amp;lt;0.001).Higher expression levels of CCL2 and CCR2 were frequently detected in NPCs compared with normal nasopharyngeal epithelium samples by IHC analyses, and both ectopic expressions were significantly associated with NPC metastasis (P = 0.039, p = 0.016, respectively) and poor overall survival (P = 0.011, p = 0.004, respectively). Overexpressions of CCL2 and CCR2 were observed in human strong-metastatic NPC cell line S18 originated from CNE2 and 58F originated from SUNE1. Correspondingly, relatively lower expressions of CCL2 and CCR2 were detected in human weak-metastatic NPC cell line S26 originated from CNE2 and SUNE1 with weaker-metastatic ability compared to 58F. Functional studies demonstrated that upregulating the expressions of CCL2 and CCR2 respectively in S26 and SUNE1 could effectively promote cell migration and invasion in vitro, and enhance tumor distant metastasis in vivo of nude mice, which could be further raised in cells overexpressing CCR2 with exogenous CCL2 in the medium. When CCL2 and CCR2 were silenced by shRNAs respectively in S18 and 58F, decrease of cell migration and invasion was showed and significantly fewer metastases were observed. However, adding exogenous CCL2 to the medium culturing CCR2-silencing cells could not rescue their weaken ability of migration and invasion and metastasis. Mechanistic investigations suggested that, dual overexpressions of CCL2/CCR2 could remarkably activate extracellular signal-regulated kinase (ERK1/2) signaling pathway, which sequentially induced matrix metalloproteinase (MMP)2/9 transcriptional activation to facilitate tumor metastasis. Taken as a whole, CCL2-CCR2 axis signaling plays a pivotal role in NPC metastasis by activating ERK1/2 pathway via upregulating MMP2 and MMP9, and this finding may help to identify novel therapeutic targets for a better clinical management of NPC. Citation Format: Jing Yang, Xing Lv, Yanqun Xiang, Xiang Guo, Jina Chen, Xinyuan Guan. CCL2/CCR2 axis signaling promotes metastasis of nasopharyngeal carcinoma by activating ERK1/2 pathway via upregulating MMP2 and MMP9. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2254. doi:10.1158/1538-7445.AM2015-2254

Can Epstein-Barr virus DNA load in nasopharyngeal brushings or whole blood predict recurrent nasopharyngeal carcinoma in a non-endemic region? A prospective nationwide study of the Dutch Head and Neck Oncology Cooperative Group.

This study estimated the value of quantitative measurements of EBV markers in the clinical management of nasopharyngeal carcinoma in a non-endemic area. The aim was to predict prognosis and detect recurrent and residual disease. In 72 patients, EBV DNA load in blood and nasopharyngeal brushes, and IgA VCA-p18 and EBNA1 in plasma were measured at different time points. At diagnosis and post-treatment, a cut-off value was used for detecting disease [positive (PPV) and negative (NPV) predictive value]. The markers were correlated as a continuous variable with tumor stage, disease-free survival (DFS) and overall survival (OS). The Cox hazard ratio model assessed hazard ratios. At diagnosis, the markers were above the COV in 45, 92, 85 and 83% of the patients, respectively. Post-treatment, DNA load test in blood and brush had the best discriminating power (blood DNA load test: PPV 39% and NPV 97%, brush for local disease: PPV 75% and NPV 99%). Post-treatment, DNA load in blood was the best predictor for OS and DFS [hazard ratio 3.2 (95% CI 1.51-3.5) and 2.3 (95% CI 1.72-5.8)]. Assessing the EBV DNA load in blood has significant prognostic value, although the clinical value is for discussion. The EBV DNA load in the brush might improve early detection of local failures post-treatment.

Epstein-Barr virus-encoded RNAs as a survival predictor in nasopharyngeal carcinoma.

Epstein-Barr virus (EBV) infection is one of the most important factors of nasopharyngeal carcinoma (NPC) endemic areas. Transcription of EBV-encoded non-polyadenylated RNAs (EBERs) are presented in most of NPC tumors. Exploring EBERs as a prognostic marker for NPC might further be informative about the biology and the progression of the disease. The aim of this study was to analyze the role of EBV latency in the clinical management of nasopharyngeal carcinoma (NPC), by detecting EBERs. RNA in situ hybridization (ISH) for detecting EBERs was carried out on 908 NPC tumor tissues. Overall survival (OS) curves were analyzed with the Kaplan-Meier method and the Cox proportional-hazards regression models. The median follow-up time was 70 months (1-120 months). Eight hundred and sixteen (89.9%) from a total of 908 consecutive NPC cases were found to be EBV-EBER positive. EBER-ISH staining revealed nuclear localization in NPC cells. In the Kaplan-Meier analysis for OS, high EBER expression levels in NPC patients were statistically significant positive prognostic factors for survival (log-rank, P = 0.022), especially in adults aged 17-40 years (P = 0.023) and in those with advanced stage disease (log-rank, P = 0.002). Cox proportional-hazards regression model analysis showed that the EBER expression level was an independent risk factor for OS (hazard ratio 0.724, P = 0.005). EBERs were frequently detected in NPC tumor tissues, and high-level EBER expression correlated with good prognosis in NPC patients, especially in adult patients and in those with advanced stage disease. EBER may serve as a potential prognostic predictor in NPC.

Selective Application of a Test Based on Risk: Using Limited Resources Wisely

Selective Application of a Test Based on Risk: Using Limited Resources Wisely

Validation of Roche LightCycler Epstein-Barr Virus Quantification Reagents in a Clinical Laboratory Setting

Epstein-Barr virus (EBV) is associated with a wide range of benign and malignant diseases, including infectious mononucleosis, lymphoma, posttransplant lymphoproliferative disorder, and nasopharyngeal carcinoma. Measurement of EBV viral load in plasma is increasingly used for rapid assessment of disease status. We evaluated the performance characteristics of an EBV polymerase chain reaction assay that uses commercial reagents and instruments from Roche Diagnostics (Indianapolis, IN). DNA was extracted from plasma using a MagNaPure instrument, and viral load was measured by real-time polymerase chain reaction on a LightCycler. Analyte-specific reagents included primers and hybridization probes targeting the EBV LMP2 gene and a spiked control sequence. Accuracy and reproducibility were established using DNA from three cell lines. The assay was sensitive to approximately 750 copies of EBV DNA per milliliter of plasma and was linear across at least four orders of magnitude. The assay detected EBV DNA in three of five samples from nasopharyngeal carcinoma patients, seven of nine infectious mononucleosis samples, and 34/34 samples from immunosuppressed patients with clinically significant EBV-related disease, whereas EBV DNA was undetectable in plasma from 21 individuals without EBV-related disease. In conclusion, this LightCycler EBV assay is rapid, sensitive, and linear for quantifying EBV viral load. The assay appears to be useful for measuring clinically significant EBV levels in immunodeficient patients.

Role of Epstein-Barr virus DNA measurement in plasma in the clinical management of nasopharyngeal carcinoma in a low risk area

Objective: To evaluate the role of quantitative measurement of Epstein-Barr virus (EBV) DNA in the clinical management of nasopharyngeal carcinoma (NPC) in a low tumour risk area (western Europe). Methods:...

Prognostic value of EBV markers in the clinical management of nasopharyngeal carcinoma (NPC): a multicenter follow-up study.

Between December 1979 and April 1982, 373 patients with untreated, undifferentiated carcinoma of the nasopharynx (NPC), 99 in Hong Kong, 120 in Tunis and 154 in Villejuif, entered a longitudinal study aimed at determining the clinical prognostic value of EBV serology after radiotherapy. A minimum of 3 years' follow-up was achieved for 319 patients (83 in Tunis, 95 in Hong Kong and 141 in Villejuif) who had regular clinical and serological testing at intervals of 6-8 months. No significant difference in initial serology (i.e., before any treatment) or variations of antibody titers at time of first follow-up was observed between patients who achieved complete remission after radiotherapy and those who did not. This included IgG and IgA antibodies to VCA, EA or EBNA. However, when patients with confirmed clinical remission 1 year after completion of radiotherapy were studied, the value of IgG/EA and mainly of IgA/EA increasing titers became highly significant for prediction of relapse, regardless of the initial titers. This demonstrated the clinical usefulness of EBV serology for NPC patients who have confirmed clinical remission after radiotherapy.