A single nucleotide polymorphism in the Epstein-Barr virus genome is strongly associated with a high risk of nasopharyngeal carcinoma.

BACKGROUND: Epstein-Barr virus (EBV) commonly infects the general population and has been associated with nasopharyngeal carcinoma (NPC), which has a high incidence in certain regions. This study aimed to address how EBV variations contribute to the risk of NPC. METHODS: Using logistic regression analysis and based on the sequence variations at EBV-encoded RPMS1, a multi-stage association study was conducted to identify EBV variations associated with NPC risk. A protein degradation assay was performed to characterize the functional relevance of the RPMS1 variations. RESULTS: Based on EBV-encoded RPMS1 variations, a single nucleotide polymorphism (SNP) in the EBV genome (locus 155391: G>A, named G155391A) was associated with NPC in 157 cases and 319 healthy controls from an NPC endemic region in South China [P < 0.001, odds ratio (OR) = 4.47, 95% confidence interval (CI) 2.71-7.37]. The results were further validated in three independent cohorts from the NPC endemic region (P < 0.001, OR = 5.20, 95% CI 3.18-8.50 in 168 cases vs. 241 controls, and P < 0.001, OR = 5.27, 95% CI 4.06-6.85 in 726 cases vs. 880 controls) and a non-endemic region (P < 0.001, OR = 7.52, 95% CI 3.69-15.32 in 58 cases vs. 612 controls). The combined analysis in 1109 cases and 2052 controls revealed that the SNP G155391A was strongly associated with NPC (P(combined) < 0.001, OR = 5.27, 95% CI 4.31-6.44). Moreover, the frequency of the SNP G155391A was associated with NPC incidence but was not associated with the incidences of other EBV-related malignancies. Furthermore, the protein degradation assay showed that this SNP decreased the degradation of the oncogenic RPMS1 protein. CONCLUSIONS: Our study identified an EBV variation specifically and significantly associated with a high risk of NPC. These findings provide insights into the pathogenesis of NPC and strategies for prevention.

Direct sequencing and characterization of a clinical isolate of Epstein-Barr virus from nasopharyngeal carcinoma tissue by using next-generation sequencing technology.

Epstein-Barr virus (EBV)-encoded molecules have been detected in the tumor tissues of several cancers, including nasopharyngeal carcinoma (NPC), suggesting that EBV plays an important role in tumorigenesis. However, the nature of EBV with respect to genome width in vivo and whether EBV undergoes clonal expansion in the tumor tissues are still poorly understood. In this study, next-generation sequencing (NGS) was used to sequence DNA extracted directly from the tumor tissue of a patient with NPC. Apart from the human sequences, a clinically isolated EBV genome 164.7 kb in size was successfully assembled and named GD2 (GenBank accession number HQ020558). Sequence and phylogenetic analyses showed that GD2 was closely related to GD1, a previously assembled variant derived from a patient with NPC. GD2 contains the most prevalent EBV variants reported in Cantonese patients with NPC, suggesting that it might be the prevalent strain in this population. Furthermore, GD2 could be grouped into a single subtype according to common classification criteria and contains only 6 heterozygous point mutations, suggesting the monoclonal expansion of GD2 in NPC. This study represents the first genome-wide analysis of a clinical isolate of EBV directly extracted from NPC tissue. Our study reveals that NGS allows the characterization of genome-wide variations of EBV in clinical tumors and provides evidence of monoclonal expansion of EBV in vivo. The pipeline could also be applied to the study of other pathogen-related malignancies. With additional NGS studies of NPC, it might be possible to uncover the potential causative EBV variant involved in NPC.

Nasopharyngeal carcinoma risk prediction via salivary detection of host and Epstein-Barr virus genetic variants.

Genetic susceptibility and Epstein-Barr virus (EBV) infection are important etiological factors in nasopharyngeal carcinoma (NPC). In this study, in southern China, where NPC is endemic, a single nucleotide polymorphism (SNP) in the EBV-encoded RPMS1 gene (locus 155391: G > A [G155391A]) and seven host SNPs (rs1412829, rs28421666, rs2860580, rs2894207, rs31489, rs6774494, and rs9510787) were confirmed to be significantly associated with NPC risk in 50 NPC cases versus 54 hospital-based controls with throat washing specimens and 1925 NPC cases versus 1947 hospital-based controls with buffy coat samples, respectively. We established a strategy to detect the NPC-associated EBV and host SNPs using saliva samples in a single test that is convenient, noninvasive, and cost-effective and displays good compliance. The potential utility of this strategy was tested by applying a risk prediction model integrating these EBV and host genetic variants to a population-based case-control study comprising 1026 incident NPC cases and 1148 controls. Receiver operating characteristic (ROC) curve analysis revealed an area under the curve of the NPC risk prediction model of 0.74 (95% CI: 0.71-0.76). Net reclassification improvement (NRI) analysis showed that inclusion of the EBV SNP significantly improved the discrimination ability of the model (NRI = 0.30, P < 0.001), suggesting the promising value of EBV characteristics for identifying high-risk NPC individuals in endemic areas. Taken together, we developed a promising NPC risk prediction model via noninvasive saliva sampling. This approach might serve as a convenient and effective method for screening the population with high-risk of NPC.

A GWAS Meta-analysis and Replication Study Identifies a Novel Locus within CLPTM1L/TERT Associated with Nasopharyngeal Carcinoma in Individuals of Chinese Ancestry.

BACKGROUND: Genetic loci within the major histocompatibility complex (MHC) have been associated with nasopharyngeal carcinoma (NPC), an Epstein-Barr virus (EBV)-associated cancer, in several GWAS. Results outside this region have varied. METHODS: We conducted a meta-analysis of four NPC GWAS among Chinese individuals (2,152 cases; 3,740 controls). Forty-three noteworthy findings outside the MHC region were identified and targeted for replication in a pooled analysis of four independent case-control studies across three regions in Asia (4,716 cases; 5,379 controls). A meta-analysis that combined results from the initial GWA and replication studies was performed. RESULTS: In the combined meta-analysis, rs31489, located within the CLPTM1L/TERT region on chromosome 5p15.33, was strongly associated with NPC (OR = 0.81; P value 6.3 × 10(-13)). Our results also provide support for associations reported from published NPC GWAS-rs6774494 (P = 1.5 × 10(-12); located in the MECOM gene region), rs9510787 (P = 5.0 × 10(-10); located in the TNFRSF19 gene region), and rs1412829/rs4977756/rs1063192 (P = 2.8 × 10(-8), P = 7.0 × 10(-7), and P = 8.4 × 10(-7), respectively; located in the CDKN2A/B gene region). CONCLUSIONS: We have identified a novel association between genetic variation in the CLPTM1L/TERT region and NPC. Supporting our finding, rs31489 and other SNPs in this region have been reported to be associated with multiple cancer sites, candidate-based studies have reported associations between polymorphisms in this region and NPC, the TERT gene has been shown to be important for telomere maintenance and has been reported to be overexpressed in NPC, and an EBV protein expressed in NPC (LMP1) has been reported to modulate TERT expression/telomerase activity. IMPACT: Our finding suggests that factors involved in telomere length maintenance are involved in NPC pathogenesis.

Nigericin selectively targets cancer stem cells in nasopharyngeal carcinoma.

Nasopharyngeal carcinoma (NPC) is prevalent in southern China, northern Africa, and Alaska. The prognosis for NPC patients at early stage is good, while it is poor for patients at late stages. Cancer stem cells (CSCs) have been proposed to be associated with tumor initiation, relapse and metastasis, and the poor prognosis of NPC likely results from residual CSCs after therapy. Study on the therapy targeting CSCs in NPC remains poor, though it received intensive attentions in other cancers. Here, we used NPC cell lines with high and low proportion of CSCs as models to explore the effect of nigericin, an antibiotic, on CSCs. We found that nigericin could selectively target CSCs and sensitize CSCs in NPC to the widely used clinical drug cisplatin both in vitro and in vivo. Moreover, downregulation of the polycomb group protein Bmi-1 may contribute to the inhibitory effect of nigericin on CSCs. Furthermore, by using the in vitro NPC cell models, we found that nigericin could significantly decrease the migration and invasion abilities, which are known to be associated with CSCs. Taken together, our results suggest that nigericin can selectively target CSCs in NPC, which could be a candidate CSCs targeting drug for clinical evaluation.