Nonendemic Nasopharyngeal Carcinoma Research Articles

24P TPD54 as a candidate biomarker for distant metastasis prediction in non-endemic nasopharyngeal carcinoma: Label-free quantitative proteomics results

24P TPD54 as a candidate biomarker for distant metastasis prediction in non-endemic nasopharyngeal carcinoma: Label-free quantitative proteomics results

Comparing BamHI-W and CE-marked assays to detect circulating Epstein-Barr Virus (EBV) DNA of nasopharyngeal cancer patients in a non-endemic area

ObjectivesPlasma Epstein-Barr Virus (EBV)-DNA is a well-established prognostic biomarker in nasopharyngeal carcinoma (NPC). Different methods for assessment include single-copy gene targeted, European Conformity (CE)-marked assays, which are mostly employed in non-endemic settings, vs multiple-copy gene targeted, in-house BamHI-W based assays, which currently represent the most widely used method for EBV-DNA quantification. To date, evidence concerning the commutability of these different assays is still limited. Materials and methodsFrom August 2016 to March 2018, 124 plasma and 124 whole blood (WB) samples from 93 NPC patients were collected at different time-points for each patient. EBV-DNA viral load was quantified in pre- (n = 12) and post-treatment (n = 9), follow-up (n = 53), and recurrent/metastatic (R/M) (n = 50) phase. For each sample, one in-house BamHI-W vs three different CE-marked plasma assays were compared; the performance of plasma vs WB matrix was also assessed. Quantitative agreement of EBV-DNA values was evaluated by linear correlation and Bland-Altman analysis. ResultsA statistically significant (p = 0.0001) agreement between all CE-marked and the BamHI-W assays was found using plasma matrix, regardless of clinical phase. The results obtained in copies/ml were comparable to those expressed in IU/ml. When using WB matrix, the number of positive detections increased in the post-treatment phase. ConclusionsOur retrospective comparison supported an agreement between Plasma BamHI-W and CE-marked assays in measuring EBV-DNA for non-endemic NPC patients. There were no significant interferences from different measurement units (IU/ml vs copies/ml). Further evaluations are needed to better clarify the role of WB.

EBV genome variations enhance clinicopathological features of nasopharyngeal carcinoma in a non-endemic region.

Nasopharyngeal carcinoma (NPC) is caused by infection with Epstein–Barr virus (EBV) and endemic in certain geographic regions. EBV lytic gene, BALF2, closely associates with viral reactivation and BALF2 gene variation, the H‐H‐H strain, causes NPC in endemic region, southern China. Here, we investigate whether such EBV variations also affect NPC in a non‐endemic region, Japan. Viral genome sequencing with 47 EBV isolates of Japanese NPC were performed and compared with those of other EBV‐associated diseases from Japan or NPC in Southern China. EBV genomes of Japanese NPC are different from those of other diseases in Japan or endemic NPC; Japanese NPC was not affected by the endemic strain (the BALF2 H‐H‐H) but frequently carried the type 2 EBV or the strain with intermediate risk of endemic NPC (the BALF2 H‐H‐L). Seven single nucleotide variations were specifically associated with Japanese NPC, of which six were present in both type 1 and 2 EBV genomes, suggesting the contribution of the type 2 EBV‐derived haplotype. This observation was supported by a higher viral titer and stronger viral reactivation in NPC with either type 2 or H‐H‐L strains. Our results highlight the importance of viral strains and viral reactivation in the pathogenesis of non‐endemic NPC.

Immunotherapy in Nonendemic Nasopharyngeal Carcinoma: Real-World Data from Two Nonendemic Regions.

Background: nasopharyngeal carcinoma (NPC) is a complex disease entity that mainly predominates in endemic regions. Real-world data with immunotherapy from nonendemic regions are limited. Methods: we collected data from patients with recurrent/metastatic (R/M) NPC treated at a center in Greece and 8 centers in Italy. Between 2016 and 2021, 46 patients who were treated with at least one cycle of immune checkpoint inhibitors (ICI) were identified. Herein, we present our results and a review of the literature. Results: assessment of response was available in 42 patients. Overall, 11 patients responded to immunotherapy (Overall Response Rate-ORR 26.2%). Three patients had complete response (CR), and 8 patients had partial response (PR). Disease control rate (DCR) was 61.9%. Median Progression Free Survival (PFS) was 5.6 months and median Overall Survival (OS) was 19.1 months. Responders to ICI improved PFS and OS as compared to that of nonresponders. A lower probability of responding to ICI was shown in patients with more than three metastatic sites (p = 0.073), metastatic disease at initial diagnosis, (p = 0.039) or EBV DNA positive before ICI initiation, (p = 0.074). Decline in EBV DNA levels was found to be statistically significant associated with best response to ICI (p = 0.049). Safety was manageable. Conclusions: among 46 patients with R/M NPC treated with immunotherapy in two nonendemic regions, ORR was 26.2% and durable responses were observed. Low disease burden could serve as a biomarker for response to ICI.

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.

Neoadjuvant chemotherapy improves survival compared with concurrent chemoradiation alone in nasopharyngeal carcinoma patients with N3 disease

Neoadjuvant chemotherapy (NAC) trials in endemic regions of nasopharyngeal carcinoma (NPC) found improved survival, but studies are lacking in nonendemic regions. We assessed whether adding NAC to concurrent chemoradiation (CRT) improves overall survival (OS), especially in high-risk nonendemic patients. Definitively treated NPC patients (n = 5424) from the National Cancer Database were analyzed for predictors of NAC and NAC effects on OS with multivariate Cox proportional hazards analysis (multivariate analysis [MVA]). Propensity score matched (1:2) survival analysis of NAC (n = 968) and CRT alone (n = 1914) was also performed. Effects on OS were stratified by risk group. On MVA, NAC-improved OS among the total cohort (hazard ratio [HR] 0.89, P = .049), particularly among stratified keratinizing histology (HR 0.82, P = .015) and N3 disease (HR 0.73, P = .046). Among propensity matched patients, NAC improved OS in patients with N3 disease (n = 336; HR 0.71, P = .046). NAC may improve OS among nonendemic NPC patients at higher risk of distant micrometastases, particularly N3 disease and those with unfavorable histology.

MRI-based radiomics nomogram may predict the response to induction chemotherapy and survival in locally advanced nasopharyngeal carcinoma.

To establish and validate a radiomics nomogram for prediction of induction chemotherapy (IC) response and survival in nasopharyngeal carcinoma (NPC) patients. One hundred twenty-three NPC patients (100 in training and 23 in validation cohort) with multi-MR images were enrolled. A radiomics nomogram was established by integrating the clinical data and radiomics signature generated by support vector machine. The radiomics signature consisting of 19 selected features from the joint T1-weighted (T1-WI), T2-weighted (T2-WI), and contrast-enhanced T1-weighted MRI images (T1-C) showed good prognostic performance in terms of evaluating IC response in two cohorts. The radiomics nomogram established by integrating the radiomics signature with clinical data outperformed clinical nomogram alone (C-index in validation cohort, 0.863 vs 0.549; p < 0.01). Decision curve analysis demonstrated the clinical utility of the radiomics nomogram. Survival analysis showed that IC responders had significant better PFS (progression-free survival) than non-responders (3-year PFS 84.81% vs 39.75%, p < 0.001). Low-risk groups defined by radiomics signature had significant better PFS than high-risk groups (3-year PFS 76.24% vs 48.04%, p < 0.05). Multiparametric MRI-based radiomics could be helpful for personalized risk stratification and treatment in NPC patients receiving IC. • MRI Radiomics can predict IC response and survival in non-endemic NPC. • Radiomics signature in combination with clinical data showed excellent predictive performance. • Radiomics signature could separate patients into two groups with different prognosis.

Expression of toll-like receptors in non-endemic nasopharyngeal carcinoma

BackgroundNasopharyngeal carcinoma (NPC) is a malignant disease with an enigmatic etiology. NPC associates with Epstein-Barr virus (EBV) and human papillomaviruses (HPVs), while immunological factors also play a role in carcinogenesis. Toll-like receptors (TLRs) are pattern recognition receptors that participate in the immunological defence against pathogens, but their functions are also linked to cancer.MethodsIn our whole population-based study, we retrieved 150 Finnish NPC cases and studied their tumour samples for TLR1, TLR2, TLR4, TLR5, TLR7, and TLR9 expressions by immunohistochemistry, and for the presence of EBV and high-risk HPVs with EBV RNA and HPV E6/E7 mRNA in situ hybridizations. In addition, we analyzed the TLR expression patterns according to age, tumour histology, EBV/HPV status, and outcome.ResultsWe found that all TLRs studied were highly expressed in NPC. Viral status of the tumours varied, and 62% of them were EBV-positive, 14% HPV-positive, and 24% virus-negative. The tumours with strong TLR2nucl or TLR5 expression were mostly virus-negative or HPV-positive keratinizing squamous cell carcinomas, and the patients with these tumours were significantly older than those with mild or negative TLR2nucl/TLR5 expression. In Kaplan-Meier analysis, the patients with strong TLR5 expression had worse survival compared to the patients with negative or mild TLR5 expression, but the results were linked to other patient and tumour characteristics. In multivariable-adjusted Cox regression analysis, the patients with positive TLR7 tumour expression had better overall survival than those with no TLR7 expression. The 5-year overall survival rates according to TLR7 expression were 66% (mild), 52% (moderate or strong), and 22% (negative).ConclusionsTLRs are highly expressed in non-endemic NPC. Intensity of TLR2 and TLR5 expressions correlate with viral status, and TLR7 seems to be an independent prognostic factor of non-endemic NPC.

Tumor-infiltrating lymphocytes associate with outcome in nonendemic nasopharyngeal carcinoma: a multicenter study

The prognostic significance of tumor-infiltrating lymphocytes (TILs) has been studied recently in many cancers. For the first time in a nonendemic region, we have evaluated the prognostic value of TILs in a whole population-based nationwide cohort of nasopharyngeal carcinoma (NPC) in Finland. A total of 115 cases from Finnish hospitals were included. TILs were analyzed using hematoxylin and eosin-stained slides according to the criteria of the International Immuno-Oncology Biomarker Working Group. TILs were evaluated separately in stromal and tumor compartments. The log-rank test and univariable and multivariable analyses were used to compare survival in patients with tumors with low and high TILs. A significant positive correlation was observed between the occurrence of intratumoral and stromal TILs (P < .001). In multivariable analysis, NPC cases with low intratumoral TILs had poor overall survival with a hazard ratio (HR) of 2.55 and 95% confidence interval (95% CI) of 1.60 to 4.05 (P < .001). Cases with low intratumoral TILs also had poor disease-specific survival (HR, 2.02; 95% CI, 1.16-3.52; P = .015). Keratinized tumors with low intratumoral TILs were associated with an even poorer overall survival (HR, 3.94; 95% CI, 2.17-7.15; P < .001) and a poor disease-specific survival (HR, 2.97; 95% CI, 1.46-6.05; P = .009). Our study demonstrates that the evaluation of TILs is simple and can be assessed routinely in NPC.

Conservation and polymorphism of EBV RPMS1 gene in EBV-associated tumors and healthy individuals from endemic and non-endemic nasopharyngeal carcinoma areas in China

As a member of the BamHI-A rightward transcripts family of the Epstein-Barr virus (EBV), RPMS1 expression has been confirmed in all EBV-associated tumors. However, few studies have investigated the single-nucleotide polymorphisms (SNPs) of RPMS1, and only one SNP site (g155391a) has been reported to be associated with nasopharyngeal carcinoma occurrence. The objective of this study was to investigate the polymorphism of RPMS1 in EBV-associated tumors (gastric carcinoma, nasopharyngeal carcinoma, and lymphoma). In this research, nested-PCR was performed to analyze DNA sequences of 420 EBV-associated samples. Phylogenetic analysis revealed four RPMS1 genotypes (RPMS1-A, RPMS1-B, RPMS1-C, and RPMS1-D). A significant difference (p < 0.05) among northern and southern China samples was observed. Furthermore, there was a significant difference between EBV-associated tumors and healthy controls for RPMS1 (p < 0.05). These findings demonstrated that RPMS1 variation was not only tumor-specific but also geographically restricted in EBV-associated samples.

The Relationship Between Environmental Factors and the Profile of Epstein-Barr Virus Antibodies in the Lytic and Latent Infection Periods in Healthy Populations from Endemic and Non-Endemic Nasopharyngeal Carcinoma Areas in China

Our previous study found that smoking was associated with an elevated level of the antibody against VCA in the Epstein-Barr virus (EBV) lytic phase, which was an important predictive marker of the risk of nasopharyngeal carcinoma (NPC). It remained unknown whether environmental factors were associated with the levels of other EBV antibodies, such as Zta-IgA, EA-IgA, EBNA1-IgA, and LMP1-IgA, in the lytic and latent infection periods. We aimed to investigate the possible environmental inducers that could affect EBV antibody levels in two independent healthy male populations from endemic NPC areas in South China (N=1498) and non-endemic NPC areas in North China (N=1961). We performed ELISA and immunoenzymatic assays to test the levels of antibodies specific to the EBV antigens. The seropositive rates of antibodies against the antigens expressed in both the EBV latent and lytic infection periods, namely, LMP1-IgA, EBNA1-IgA, and Zta-IgA, in endemic areas (28.65%, 5.43% and 14.49%, respectively) were significantly higher than those in non-endemic areas (14.43%, 1.07% and 6.32%, respectively). Smoking was associated with higher seropositivity for EBNA1-IgA (OR=1.47, 95% CI=1.12–1.93) and Zta-IgA (OR=1.28, 95% CI=0.99–1.66), with dose-response effects, while not associated with the levels of LMP1-IgA. In conclusion, smoking was an important environmental factor, which associated with increased levels of EBNA1-IgA, and Zta-IgA.

Association of Human Papillomavirus Infection and p16 status in Nonendemic Nasopharyngeal Carcinoma: Incidence, Prevalence, and Prognostic Role

Association of Human Papillomavirus Infection and p16 status in Nonendemic Nasopharyngeal Carcinoma: Incidence, Prevalence, and Prognostic Role

Induction chemotherapy with docetaxel, cisplatin and fluorouracil followed by concurrent chemoradiotherapy or chemoradiotherapy alone in locally advanced non-endemic nasopharyngeal carcinoma

ObjectivesTo evaluate the efficacy of induction chemotherapy with docetaxel, cisplatin and fluorouracil (TPF) followed by concurrent chemoradiotherapy (IC+CCRT) or CCRT alone in non-endemic locally advanced nasopharyngeal carcinoma (NPC) patients. Materials and methodsData of 106 patients with NPC treated from January 1999 to June 2012 with IC+CCRT (n=58) or CCRT alone (n=48) were retrospectively reviewed. ResultsMedian follow-up was 6.4years. Distribution of age, performance status, stage and concurrent chemotherapy regimen were imbalanced between the two groups. The 5-year overall survival (OS) and progression-free survival (PFS) were not significantly different between IC+CCRT and CCRT groups (OS: 78.3% vs. 82.7%, p=0.77; PFS: 72.5% vs. 68.2%, p=0.81, respectively). There were less total cumulative incidence of grade 3–4 late radiation morbidity in the IC+CCRT group (44.8% vs. 70.8%, p=0.01). Five-year OS for patients with post-IC complete response (CR), partial response (PR) and stable disease (SD) sub-groups were 100%, 79.4% and 60%, respectively. ConclusionCompared with CCRT alone, IC (TPF regimen)+CCRT did not improve OS or PFS in patients with NPC, but less grade 3–4 late toxicities were observed. Responsiveness of IC may provide additional prognostic information.

Prognostic Model of Death and Distant Metastasis for Nasopharyngeal Carcinoma Patients Receiving 3DCRT/IMRT in Nonendemic Area of China.

Few studies were conducted to explore the prognostic factors for nonendemic nasopharyngeal carcinoma (NPC) in the era of 3-dimensional conformal radiation therapy (3DCRT)/intensity-modulated radiation therapy (IMRT). The aim of this study was to evaluate the potential prognostic factors for nonendemic NPC.Between January 2004 and December 2011, a total of 393 nonendemic NPC patients receiving 3DCRT/IMRT were reviewed according to the inclusion and exclusion criteria. The prognostic factors we analyzed included age, T stage, N stage, lymph node diameter, primary tumor volume, WHO histology types, and cranial nerve related symptoms. All patients were staged according to the 7th edition of the American Joint Committee on Cancer (AJCC) system. The factors found to be associated with the endpoints by univariate analyses were then entered into multivariate Cox proportional hazards regression analysis.The median follow-up time was 61.4 months (range: 4–130 months). The 5-year local recurrent-free survival (LRFS), nodal relapse-free survival (NRFS), distant metastasis free survival (DMFS), and disease-specific survival (DSS) for all patients were 89.3%, 96.4%, 73.5%, and 74.3%, respectively. Multivariate analysis indicated that N stage (N2–3), WHO pathologic type II, and primary tumor volume (>23 mL) were 3 independent prognostic factors for DSS and DMFS. According to the number of prognostic factors, patients were divided into 3 risk groups: low-risk group (patients without any risk factors); intermediate-risk group (patients with only 1 risk factor); and high-risk group (patients with more than 2 risk factors). The 5-year DSS for low, intermediate, and high-risk groups were 91.5%, 75.2%, and 49.3%, respectively (P < 0.001). The 5-year DMFS for low, intermediate, and high-risk groups were 89.4%, 77.9%, and 49.4%, respectively (P < 0.001).Advanced N stage (N2–3), larger tumor volume (>23 mL), and histological WHO type II are independently prognostic factors for nonendemic NPC patients in China.

Characterization of the Variability of Epstein-Barr Virus Genes in Nasopharyngeal Biopsies: Potential Predictors for Carcinoma Progression.

Epstein-Barr virus (EBV) infection is a significant factor in the pathogenesis of nasopharyngeal carcinoma, especially in the undifferentiated carcinoma of nasopharyngeal type (UCNT, World Health Organization type III), which is the dominant histopathological type in high-risk areas. The major EBV oncogene is latent membrane protein 1 (LMP1). LMP1 gene shows variability with different tumorigenic and immunogenic potentials. EBV nuclear antigen 1 (EBNA1) regulates progression of EBV-related tumors; however, the influence of EBNA1 sequence variability on tumor pathogenesis is controversial. The aims of this study were to characterize polymorphisms of EBV genes in non-endemic nasopharyngeal carcinoma biopsies and to investigate potential sequence patterns that correlate with the clinical presentation of nasopharyngeal carcinoma. In total, 116 tumor biopsies of undifferentiated carcinoma of nasopharyngeal type (UCNT), collected from 2008 to 2014, were evaluated in this study. The genes EBNA2, LMP1, and EBNA1 were amplified using nested-PCR. EBNA2 genotyping was performed by visualization of PCR products using gel electrophoresis. Investigation of LMP1 and EBNA1 included sequence, phylogenetic, and statistical analyses. The presence of EBV DNA was significantly distributed between TNM stages. LMP1 variability showed six variants, with the detection of the first China1 and North Carolina variants in European nasopharyngeal carcinoma biopsies. Newly discovered variants Srb1 and Srb2 were UCNT-specific LMP1 polymorphisms. The B95-8 and North Carolina variants are possible predictors for favorable TNM stages. In contrast, deletions in LMP1 are possible risk factors for the most disfavorable TNM stage, independent of EBNA2 or EBNA1 variability. A newly discovered EBNA1 subvariant, P-thr-sv-5, could be a potential diagnostic marker, as it represented a UCNT-specific EBNA1 subvariant. A particular combination of EBNA2, LMP1, and EBNA1 polymorphisms, type 1/Med/P-thr was identified as a possible risk factor for TNM stage IVB or progression to the N3 stage.

Long-term patterns of relapse and survival following definitive intensity-modulated radiotherapy for non-endemic nasopharyngeal carcinoma

We report treatment outcomes for a large non-endemic cohort of patients with nasopharyngeal carcinoma treated with intensity-modulated radiotherapy (IMRT) and chemotherapy. We identified 177 consecutive patients with newly diagnosed, non-metastatic nasopharyngeal cancer treated with definitive IMRT between 1998 and 2011. Endpoints included local, regional, distant control, and overall survival. Median follow-up was 52months. The 3-/5-year actuarial rates of local control, regional control, distant control, and overall survival were 92%/83%, 93%/91%, 86%/83%, and 87%/74%, respectively. The median time to local recurrence was 30months; the annual hazard of local recurrence did not diminish until the 6th year of follow-up. Overall, we observed excellent rates of disease control and survival consistent with initially reported results from our institution. Attaining locoregional control in patients with extensive primary tumors remains a significant clinical challenge. With mature follow-up we observed that more than half of observed local relapses occurred after 2years, a pattern distinct from that of carcinomas arising from other head and neck sites. These findings raise the possibility that patients with NPC may benefit from close follow-up during post-treatment years 3-5.

High prevalence of the EBER variant EB-8m in endemic nasopharyngeal carcinomas.

Epstein-Barr virus (EBV)-encoded small RNAs (EBERs) are the most highly expressed transcripts in all EBV-associated tumors and are involved in both lymphoid and epithelioid carcinogenesis. Our previous study on Chinese isolates from non-endemic area of nasopharyngeal carcinoma (NPC) identified new EBER variants (EB-8m and EB-10m) which were less common but relatively more frequent in NPC cases than healthy donors. In the present study, we determined the EBER variants in NPC cases and healthy donors from endemic and non-endemic areas of NPC within China and compared the EBER variants, in relation to the genotypes at BamHI F region (prototype F and f variant), between population groups and between two areas. According to the phylogenetic tree, four EBER variants (EB-6m, EB-8m, EB-10m and B95-8) were identified. EB-6m was dominant in all population groups except for endemic NPC group, in which EB-8m was dominant. EB-8m was more common in endemic NPC cases (82.0%, 41/50) than non-endemic NPC cases (33.7%, 32/95) (p<0.0001), and it was also more frequent in healthy donors from endemic area (32.4%, 24/74) than healthy donors from non-endemic area (1.1%, 1/92) (p<0.0001). More importantly, the EB-8m was more prevalent in NPC cases than healthy donors in both areas (p<0.0001). The f variant, which has been suggested to associate with endemic NPC, demonstrated preferential linkage with EB-8m in endemic isolates, however, the EB-8m variant seemed to be more specific to NPC isolates than f variant. These results reveal high prevalence of EBER EB-8m variant in endemic NPC cases, suggesting an association between NPC development and EBV isolates carrying EB-8m variant. Our finding identified a small healthy population group that shares the same viral strain which predominates in NPC cases. It could be interesting to carry extensive cohort studies following these individuals to evaluate the risk to develop NPC.

Expression of Epstein-Barr Virus-encoded Latent Membrane Protein (LMP-1), p16 and p53 Proteins in Nonendemic Nasopharyngeal Carcinoma (NPC): A Clinicopathological Study

Although the latent membrane protein type 1 (LMP1) is frequently expressed in Epstein-Barr virus (EBV) malignancies, its contribution to the pathogenesis of nasopharyngeal carcinoma (NPC) is not fully defined. LMP1 functions as a viral mimic of the TNFR family member engaging a number of signaling pathways that induce morphological and phenotypic alterations. This study aimed to investigate the LMP1 expression and EBV infection in relation to clinical outcome and survival in a series of Mexican NPC patients. We also studied expression of p16 and p53 proteins. We analyzed in 25 tumor specimens the expression of LMP1, p16 and p53 by immunohistochemistry (IHC) and EBV presence by IHC/in situ hybridization. Differences in clinical outcome and survival in relation to protein expression were correlated through χ(2) statistics and Kaplan-Meier survival curves. Our results showed a rate of 92% (23/25) of EBV infection. The expressions of LMP-1, p16 and p53 proteins were 40.0, 44.0 and 40.0%, respectively. LMP-1 immunoexpression was more common in older patients (>50 vs. <50 years old, p = 0.02) and with parapharyngeal space invasion (p = 0.02). The presence of metastatic disease at diagnosis (p = 0.03), distant recurrence disease (p = 0.006) and shorter distance recurrence-free survival (p = 0.05) was associated with lack of p16. In our series, EBV infection rates are particularly high for nonendemic NPC, although without a statistically significant difference in overall survival, LMP1 and p16 expression was correlated with poorer clinical prognosis. Probably, LMP1 and p16 detection identify a worse clinical prognosis in NPC patient subgroup.

Nonendemic HPV-Positive Nasopharyngeal Carcinoma: Association With Poor Prognosis

To investigate the relationship between human papillomavirus (HPV) and Epstein-Barr virus (EBV) in nonendemic nasopharyngeal carcinoma (NPC) and assess the prognostic implications of viral status. Paraffin-embedded tumor specimens from 62 patients with primary NPC diagnosed between 1985 and 2011 were analyzed for EBV and high-risk HPV. EBV status was determined by the use of in situ hybridization for EBV encoded RNA. HPV status was assessed with p16 immunohistochemistry and multiplex polymerase chain reaction MassArray for determination of HPV type. Proportional hazards models were used to compare the risk of death among patients as stratified by viral status. Of 61 evaluable tumors, 26 (43%) were EBV-positive/HPV-negative, 18 (30%) were HPV-positive/EBV-negative, and 17 (28%) were EBV/HPV-negative. EBV and HPV infection was mutually exclusive. HPV positivity was significantly correlated with World Health Organization grade 2 tumors, older age, and smoking (all P<.001). The racial distribution of the study population was 74% white, 15% African American, and 11% Asian/Middle Eastern. Among HPV-positive patients, 94% were white. At a median follow-up time of 7 years, HPV-positive and EBV/HPV-negative tumors exhibited worse outcomes than did EBV-positive tumors, including decreased overall survival (hazard ratio [HR] 2.98, P=.01; and HR 3.89, P=.002), progression-free survival (HR 2.55, P=.02; and HR 4.04, P<.001), and locoregional control (HR 4.01, P=.03; and HR 6.87, P=.001). In our Midwestern population, high-risk HPV infection may play an etiologic role in the development of nonendemic, EBV-negative NPC. Compared with EBV-positive NPC, HPV-positive and EBV/HPV-negative NPC are associated with worse outcomes. A larger confirmatory study is needed to validate these findings.

Expression modes and clinical manifestations of latent membrane protein 1, Ki‐67, cyclin‐B1, and epidermal growth factor receptor in nonendemic nasopharyngeal carcinoma

We aimed to identify clinical significance of latent membrane protein 1 (LMP1), Ki-67, cyclin-B1, and epidermal growth factor receptor (EGFR), in nonendemic nasopharyngeal carcinoma (NPC). The relation between expression of the markers in 45 NPC specimens and clinicopathological and survival variables was statistically analyzed. LMP1 was present in 33% of the tumors, and its presence was associated with advanced nodal and disease stages. Overexpression was defined as labeling index > or = median value for Ki-67, > or = 15% for cyclin-B1, and > or =50% for EGFR, and it was displayed in 50%, 55%, and 80% of the specimens, respectively. Strong EGFR staining intensity and not overexpression of the 3 markers was the variable with statistically significant impact on treatment outcomes in terms of worse local and locoregional tumor control rates. Our results suggest that the evaluation of EGFR staining intensity in patients with NPC may identify a subgroup of patients with poor prognosis.