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Epstein-Barr virus (EBV) C promoter (Cp) hypermethylation, a crucial factor for EBV latent infection of nasopharyngeal epithelial cells, has been recognized as a promising biomarker for nasopharyngeal carcinoma (NPC) detection. In this study, we develop a novel EBV Cp methylation quantification (E-CpMQ) assay and evaluate its diagnostic performance for NPC detection. A novel qPCR assay for simultaneous quantification of methylated- and unmethylated EBV Cp was developed by the combinational modification of MethyLight and QASM, with an innovative calibrator to improve the detection accuracy and consistency. The NP swab samples and synthetic standards were used for the analytical validation of the E-CpMQ. The diagnostic efficacy of the developed E-CpMQ assay was validated in 137 NPC patients and 137 non-NPC controls. The E-CpMQ assay can detect the EBV Cp methylation ratio in one reaction system under 10 copies with 100 % recognition specificity, which is highly correlated to pyrosequencing with a correlation coefficient over 0.99. The calibrated E-CpMQ assay reduces the coefficient of variation by an average of 55.5 % with a total variance of less than 0.06 units standard deviation (SD). Linear methylation ratio detection range from 4.76 to 99.01 %. The sensitivity and specificity of the E-CpMQ respectively are 96.4 % (95 % CI: 91.7–98.8 %), 89.8 % (95 % CI: 83.5–94.3 %). The developed E-CpMQ assay with a calibrator enables accurate and reproducible EBV Cp methylation ratio quantification and offers a sensitive, specific, cost-effective method for NPC early detection. [ABSTRACT FROM AUTHOR]

Previous studies have demonstrated strong associations between host genetic factors and Epstein–Barr virus (EBV) VCA‐IgA with the risk of nasopharyngeal carcinoma (NPC). However, the specific interplay between host genetics and EBV VCA‐IgA on NPC risk is not well understood. In this two‐stage case–control study (N = 4804), we utilized interaction and mediation analysis to investigate the interplay between host genetics (genome‐wide association study‐derived polygenic risk score [PRS]) and EBV VCA‐IgA antibody level in the NPC risk. We employed a four‐way decomposition analysis to assess the extent to which the genetic effect on NPC risk is mediated by or interacts with EBV VCA‐IgA. We consistently found a significant interaction between the PRS and EBV VCA‐IgA on NPC risk (discovery population: synergy index [SI] = 2.39, 95% confidence interval [CI] = 1.85–3.10; replication population: SI = 3.10, 95% CI = 2.17–4.44; all pinteraction < 0.001). Moreover, the genetic variants included in the PRS demonstrated similar interactions with EBV VCA‐IgA antibody. We also observed an obvious dose–response relationship between the PRS and EBV VCA‐IgA antibody on NPC risk (all ptrend < 0.001). Furthermore, our decomposition analysis revealed that a substantial proportion (approximately 90%) of the genetic effects on NPC risk could be attributed to host genetic‐EBV interaction, while the risk effects mediated by EBV VCA‐IgA antibody were weak and statistically insignificant. Our study provides compelling evidence for an interaction between host genetics and EBV VCA‐IgA antibody in the development of NPC. These findings emphasize the importance of implementing measures to control EBV infection as a crucial strategy for effectively preventing NPC, particularly in individuals at high genetic risk. [ABSTRACT FROM AUTHOR]

Background Epstein-Barr virus (EBV) reactivation from latent to lytic infection has been considered as a key step in nasopharyngeal carcinoma oncogenesis. However, epidemiological evidence regarding environmental risk factors for EBV reactivation on a population level remains largely lacking. Methods We enrolled 1916 randomly selected adults from the general population of Guangdong and Guangxi, China, from 2010 to 2014. Information on environmental factors was collected via a structured interview. Serum immunoglobulin A antibodies against EBV viral capsid antigen and nuclear antigen 1 were measured by enzyme-linked immunosorbent assay to evaluate EBV reactivation status. We used logistic regression to calculate odds ratios (ORs) with 95% confidence intervals (CIs) for the associations of EBV reactivation with various environmental factors. Results No associations were observed between EBV reactivation and extensive environmental factors, including alcohol or tea drinking, a history of chronic ear/nose/throat diseases, use of medications or herbs, consumption of salted fish or preserved foods, oral hygiene, sibship structure, and various residential and occupational exposures. Only cigarette smoking was associated with EBV reactivation (current smokers vs never smokers; OR = 1.37; 95% CI = 1.02–1.83), with positive exposure-response trends with increasing intensity, duration, and pack-years of smoking. Conclusions Consistent with previous studies, we found an association between cigarette smoking and EBV reactivation. Other examined exposures were not associated with EBV reactivation. These null results could suggest either more complex interactions between exposures and EBV reactivation or a predominant role of host and/or viral genetic variation. [ABSTRACT FROM AUTHOR]

Introduction: Prospective evidence for herbal diet and nasopharyngeal carcinoma (NPC) development is absent. We therefore evaluated the associations of herbal soup and herbal tea with NPC in a prospective cohort study in southern China. Methods: Based on an NPC screening cohort established in 2008–2015, information on herbal diet consumption, potential confounding factors, and Epstein-Barr virus (EBV) antibody levels were collected from 10,179 individuals aged 30–69 years in Sihui city, southern China. Cox regression models were performed to examine herbal diet with NPC risk, and logistic regression models were used to examine herbal diet with EBV reactivation. Results: During a median of 7.54 years of follow-up, 69 participants developed NPC. Herbal soup consumption was associated with decreased NPC risk, with HRs of 0.31 (95% confidence interval (CI): 0.15–0.62) for the highest intake frequency and 0.29 (95% CI: 0.16–0.51) for a longer duration. However, herbal tea was not significantly associated. Moreover, we identified herbal soup was inversely associated with EBV seropositivity among all the participants at baseline, with the adjusted ORs being 0.78 (95% CI: 0.65–0.93) for immunoglobulin A antibodies against EBV capsid antigens (VCA-IgA) and 0.76 (95% CI: 0.64–0.91) for nuclear antigen 1 (EBNA1-IgA) in those with the highest frequency and 0.70 (95% CI: 0.59–0.84) for VCA-IgA and 0.64 (95% CI: 0.54–0.77) for EBNA1-IgA in those with the longer duration. Inverse associations were also observed in non-NPC individuals. Conclusions: With inhibition of EBV reactivation by plants, herbal soup could significantly decrease the risk of NPC in endemic areas. [ABSTRACT FROM AUTHOR]

Using Epstein-Barr virus (EBV)-based markers to screen populations at high risk for nasopharyngeal carcinoma (NPC) is an attractive preventive approach. Here, we develop a comprehensive risk score (CRS) that combines risk effects of EBV and human genetics for NPC risk stratification and validate this CRS within an independent, population-based dataset. Comparing the top decile with the bottom quintile of CRSs, the odds ratio of developing NPC is 21 (95% confidence interval: 12–37) in the validation dataset. When combining the top quintile of CRS with EBV serology tests currently used for NPC screening in southern China, the positive prediction value of screening increases from 4.70% (serology test alone) to 43.24% (CRS plus serology test). By identifying individuals at a monogenic level of NPC risk, this CRS approach provides opportunities for personalized risk prediction and population screening in endemic areas for the early diagnosis and secondary prevention of NPC. Epstein-Barr virus (EBV) is associated with high risk for nasopharyngeal carcinoma (NPC). Here, the authors develop a comprehensive risk score that combines risk effects of EBV and human genetics for NPC risk stratification. [ABSTRACT FROM AUTHOR]

Objective: Plasma Epstein-Barr virus (EBV) DNA is considered a biomarker for nasopharyngeal carcinoma (NPC). However, its long-term role in NPC development is unclear.Materials and Methods: A total of 1363 participants seropositive for EBV VCA-IgA and EBNA1-IgA in a community-based NPC screening program in southern China were tested for plasma EBV DNA levels by real-time qPCR between 2008 and 2015. New NPC cases were confirmed by active follow-up approach and linkage to local cancer registry through the end of 2016. Cox proportional hazards regression analysis was performed to calculate the hazard ratios (HRs) for NPC risk with plasma EBV DNA.Results: Thirty patients were newly diagnosed during a median 7.5 years follow-up. NPC incidence increased with the plasma EBV DNA load ranging from 281.46 to 10,074.47 per 100,000 person-years in participants with undetectable and ≥ 1000 copies/ml levels; the corresponding cumulative incidence rates were 1.73 and 50%. Furthermore, plasma EBV DNA loads conferred an independent risk for NPC development after adjustment for other risk factors, with HRs of 7.63 for > 3-999 copies/ml and 39.79 for ≥1000 copies/ml. However, the HRs decreased gradually after excluding NPC cases detected in the first 2 to 3 years and became statistically nonsignificant by excluding cases detected during the first 4 years.Conclusion: Elevated plasma EBV DNA can predict NPC risk over 3 years. Monitoring plasma EBV DNA can be used as a complementary approach to EBV serological antibody-based screening for NPC. [ABSTRACT FROM AUTHOR]

Background: The association between smoking and nasopharyngeal carcinoma (NPC) is still uncertain. The aim of this study was to validate smoking effect on NPC and explore if smoking can induce NPC by persistently reactivating EBV in long‐term based on a prospective cohort design. Methods: A NPC screening cohort with 10 181 eligible residents in Sihui city, southern China was conducted from 2008 to 2015. The smoking habit was investigated through the trained interviewers and EBV antibodies (VCA‐IgA, EBNA1‐IgA) as screening markers were tested periodically. New NPC cases were identified through local cancer registry. Cox's regression model was used to estimate the adjusted hazard ratios (aHRs) of smoking on NPC incidence. In the non‐NPC participants, the associations between smoking and EBV seropositivity in different periods were assessed by logistic regression and generalized estimating equations (GEE). Results: With a median of 7.54 years, 71 NPCs were diagnosed ≥1 year after recruitment. Compared with never smokers, the aHRs of developing NPC among ever smokers were 3.00 (95%CI: 1.46‐6.16). Stratified by sex, the HRs of ever smoking were 2.59 (95%CI: 1.07‐6.23) for male and 3.75 (95%CI: 1.25‐11.20) for female, respectively. Among the non‐NPC individuals, ever smoking was not only associated with EBV seropositivity at baseline, but also in the 3‐5 years of follow up, with adjusted odds ratios (aORs) of 1.68 (95%CI: 1.29‐2.18) for VCA‐IgA and 1.92 (95%CI: 1.42‐2.59) for EBNA1‐IgA. Among the smokers who were tested EBV antibodies at least twice, the similar results were obtained using GEE. Conclusion: Smoking could significantly increase the long‐term risk of NPC in southern China, partly by persistently reactivating EBV. [ABSTRACT FROM AUTHOR]

Background: Oral Epstein-Barr virus (EBV) status reflects host EBV activity and potentially links to EBV-associated diseases, however, factors influencing oral EBV loads or reactivation, such as environmental exposures or host factors, are not fully understood.Methods: A 2-stage, multicenter, cross-sectional study of 6558 subjects from 21 administrative cities of southern China and 3 populations from representative geographical areas in China (referred to as the south, north, and northeastern populations) was performed. The relationships between demographical factors and environmental exposures to EBV loads were analyzed by logistic regression models.Results: Current smoking, with a dose-response effect, was found to be strongly associated with higher oral EBV loads in the pooled data, with an odds ratio of 1.58 (95% confidence interval, 1.39-1.79), as well as in each of the separate populations. The odds ratio increased to 3.06 when current smokers in southern China were compared to never smokers in northern China. Additionally, higher oral EBV loads tended to be detected in older participants, male participants, and participants in southern China.Conclusions: This study provided evidence linking the effect of host-environmental factors, particularly smoking, to oral EBV activity. It could strengthen our understanding of the possible causal roles of EBV-related diseases, which may help to prevent or mitigate EBV-associated diseases. [ABSTRACT FROM AUTHOR]

Abstract: The link of nasopharyngeal carcinoma (NPC) with Epstein‐Barr virus (EBV) has been established for decades. Although an abnormal high level of EBV sero‐antibody spectrum and cell‐free circulating EBV DNA loads were exhibited in NPC patients, oral EBV DNA loads, which are primarily responsible for the EBV transmission, has not been previously studied in NPC patients. We conducted an epidemiological study to measure the oral EBV loads, viral components, and the relationship with the serum antibody titers in a large case‐control population (968 cases and 1656 controls) and a family‐based population (91 cases and 165 unaffected family members). EBV DNA loads were detected by quantitative PCR approach targeting the BamHI‐W region. Although a large individualized variation existed, we still observed a decreased oral EBV DNA loads in the population of NPC patients compared to that of healthy controls (ORs were 1.00, 0.69, 0.62, 0.33 classified by the quartiles of viral loads, Ptrend < .001) and family members. In contrast, the elevated levels of oral EBV loads were present in asymptomatic males and elders, suggesting a different important source for EBV transmission. Notably, oral EBV loads were inversely associated with serum antibody titers of VCA‐IgA, EA‐IgA (All Ptrend < .001) in the cases but not in the controls. Our study provides the first epidemiological data of oral EBV loads and viral components in NPC patients and controls in the highest risk area of Southern China, indicating that NPC status is unlikely to be an important determinant of EBV transmission. [ABSTRACT FROM AUTHOR]

Epstein-Barr virus (EBV) is one of the risk factors of nasopharyngeal carcinoma (NPC), and understanding the modifiable risk factors of EBV activation is crucial in the prevention of NPC. In this study, we aimed to investigate the association between solid fuel use and EBV seropositivity in a high-risk area of NPC. Our study was based on the baseline findings from an ongoing population-based prospective cohort in Sihui county in Southern China. We explored the association between current use of solid fuel in cooking and EBV seropositivity, and NPC-related EBV activation, using logistic regression models. Stratification analyses were further conducted to assess potential effect modifiers. We also examined the impact of frequency and duration of solid fuel use, and switch in fuel types, on EBV seropositivity among ever users. Of the 12,579 participants included in our analysis, 4088 (32.5%) were EBV seropositive and 421 (3.3%) were high risk for NPC-related EBV activation. Solid fuel use was associated with a higher risk of EBV seropositivity and NPC-related EBV activation, with odds ratios (ORs) of 1.33 (95%CI: 1.01, 1.76) and 1.81 (95%CI: 1.03, 3.18), respectively. Higher risk of EBV seropositivity was observed for those who did not use ventilation apparatus and those who consumed salted food. Among ever users, OR was highest for participants with more than 40 years of solid fuel exposure (1.17, 95%CI: 1.00–1.37) and who have been constantly using solid fuel (1.30, 95%CI: 0.96–1.75). We did not find a statistically significant impact of cooking frequency on EBV seropositivity. The identification of solid fuel as a risk factor for EBV activation is of great value for understanding the etiology of NPC. Our findings also have important public health implications given the fact that a third of the global population still lack access to clean cooking, especially in low resource settings. [Display omitted] • Solid fuel is associated with increased risk of EBV seropositivity and activation. • For ever-users, largest effect in those using solid fuel constantly over 40 years. • High risk of solid fuel among ventilator nonusers and those who consume salted food. [ABSTRACT FROM AUTHOR]

BACKGROUND The utility of circulating Epstein-Barr Virus (EBV) DNA as a tumor marker for nasopharyngeal carcinoma (NPC) detection suggests that it might improve the diagnostic performance of anti-EBV antibody markers in NPC screening. In this study, the authors evaluated whether circulating EBV DNA load is capable of distinguishing NPC patients from high-risk individuals who have positive anti-EBV antibodies. METHODS In a population-based NPC screening trial in Sihui City and Zhongshan City, Guangdong Province, China, the authors previously identified 862 high-risk participants with 2 screening markers, immunoglobulin A (IgA) antibodies to EBV capsid antigen (VCA/IgA) and nuclear antigen-1 (EBNA1/IgA). In the current study, real-time polymerase chain reaction was used to measure the baseline plasma EBV DNA load among 825 participants (97%). Follow-up was extended to the end of 2011 to evaluate the diagnostic and predictive values of plasma EBV DNA load. RESULTS By using 0 copies/mL as the cutoff value, plasma EBV DNA had a sensitivity of 86.8% (33 of 38 patients) for NPC detected within the first year of follow-up, yielding a positive predictive value of 30% (33 of 110 participants) and a negative predictive value of 99.3% (696 of 701 participants). The patients who had early stage NPC had lower sensitivity (81.5%; 22 of 27 patients) than those who had advanced NPC (100%; 11 of 11 patients). For the 14 patients who had NPC detected after 1 year of follow-up, only 50% (7 of 14 patients) tested positive for EBV DNA at baseline. CONCLUSIONS The plasma EBV DNA load may improve the accuracy of diagnosing NPC in high-risk individuals, but it appears to have limited value in screening patients who have early stage NPC and predicting NPC development. Cancer 2014;120:1353-1360. © 2014 American Cancer Society. [ABSTRACT FROM AUTHOR]

Background Elevated levels of antibodies against antigens in the Epstein–Barr virus (EBV) lytic phase are important predictive markers for nasopharyngeal carcinoma (NPC) risk. Several lifestyle factors, including smoking, have also been associated with NPC risk. We hypothesized that some specific lifestyle factors induce transformation of EBV from the latent to the lytic stage and contribute to NPC occurrence. Methods We conducted a case–control study using data from male case patients (n = 1316) and control subjects (n = 1571) living in Guangdong Province, an area in China at high risk for NPC, to study potential NPC risk factors and EBV inducers. Two independent healthy male populations from a second high-risk area (n = 1657) and a low-risk area (n = 1961) were also included in the analysis of potential EBV inducers using logistic regression models. In vitro assays were performed to investigate the effect of cigarette smoke extract on EBV activation in two EBV-positive cell lines. All statistical tests were two-sided. Results Smoking was associated with an increased risk of NPC among the Guangdong participants with 20–40 and 40 or more pack-years vs never smokers (OR = 1.52, 95% CI = 1.22 to 1.88 and OR = 1.76, 95% CI = 1.34 to 2.32, respectively; Ptrend < .001). Smoking was the only factor linked to EBV seropositivity among the expanded control group and the independent low-risk population. In vitro experiments showed that cigarette smoke extract promoted EBV replication, induced the expression of the immediate-early transcriptional activators Zta and Rta, and increased transcriptional expression levels of BFRF3 and gp350 in the lytic phase. Conclusion Smoking is not only associated with NPC risk in individuals from China but is also associated with EBV seropositivity in healthy males and is involved in EBV activation. [ABSTRACT FROM PUBLISHER]

The article presents Liu Z, Ji MF, Huang QH's response to the invitory commentary on the article 'Two Epstein-Barr virus–related serologic antibody tests in nasopharyngeal carcinoma (NPC) screening: results from the initial phase of a cluster randomized controlled trial in southern China' by Dr. Hildesheim. The article also discusses risk prediction model incorporating other risk factors can be used to improve the efficiency of NPC screening in both endemic and nonendemic areas.