A comparison of EBV serology and serum cell-free DNA as screening tools for nasopharyngeal cancer: Results of the Singapore NPC screening cohort

We aimed to evaluate the effectiveness of nasopharyngeal cancer (NPC) screening by comprehensive clinical follow-up and adjunctive Epstein–Barr virus (EBV) testing. In a prospective cohort study, 524 individuals with a first-degree family history of NPC were recruited at a university clinical center in Singapore. The cohort was evaluated at baseline and at 6 monthly intervals, with a complete head and neck examination including nasopharyngeal endoscopy. Blood was taken at baseline and at yearly intervals for EBV Viral Capsid Antigen (VCA) IgA, EBV Early Antigen (EA) IgA serology and serum cell-free EBV DNA. Nasopharyngeal biopsy was performed when any irregularity in the nasopharynx was observed, or when EBV markers were elevated. The mean duration of follow-up was 57.7 months, with an average of 8.6 clinical visits per participant. Five participants (0.96%) were identified to have NPC, giving a prevalence of 199 per 100,000 person-years of screening. Four of the five NPC cases identified had asymptomatic T1 disease, at an earlier stage compared to NPC patients diagnosed in the clinic during the same time period (p = 0.0297). All NPC cases identified had elevated EBV-EA IgA titers ≥1:10, with a specificity of 94.6% and a positive predictive value of 15.2%, outperforming EBV-VCA IgA and serum EBV DNA. Two NPC cases were biopsied only because of elevated EBV serology titers, with increasing EBV-EA IgA titers preceding the diagnosis of NPC. In conclusion, screening for NPC is effective in identifying early-stage disease. Adjunctive EBV-EA IgA testing improved the effectiveness of screening.     

Evaluation of nonviral risk factors for nasopharyngeal carcinoma in a high‐risk population of Southern China

To understand the role of environmental and genetic influences on nasopharyngeal carcinoma (NPC) in populations at high risk of NPC, we have performed a case‐control study in Guangxi Province of Southern China in 2004–2005. NPC cases (n = 1,049) were compared with 785 NPC‐free matched controls who were seropositive for IgA antibodies (IgA) to Epstein‐Barr virus (EBV) capsid antigen (VCA)—a predictive marker for NPC in Chinese populations. A questionnaire was used to capture exposure and NPC family history data. Risk factors associated with NPC in a multivariant analysis model were the following: (i) a first, second or third degree relative with NPC [attributable risk (AR)= 6%, odds ratio (OR) = 3.1, 95% confidence interval (CI) = 2.0–4.9, p < 0.001]; (ii) consumption of salted fish 3 or more than 3 times per month (AR = 3%, OR = 1.9, 95% CI = 1.1–3.5, p = 0.035); (iii) exposure to domestic wood cooking fires for more than 10 years (AR = 69%, OR = 5.8, 95% CI = 2.5–13.6, p < 0.001); and (iv) exposure to occupational solvents for 10 or less years (AR = 4%, OR = 2.6, 95% CI = 1.4–4.8, p = 0.002). Consumption of preserved meats or a history of tobacco smoking were not associated with NPC (p > 0.05). We also assessed the contribution of EBV/IgA/VCA antibody serostatus to NPC risk—32.2% of NPC can be explained by IgA+ status. However, family history and environmental risk factors cumulatively explained only 2.7% of NPC development in NPC high risk population. These findings should have important public health implications for NPC risk reduction in endemic regions. © 2009 UICC     

Early detection of nasopharyngeal carcinoma by plasma Epstein‐Barr virus DNA analysis in a surveillance program

BACKGROUND:

Nasopharyngeal carcinoma (NPC) is prevalent in Southeast Asia. Over the last decade, plasma Epstein‐Barr virus (EBV) DNA has been developed as a tumor marker for NPC. In this study, the authors investigated whether plasma EBV DNA analysis is useful for NPC surveillance.

METHODS:

In total, 1318 volunteers ages 40 to 60 years were prospectively recruited. Plasma EBV DNA and serology for viral capsid antigen immunoglobulin A (IgA) were measured. Participants who had detectable plasma EBV DNA or positive IgA serology underwent nasal endoscopic examination and a follow‐up plasma EBV DNA analysis in approximately 2 weeks. All participants were followed for 2 years to record the development of NPC.

RESULTS:

Three individuals with NPC were identified at enrolment. All of them were positive for EBV DNA and remained positive in follow‐up analysis. Only 1 of those patients was positive for EBV serology. In 1 patient who had NPC with a small tumor confined to the mucosa, the tumor was not detectable on endoscopic examination. Because of a 2‐fold increase in plasma EBV DNA on the follow‐up analysis, that patient underwent magnetic resonance imaging, which revealed the tumor. Among the participants who did not have NPC but had initially positive plasma EBV DNA results, approximately 66% had negative EBV DNA results after a median of 2 weeks.

CONCLUSIONS:

Plasma EBV DNA analysis proved useful for detecting early NPC in individuals without a clinical suspicion of NPC. Repeating the test in those who had initially positive results differentiated those with NPC from those who had false‐positive results. Cancer 2013. © 2013 American Cancer Society.     

Epstein‐Barr virus strain variation and cancer

Epstein‐Barr virus (EBV) is a human tumor virus and is etiologically linked to various malignancies. Certain EBV‐associated diseases, such as Burkitt lymphomas and nasopharyngeal carcinomas, are endemic and exhibit biased geographic distribution worldwide. Recent advances in deep sequencing technology enabled high‐throughput sequencing of the EBV genome from clinical samples. Rapid cloning and sequencing of cancer‐derived EBV genomes, followed by reconstitution of infectious virus, have also become possible. These developments have revealed that various EBV strains are differentially distributed throughout the world, and that the behavior of cancer‐derived EBV strains is different from that of the prototype EBV strain of non‐cancerous origin. In this review, we summarize recent progress and future perspectives regarding the association between EBV strain variation and cancer.     

Modulation of the tumor microenvironment by Epstein‐Barr virus latent membrane protein 1 in nasopharyngeal carcinoma

Latent membrane protein 1 (LMP1) is a primary oncogene encoded by the Epstein‐Barr virus, and various portions of LMP1 are detected in nasopharyngeal carcinoma (NPC) tumor cells. LMP1 has been extensively studied since the discovery of its transforming property in 1985. LMP1 promotes cancer cell growth during NPC development and facilitates the interaction of cancer cells with surrounding stromal cells for invasion, angiogenesis, and immune modulation. LMP1 is detected in 100% of pre‐invasive NPC tumors and in approximately 50% of advanced NPC tumors. Moreover, a small population of LMP1‐expressing cells in advanced NPC tumor tissue is proposed to orchestrate NPC tumor tissue maintenance and development through cancer stem cells and progenitor cells. Recent studies suggest that LMP1 activity shifts according to tumor development stage, but it still has a pivotal role during all stages of NPC development.     

Lung cancer mortality trends in 36 European countries: secular trends and birth cohort patterns by sex and region 1970–2007

Smoking is a major contributor to all‐cause mortality in Europe and accounts for one‐fifth of the cancer‐related deaths. Monitoring the tobacco epidemic via an analysis of lung cancer trends is essential in helping countries arrest the effects of tobacco epidemic in the region. The study aims to provide a comprehensive and up‐to‐date overview of the temporal patterns of lung cancer mortality in Europe, emphasizing country‐ and sex‐specific differences. National lung cancer mortality data were extracted from the WHO mortality databank by age, sex, year of death (1970–2007) for 36 countries in Europe. Trends in lung cancer mortality in men have tended to decrease in many European countries during the last two decades, particularly in North and Western Europe. Among women, mortality rates are still increasing in many countries, although in a few populations, rates are beginning to stabilize, notably in the high‐risk countries within Eastern Europe (Hungary, Poland and the Czech Republic), and in Northern Europe (Denmark, Iceland and the United Kingdom). Men and women are clearly in very different phases of the smoking epidemic, and, as reflected in the mortality rates by birth cohort, the stage varies widely by country within each European region. That lung cancer mortality trends in men are on a downwards path in most European countries while female rates continue to rise, points to an urgent need for national and European prevention strategies that target tobacco cessation and prevention among European women.     

Spontaneous T cell responses to Epstein-Barr virus-encoded BARF1 protein and derived peptides in patients with nasopharyngeal carcinoma: bases for improved immunotherapy

Immunotherapy approaches targeting Epstein-Barr virus (EBV)-encoded antigens induce objective clinical responses only in a fraction of patients with undifferentiated nasopharyngeal carcinoma (UNPC). In the present study, we have characterized the immunogenicity of the EBV-encoded BARF1 oncogene with the aim to assess whether this protein could constitute a new target antigen for immunotherapy in this setting. Spontaneous CD4+ and CD8+ T cell responses specific for the recombinant p29 BARF1 protein were detected by IFNgamma-ELISPOT in both EBV-seropositive donors and UNPC patients, but not in EBV-seronegative individuals. Using immunoinformatic prediction tools, we have selected 5 different candidate BARF1 T cell epitopes presented by HLA-A*0201. Although only one of these peptides was able to bind HLA-A2 with low affinity in the T2 stabilization assay, all 5 BARF1 nonamers readily elicited specific CD8+ T cell responses in EBV-seropositive HLA-A*0201+ donors and UNPC patients. Notably, the magnitude of CD8+ T cell responses to the whole BARF1 protein and derived A*0201 peptides was significantly higher in UNPC patients than in healthy donors. Moreover, cytotoxic T lymphocytes specific for the p2-10, p23-31, or p49-57 BARF1 peptides were easily obtained from HLA-A*0201+ donors. These cultures were not only able to lyse autologous targets loaded with the antigenic peptide, but also recognized tumor cells endogenously expressing BARF1 in an antigen-specific and HLA-A2-restricted manner. These findings, indicate that BARF1 is a particularly attractive antigen with immunogenic properties in most UNPC patients and provide valuable information to develop new strategies to improve the efficacy of EBV-targeting immunotherapy of UNPC patients.     

EBV-associated gastric carcinoma in high- and low-incidence areas for nasopharyngeal carcinoma

Background:

Approximately 10% of gastric carcinomas are associated with Epstein–Barr virus (EBV). The Inuit in Greenland have a high incidence of EBV-associated nasopharyngeal carcinoma.

Methods:

We conducted a population-based case–control study comparing gastric carcinomas in Greenland and in Denmark.

Results:

The prevalence rate of EBV-associated gastric carcinomas was 8.5% in both populations.

Conclusion:

The findings of this study argue against a general susceptibility to EBV-associated carcinomas among the Inuit.     

Vesicle-bound EBV-BART13-3p miRNA in circulation distinguishes nasopharyngeal from other head and neck cancer and asymptomatic EBV-infections

Cell-free microRNA (miRNA) in biofluids released by tumors in either protein or vesicle-bound form, represent promising minimally-invasive cancer biomarkers. However, a highly abundant non-tumor background in human plasma and serum complicates the discovery and detection of tumor-selective circulating miRNAs. We performed small RNA sequencing on serum and plasma RNA from Nasopharyngeal Carcinoma (NPC) patients. Collectively, Epstein Barr virus-encoded miRNAs, more so than endogenous miRNAs, signify presence of NPC. However, RNAseq-based EBV miRNA profiles differ between NPC patients, suggesting inter-tumor heterogeneity or divergent secretory characteristics. We determined with sensitive qRT-PCR assays that EBV miRNAs BART7-3p, BART9-3p and BART13-3p are actively secreted by C666.1 NPC cells bound to extracellular vesicles (EVs) and soluble ribonucleoprotein complexes. Importantly, these miRNAs are expressed in all primary NPC tumor biopsies and readily detected in nasopharyngeal brushings from both early and late-stage NPC patients. Increased levels of BART7-3p, BART9-3p and particularly BART13-3p, distinguish NPC patient sera from healthy controls. Receiver operating characteristic curve analysis using sera from endemic NPC patients, other head and neck cancers and individuals with asymptomatic EBV-infections reveals a superior diagnostic performance of EBV miRNAs over anti-EBNA1 IgA serology and EBV-DNA load (AUC 0.87–0.96 vs 0.86 and 0.66 respectively). The high specificity of circulating EBV-BART13-3p (97%) for NPC detection is in agreement with active secretion from NPC tumor cells. We conclude EV-bound BART13-3p in circulation is a promising, NPC-selective, biomarker that should be considered as part of a screening strategy to identify NPC in endemic regions.     

Laboratory markers of tumor burden in nasopharyngeal carcinoma: a comparison of viral load and serologic tests for Epstein-Barr virus

Epstein-Barr virus (EBV) is present within the tumor cells of most cases of nasopharyngeal carcinoma (NPC). Recent studies suggest that tumor burden is proportional to the level of EBV DNA in blood and that rapid blood testing can be used to guide therapeutic intervention. The relative utility of viral load vs. serology has been insufficiently studied. In our study, EBV viral load was measured by quantitative PCR using either real-time or end-point detection systems in serum samples from 124 NPC patients (93 pretreatment, 13 relapsed, 18 in remission) and 40 controls. Serologic titers against EBV early antigen were measured in the same serum samples. EBV DNA was detectable in 64 of 93 untreated NPC patients (69%; mean viral load 11,211 copies/ml), 11 of 13 relapsed NPC patients (85%; mean 53,039 copies/ml) and 0 of 18 remission patients. EBV DNA was detectable in only 1 of 40 non-NPC controls (3%). In 34 instances where paired plasma and serum samples were available for testing, both were effective sample types, and there was no significant difference between end-point and real-time methods for measuring viral load. Early antigen (EA) IgA and IgG titers were elevated in most NPC patients regardless of whether their disease was active or in remission. EBV viral load was more informative than was EA serology for distinguishing remission from relapsed disease. EBV DNA measurement appears to be a noninvasive way to monitor tumor burden after therapy.     

Global trends in colorectal cancer mortality: projections to the year 2035

Colorectal cancer (CRC) is the third most common cancer worldwide and the fourth most common cause of cancer death. Predictions of the future burden of the disease inform health planners and raise awareness of the need for cancer control action. Data from the World Health Organization (WHO) mortality database for 1989–2016 were used to project colon and rectal cancer mortality rates and number of deaths in 42 countries up to the year 2035, using age-period-cohort (APC) modelling. Mortality rates for colon cancer are predicted to continue decreasing in the majority of included countries from Asia, Europe, North America and Oceania, except Latin America and Caribbean countries. Mortality rates from rectal cancer in general followed those of colon cancer, however rates are predicted to increase substantially in Costa Rica (+73.6%), Australia (+59.2%), United States (+27.8%), Ireland (+24.2%) and Canada (+24.1%). Despite heterogeneous trends in rates, the number of deaths is expected to rise in all countries for both colon and rectal cancer by 60.0% and 71.5% until 2035, respectively, due to population growth and ageing. Reductions in colon and rectal cancer mortality rates are probably due to better accessibility to early detection services and improved specialized care. The expected increase in rectal cancer mortality rates in some countries is worrisome and warrants further investigations.     

Epstein‐Barr virus and risk of non‐Hodgkin lymphoma in the cancer prevention study‐II and a meta‐analysis of serologic studies

Epstein‐Barr virus (EBV) causes rare, malignant lymphomas. The role of EBV in other non‐Hodgkin lymphomas (NHLs) remains unclear, but mildly reduced immune function could lead to reactivation of EBV and subsequent NHL. We examined the association between prospectively‐collected plasma EBV antibodies and NHL risk in the Cancer Prevention Study‐II (CPS‐II) Nutrition Cohort and conducted a meta‐analysis of our and published results. The CPS‐II study included 225 NHL cases and 2:1 matched controls. No associations were observed between EBV serostatus or antibody levels and risk of NHL overall. However, when including only the three most common types of NHL (diffuse large B‐cell lymphoma, follicular lymphoma, chronic lymphocytic leukemia/small lymphocytic lymphoma), high compared to low early antigen (EA‐D) diffuse and BZLF1‐encoded replication activator antibodies were associated with approximately 60% higher risk of NHL. Odds ratios (ORs) for EBV nuclear antigen‐1 and viral capsid antigen (VCA)‐p18 were elevated but not statistically significant. In the meta‐analysis, both EA (summary OR = 1.52, 95% confidence interval (CI): 1.16–2.00) and VCA (summary OR = 1.20, 95% CI: 1.00–1.44) were positively associated with NHL risk. These results suggest EBV may be associated with a wider spectrum of NHL subtypes, but further study is needed to confirm and fully understand these associations.     

A population‐based prospective study of energy‐providing nutrients in relation to all‐cause cancer mortality and cancers of digestive organs mortality

The effect of dietary composition on mortality in low‐income countries is largely unknown. We evaluated whether percentages of dietary energy derived from protein, fat and carbohydrates were associated with all‐cause and cancer mortalities in a Bangladeshi population. Data from a prospective population‐based cohort study of 17,244 men and women were used. Percentages of dietary energy derived from protein, fat and carbohydrates, assessed using a validated food‐frequency questionnaire at baseline, were analyzed in relation to mortality over an average of 9 years (155,126 person‐years) of follow‐up. Cox proportional hazards regression models were used to estimate hazard ratios for all cause, all cancer and cancers of the digestive organs mortalities. Percentage of dietary energy from protein appeared to be significantly associated with cancer mortality. Fully adjusted hazard ratios for cancer mortality in increasing tertiles of percentage of dietary energy from protein were 1.0 (reference), 1.21 (0.73, 2.00) and 1.84 (1.08, 3.15) (p for trend = 0.023). These associations were much stronger for deaths from cancers of the digestive organs with fully adjusted hazard ratios in increasing tertiles of percentage of dietary energy from protein being 1.0 (reference), 2.25 (0.91, 5.59) and 4.85 (1.88, 12.51) (p for trend = 0.001). No significant associations in relation to cancer‐related mortality were observed for percentage of dietary energy from fat. Novel findings from this prospective study show protein is an important risk factor or proxy to an important risk factor for cancer mortality especially from digestive organ cancers in Bangladesh. © 2013 UICC     

Global trends in oral and pharyngeal cancer incidence and mortality

To provide an up-to-date overview of recent trends in mortality from oral and pharyngeal cancer, we analyzed death certification data for 61 countries worldwide provided by the World Health Organization in 2010–2015, and, for selected most populous countries, over the period 1970–2016. For 12 largest countries, we analyzed incidence derived from Cancer Incidence in Five Continents in 1960–2012 for all oral and pharyngeal cancers and by subsites. In 2015, male age-standardized (world population) death rates per 100,000 were 5.03 in the European Union (EU), 8.33 in the Russian Federation, 2.53 in the United States (USA), and 3.04 in Japan; corresponding rates in women were 1.23, 1.23, 0.82, and 0.76. Male mortality decreased over the last decades in several European countries, with earlier and sharper declines in southern Europe; conversely, mortality was still increasing in a few eastern European countries and the United Kingdom. Mortality in men also decreased in Argentina, Australia, and Hong Kong, while it leveled off over more recent calendar years in Brazil, Japan, Mexico, the Republic of Korea, as well as in Australia and the USA. Female mortality slightly rose in various European countries. Overall incidence trends in the largest countries were broadly consistent with mortality ones, but oropharyngeal cancer incidence rose in many countries. Changes in tobacco and alcohol exposure in men over the last decades likely explain the favorable trends in oral and pharyngeal cancer mortality and incidence observed in selected countries worldwide, while increased human papillomavirus infection is likely responsible for the rise in oropharyngeal cancer incidence.