**Abstract**

Nasopharyngeal carcinoma (NPC) is one of head and neck cancer. It has a complicated etiology involving Epstein–Barr virus (EBV) infection, environment changes, and genetic susceptibility. Early symptoms of NPC are unspecific, so most NPC patients are diagnosed at a late stage. An effective population screening strategy could increase the early detection and early diagnosis rate. As far, serological detection of EBV antibodies such as VCA-IgA, EA-IgA, and EBNA1-IgA, are widely used in NPC mass screening; EBV DNA load detection in plasma or nasopharyngeal swab was applied to screen in endemic populations for assessing the feasibility. However, the current screening schemes still have disadvantages such as lowly positive predictive value, unclear effectiveness of screening and cost-effectiveness. In the present chapter, we try to review the different screening strategies for NPC to understand the future direction of development.

**Keywords:** Nasopharyngeal carcinoma, Screening, Early diagnosis, Biomarker, Epstein–Barr virus, Antibody, DNA load

## **1. Introduction**

Screening is primary measure of secondary prevention of cancer. It mainly regularly monitor the asymptomatic high-risk population to achieve the purpose of early detection and early treatment of malignant tumors. Cancer carring out the secondary prevention should have the following conditions: 1) great harm to the health and life of the population; 2) long enough pre-clinical period; 3) better effect of both the early treatment and intervention of precancerous lesions; 4) screening methods with effective sensitivity and specificity.

Nasopharyngeal carcinoma (NPC) is a one of head and neck cancer. At present, the etiology of NPC has not remained completely elucidated, generally considered involving Epstein–Barr virus (EBV) infection, environment changes, and genetic susceptibility. There are no feasible preventive measures for NPC. However, it has secondary prevention. First, NPC exhibiting marked racial and geographical differences, is epidemic in the population of Southern China, Southeast Asia, and North Africa, which is extremely harmful to human life and health [1]. In 2014, the incidence and mortality of NPC were 2.48 per 100,000 and 1.23 per 100,000 in China respectively [2]. At the same time, the morbidity and mortality of males were higher than females. The morbidity was mainly in the young while the mortality was mainly in middle and old age. Besides, the occurrence and development of NPC is a multi-stage process, which includes initiation, promotion, malignant transformation, and advanced stage of disease [3]. It takes a certain amount of

time to develop into a malignant tumor. In addition, the clinical stage of NPC is an important factor affecting therapeutic outcome. The 10-year survival rate for NPC with stages I and II can reach up to over 90%, whereas for patients with stage III and IV is less than 50% [4]. At last, there are effective methods to detect pre-clinical patients, and the level of antibody against EBV antigens is significantly related to the risk of NPC [5, 6].

NPC first occurs in the epithelium of the nasopharynx, which can invade the base of the skull and metastasize to the cervical lymph nodes. It has the characteristics of complex manifestations, hidden onset, non-specificity of the initial symptoms, and difficulty in early diagnosis. According to statistics, patients with NPC at an early stage who came to the hospital accounted for only about 20% of the total [7]. Strengthening secondary prevention - early detection, early diagnosis, and early treatment - is an important part of the prevention and treatment of NPC. At the same time it is the key to improve the cure rate and obtain a better prognosis of NPC patients.

## **2. EBV specific antibodies-based serologic testing**

EBV belonging to γ-herpesvirus is a human herpesvirus with B lymphocytes. Nearly 95% of adults worldwide are infected with this virus. EBV in infected cells can be divided into two states: EBV latent infection and EBV lytic infection. Only a few virus genes are expressed in EBV latent infection, which can ensure the basic replication function of the virus but lossing infection ability. In EBV lytic infection, EBV needs to be activated about 80 ~ 100 viral genes to complete host-to-host propagation, and finally produce and form infectious virions (or viral particles). After the initial infection, EBV can establish a lifelong latent infection in the host, and persistent EBV lysis replication state infection can lead to a series of human malignant tumors [8].

EBV infection is closely related to the occurrence and development of NPC. EBV latent infection of nasopharyngeal epithelial cells is considered to be a key step in the carcinogenesis of epithelial cells. After EBV infection, the expressed virus genes can produce different antigens, such as EBV nuclear antigen (EBNA), membrane antigen (MA), early antigen (EA), viral capsid antigen (VCA), BZLF1 transcription activator protein (Zta), BRLF1 transcription activator protein (Rta), etc. [9].

The detection of antibodies against EBV antigens in the sera of NPC patients was reported as early as 1966. Helen W et al. first proposed the view that immunoglobulin A (IgA) antibodies against EBV can be used for the diagnosis of NPC [10]. Studies also confirmed that the expression of IgA antibodies against VCA (VCA-IgA) in NPC patients was higher than in healthy people and the antibody titer was related to the stage of NPC. The idea of using this antibody for NPC screening also was proposed [11]. In 1977, Y Zeng et al. established a prospective prevention and treatment site for NPC in Cangwu, Guangxi province China, in order to carry out research on early diagnosis and etiology analysis. The first NPC mass screening was carried out in Cangwu County by the application of the immunoenzymatic (IE) method to detect VCA-IgA and EA-IgA [12]. Therefore, the NPC screening model suitable for the population in the high-risk areas was established by Zeng's team. Then this mass screening model was promoted to three high-risk areas in China, including Guangxi, Guangdong, and Hainan province; and more than four hundred thousand people were screened for NPC [13–16]. In the 1980s, similar methods were used to screen and follow up large populations in Guangdong and Taiwan provinces [17, 18]. SM Cao et al. performed a prospective screening study of 18,986 subjects with a 20-year follow-up in Guangdong province using the same method [6].

#### *Screening of Nasopharyngeal Carcinoma DOI: http://dx.doi.org/10.5772/intechopen.97398*

This study showed that both VCA-IgA and EA-IgA antibodies were effective serum markers for NPC screening in high-risk areas. Thus, this method was considered as the standard tool for NPC mass screening in China. However, the IE method for detecting EBV antibodies also has disadvantages, such as tedious operation, long time consuming, no quality control standard, and subjective influence on manual interpretation results. These characteristics made it difficult to consistently perform in a large population.

Enzyme-linked immunoassay (ELISA) with many advantages compared with the IE method, such as simple operation, automatic detection, and interpretation of results by a microplate reader, which have subsequently been applied in NPC screening. Many studies about using ELISA for NPC screening were reported. The detection of ZEBRA-IgG by ELISA was applied to screen NPC, but it's specificity and sensitivity were lower compared with the detection of VCA-IgA based on IE [19]. ELISA-based detection of EBV-related antibodies, such as VCA-IgA, EBNA1- IgA have also found to be a marker for NPC screening [20–22]. The detection rate of one single marker was found to be not ideal, and issues such as the combination of indicators for joint detection, the setting of thresholds, and the strategy of screening intervals were discussed.

In Indonesia, the two-step approach employed the EBV IgA ELISA based on a combination of VCA p18- and EBNA1-derived synthetic peptides as an initial screening test and the EA-IgA ELISA as a confirmation test. The sensitivity and specificity for diagnosing NPC using it significantly increased, as well as positive predictive value and negative predictive value [23].

JY Guo et al. evaluated the diagnostic effect of VCA-IgA, EA-IgA and Rta-IgG antibody detection alone or Combiningly in NPC. The triple-positive of VCA-IgA, EA-IgA and Rta-IgG antibodies suggested the highest risk of NPC, and the triplenegative of them showed the lowest risk [24].

In Taiwan, the ability of anti-EBV-IgA antibody to detect NPC in a high-risk population was evaluated. These markers targeted at the following EBV peptides including EBNA1, VCAp18, EAp138, Ead\_p47 and VCAp18 + EBNA1 peptide mixture. The result showed that EBNA1-IgA was a sensitive biomarker for differential diagnosis of NPC. At the same time they identified 80% of the highrisk individuals who developed to NPC during follow-up (80% sensitivity) during measuring at baseline [25].

SM Cao's team developed a prediction formula to calculate Logit P-value with VCA-IgA and EBNA1-IgA as variables (Logit P = −3.934 + 2.203 × VCA/ IgA + 4.797 × EBNA1/IgA). The specificity of the new screening scheme is equivalent to traditional screening scheme with the IE method (estimated at 98.5%), but the sensitivity of former (75.0%) is significantly higher than the latter (25.0%) [26]. A total of 28,688 Guangdong residents aged 30–59 years were screened by the combination of two EBV antibodies tests in addition to indirect mirror examination in the nasopharynx and/or lymphatic palpation (IMLP) in Sihui and Zhongshan, Guangdong province China. After one year of follow-up, the total detection rate of NPC was 0.14% (41/28,688), and the early diagnosis rate was as high as 68.3% (28/41) [27]. After six-year follow-up, the sensitivity of the new scheme was 95.7%, with AUC = 0.926 (95% CI: 0.885–0.966). The new screening scheme for NPC is verified to be the preferred serum diagnostic strategy for long-term screening in high-incidence areas of NPC [28]. For the best interval, studies have shown that the incidence of NPC was low in the first few years after the negative screening and then it would increase to the general population level. Therefore, the screening interval of 4–5 years may be more appropriate than 9–10 years after VCA-IgA negative detection in NPC screening [29]. The above research results were adopted by the Chinese Technical Program of Cancer Early

Diagnosis and Early Treatment --Technical Scheme of NPC Screening to guide the annual routine population screening in NPC high-risk areas (**Figure 1**).

However, there are still limitations in NPC screening using EBV antibodies as tumor markers. The false-positive rate of EBV serological screening is relatively high. The positive rate of EBV antibody in the high-risk areas of NPC is 3% ~ 10%. High-risk groups require further examinations, such as nasopharyngeal fibroscopy

#### **Figure 1.**

*Flowchart of NPC screening procedure for nasopharyngeal carcinoma (NPC). EBV, Epstein–Barr virus; ab, antibody. Cited by expert group of NPC screening project, expert committee on early diagnosis and treatment of cancer project, China. The technical scheme of NPC screening, early diagnosis and early treatment (2015 edition).*

and nasopharyngeal tissue biopsy. Pathologically diagnosed NPC only accounts for 1.5% ~ 3.3% of the high-risk population at the initial screening, which further leads to a higher false positive rate [27, 30, 31].
