**4.1 Screening in high risk group with positive family history**

The incidence of NPC is relative low through most of the world (less 1/100,000). However, familial clustering of NPC has been widely documented in Chinese population (Chen and Huang, 1997) and individual with family history are at a high risk of developing the disease. Both family history and anti-EBV seropositivity are determinants in subsequent NPC development. Based on mass screening with positive family history conducted in Taiwan and Hong-Kong (Choi et al., 2011; Hsu et al., 2011; Ng et al., 2010; Yu et al., 2011), subjected with positive family history will increase risk of subsequent NPC development. Screening this high risk group significantly shifted to earlier stage (Ng et al., 2010), which mean it is a screening modality to be worth.

The etiology of NPC is not completely understood now; approaches to primary prevention of NPC remain inconclusive. This situation highlights the need of secondary prevention for early detection, diagnosis, and treatment of NPC. The effectiveness of a screening program rests on several key factors: "the importance of the disease; a welldefined population at risk; the availability of noninvasive, low-cost screening tests that can detect disease at an early stage; and effective treatments resulting in long term survival."(Choi et al., 2011) Screening for NPC for positive family history with EBV serology meets all of these criteria.

Chen first reported an evaluation of screening for NPC using Markov chain models (Chen et al., 1999). Average duration of the PCDP is estimated as 3.1 years was estimated; therefore it is possible for massive screening in NPC. The stage distribution for EBV-IgA antibodies is higher (68.7%) than in the non-screened (25%) group for stage I and II. Based on these findings, they suggest design a randomized trial in a high incidence area such as Hong Kong.

In another cohort study from Taiwan (Hsu et al., 2011) comparing different covariates, the authors compared the long-term risk NPC of male participants in a NPC cohort after adjusting for anti- EBV seromarkers and cigarette smoking. The adjusted hazard ratio was 6.8 (95% CI: 2.3, 20.1) for the multiplex family cohort compared with the community cohort. In the evaluation of anti-EBV VCA IgA and anti-EBV DNase, the adjusted hazard ratios were 2.8 (95% CI: 1.3, 6.0) and 15.1 (95% CI: 4.2, 54.1) for those positive for 1 EBV seromarker and positive for both seromarkers, respectively, compared with those negative for both EBV seromarkers. The adjusted hazard ratio was 31.0 (95% CI: 9.7, 98.7) for participants who reported a family history of NPC and who were anti-EBV-seropositive compared with individuals without such a history who were anti-EBV-seronegative. These findings suggest that both family history of NPC and anti-EBV seropositivity are important determinants of subsequent NPC development. Screening for high risk multiplex family may be more costeffective.

Screening in Hong-Kong with positive NPC family history was conducted since 1994 (Lee et al., 2005; Ng et al., 2010; Ng et al., 2005). Participants in this screening program for NPC were all first degree relatives of patients with NPC and they were all 18 years of age or older. Participants were offered annual assessment serological test of EBV and endoscopic examination of the nasopharynx. Between 1994 and 2005, total 1,199 asymptomatic family members of NPC patients were recruited and reported (Ng et al., 2010). Eighteen participants of the screening program developed NPC; 16 of them were detected in the screening. Stage distributions and survival outcomes of the 17 cases were compared with that of 1,185 consecutive symptomatic patients diagnosed in the same period through general referral. It was found that the screening program resulted in early detection of cancer (Table 2), with 59% presenting at early stage (stage I: 41%, stage II: 18%) compared to 24% (stage I: 1%, stage II: 23%) of symptomatic cancers (P=0.001), and a significant improvement in disease-free survival (P = 0.04). The cancer specific survival and overall survival rate at 5-year are also higher (92 vs. 77% and 92 vs. 70%, respectively).

#### **4.2 Screening strategies of NPC among individual from high risk NPC families**

The efficacy of any screening strategy should be evaluated before putting it into practice. Based on Markov chain models, Choi simulated and compared the outcomes of 4 screening strategies over a period of 12 years: (A) Annual screening, (B) biennial screening, (C) triennial screening, and (D) triennial screening for participants tested EBV negative and annual screening once the participants are tested EBV positive (Choi et al., 2011) . The result is summarized at **Table 3**, strategy A (screening annually) yields the maximum disease pickup rate, but strategy D (triennial screening for participants tested EBV negative and annual screening once the participants are tested EBV positive) offered the highest efficacy for NPC screening of family members of NPC patients.


\* Relative to strategy A

90 Carcinogenesis, Diagnosis, and Molecular Targeted Treatment for Nasopharyngeal Carcinoma

PCR can be performed in a series fashion (O et al., 2007). If this work-up is positive; then the patient should be referred to a specialist for further work-up including fiberoptic nasopharyngoscopy or MRI examination. The economic cost of providing this screening is reduce by prescreening to exclude the low-risk patients and performing series testing on the

**4. Applications of EBV antibodies and EBV DNA as markers of NPC among** 

The incidence of NPC is relative low through most of the world (less 1/100,000). However, familial clustering of NPC has been widely documented in Chinese population (Chen and Huang, 1997) and individual with family history are at a high risk of developing the disease. Both family history and anti-EBV seropositivity are determinants in subsequent NPC development. Based on mass screening with positive family history conducted in Taiwan and Hong-Kong (Choi et al., 2011; Hsu et al., 2011; Ng et al., 2010; Yu et al., 2011), subjected with positive family history will increase risk of subsequent NPC development. Screening this high risk group significantly shifted to earlier stage (Ng et al., 2010), which mean it is a

The etiology of NPC is not completely understood now; approaches to primary prevention of NPC remain inconclusive. This situation highlights the need of secondary prevention for early detection, diagnosis, and treatment of NPC. The effectiveness of a screening program rests on several key factors: "the importance of the disease; a welldefined population at risk; the availability of noninvasive, low-cost screening tests that can detect disease at an early stage; and effective treatments resulting in long term survival."(Choi et al., 2011) Screening for NPC for positive family history with EBV

Chen first reported an evaluation of screening for NPC using Markov chain models (Chen et al., 1999). Average duration of the PCDP is estimated as 3.1 years was estimated; therefore it is possible for massive screening in NPC. The stage distribution for EBV-IgA antibodies is higher (68.7%) than in the non-screened (25%) group for stage I and II. Based on these findings, they suggest design a randomized trial in a high incidence area such as Hong

In another cohort study from Taiwan (Hsu et al., 2011) comparing different covariates, the authors compared the long-term risk NPC of male participants in a NPC cohort after adjusting for anti- EBV seromarkers and cigarette smoking. The adjusted hazard ratio was 6.8 (95% CI: 2.3, 20.1) for the multiplex family cohort compared with the community cohort. In the evaluation of anti-EBV VCA IgA and anti-EBV DNase, the adjusted hazard ratios were 2.8 (95% CI: 1.3, 6.0) and 15.1 (95% CI: 4.2, 54.1) for those positive for 1 EBV seromarker and positive for both seromarkers, respectively, compared with those negative for both EBV seromarkers. The adjusted hazard ratio was 31.0 (95% CI: 9.7, 98.7) for participants who reported a family history of NPC and who were anti-EBV-seropositive compared with individuals without such a history who were anti-EBV-seronegative. These findings suggest that both family history of NPC and anti-EBV seropositivity are important determinants of

at-risk patients.

**individual from high risk NPC families** 

screening modality to be worth.

serology meets all of these criteria.

Kong.

**4.1 Screening in high risk group with positive family history** 

Strategy A. Annual screening

Strategy B. Biennial screening

Strategy C. Triennial screening

Strategy D. Triennial screening for participants tested EBV negative and annual screening once the participants are tested EBV positive

Table 3. Simulated screening result with the four strategies based on an imaginary population of 6,000 participants follow up for 12 years with family history of NPC (Choi et al., 2011).

Epstein-Barr Virus Serology in the Detection and Screening of Nasopharyngeal Carcinoma 93

I am heartily thankful for Miss Wan-Lun Hsu and Miss Yu-Ping Cheng for data collection,

Bureau of health promotion, Executive Yuan, Taiwan, (2010). *Cancer registry annual report*.

Chan, K. C., and Lo, Y. M. (2002). Circulating EBV DNA as a tumor marker for

Chang, K. P., Hsu, C. L., Chang, Y. L., Tsang, N. M., Chen, C. K., Lee, T. J., Tsao, K. C.,

Chen, D. L., and Huang, T. B. (1997). A case-control study of risk factors of nasopharyngeal

Chen, H. H., Prevost, T. C., and Duffy, S. W. (1999). Evaluation of screening for

Chien, Y. C., Chen, J. Y., Liu, M. Y., Yang, H. I., Hsu, M. M., Chen, C. J., and Yang, C. S.

Choi, C. W., Lee, M. C., Ng, W. T., Law, L. Y., Yau, T. K., and Lee, A. W. (2011). An analysis

Gulley, M. L. (2001). Molecular diagnosis of Epstein-Barr virus-related diseases. J Mol Diagn

Hsu, W. L., Yu, K. J., Chien, Y. C., Chiang, C. J., Cheng, Y. J., Chen, J. Y., Liu, M. Y., Chou, S.

Ji, M. F., Wang, D. K., Yu, Y. L., Guo, Y. Q., Liang, J. S., Cheng, W. M., Zong, Y. S., Chan, K.

Lee, A. W., Sze, W. M., Au, J. S., Leung, S. F., Leung, T. W., Chua, D. T., Zee, B. C., Law, S.

Li, S., Deng, Y., Li, X., Chen, Q. P., Liao, X. C., and Qin, X. (2010). Diagnostic value of

Liu, Y., Fang, Z., Liu, L., Yang, S., and Zhang, L. (2011). Detection of epstein-barr virus DNA

Lo, Y. M., Chan, L. Y., Chan, A. T., Leung, S. F., Lo, K. W., Zhang, J., Lee, J. C., Hjelm, N. M.,

Huang, C. G., Chang, Y. S., Yu, J. S., and Hao, S. P. (2008). Complementary serum test of antibodies to Epstein-Barr virus nuclear antigen-1 and early antigen: a possible alternative for primary screening of nasopharyngeal carcinoma. *Oral Oncol*

nasopharyngeal carcinoma: trial design using Markov chain models. *Br J Cancer* 79,

(2001). Serologic markers of Epstein-Barr virus infection and nasopharyngeal

of the efficacy of serial screening for familial nasopharyngeal carcinoma based on

P., You, S. L., Hsu, M. M., et al. (2011). Familial tendency and risk of nasopharyngeal carcinoma in taiwan: effects of covariates on risk. *Am J Epidemiol* 

H., Ng, S. P., Wei, W. I., et al. (2007). Sustained elevation of Epstein-Barr virus antibody levels preceding clinical onset of nasopharyngeal carcinoma. *Br J Cancer* 

C., Teo, P. M., Tung, S. Y., et al. (2005). Treatment results for nasopharyngeal carcinoma in the modern era: the Hong Kong experience. *Int J Radiat Oncol Biol* 

Epstein-Barr virus capsid antigen-IgA in nasopharyngeal carcinoma: a meta-

in serum or plasma for nasopharyngeal cancer: a meta-analysis. *Genet Test Mol* 

Johnson, P. J., and Huang, D. P. (1999). Quantitative and temporal correlation

nasopharyngeal carcinoma. *Semin Cancer Biol* 12, 489-496.

carcinoma in Taiwanese men. *N Engl J Med* 345, 1877-1882.

Markov chain models. *Fam Cancer* 10, 133-139.

**6. Acknowledgment** 

**7. References** 

28.

44, 784-792.

1894-1900.

3, 1-10.

173, 292-299.

96, 623-630.

*Phys* 61, 1107-1116.

*Biomarkers* 15, 495-502.

analysis. *Chin Med J* 123, 1201-1205.

carcinoma. *Cancer Lett* 117, 17-22.

manuscript reviewing and modification.

Various EBV screening seromarkers were ever been reported for massive screening in high risk group with positive family NPC history (**Table 4.**) (Ng et al., 2010; Yu et al., 2011). The positive predicative value (PPV) of EBV seromarkers was below 10% and the negative predicative value (NPV) was higher than 99%. Because the average duration of the preclinical screen-detectable phase is estimated as 3.1 years, annual check-up once the participants are tested EBV positive is necessary. For the high NPV, triennial screening is reasonable for NPC family history with negative EBV seromarkers.


Table 4. Comparing different screening seromarkers in high risk NPC screening with positive family history.

#### **5. Conclusion**

EBV is associated with the development of NPC and the infection occurs in the early phases of carcinogenesis. A long period of pre-clinical detectable phase offers the opportunity for screening and early diagnosis of NPC. EBV antibodies and EBV DNA have the potential for screening, diagnosis, monitoring and prognosis of NPC. NPC screening in a high-risk, endemic population using EBV-specific serologic markers seems effective. Conduction of prospective randomized controlled screening trials is necessary to validate the costeffectiveness.

#### **6. Acknowledgment**

I am heartily thankful for Miss Wan-Lun Hsu and Miss Yu-Ping Cheng for data collection, manuscript reviewing and modification.

#### **7. References**

92 Carcinogenesis, Diagnosis, and Molecular Targeted Treatment for Nasopharyngeal Carcinoma

Various EBV screening seromarkers were ever been reported for massive screening in high risk group with positive family NPC history (**Table 4.**) (Ng et al., 2010; Yu et al., 2011). The positive predicative value (PPV) of EBV seromarkers was below 10% and the negative predicative value (NPV) was higher than 99%. Because the average duration of the preclinical screen-detectable phase is estimated as 3.1 years, annual check-up once the participants are tested EBV positive is necessary. For the high NPV, triennial screening is

Specificity

Positive% NPV PPV

83.3%/87.0% 14.0% 99.7% 8.9%

88.9%/87.0% 15.1% 99.8% 9.5%

36.4%/73.8% 26.2% 99.5% 0.8%

7.1%/95.7% 4.3% 99.4% 1.0%

92.9%/16.0% 84.0% 99.7% 0.6%

50.0%/84.3% 15.9% 99.7% 1.8%

85.7%/51.2% 49.0% 99.8% 1.0%

23.1%/88.1% 12.0% 99.5% 1.1%

84.6%/92.3% 8.1% 99.9% 5.6%

reasonable for NPC family history with negative EBV seromarkers.

VCA IgA (IF) Positive (>1:10 dilution of serum)

Positive (OD405 > 0.50)

Positive (OD405 > 0.10)

Positive (OD405 > 0.20)

Positive (OD405 > 0.10)

(>400 neutralizing units)

(>160 neutralizing units)

Table 4. Comparing different screening seromarkers in high risk NPC screening with

EBV is associated with the development of NPC and the infection occurs in the early phases of carcinogenesis. A long period of pre-clinical detectable phase offers the opportunity for screening and early diagnosis of NPC. EBV antibodies and EBV DNA have the potential for screening, diagnosis, monitoring and prognosis of NPC. NPC screening in a high-risk, endemic population using EBV-specific serologic markers seems effective. Conduction of prospective randomized controlled screening trials is necessary to validate the cost-

Author Journal Modality Sensitivity/

Anti-EBV VCA IgA

 Annual anti-EBV serology and nasopharyngoscopy

VCA IgA (ELISA)

VCA IgA (ELISA)

EBNA1 IgA

EBNA1 IgA

Dnase Positive

Dnase Positive

Wai *el al.* (Ng et al., 2010) (Hong-Kong)

Yu *et al. (Yu et al., 2011)* (Taiwan)

Fam Cancer (2010)

Clin Cancer Res (2011)

positive family history.

**5. Conclusion** 

effectiveness.


between circulating cell-free Epstein-Barr virus DNA and tumor recurrence in nasopharyngeal carcinoma. *Cancer Res* 59, 5452-5455.

**6** 

**Imaging of Nasopharyngeal Carcinoma** 

*2Department of Medical Imaging and Otolaryngology-Head and Neck Surgery,* 

*5Department of Medical Imaging and Otolaryngology-Head and Neck Surgery,* 

Nasopharyngeal carcinoma (NPC) is the most common neoplasm to affect the nasopharynx (NP). Arising from the epithelial lining of the nasopharyngeal mucosa, NPC is distinct from squamous cell carcinoma affecting other sites of the pharyngeal space. NPC typically originates in the lateral wall of the nasopharynx and is noted as a locally aggressive neoplasm with a high incidence of metastases to cervical lymph nodes. The primary tumour can extend within the nasopharynx and/or to the base of the skull, palate, nasal cavity or oropharynx. Distant metastases can arise in bone, lung, mediastinum, and, more rarely, the

The etiology of NPC is multifactorial and involves many environmental and genetic risk factors (Henderson *et al.*, 1976). In particular, diets high in salt-preserved foods – such as salted fish, meat, eggs, fruits and vegetables in the Southern Asian diet – have been identified as possible causative factors acting through the carcinogen, *N*nitrosodimethylamine (Yu *et al.*, 1988). In contrast, frequent consumption of fresh fruits and/or vegetables – especially during childhood – is associated with a lower risk of NPC. In addition, studies have indicated a causal role for the Epstein-Barr virus (EBV) in the development of NPC (Chang & Adami, 2006). EBV is a DNA virus responsible for infectious mononucleosis, post-transplantation lymphoproliferative disease, and Burkitt's lymphoma. Elevated IgG and IgA antibody titres against viral capsid antigen, early antigen, and latent viral nuclear antigens have been noted in NPC patients. Furthermore, these antibodies have been shown to precede tumour development by several years. EBV DNA, RNA, and gene products have also been detected in tumour cells (Chang & Adami, 2006). EBV serology is currently used as a screening tool for high-risk populations in southern China (Glastonbury, 2007). Certain genetic haplotypes – HLA A2, Bsn 2, B46, and B58 – are also associated with

**1. Introduction** 

liver (Brennan, 2006).

**1.1 Imaging of nasopharyngeal carcinoma** 

Michael Chan1, Eric Bartlett2, Arjun Sahgal3,

*3Department of Radiation Oncology, University of Toronto 4Division of Life Sciences, University of Western Ontario* 

Stephen Chan4 and Eugene Yu5 *1Faculty of Medicine, University of Toronto* 

*University of Toronto* 

*University of Toronto* 

*Canada* 

