U. Prasad

Elevated Serum Transforming Growth Factor β1 Levels in Epstein-Barr Virus-Associated Diseases and Their Correlation with Virus-Specific Immunoglobulin A (IgA) and IgM

ABSTRACT Transforming growth factor β (TGF-β) is an immunosuppressive cytokine which can induce immunoglobulin A (IgA) switch and Epstein-Barr virus (EBV) replication in latently infected cells. Here we report elevated serum levels of TGF-β in various EBV-associated diseases correlating positively with EBV-specific IgA titers and negatively with IgM titers, suggesting a role for this cytokine in the pathogenesis of these diseases.

A high incidence of serum IgG antibodies to the Epstein-Barr virus replication activator protein in nasopharyngeal carcinoma

TheBamHI Z EBV replication activator (ZEBRA) protein is involved in the switch from latency to productive cycle of Epstein-Barr virus. A recombinant ZEBRA protein was synthesized and assessed in enzymelinked immunosorbent assay (ELISA) for serum IgG response in nasopharyngeal carcinoma (NPC) patients. In 100 NPC serum samples that were positive for IgA to the EBV viral capsid antigen (VCA), 75% had IgG anti-ZEBRA antibodies. In contrast, only 3/83 (3.6%) serum samples from healthy donors and 2/50 (4%) from other cancers were positive for IgG to ZEBRA. Interestingly, in a selected group of 100 NPC sera negative for IgA to VCA, 25% contained IgG anti-ZEBRA antibodies. This suggests that the ELISA for IgG anti-ZEBRA may also identify earlier cases of NPC not detected by the conventional immunofluorescence test for IgA to VCA.

The Epstein‐Barr Virus (EBV) major envelope glycoprotein gp350/220‐specific antibody reactivities in the sera of patients with different EBV‐associated diseases

gp350 of Epstein‐Barr virus (EBV) induces a strong immune response in EBV‐infected individuals, but relatively little is known about the clinical relevance of this response in patients with different EBV‐associated malignancies and other diseases. Using our gp350‐expressing cell clones, we studied gp350‐specific humoral immune responses in the sera of individuals with nasopharyngeal carcinoma (NPC), chronic symptomatic EBV infection (CEI), Hodgkins disease (HD), acute infectious mononucleosis (IM) and healthy EBV‐seropositive individuals (HI). The titres of antibody‐dependent cellular cytotoxicity (ADCC) antibodies were highest in HI followed by CEI, HD and NPC. EBV‐neutralizing (NA) and gp350‐specific IgG antibody profiles in these conditions were: CEI > HI > NPC > HD, whereas IgA titres were the highest in NPC sera followed by CEI and HD. The sera from IM patients were found to be negative for gp350‐specific ADCC and IgA activities. Sera from HI were also negative for gp350‐specific IgA. A significant positive correlation was found between serum gp350 IgA and viral capsid antigen IgA and a significant negative one between IgM and ADCC titres. High IgA titres were also found in CEI and EBV‐genome positive HD in addition to NPC. Importantly, gp350‐specific IgA titres were of prognostic value in NPC patients. Our data provide new insights about the clinical relevance of gp350‐specific immune responses in these diseases. Int. J. Cancer (Pred. Oncol.) 79:481–486, 1998.© 1998 Wiley‐Liss, Inc.

Screening for nasopharyngeal carcinoma with an ELISA using the Epstein-Barr virus nuclear antigen, EBNA 1 : a complementary test to the IgA/VCA immunofluorescence assay

An ELISA using the Epstein-Barr virus nuclear antigen 1 (EBNA 1) was found to detect selectively specific IgA in sera from patients with nasopharyngeal carcinoma (NPC). The antigen, p107, was a 20-amino acid synthetic peptide, representing a major epitope of EBNA 1.267/294 (90.8%) of NPC patients had IgA antibodies to p107 but in normal individuals, only 41/577 (7.1%) had IgA/p107. In sera from patients with other cancers, 11/77 (14.3%) had IgA/p107 reactivity. 124 IgA/VCA positive and 86 IgA/VCA negative NPC sera were also tested for IgA/p107 binding in ELISA. The majority of IgA/VCA positive sera (117) also contained IgA/p107 antibodies. Of interest was the detection of 74/86 IgA/p107 reactive sera in the IgA/VCA negative group. The results suggest that the IgA/p107 ELISA could become a useful, complementary screening assay to the IgA/VCA immunofluorescence test for detection of NPC.

Transfer factor with anti-EBV activity as an adjuvant therapy for nasopharyngeal carcinoma: A pilot study

Overall survival of nasopharyngeal carcinoma (NPC) at UICC stage IV still remains unsatisfactory even with combination chemotherapy (CT) and radio-therapy (RT). In view of the association of reactivation of Epstein-Barr virus (EBV) with the development and recurrence of NPC, immunotherapy in the form of transfer factor (TF) with specific activity against EBV (TF-B1) was suggested as an adjuvant to a combination of CT and RT in order to improve survival. In the present study, 6 UICC Stage IV patients received TF-B1 and another 6 patients matched for disease stage were given TF prepared from peripheral blood leucocytes (TF-PBL). Results were compared with another 18 patients matched by age, sex, and stage of disease who received standard therapy without TF during the same period (C group). After a median follow up of 47.5 months, the survival for the TF-B1 group was found to be significantly better (P=<0.05) than the PBL and C group. While the 8 patients with distant metastasis (DM), not treated with TF-B1 (6 in the control and 2 in the PBL group), died due to progressive disease (average survival being 14.3 months), both patients with DM in the TF-B1 group had complete remission: one died of tuberculosis after surviving for 3.5 years and another is still alive, disease free, after 4.2 years. Although the series involved a small number of cases, the apparent effect of adjuvant immunotherapy in the form of TF with anti-EBV activity is of considerable interest.

Long-term survival of nasopharyngeal carcinoma patients treated with adjuvant chemotherapy subsequent to conventional radical radiotherapy

PURPOSE To assess the long-term survival of patients with nasopharyngeal carcinoma (NPC) who were treated with conventional radical radiotherapy (RT) followed by adjuvant chemotherapy. METHODS AND MATERIALS Ninety-one newly diagnosed patients with Stage III and IV (American Joint Committee on Cancer, 1988) NPC, seen at the University of Malaya Medical Center, Kuala Lumpur, Malaysia between January 1992 and May 1997, were treated with RT followed by adjuvant chemotherapy. The tumor dose was 70 Gy delivered in 35 fractions, 5 fractions weekly. Three cycles of chemotherapy, each consisting of 5-fluorouracil, 1 g/m(2)/d on Days 1-4 and cisplatin 100 mg/m(2) on Day 1, were administered 3 weeks after RT completion. Thirty-six patients had Stage II, 10 had Stage III, and 45 had Stage IV disease (AJCC 1997 staging system). RESULTS After a median follow-up of 61 months, the 5-year overall survival rate for all 91 patients was 80.1%, the disease-free survival rate was 76%, and the locoregional control rate was 85%. The 3-year overall survival rate for Stage II was 94.3%; it was 80% for Stage III and 79.8% for Stage IV (p = 0.0108). The 3-year DFS rate for Stage II was 90%; it was 80% for Stage II and 65% for Stage IV. The rate of distant failure for Stage IV was 8.9%. CONCLUSION Radical RT followed by adjuvant chemotherapy was effective in our patients with locoregionally advanced NPC. The long-term results appear encouraging, even for patients with Stage IV disease. This single institution experience deserves further investigation in prospective trials.

Detection of Epstein-Barr virus DNA in nasopharyngeal carcinoma using a non-radioactive digoxigenin-labelled probe

The presence of Epstein Barr virus (EBV) DNA in biopsies from the post-nasal space (PNS) of patients suspected of nasopharyngeal carcinoma (NPC) was detected by in situ cytohybridization with an EBV DNA probe labelled with the novel labelling compound digoxigenin. The digoxigenin probe was hybridised to cryostat sections of NPC biopsies and subsequently detected by an enzyme immunoassay procedure. It was found that in situ cytohybridization using the digoxigenin probe was much more rapid and sensitive (96 h compared to five weeks) than the current method of using 3H-labelled probe. Using the digoxigenin EBV probe, it was found that in all the eighteen NPC biopsies tested, EBV DNA was detected in malignant epithelial cells and infiltrating lymphocytes. EBV DNA was also detected in some normal epithelial cells in these NPC biopsies. EBV DNA was not detected in epithelial cells of non-malignant biopsies.

Linear epitopes of the replication-activator protein of Epstein-Barr virus recognised by specific serum IgG in nasopharyngeal carcinoma.

The linear antigenic epitopes of the Epstein-Barr virus replication activator protein (ZEBRA), recognised by specific serum IgG in nasopharyngeal carcinoma (NPC), were determined. This was achieved by synthesizing the entire amino acid sequence of ZEBRA as a set of 29, 22-residue peptides with an overlap of 14 amino acids. The ZEBRA peptides were tested in enzyme-linked immunosorbent assay (ELISA) for IgG binding in sera from 37 selected NPC patients who had IgG antibodies to the native ZEBRA protein. The most immunogenic epitope was peptide 1 at the amino-terminal end with 36 of the sera reactive against it. Further analysis of peptide 1, using the multipin peptide-scanning technique, defined a 10-amino-acid sequence FTPDPYQVPF, which was strongly bound by IgG. Two other regions of ZEBRA were also identified as immunodominant IgG epitopes, namely peptide 11 (amino acids 82–103) and peptide 19/20 (amino acids 146–175) with 8–13 of the NPC sera reactive against the peptides. The number of peptides reactive with individual NPC serum varies from 1 to 6 or more and there is some correlation between a greater number of peptide (at least 4) bound and a higher (at least 1:40) titre of serum IgA to viral capsid antigen. The immunodominant ZEBRA peptide 1 could be utilised in IgG ELISA for the detection of NPC.

In Situ Analysis of Epstein-Barr Viral Expression in Nasopharyngeal Carcinoma

Epstein-Barr virus (EBV) has a known association with several malignancies including African Burkitt’s lymphoma (BL) 1, nasopharyngeal carcinoma (NPC)2 and diffuse polyclonal non-Burkitt lymphomas in transplant patients3.

Early Diagnosis of Nasopharyngeal Carcinoma: A Multi-Pronged Approach

Nasopharyngeal carcinoma (NPC) is one of the highly malignant tumours of the head and neck, affecting most commonly the ethnic Chinese, living in the Southeast Asian countries—South China, Taiwan and Hong Kong. It is reputed for delayed diagnosis and as such poor prognosis (1). Not infrequently NPC patients with obvious clinical features, at times with extensive disease, are not diagnosed early due to inability on the part of cliniciansto obtain representative biopsy specimens for histopathological confirmation. Over a period of time this problem of late diagnosis has partly been resolved due to increased awareness on the part of the patients and the doctors in these regions, utilization of modern facilities to visualise the nasopharynx [in particular the fossa of Rosenmuller (FOR)] and by the detection of elevated titres of IgA anti Epstein Barr virus capsid antigen (IgA/VCA test).