Viral attributes and host factors in carcinogenesis: herpesviruses.

This paper describes two different experiments of nature: 1) the persistence of unusual virus strains of Epstein-Barr virus (EBV) (which proved oncogenic in vitro) and cytomegalovirus (CMV) in lymphoid cells following congenital or early acquired infection; 2) the occurence of multiple cases of Burkitt's lymphoma and nasopharyngeal carcinoma in one family. All the members of this family were EBV viral capsid antigen (VCA) and nuclear antigen (EBNA) antibody positive. The two patients with nasopharyngeal carcinoma had high titers of EBV-VCA, EA, and EBNA antibodies. The only member of this family having EBV early antigen (EA antibodies in addition to the patients with tumors was the mother. Borderline IgA deficiency was documented in 3 members of this family. These findings illustrate the importance of host factors (intracellular resistance to transformation and secondarily, immunological surveillance) in the outcome of the host-virus challenge whether cancer or infectious disease is the outcome. Extensive studies of these cases may provide the best insight into the mysteries of viral oncogenesis.     

Abnormal anti-Epstein Barr virus antibodies in carriers of the X-linked lymphoproliferative syndrome and in females at risk

The asymptomatic hemizygous female carriers of the X-linked lymphoproliferative syndrome (XLP) have abnormal antibody responses to EBV. This suggests partial expression of the defect that leads to EBV-provoked life-threatening diseases in their affected sons. EBV specific antibodies were measured in 65 serum samples of 12 obligate carrier females and seven of their daughters (females at risk) during periods ranging from 1 to 5 yr. Abnormal qualitative antiviral capsid antigen (VCA) IgG titers were nearly fourfold higher than normal controls, two carriers had persistent IgM anti-VCA antibody, two-thirds had persistent IgA anti-VCA antibody, and half of the women had titers to early antigen (EA). Five of seven females exhibited a similar persistent pattern. In contrast, none of the unaffected family members nor 23 normal controls expressed IgA or IgM titers to VCA even with high exposure to the virus, and anti-EA was detected in only one control. Therefore, these findings may prove useful for detecting carriers of the syndrome. Abnormal anti-EBV titers similar to the carrier pattern have been reported in patients and other immunosuppressed individuals, and are indicative of active viral infection.     

Human immune responses to synthetic peptides from the Epstein-Barr nuclear antigen

Humans infected with Epstein-Barr virus (EBV), the causative agent of infectious mononucleosis, develop antibodies against a nuclear antigen (EBNA) that is present in virally transformed B lymphocytes. The EBNA protein contains a unique glycine-alanine repeating sequence. We have synthesized peptides corresponding to various regions of the EBNA molecule within and near this sequence. Rabbit antibodies against the peptides within the sequence reacted directly with the EBNA protein, as detected by Western blotting. The sera of individuals with antibodies against Epstein-Barr virus contained abundant antibodies also reactive with one or several of the synthetic peptides within the sequence. Moreover, human antibodies against these simple peptides were induced specifically early in the course of infectious mononucleosis. When compared with normal controls, antibody levels to the glycine-alanine peptides were significantly higher in patients with rheumatoid arthritis and progressive systemic sclerosis, but not in patients with two other autoimmune diseases. These results document that i) antibodies against the peptides detect the EBNA protein, ii) humans infected with EBV produce high titers of antibodies reactive with these synthetic antigens, and iii) antibody titers against the peptides are abnormally elevated in certain autoimmune diseases.     

Evaluation of purified H and M antigens of histoplasmin as reagents in the complement fixation test.

Complement-fixation (CF) tests were performed with purified H and M antigens, histoplasmin, and Histoplasma capsulatum whole cell yeast phase antigen using sera of 126 patients with proven or suspected histoplasmosis. Specific titers for either H or for M antibody were obtained with the individual purified antigens; the highest titers were comparable to those obtained with histoplasmin. However, in sera containing only anti-M antibody, the titers obtained with the purified M antigen were 2 to 16 times those obtained with the histoplasmin or yeast phase antigens. The CF test for either H or M antibody was 4 to 32 times as reactive as the agar-gel microimmunodiffusion test; in general precipitin lines were obtained with either H or M antigens from sera with CF titers greater than or equal to 8. With sera containing H antibody, there was an excellent correlation between the CF titers obtained with purified M antigen and histoplasmin. The correlations of CF titers with H antigen and either histoplasmin or yeast phase antigen were very low.     

Epstein-barr virus (EBV)-associated antibodies in a case of Burkitt's lymphoma during 14 years of sustained remission

Summary A Burkitt's lymphoma (BL) patient who sustained remission for more than 14 years after chemotherapy was monitored by means of serial serum samplings. The sera were titrated for antibodies against the Epstein-Barr virus (EBV)-associated cell membrane antigen (MA), viral capsid antigens (VCA), early antigen complex (EA R/D), and nuclear antigen (EBNA), and also for reactivity in the antibody-dependent cellular cytotoxicity (ADCC) test. The initial serological profile corresponded to that of most BL patients with active disease. During remission, it changed to resemble that of normal persons with persistent, latent EBV infection, at least qualitatively. The prognostic and biological implications of the titer levels and their changes are discussed.Abbreviations ADCC antibody-dependent cellular cytotoxicity - BCG Bacillus Calmette-Guérin - BL Burkitt's lymphoma - D diffuse component of EA - EA Epstein-Barr virus-induced early antigens - EBNA Epstein-Barr virus-specific nuclear antigen - EBV Epstein-Barr virus - KCC Kenya Cancer Council Registry - MA EBV-associated cell membrane antigen complex - NK natural killer - R restricted component of EA - VCA EB viral capsid antigen complex     

Fluctuations of epstein-barr virus serological antibodies and risk for nasopharyngeal carcinoma: a prospective screening study with a 20-year follow-up

Background

The impact of variation of Epstein-Barr virus (EBV) antibody titers before the development of nasopharyngeal carcinoma (NPC) is still unclear. We analyzed the fluctuations of antibodies against EBV before histopathological diagnosis to assess the risk of NPC and aimed to provide a reliable basis for screening in high risk populations.

Methods

This study was based on a population-based screening program in Sihui County in Guangdong Province of China. A total of 18,986 subjects were recruited in 1987 and 1992, respectively. Baseline and repeated serological tests were performed for IgA antibodies against EBV capsid antigen (VCA/IgA) and early antigen (EA/IgA). Follow-up until the end of 2007 was accomplished through linkage with population and health registers. Cox proportional hazards regression model was used to estimate the relative risk of NPC in association with EBV antibodies. Time-dependent receiver operating characteristic curve (ROC) analysis was used to further evaluate the predictive ability.

Results

A total of 125 NPCs occurred during an average of 16.9 years of follow-up. Using baseline information alone or together with repeated measurements, serological levels of VCA/IgA and EA/IgA were significantly associated with increased risks for NPC, with a striking dose-response relationship and most prominent during the first 5 years of follow-up. Considering the fluctuant types of serological titers observed during the first three tests, relative risk was highest among participants with ascending titers of EBV VCA/IgA antibodies with an adjusted hazard ratio (HR) of 21.3 (95% confidence interval [CI] 7.1 to 64.1), and lowest for those with decreasing titers (HR = 1.5, 95% CI 0.2 to 11.4), during the first 5 years of follow-up. Time-dependent ROC analysis showed that VCA/IgA had better predictive performance for NPC incidence than EA/IgA.

Conclusion

Our study documents that elevated EBV antibodies, particularly with ascending titers, are strongly associated with an increased risk for NPC.     

Naturally occurring plasmodium-specific IgA antibody in humans from a malaria endemic area

Blood samples collected from individuals belonging to an endemic area in Uttar Pradesh, were tested for plasmodial antigen specific immunoglobulin A (IgA) by enzyme immuno assay using soluble extract ofPlasmodium falciparum from culture. Among 773 (20.18%,P < 0.0001) samples 156 sera demonstrated a detectable seropositivity for antigen specific IgA. IgA levels were higher among individuals who experienced repeated attacks of malaria compared to acute infected patients. Among seropositive individuals the IgA titers were found increased with the age. Immunoglobulin isolated from sera having high level of IgA showed growth inhibitory effect inPlasmodium falciparum in vitro. A group of sera with high IgA antibody againstPlasmodium falciparum crude antigen showed seronegativity with specific peptides. Statistically, no positive or negative correlations were observed between antigen specific IgG and IgA. However, there was a tendency towards negative correlation between IgA and IgM. Mechanisms for the parasite specific IgA production remain to be established.     

Changes in Epstein-Barr virus antibody titers associated with aging

Antibody titers to the Epstein-Barr virus (EBV), early antigen (EA) IgG, and virus capsid antigen (VCA) IgG and IgA, were measured in 44 geriatric subjects to determine if the depression in cellular immunity known to be associated with aging affects the expression of latent EBV. Similar assays were performed on plasma obtained from a young adult (medical student) population as a control group. We found that 89% of the geriatric samples were positive for EA IgG, and 83% of the plasma obtained from medical students were positive for EA IgG. One hundred percent of the geriatric subjects were positive for VCA IgG, and 87% of the medical students were positive for VCA IgG. Seven percent of the medical student blood samples were positive for VCA IgA; in contrast, 36% of the blood samples obtained from the geriatrics subjects were positive. Significant differences were also found in the geometric mean titers (GMT) of antibodies to EBV antigens; the GMT to EBV EA and VCA were significantly higher in the geriatric group. The data suggest that there may be some loss of control over latent EBV by the cellular immune response in geriatric individuals.     

用转染EB病毒基因片段的真核细胞检测鼻咽癌病人的抗EB病毒核抗原-1(EBNA-1)抗体

用转染EBV DNA Bam HI K片段后稳定表达EBNA-1的K_4细胞作为靶细胞,检测50份鼻咽癌病人和38份健康对照者血清中的IgG/EBNA-1抗体;阳性率分别为100％和92％。前者的平均几何滴度为89.4,后者为18.3,前者约为后者的5倍。同一批被检血清经SPA吸收去除IgG竞争性抑制后,鼻咽癌病人的IgA/EBNA-1抗体阳性率达78％(GMT 20.9),健康人的IgA/EBNA-1抗体阳性率仅5.3％,效价亦很低(GMT 5.2)。表明IgA/EBNA-1抗体对鼻咽癌是比较特异的,可考虑作为鼻咽癌血清学诊断的指标之一。     

Epstein-Barr virus antibodies in patients with ataxia-telangiectasia and other immunodeficiency diseases

An unusual antibody response to the Epstein—Barr virus (EBV) has been noted in patients with ataxia—telangiectasia. Of a group of 16 such patients 8 were found to have antibodies in their serum to the EBV viral capsid antigen (VCA), and 4 of them also had antibodies to the EBV early antigen (EA); antibodies to the nuclear antigen (EBNA), however, were absent in 3 of the 8. The antibody pattern persisted for more than 2 years in the patients available for follow-up study. In comparison, of 24 patients with various other immunodeficiency syndromes 9 were found to have EBV-VCA antibodies in their serum, but none of the 9 had EA antibodies and 3 lacked EBNA antibodies. Two other groups of subjects, all of whom had EBV-VCA and EBNA antibodies in their serum late after an EBV infection, were also studied; 82 had infectious mononucleosis and 55 were healthy and had no such history. EA antibodies were detected in 45 of the first group during the acute phase of the illness but persisted in only 6 of the 68 who were followed up for more than 2 years, and they were detected in only 7 of the second group.All eight lymphoblastoid cell lines established from the peripheral blood of the four patients with ataxia—telangiectasia who are still available for follow-up study express EBV-VCA, whereas most similar cell lines established from normal individuals express only EBNA. In two of these patients cell-mediated immunity, as assessed from lymphocyte transformation induced by mitogens, was markedly decreased but autologous cell-mediated immune regression of EBV-induced transformation of B (bone-marrow-derived)-lymphocytes was normal. The percentage of T (thymus-derived)-helper cells was greatly decreased in two of the three patients in whom it was measured, and the percentage of T-suppressor cells was greatly increased in one of them, but the percentage of total T-lymphocytes was within normal limits in all three.The possible significance of these findings — in particular, the persistence of EA antibodies and the diminished restriction of expression of EA — in the late development of tumours after an EBV infection in patients with ataxia-telangiectasia deserves careful attention. Finally, the apparent correlation between immunoglobulin deficiency and poor or absent EBNA antibody response warrants further study.     

Epstein-Barr virus-positive Burkitt's lymphoma in a German woman during pregnancy

A fatal case of a Burkitt's lymphoma which occurred in a 34-year-old German woman during pregnancy is described. Nearly all organs showed either diffuse or nodular infiltration by tumor cells. Placenta and fetus were free of detectable tumor tissue. The patient had extremely high antibody titers (1 : 2056), both against Epstein-Barr virus capsid antigen (VCA) and the early antigen complex (EA). Within the tumor cells the Epstein-Barr virus-specific nuclear antigen EBNA and viral DNA was detected. A cell line established from a tumor biopsy displayed a translocation involving chromosomes 2 and 8. The role of Epstein-Barr virus in the development of Burkitt's lymphoma is discussed.     

用重组Epstein-Barr病毒早期蛋白P83为抗原检测鼻咽癌病人血清中IgA抗体

以重组EB病毒早期蛋白P83为抗原,用Western blot检测了135份鼻咽癌病人和100份正常人血清中IgA/P83抗体,阳性率分别为96％和0,以常规的间接免疫酶法检测这两种血清中IgA/EA抗体,阳性率分别为71％和0。IgA/P83抗体的几何平均滴度较IgA/EA高4倍以上。这表明,用Western blot查IgA/P83敏感、特异,可替代用间接免疫酶法查IgA/EA用于鼻咽癌的早期诊断。     

Comparison of 4 serological tests--complement fixation, neutralization, fluorescent antibody to membrane antigen and immune adherence hemagglutination--for assay of antibody to varicella-zoster (V-Z) virus.

Four tests for antibody to varicella-zoster (V-Z) virus were compared; these were tests of complement fixation (CF), neutralization (NT), fluorescent antibody to membrane antigen (FAMA) and immune adherence hemagglutination (IAHA). Fifty-two sera from patients with varicella and zoster and from recipients of live varicella vaccine were examined by the 4 tests. The CF test was least sensitive, but the antibody titers by the NT, FAMA and IAHA tests were roughly comparable. The IAHA test was the simplest and fastest to perform, and appeared suitable for routine serological assay to V-Z virus. The correlation between the IAHA antibody titer and susceptibility of individuals to clinical varicella was investigated retrospectively using sera obtained during 2 outbreaks of varicella in an institution for children, where all the unvaccinated children had developed varicella symptoms. Most of the 25 pre-exposure sera from unvaccinated children examined by the IAHA test had tiers of less than 1:2. In contrast, all the 23 sera from vaccinated children who did not develop varicella had detectable antibody titers of 1:2 to 1:64. These results indicate that the IAHA titer reflects the susceptibility or resistance of individuals to clinical varicella.     

Anticomplement immunofluorescence for the titration of antibody to varicella-zoster virus

Anticomplement immunofluorescence (ACIF) was tested for its use for the titration of antibody against varicella-zoster virus (VZV). ACIF antibody responses of patients with VZV infection were specific for VZV antigen and heterotypic responses to herpes simplex virus type-1 and cytomegalovirus antigens were not observed. Comparative studies of ACIF, membrane immunofluorescence (MIF) and indirect immunofluorescence (IF), using acetone-fixed antigen, were carried out with nonimmune sera and convalescent sera of patients who had recovered from varicella, herpes zoster and Rumsey Hunt disease. Nonspecific staining occurred with some nonimmune sera at a 1:4 dilution in the MIF and IF tests, after freezing and thawing of the serum, but not in the ACIF test. The antibody titers in convalescent sera agreed well in these three methods and the highest titer was obtained by MIF. The titers in ACIF and IF were similar but the ACIF antibody decreased earlier than the IF antibody during convalescence. On the other hand there was a discrepancy between the titers of ACIF and those of MIF and IF antibody in the sera of healthy adults, all sera with titers higher than 10 in the MIF and IF tests had titers below 10 in the ACIF test. The average titer of ACIF antibody declined to less than 10 with increasing age (13 to more than 20 years), whereas the MIF antibody increased during the same period of life.     

Impact of ELISA and immunoblot as diagnostic tools one year after eradication of Helicobacter pylori in a multicentre treatment study

The performance of serological tests for Helicobacter pylori infections is hampered by the persistence of antibodies after eradication therapy or spontaneous healing. Detection of different antigens or immunoglobulin classes might have an impact on the validity of serodiagnosis. The aim of this study was to assess the decrease in IgA and IgG antibody levels after eradication of H. pylori. Serum samples of 242 patients with active duodenal ulcer were tested with the ELISA and the immunoblot (IB) techniques for H. pylori-specific IgA and IgG antibodies before therapy and 1 year after successful eradication. From a total of 81 patients paired sera were available. At the end of the follow-up period ELISA antibody titres from the IgA class had decreased from a mean value of 6.69 to 4.26 units (P = 0.0001), and IgG class antibody titres from a mean value of 21.9 to 12.1 units (P = 0.0001). Regarding seroreversion, from 34 initially IgA positive sera 16 (47%), and from 74 IgG positive sera 18 (24%), had definitively reverted to 'negative'. One year after eradication, when tested with the immunoblot, the antibody responses against specific antigens of 37% IgA-positive sera (23/62) and 8% IgG-positive sera (6/78) reverted to 'negative', compared to a seroreversion rate of 27% of the anti-CagA IgA-positive sera (18/67) and of 9% of the anti-CagA IgG-positive sera (7/79). In conclusion, despite an overall significant decrease of H. pylori antibodies, both tests cannot be recommended for monitoring treatment success.     

CEA-containing immune complexes in sera of patients with colorectal and breast cancer — analysis of complexed immunoglobulin classes

Summary A sandwich enzyme immunoassay was developed to detect circulating immune complexes containing carcinoembryonic antigen (CEA) and immunoglobulin (Ig) G, IgA, or IgM using a nitrocellulose-bound anti-CEA antibody as the solid phase reagent. Elevated levels of CEA-containing circulating immune complexes (CEA-IC) were found in 15.4% of 117 sera from patients with colorectal cancer in a postsurgery follow-up study. Also in 24.5% of 102 sera from patients with breast cancer in different states of disease CEA-IC were found. The predominant Ig determined in CEA-IC of colorectal cancer patients was IgA, followed by IgG and IgM, whereas IgG and IgM were the most frequent Igs in CEA-IC of breast cancer patients. Elevated CEA levels were found in 12.0% of the colorectal cancer patients and in 25.4% of sera from breast cancer patients. No significance for the coincidence of elevated CEA levels and CEA-IC was recorded in all patients sera tested. In sera of patients with disease recurrence, however, both parameters were shown to be elevated (CEA 80.7% and CEA-IC 42.3%). The data presented indicate the detection of CEA-IC as an additional parameter for the identification of patients at increased risk for disease recurrence.     

Allergic bronchopulmonary aspergillosis (ABPA): Studies on the general and specific humoral response

Serum specimens from 138 patients suffering from chronic respiratory disorders including 63 with allergic bronchopulmonary aspergillosis (ABPA), 20 with suspected ABPA, 25 with pulmonary tuberculosis, 14 with bronchial asthma, 10 with chronic bronchitis and 6 with miscellaneous pulmonary conditions were studied for circulating antibodies to Aspergillus. The ammonium sulfate test was employed with an iodine-125 labeled mycelial component derived from Aspergillus fumigatus. When compared to normal controls from the same area, this test indicated that sera from 82 per cent of patients with ABPA had elevated binding titers to the radiolabeled antigenic component. Immunodiffusion using a culture filtrate antigen from A. fumigatus, revealed precipitating antibody to this fungus in 89 per cent of sera from ABPA patients. The majority of patients with ABPA demonstrated marked elevations of total serum IgE, moderate elevations of serum IgA and IgD and slightly increased levels of IgG and IgM.This study was supported in part by Research Grant AI 10940 from the National Institutes of Health and by NHLI Contract N01-HL-3-2942(B), and forms a part of the Ph.D. thesis submitted by Z.U.K. to the University of Delhi.     

Clinical and immunological effect of intravesical interleukin-2 on superficial bladder cancer

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.     

血清中Epstein-Barr病毒膜抗原IgA抗体检测法的改进及应用

用萄葡球菌菌体A蛋白(SPA)预先处理被检血清,以去除抗体IgG部份的竞争性结合,提高了间接免疫荧光法检查鼻咽癌病人血清中EB病毒膜抗原IgA(IgA/MA)抗体的敏感性及特异性,检查48例鼻咽癌病人血清IgA/MA抗体,阳性率为100％,血清几何平均滴度为141;40例其它恶性肿瘤病人和46例正常人都检不出IgA/MA抗体,免疫荧光法测得IgA/MA抗体阴性的6例鼻咽癌病人血清,SPA吸收后呈阳性反应,此改进方法可用以追踪观察鼻咽癌病的病程及预后。