Nonvirion Antigens Research Articles

Cytotoxic T Lymphocytes Directed against MAIDS-Associated Tumors and Cells from Mice Infected by the LP-BM5 MAIDS Defective Retrovirus

The LP-BM5 retrovirus, a complex containing ecotropic helper, recombinant MCF, and defective retroviruses, causes an immunodeficiency-termed mouse AIDS (MAIDS). Many disease features of MAIDS resemble those of AIDS, including terminal B cell lymphomas. Previously we generated from MAIDS-susceptible C57BL/6 mice cytolytic T lymphocytes (CTL) specific for MAIDS-associated B cell lymphomas. Data of the present study (1) exclude MCF and establish a role for defective virus in generating C57BL/6 CTL to MAIDS-associated tumors by experiments involving in vitro stimulation with cells from LP-BM5, ecotropic, or ecotropic-rescued defective virus-infected mice and (2) confirm that such CTL are specific for tumors of MAIDS origin. Several approaches testing for direct involvement of defective virus or its gag-encoded polyprotein, however, did not provide evidence that MAIDS tumor-specific CTL were directed to structural virion proteins, suggesting the possibility that such CTL are specific for nonvirion antigens whose expression depends on the action of the defective genome in the MAIDS disease process.

Study of antibodies to adenoviruses in patients with tumors of the urogenital system.

The presumable relationship between viruses and malignant disease has been studied. Approximately 4000 assays for antibodies to adenovirus were carried out with sera of patients with malignant and nonmalignant diseases of the urogenital organs and other organic diseases. The search was directed in the first place at antibodies to the early non-virion antigens of adenovirus-12 of oncogenic properties. On the evidence of the complement fixation reaction antibodies to adenoviruses were found to be less frequent in malignant tumors and in hypertrophy of the prostate than in the control group. Antibody to the non-virion antigen of adenoviruses was found in 53 per cent of patients with neoplastic diseases and prostatic hypertrophy, in 18 per cent of patients with urogenital diseases other than tumors and in 4 per cent of those with other organic diseases. The results point to a possible role of adenoviruses in tumors of the urogenital organs.

Immunocytochemical approach to virus-induced brain tumors - from the aspect of T (tumor) antigen.

Recently isolated human papovaviruses such as JC and BK viruses are not only antigenically related to SV40, but also neuro-oncogenic in experimental animals. Cells transformed in vitro or brain tumors induced by these viruses are usually free of infectious virus. However, the causual relationship between the tumor or transformed cells and the viruses can be immunocytochemically demonstrated by the presence of virus-specific non-virion antigens such as T (tumor) antigen. In the present study, the authors successfully applied a sensitive enzyme-labelled antibody method using Fab' fragment to localizing T antigens in SV40 or BK virus-induced brain tumors and infected cells. T antigens of SV40 and BK virus are immunocytochemically indistinguishable. Positive staining for T antigen is seen as a granular pattern in the tumor cell nuclei, but not the nucleoli, in the interphase and it appears as a loose network of electron-dense reaction precipitates associated with nuclear chromatin, whereas the chromosomes in mitosis do not seem to be tagged with immunoperoxidase reaction products for T antigens. On the other hand, T antigen is first observed in the cytoplasm of SV40-infected cells as early as 3 hours postinfection with subsequent transport to the nuclei within 24 hours. The intensity of staining of T antigen is markedly reduced because of maturation of virions in the infected cell nuclei 24 hours after infection. The authors believe that extensive studies of papovavirus T antigens are necessary to elucidate papovavirus oncology and may provide more information regarding possible viral etiology of human tumors of the central nervous system.

Immune response in rabbits to virion and nonvirion antigens of the Flavivirus kunjin.

The nature of the antibodies formed in rabbits in response to the following Kunjin virus antigens was examined: infectious suckling mouse brain (SMB), purified virion or rapidly sedimenting hemagglutinin (RHA), slowly sedimenting hemagglutinin (SHA), and envelope fragments prepared from RHA disrupted by 0.1 or 0.2% sodium deoxycholate (DOC). The hemagglutination-inhibiting (HI) and neutralizing antibody responses to SMB, RHA, and large envelope fragments (0.1% DOC) were remarkably uniform, antibodies appearing at the same time, attaining similar HI titers (lowest to envelope), and being of similar avidity early and late in the respone. The 19S (immunoglobulin M) antibodies to all antigens were always relatively type-specific, whereas the 7S (immunoglobulin G) antibodies were always broadly cross-reactive in HI tests. These results confirm that the envelope antigen is the principal antigen involved in the stimulation of protective neutralizing antibodies and contains both type- and group-specific antigenic determinants. The results also establish that there is no significant advantage in using purified RHA or SHA either for immunization or as hemagglutinin antigens in attempts to obtain greater specificity in the HI test. No differences were detected in the antibody responses to infective Kunjin virus, within the range 1,400 to 10(9) plaque-forming units (PFU). Below 1,400 PFU, there was no detectable response. Inactivated virus (10(6) PFU) also stimulated the normal antibody response. In contrast, small envelope fragments (derived with 0.2% DOC) and a detergent-solubilized extract of infected cells were unable to stimulate a detectable antibody response and the small envelope fragments may have induced low dose tolerance in one of two rabbits.

Distribution of dengue-2 antigens by electron immunocytochemistry

The distribution of dengue-2 antigens was studied in infected monkey kidney cells (LLC MK2) using an indirect, horseradish peroxidase-conjugated immunoglobulin technique. This procedure allowed both light and electron microscopic examination of serial-step sections of individual cells cut in a plane perpendicular to the monolayer. Both virion and nonvirion antigens were identified on the plasma membrane with this technique. A diffuse cytoplasmic reaction product was also present. The intensity and distribution of the cytoplasmic reaction product was related to disruption of the plasma membrane.

Cell-mediated immunity to herpes simplex virus types 1 and 2 antigens in leukoplakia and carcinoma in man.

Cell-mediated immune responses to herpes simplex virus 1 and type 2 virion and non-virion antigens were assessed in patients and in controls with oral leukoplakia, carcinoma and recurrent herpes labialis. Enhanced proliferation was found in lymphocytes from patients with recurrent herpes labialis or with leukoplakia showing epithelial atypia, and depressed responses were found in carcinoma. Very significantly positive correlations were shown between the responses to each of the herpes virus antigens. A specific increase in cell-mediated immunity to herpes virus in epithelial atypia was confirmed, but separation of the nonvirion from virion antigen is necessary before specific cell-mediated immune responses to the non-virion antigen can be assessed.

Antibodies to herpesvirus nonvirion antigens in squamous carcinomas.

Serums from tumor-bearing patients, cured patients, and normal subjects were examined for antibodies to the separated complement-fixing reactive components of nonvirion antigens of herpesvirus type 1 and type 2. The occurrence of antibodies to the antigens was similar in serums from tumor-bearing patients and cured patients. Antibodies to the antigens were observed among 21 of 24 (87 percent) cervical cancer cases, 44 of 49 (90 percent) laryngeal cancer cases, 15 of 24 (62 percent) cases of squamous cell carcinomas of the head and neck excluding the larynx, 2 of 24 (8 percent) nonsquamous cell cancer cases, and 3 of 51 (6 percent) normal subjects. By contrast, no differences were found in the titers of neutralizing antibodies to the virus in serums from laryngeal cancer patients and controls. The observations support an etiologic role of herpesviruses in cervical cancer and in laryngeal cancer, and possibly other squamous cell cancers of the head and neck.

Herpes Simplex and Herpes Genitalis Viruses in Etiology of Some Human Cancers

The results of complement fixation tests on 202 sera from people without cancer and from patients with cancer in 29 different areas of the body indicated that only those with nine varieties of advanced cancer (lip, mouth, oropharynx, nasopharynx, kidney, urinary bladder, prostate, cervix uteri, and vulva-all of 56 tested) gave positive specific reactions with nonvirion antigens induced by the DNA herpes simplex (HSV 1) and herpes genitalis (HSV 2) viruses. None of 57 people without cancer (including 10 with current and 18 with recurrent HSV 1 or HSV 2 infections), none of 81 patients with 20 other varieties of advanced cancer (gum, tongue, tonsil, salivary gland, accessory sinus, epiglottis, lung-bronchus, stomach, colon, breast, corpus uteri, ovary, testis, liver, thyroid, Wilms' embryonal kidney, melanoma, Hodgkin's disease, acute lymphocytic leukemia, and acute myelocytic leukemia), and none of four women with early malignant changes in the cervix uteri gave positive results. The seven patients with advanced cancer of the lip or oropharynx gave positive reactions with HSV 1 but not with HSV 2 nonvirion antigens (compatible with involvement of only HSV 1), all of the 13 women with advanced cancer of the cervix uteri and the one woman with advanced cancer of the vulva gave positive reactions with both HSV 1 and HSV 2 nonvirion antigens (compatible with involvement of only HSV 2), while among the 35 other positive patients only two (one with cancer of the kidney and one with cancer of the bladder) reacted with HSV 1 and not at all with HSV 2 nonvirion antigens. Positive sera failed to react with cells harvested at different times after high-multiplicity infection with the DNA vaccinia virus. Massive absorption of positive sera with trypsinized, uninfected human embryonic kidney cells failed to remove, or lower the titer of, the HSV 1 and HSV 2 nonvirion antibodies. All of these data taken together are interpreted as indicating that HSV 1 and HSV 2 play an etiologic role in certain human cancers, because they provide the kind of evidence by which virus-free experimental cancers can be proved to have been originally induced by such DNA viruses as polyoma, Simian Virus 40, or certain types of adenovirus.

Cytological localization of Dengue-2 antigens: an immunological study with ultrastructural correlation.

Immunofluorescence and electron microscopy were used to study the development and cytological distribution of dengue-2 virion and nonvirion antigens in monkey kidney cells. The type and combination of fixatives were found to affect the intensity of fluorescence. Paraformaldehyde fixation alone resulted in a low level of fluorescence, but an additional fixation step with ethanol or acetone resulted in maximum staining. No fluorescence was obtained after fixation with glutaraldehyde alone or in combination with other solvents. Methanol caused selective ablation of fluorescence by antibody to purified virions. Dengue-specific fluorescence that was most intense in the perinuclear area radiated in a granular pattern of decreasing intensity into the cytoplasm. The perinuclear fluorescence was associated with nonvirion antigens and the cytoplasmic fluorescence was associated with virion antigens. Electron micrographs of infected cells revealed vesicular bodies with reticular electron-dense centers, clusters of structurally complete virions deep within the endoplasmic reticulum, and single virus particles in vacuoles near the periphery of the cell.

Nonvirion antigens produced by herpes simplex viruses 1 and 2.

Of nine herpes simplex virus 1 strains (from lip, mouth, throat, cornea, or brain) only five produced enough nonvirion antigen (i.e., not a structural component of the virus) to be detected by complement fixation with specially prepared, virion-absorbed, type-1 guinea pig antisera, while the remaining four strains produced only enough of the same antigen to induce specific antibody in hyperimmunized guinea pigs. While the type 1 virion antiserum used reacted equally well by complement fixation with the type 1 and type 2 strains, the type 1 nonvirion antisera failed to react with nonvirion antigens produced by three type-2 (genital) strains. However, type 2 nonvirion antiserum reacted equally well with the three type 2 and four type 1 nonvirion antigens that were tested. It appears, therefore, that while herpes simplex virus 1 codes only for type 1 nonvirion antigen, herpes simplex 2 codes not only for an immunologically distinct type 2 nonvirion antigen but also for enough type 1 nonvirion antigen to stimulate antibody production for it. Herpes simplex 2 nonvirion antigen exhibited the same properties as type 1, i.e., its activity was lost on storage at 4 degrees for 15 days, it was sedimented by centrifugation at 33,360 x g for 1 hr, and the maximum concentration was found at 3 hr in guinea pig kidney culture cells, but at 24 hr in HEp 2 and rabbit kidney culture cells. Sera from patients with genital lesions caused by herpes simplex virus 2, as well as from randomly selected adults, failed to react with either type 2 or type 1 nonvirion antigens. Accordingly, the basic information is now available to permit the use of these nonvirion antigens to determine the possible role of the herpes simplex viruses in certain human cancers.

Soluble membrane antigens of lip and cervical carcinomas: reactivity with antibody for herpesvirus nonvirion antigens.

With the use of antibody for herpesvirus nonvirion antigens (not structural components of the virus) complement fixing reactivity has been shown for soluble membrane antigens separated from lip and cervical carcinomas but not for similar extracts from normal vaginal tissue or intestinal carcinoma. Neither the serum obtained from the guinea pig before hyperimmunization with the herpesvirus nonvirion antigen nor the antiserum of guinea pigs immunized with comparable uninfected cell extracts reacted with these tumor soluble membrane antigens. Since the above soluble membrane antigens could be specific markers for the presence of virus genome within the tumor cells, the findings could support an etiological role of herpesvirus in selected human malignancies.