Discrimination of leukemias, lymphomas, and non-neoplastic controls by retroviral serum markers.
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Biomedical subjects
Publications and source records attributed to V Erfle.
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In a first seroepidemiological study on the prevalence of the human T-lymphotropic retrovirus HTLV-III in West Germany, sera of 26 patients with acquired immunodeficiency syndrome (AIDS), 33 patients with lymphadenopathy syndrome (LAS) or AIDS related complex (ARC), and 113 homosexual men at risk of AIDS were screened for IgG antibodies to HTLV-III by an enzyme linked immunosorbent assay (ELISA). 22 out of 26 AIDS-patients (84.6%), 24 out of 33 LAS-patients (72.7%), and 44 out of 113 healthy homosexual men with increased risk of AIDS (38.9%) were found positive for antibodies to HTLV-III. Heterosexual controls including healthy laboratory workers and medical personnel with contact to AIDS patients did not show antibodies to HTLV-III. The HTLV-III antibodies analyzed predominantly recognize a protein of molecular weight 41,000 (p41).
The prevalence of antibodies to human T-lymphotropic virus III (HTLV-III) was determined in a total of 140 hemophiliacs and 36 polytransfused patients from three medical centers by an enzyme linked immunosorbent assay (ELISA) and confirmatory tests. 58 hemophiliacs (41.4%) were seropositive. In all instances where the origin of the coagulation factors given to these patients could be determined, blood products came from the United States. In addition, 2 of 36 polytransfused patients, mostly with acute leukemias, who were transfused with blood products from local donors were positive for HTLV-III antibodies. No HTLV-III antibodies were detected in 237 blood donors selected in part from the donor pool of the polytransfused patients.
Sensitive cocultivation techniques were applied to study the radiation-induced activation of endogenous retroviral genomes in different mouse strains by the alpha-emitting radionuclide 224Radium. Activated infectious C-type retroviruses were detected in spleen, bone marrow and bone tissues of C57BL/6-, BALB/c- and NMRI mice. The titres of high-dose-irradiated animals were higher than those found in low-dose-irradiated animals. Infectious retrovirus could be detected with a dose of 13.2 rad (maximum dose rate 0.9 rad/day) in the skeleton, and a dose of 4.2 rad (maximum dose rate 0.3 rad/day) in the spleen. The virus activation pattern was different in the three mouse strains. These data indicate that activation of endogenous retroviral genomes by alpha-irradiation shows a dose-effect relationship and a dependence on the genetic background of the mouse.
We have shown previously that antigens and also circulating immune complexes related to the primate retroviral envelope glycoprotein SiSV gp70 correlate with early mortality and survival of 56 patients with acute leukemias and chronic myelogenous leukemia in blast crisis. The prevalence and general distribution of these antigens and immune complexes in human sera was therefore of obvious interest. We now report an analysis of a total of 200 sera from 166 patients. Of these 113 sera were obtained from 84 patients with acute or chronic leukemias and 87 from 82 non-leukemic subjects, including laboratory workers and patients with non-leukemic neoplasias. Antigens and immune complexes were determined by enzyme-linked immunosorbent assays (ELISA). The anti-SiSV gp70 antiserum used predominantly recognized the protein moieties of the glycoproteins. The distribution of SiSV gp70-related antigens and immune complexes was similar among leukemic and non-leukemic sera. The prevalence of SiSV gp70-related antigens was 53% and of SiSV gp70-related immune complexes 49% in all sera. SiSV gp70-related antigens were detected in a somewhat higher proportion of non-leukemic (69%) than leukemic sera (40%), whereas SiSV gp70-related immune complexes and cross-reactive antibodies were more evenly distributed in leukemic and non-leukemic sera (in 46 and 51% of leukemic and 54 and 51% of non-leukemic sera). Presence of antigens correlated with presence of SiSV gp70-related immune complexes in 71% of all sera, but in 13% of all sera antigens were detectable only by determining SiSV gp70-related immune complexes. Total circulating immune complexes did not correlate with SiSV gp70-related immune complexes. The origin and pathophysiological role of the antigens are discussed.
The prevalence of antibodies to human T-lymphotropic virus III was determined in acquired immunodeficiency syndrome (AIDS) risk groups by an enzyme linked immunosorbent assay and confirmatory tests in four different areas in West Germany. Twenty-four of 28 homosexual AIDS patients (86%), 24 of 33 homosexual patients with lymphadenopathy syndrome or AIDS related complex (73%), and 44 of 113 asymptomatic homosexuals at risk for AIDS (39%) were seropositive. In three groups of hemophiliacs, 8 of 35 in 1983 (23%), 25 of 65 in early 1984 (39%), and 19 of 23 in late 1984 (83%) showed positive results. Two sera from 36 polytransfused patients were also positive, whereas 36 selected blood donors, and 32 healthy laboratory and clinical personnel were all negative. Also no human T-lymphotropic virus III antibodies were detected in sera of 187 prostitutes in the Munich area.
Human sera contain antigens and also circulating immune complexes that are related to the primate retroviral envelope glycoprotein gp70 of simian sarcoma/simian sarcoma associated virus (SiSV) and of gibbon ape leukemia virus (GaLV). SiSVgp70 related antigens (AG) and immune complexes (IC) are detected both in leukemic and in nonleukemic sera. In a further analysis of these data, the prognostic significance of SiSVgp70 related AG and IC in leukemic patients was examined. The data show that the presence of SiSVgp70 related AG and IC indicates an unfavorable clinical course and a shorter survival time in acute leukemias (AL) and in chronic myelogenous leukemia in blast crisis (CML-BC). Survival data of 56 of 64 patients tested were analyzed (38 patients with AL and 18 patients with CML-BC). Patients with AL whose sera were positive for SiSVgp70 related AG and IC had a median survival time of 9.5 months after diagnosis versus 16 months for patients negative for such AG and IC. This difference in survival time was more pronounced for patients with acute nonlymphocytic leukemia (ANLL) (6.5 versus 19 months). The difference in survival between SiSVgp70 related AG- and IC-negative and positive groups as tested by life table analysis (log-rank test) is significant (P less than 0.05). Patients with AL of the AG- and/or IC-positive group had fewer complete remissions. Patients who had no remissions belong to the AG- and/or IC-positive group (P = 0.06). Patients with CML-BC whose sera were positive for SiSVgp70 related AG and/or IC had a median survival time of 2 months after diagnosis versus 7 months for patients with sera negative for such AG and IC. As tested by log-rank test, survival curves between the two groups are significantly different (P less than 0.05). These findings suggest that SiSVgp70 related AG and IC may play an important role in the course of acute leukemia and can provide useful prognostic information.
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Several antibiotics were examined for their potential to eliminate mycoplasmas from contaminated cell cultures. Acholeplasma laidlawii, Mycoplasma arginini, Mycoplasma hyorhinis and Mycoplasma orale were effectively eliminated from experimentally contaminated mouse fibroblasts and mink epithelial cells by the use of the antibiotics minocycline and tiamutin . An elimination procedure was established, which involved the consecutive treatment of the cultures over a period of 3 weeks, followed by cell cloning. This procedure was effective when applied to cell lines which had been contaminated with unidentified and partially non-cultivable strains of mycoplasmas.
Spontaneous osteomas in strain 101 mice, a strain which has a high incidence of benign bone tumours, harbour numerous C-type virus-like particles with pleomorphic characteristics. A cell-free extract from osteomas from two mice induced bone tumours, together with osteopetrosis and lymphomas, in newborn mice of the low incidence NMRI strain after a latent period of 12 to 15 months. When C3H embryo fibroblasts were infected with the osteoma extract, the resulting cell line produced virus (OA MuLVC) with a high titre. OA MuLVC was cloned by serial endpoint dilution and NIH 3T3 cells were productively infected. The resulting virus was named OA MuLVN. OA MuLVC and OA MuLVN also induced bone tumours, osteopetrosis and lymphomas 12 to 15 months after injection into newborn NMRI mice. The isolated virus showed typical characteristics of the murine retrovirus group. Fv-1 host range restriction assays classified the viruses as N-ecotropic and XC-positive. Tryptic p30 peptide analysis and RNase T1 fingerprint analysis of OA MuLVC and OA MuLVN indicated that OA MuLVC contains an Akv-like virus as well as additional components, whereas OA MuLVN is closely related to Akv, but not identical to it. Serological analysis of the envelope proteins using monoclonal antibodies also showed the virus to be similar, but not identical, to Akv virus.
The detection and characterization of oncogenes via RNA tumor viruses (or retroviruses) and the recognition of their location at breakpoints of chromosomal translocations which are frequently found in certain human neoplasms has promoted present understanding of molecular mechanisms underlying carcinogenesis. Oncogenes are cellular genes which can be transduced by RNA tumorviruses and induce malignant transformation under experimental conditions in vivo and in vitro. A role of retroviruses in human leukemogenesis is suggested by epidemiological observations and by the isolation of such viruses from several human T-cell leukemias and lymphomas (human T-cell leukemia/lymphoma virus or HTLV) as well as by biochemical association of retroviral markers with human leukemias. A role of HTLV has been suggested also in a human immune deficiency syndrome (AIDS). In view of the well known role of many factors in carcinogenesis the concept of carcinogenesis as a multistep process as well as the concept of cocarcinogenesis and the role of cofactors other than viruses, such as radiation and chemicals, aging, hormones, graft vs host reaction, environmental factors etc., will have to be carefully considered.
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Antigens have been detected in 35 to 40% of sera from patients with leukemia that cross-react with the Mr 30,000 core proteins (p30) of baboon endogenous virus (BaEV) and/or of simian sarcoma-simian sarcoma-associated virus (SiSV) in a solid-phase enzyme immunoassay using anti-SiSV p30 and anti-BaEV p30 antisera. These antigens could not be found in sera from nonleukemic persons. Fetal calf serum; normal chicken, goat, rabbit, and rhesus sera; and normal human serum components like albumin, immunoglobulins, and transferrin did not react with the anti-SiSV p30 and anti-BaEV p30 antisera. The reactivity in the leukemic sera was abolished by treatment with protease, but not with glycosidases. The antigens purified by immunoaffinity chromatography showed essentially one band with an apparent molecular weight of 70,000 on sodium dodecyl sulfate: polyacrylamide gel electrophoresis. In competition enzyme-linked immunosorbent assays the leukemia-associated antigens competed out SiSV p30 in the anti-SiSV p30 system. Peptide mapping experiments with antigens from sera of two different leukemic patients showed that the two antigens were identical concerning numbers of peptides and their position. Eleven of 21 major peptides of SiSV p30 and 10 of 20 major peptides of BaEV p30 (50 to 60% of major peptides) showed mobilities identical with those of the major peptides of the human antigens. The data suggest the presence in human sera of retroviral antigens closely associated with leukemia.
The applicability of the enzyme immunoassay technique to the detection of group and interspecies determinants of C-type retroviral proteins was tested. For this purpose four groups of C-type retroviruses (MuLV, FeLV, SiSV/GaLV, and BaEV/RD114) were assayed for group-specific and interspecies reactivities of their p30 proteins by an enzyme-linked immunosorbent assay (ELISA). We found that the ELISA can detect group-specific as well as interspecies determinants with sensitivity and reproducibility in purified p30 proteins, disrupted viruses, and cell extracts if an anti-p30 interspecies antiserum is used. If monospecific antisera against MuLV p30, SiSV p30, or BaEV p30 were used, only group-specific reactivities were detected reproducibly, whereas the detectability of interspecies determinants depended on the antisera used and varied even with the same antisera. In assays in which the reactivity of native and denatured p30 proteins was compared the detectability of sodium dodecyl sulphate-denatured MuLV p30 was better than that of native MuLV p30 suggesting that some of the most broadly cross-reactive sequences are localized inside the protein molecule and are freed by the denaturation process. Antisera raised against native and denatured p30 proteins showed identical spectra of reactivity.
Activation of endogenous C-type retroviruses by nuclear polyhedrosis viruses (NPV) was investigated in "in vitro" cell culture systems of four mammalian species: mouse, rat, monkey and man. Cells were treated with NPV, (isolated from larvae and insect cell cultures), NPV-DNA, C-type virus-activating chemicals (halogenated pyrimidine analogue) and chemical insecticides alone and in combination. C-type retroviruses were activated only in mouse cells and transformed rat cells by the halogenated pyrimidine analogue. In NPV-treated cell cultures no C-type retrovirus activation was detectable. In simultaneous treatments of the cells with nuclear polyhedrosis viruses and chemicals no potentiating effects by the NPV could be detected. Virions of NPV in cell cultures showed unaltered morphology and upon reisolation were infectious in homologous insect cell cultures for a long time. No influence upon growth or morphology of the treated mammalian cells could be observed. From our experiments we conclude that the application of nuclear polyhedrosis viruses for pest control is uncritical with respect to their retrovirus activating potential in mammalian cells.