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I Hellström

Publications and source records attributed to I Hellström.

At least 163 records · Page 9Linked to original sources
PubMed1981

Cell-mediated suppression of tumor immunity has a non-specific component. I. Evidence from transplantation tests.

We have first demonstrated that the growth of a transplanted, 3-methyl-cholanthrene-induced BALB/c sarcoma, MCA-1425, in specifically immune syngeneic mice, was facilitated when the MCA-1425 tumor cells were injected together with either a mixture of irradiated MCA-1425 cells and thymus cells from mice immune to MCA-1425 or a mixture of irradiated cells from an antigenically different BALB/c sarcoma, MCA-1460, and thymus cells from mice immune to MCA-1460. We then performed Winn assays to study the induction of an immune response that could prevent the outgrowth of MCA-1425 cells in irradiated syngeneic recipients. Tumor immunity was found to be suppressed, if the mice immunized by a subcutaneous transplant of MCA-1425 cells and used as lymphocyte donors, were injected intraperitoneally with cells from either a different sarcoma, MCA-1460, or from 12-day BALB/c embryos. We tentatively attribute our finding to antigen-non-specific suppressor effector cells and postulate that an antigen-specific event is often involved in their induction.

Animals↗
PubMed1981

Cell-mediated suppression of tumor immunity has a non-specific component. II. Evidence from cell culture experiments.

In vitro studies were performed to search for cells in the thymus of BALB/c mice with MCA-induced sarcomas which could suppress either a secondary immune response to MuLV-associated tumor antigens present on a Moloney leukemia virus-induced lymphoma, LSTRA, or a primary cytotoxic response to C57BL/6 alloantigens. They showed that thymocytes from mice with sarcomas were suppressive in both systems, and that suppression was independent of whether or not the sarcomas expressed MuLV antigen gp70. Furthermore, normal thymocytes, when combined with mitomycin-C-treated cells from any of several different MCA-induced sarcomas, suppressed the in vitro generation of a secondary cytolytic response to LSTRA. We postulate that exposure of mice to certain tumor antigens induces cells which, in the presence of the same tumor antigens which are not detectably present on the tumor cells inducing the response.

Animals↗
PubMed1981

Quantitative analysis of melanoma-associated antigen p97 in normal and neoplastic tissues.

We have used two highly sensitive assays to quantitate p97, a protein associated with human melanoma, in cultured cells and normal adult, fetal, and neoplastic tissues. To measure p97 at the surface of intact cells, radiolabeled Fab fragments of a monoclonal antibody specific for p97 were used in a binding assay. To measure p97 in detergent-solubilized membrane preparations, we used a novel double-determinant immunoassay that uses two monoclonal antibodies to two distinct antigenic determinants of p97. These assays revealed that although p97 is present in small amounts in normal adult tissues, it is present in much larger amounts in most melanomas, in some other tumors (both benign and malignant), and in certain fetal tissues. We conclude that monoclonal antibodies to p97 may prove to be of value for the diagnosis and therapy of melanoma.

Animals↗
PubMed1981

Structural characterization of human melanoma-associated antigen p97 with monoclonal antibodies.

Monoclonal antibodies were used to study p97, a human melanoma-associated antigen (MAA). Four hybridomas, designated 4.1, 96.5, 118.1, and 8.2, were obtained by fusing mouse myeloma cells with spleen cells from mice immunized with human melanoma cells. Antibodies 4.1 and 8.2 were IgG1; antibodies 96.5 and 118.1 were IgG2a. Sequential immunoprecipitation (IP) and sodium-dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) showed that all 4 antibodies recognized the same 97 kilodalton (kD) protein. Binding studies with 125I-labeled antibody showed that antibodies 4.1 and 96.5 bound the same epitope, p97a. Antibodies 118.1 and 8.2 defined epitopes p97b and p97c, respectively. Six monoclonal antibodies (M17, L1, L10, R10, I12, and K5) specific for gp95, a kD melanoma cell surface glycoprotein were also tested. Sequential IP showed that these antibodies bound p97; p97 and gp95 are thus identical. Binding studies showed that antibody m17 bound epitope p971, and antibodies L1, L10, and R19 bound epitope p97c. Antibodies I12 and K5 defined 2 other epitopes, p97d and p97e, respectively. SDS-PAGE under nonreducing conditions indicated that p97 is monomeric, probably with intrachain disulfide bonds. Cell-surface labeling of sialic acid residues and neuraminidase digestion showed that p97 is a sialoglycoprotein. Digestion of p97 with papain or trypsin produced a stable 40 kD fragment, which expressed epitopes p971, p97b, and p97c, but not p97d or p97e.

Animals↗
PubMed1981

Monoclonal antibodies to two determinants of melanoma-antigen p97 act synergistically in complement-dependent cytotoxicity.

Microcytotoxicity tests were used to study complement-dependent cytotoxicity of 2 monoclonal IgG2a antibodies, 96.5 and 118.1, both alone and in combination. These antibodies are specific for 2 antigenic determinants, p97a and p97b, respectively, of p97, a cell-surface glycoprotein present on most human melanomas and, in smaller amounts, on other tumors and normal tissues. For 10 of 12 melanoma lines, complement-dependent cytotoxicity was much greater in the presence of both antibodies than with either antibody alone. Synergism was not observed when the antibodies were tested in combination with an IgG2a antibody to a different cell-surface glycoprotein, nor was it detected in tests of 4 non-melanoma cell lines.

Antibodies↗
PubMed1981

Clonal variation in expression of a human melanoma antigen defined by a monoclonal antibody.

Previous work established 3 monoclonal mouse antibodies, 3.1, 3.2, and 3.3, which define an antigenic determinant that is expressed by a human melanoma, M1804, and, in smaller amounts, by another melanoma, M1801. In the present study we first investigated the expression of this determinant, which we refer to as 3.1, by cells from different melanomas, using membrane immunofluorescence techniques. We found that some tumors, such as M1801, appeared to be mixtures of cells that varied in their expression of 3.1. Clones were established from cultures of M1801 and M1804, and their expression of 3.1 was studied by a variety of serologic techniques, including membrane immunofluorescence, complement-dependent cytotoxicity, and absorption. Some of the clones expressed determinant 3.1, and others did not. By recloning a late passage of a 3.1-positive clone from M1801, a negative subclone was established, proving that 3.1-positive cells can yield 3.1-negative progeny. A clone that had lost 3.1 was tested and found to still express p97, an antigen defined by a different monoclonal antibody. We conclude that since some melanomas are heterogeneous with respect to the expression of antigenic determinant 3.1, and since 3.1 negative progeny can be derived from 3.1-positive cells, there is the potential for selection of 3.1-negative cells. The implications of this, for antigenic determinant 3.1 and possibly for other tumor antigens, must be taken into account when considering using monoclonal antibodies for tumor therapy.

Animals↗
PubMed1981

Use of monoclonal antibodies for quantitative analysis of antigens in normal and neoplastic tissues.

We describe two methods of analyzing tissues for cell-surface antigens identified by monoclonal antibodies. The first method is a binding assay, in which a membrane fraction prepared by differential centrifugation of a tissue homogenate is incubated with 125I-labeled antibody. The specificity of the observed binding is established by competition with unlabeled antibody. The second method, double-determinant immunoassay, involves two monoclonal antibodies that recognize two distinct epitopes of the antigen molecule. An immunoabsorbent prepared from one of the antibodies is incubated with a detergent lysate of the tissue. The immunoabsorbent is then washed and incubated with the 125I-labeled second antibody. In both assays the amount of radiolabeled antibody bound is proportional to the amount of antigen in the test sample, and the limit of detection is approximately 10 pg of antibody, corresponding to less than 1 fmol of antigen.

Antibodies↗
PubMed1980

Protein antigens of normal and malignant human cells identified by immunoprecipitation with monoclonal antibodies.

Spleen cells from a mouse immunized with human melanoma cells were fused with mouse myeloma cells, and somatic cell hybrids were grown in selective medium. Eight hybrids, which secreted antibodies to protein antigens of the melanoma cell line, were identified by immunoprecipitation of a 125I-labeled melanoma cell lysate followed by sodium dodecyl sulfate-gel electrophoresis of the immunoprecipitates and autoradiography. Seven of the eight melanoma proteins identified in this way were present at the cell surface. Two of the cell surface proteins, p80 and p97, were not detected in autologous fibroblasts.

Animals↗
PubMed1980

Cells from chemically induced mouse sarcomas retain tumor-specific transplantation antigenicity during prolonged passage in vitro.

Tumor-specific transplantation antigens (TSTA) expressed by cells from transplantable lines of 3-methylcholanthrene-induced BALB/c sarcomas were maintained over a 26-week culture period, which represented 2-32 passages in vitro. Cells from in vitro-passaged sarcomas were immunogenic in that they induced specific resistance in syngeneic mice to challenge with cells from the same sarcoma. They were also immunosensitive in that their outgrowth in syngeneic mice was prevented when they were mixed with lymph-node cells from mice immunized with in vivo-passaged cells from the respective sarcoma.

Animals↗
PubMed1980

Identification of a cell surface protein, p97, in human melanomas and certain other neoplasms.

BALB/c mice were immunized with a human melanoma cell line, SK-MEL 28, and their spleen cells were fused with mouse NS-1 myeloma cells. Hybrid cells were tested in an indirect 125I-labeled protein A assay for production of antibodies that bound to surface antigens of SK-MEL 28 melanoma cells but not to autologous skin fibroblasts. One hybridoma, designated 4.1, had the required specificity. It was cloned and grown in mice as an ascites tumor. The monoclonal IgG1 antibody produced by the hybridoma was purified from the ascites fluid and labeled with 125I. The labeled antibody bound, at significant levels, to approximately 90% of the melanomas tested and to approximately 55% of other tumor cells, but not to three B-lymphoblastoid cell lines or to cultivated fibroblasts from 15 donors. Immunoprecipitation and sodium dodecyl sulfate gel electrophoresis were used to detect the target antigen in 125I-labeled cell membranes of both cultivated cells and tumor biopsy samples. A protein with a molecular weight of 97,000 was identified. This protein, designated p97, was present in both cultured cells and biopsy material from melanomas and certain other tumors, but it was not detected in eight different samples of normal adult epithelial or mesenchymal tissues obtained from five donors.

Antibodies, Neoplasm↗
PubMed1980

T-T hybridoma product specifically suppresses tumor immunity.

Culture supernatants of spleen or thymus cells from BALB/c mice bearing transplanted, syngeneic, methylcholanthrene-induced sarcomas suppress T lymphocyte-mediated lysis of cells from the tumor borne by the donor of the spleen or thymus cells. On this basis, we hybridized thymus cells from mice bearing sarcoma MCA-1490 with cells from the T lymphoma BW5147. The hybrids (hybridomas) formed were tested for production of factors that could suppress T lymphocyte-mediated lysis of MCA-1490 cells. One hybridoma, and a clone derived from it, produced factors that suppressed the lysis of MCA-1490 cells in vitro. In addition, these factors enhanced the growth of MCA-1490 in immune mice and prevented the destruction of MCA-1490 cells by immune lymphocytes in tumor neutralization (Winn) assays. In vitro lysis of cells from another MCA-induced sarcoma by immune lymphocytes was not suppressed. The suppressor factors did not affect the proliferative response of BALB/c lymphocytes to mitogens or the generation of a cytotoxic response to C57BL/6 alloantigens. Neither did they inhibit the generation of primary or secondary cytotoxic responses to murine leukemia virus-related antigens present on a BALB/c lymphoma line, LSTRA. Although our findings suggest that these suppressor factors are specific for MCA-1490, their specificity for antigens restricted to this tumor needs further definition.

Animals↗
PubMed1980

Antibodies to murine leukemia virus gp70 and p15(E) in sera of BALB/c mice immunized with syngeneic chemically induced sarcomas.

The sera of BALB/c mice immunized with syngeneic methylcholanthrene-induced sarcomas commonly contain antibodies that react with the immunizing tumor line and also with other chemically induced sarcomas. Three lines of evidence suggest that the cross-reactive antibodies are directed against antigenic determinants of gp70 and p15(E), envelope proteins of endogenous murine leukemia virus (MuLV). 1) The antibodies bind only to sarcomas expressing MuLV antigens, 2) they are removed by absorption with purified MuLV antigens, and 3) they precipitate proteins identified as gp70 and p15(E) from 125I-labeled sarcoma cell lysates.

Absorption↗
PubMed1979

Cell-mediated reactivity to antigens shared by Moloney-virus-induced lymphomas (LSTRA) and certain 3-methylcholanthrene-induced mouse sarcomas.

Spleen cells (SC) both from BALB/c mice whose primary Moloney sarcoma virus (MSV)-induced sarcomas had spontaneously regressed and from normal, untreated BALB/c mice, were co-cultivated for 5 days with mitomycin-C-treated LSTRA cells; LSTRA is a BALB/c Moloney lymphoma which shares cell surface antigens with MSV-indiced sarcomas. These SC, referred to as CMR and CU cells, respectively, were shown to be cytotoxic to LSTRA cells in 3 h 51Cr-release assays; CMR cells showed, in most cases, the greatest lytic activity against LSTRA targets. The same SC were also reactive, in 20-h microcytotoxicity and 51Crassays, against target cells from a variety of transplanted sarcomas indiced by 3-methylcholanthrene (MCA) in Balb/c mice. The highest reactivity was seen when CMR or CU cells were tested against target cells from sarcoma lines that expressed an NB-ecotropic MuLV cross-reacting serologically with Moloney virus. Reactivity against isotope-labelled tumor cells expressing MuLV-associated cell surface antigens could be competititively inhibited by adding unlabelled tumor cells expressing such antigens. Finally, Winn assays were performed in which CMR cells strongly inhibited the outgrowth of cells from three sarcoma lines that express the NB-ecotropic MuLV. There was less but significant inhibition of cells from some other MCA sarcomas, either negative for the expression of MuLV-associated antigens or expressing the N-ecotropic endogenous BALB/c MuLV. CU cells enhanced tumor outgrowth in Winn assays at least as often as they inhibited it.

Animals↗
PubMed1979

Enhancement of tumor outgrowth by tumor-associated blocking factors.

Transplanted lines of 3-methylcholanthrene (MCA)-induced BALB/c sarcomas were used for experiments in which tumors were mechanically suspended in Waymouth's culture medium, the suspensions centrifuged and filtered and the remaining cell-free "tumor fluids" (TF) tested for activity in vitro and in vivo. TF were found to inhibit ("block") cell-mediated cytotoxicity to tumor target cells in vitro with specificity for each tumor tested and even at high dilutions (1:1,000-1:10,000). In vivo, TF enhances the outgrowth of subcutaneously transplanted tumors in non-immune syngeneic mice. This enhancement had both specific and non-specific components, with the non-specific component being most apparent. Enhancement was best demonstrated when the TF were inoculated together with the tumor inocula. In one of two experiments in which TF were given intraperitoneally, specific enhancement was seen, whereas in the other, inhibition of tumor outgrowth was observed.

Animals↗
PubMed1979

Indirect 125I-labeled protein A assay for monoclonal antibodies to cell surface antigens.

An assay for detection of monoclonal hybridoma antibodies against cell surface antigens is described. Samples of spent medium from the hybridoma cultures are incubated in microtest wells with cells, either as adherent monolayers or in suspension. Antibodies bound to surface antigens are detected by successive incubations with rabbit anti-immunoglobulin serum and 125I-labeled protein A from Staphylococcus aureus, followed by autoradiography of the microtest plate or scintillation counting of the individual wells. Particular advantages of this assay for screening hybridomas are: (1) commerically available reagents are used, (2) antibodies of any species and of any immunoglobulin class or subclass can be detected, and (3) large numbers of samples can be screened rapidly and inexpensively. We have used the assay to select hybridomas producing monoclonal antibodies to surface antigens of human melanomas and mouse sarcomas.

Animals↗