Cross-Reactivity of Murine Anti-Human High Molecular Weight-Melanoma Associated Antigen Monoclonal Antibodies with Guinea Pig Melanoma Cells1 (original) (raw)
Related papers
Molecular Immunology, 1983
Normal human sera were analyzed for the presence and molecular form of two human melanoma-associated antigens (MAAs), the 250 "melanoma-specific" glycoprotein and the 100 K "common tumor antigen". The 250K MAA was not synthesized by any cultures other than human melanoma and was not detectable in normal human serum. In contrast, the 100 K MAA, which is present in spent medium of cultured human melanoma, carcinoma and fetal melanocytes but not of adult normal cells, was found in normal human serum in nanogram quantities. This serum form of the 100 K MAA was also found in pooled sera of higher apes but not of lower species. The cell-derived form of the 100 K MAA, present in spent culture medium, had a similar phylogenetic distribution. The molecule was produced by cultured brain glia from gorilla, but not by melanoma cells from miniature swine or dog. The 100 K MAA from gorilla glia had a mol. wt identical to the molecule produced by human melanoma cells. Molecular characterization of this MAA in normal human serum showed that it was heterogeneous in size and was present in fractions > 100 kd after analytical HPLC gel sieving under non-denaturing conditions. In contrast, MAA from spent culture medium of melanoma cells was 100 kd or less in chemically defined medium (CDM) with no protein supplement, but had a higher mol. wt in CDM with BSA or fetal calf serum supplement, simila; to the se&m form of the mole&le. An association of the 100 K MAA with albumin was demonstrated bv analvtical HPLC gel sieving and SDS-PAGE analvsis of monoclonal antibody immunoprecipitates. Tie 100-K MAA wasldissociated from albumin in nor&al human serum by treatment with SDS and fractionation by gel sieving. Under these conditions 100 K MAA from serum co-migrated with similarly treated 100 K from melanoma cells. These results indicate that the 100 K MAA is a normal serum constituent which forms a strong, non-covalent association with albumin and is evolutionarily restricted to higher apes or humans.
Cancer research, 1979
To characterize the antigen present on the surface of cultured human malignant melanoma, three monkey xeno geneic antisera were raised. After appropriate absorption with pooled human erythrocytes, peripheral blood leuko cytes, and liven homogenate, no blood group or HLA neac tivity was detectable. Analysis of melanoma antigenic spec ificity was performed by the mixed hemadsonption microas say on live monolayer cells in conjunction with quantitative microabsorption analysis. To eliminate reactivity against nonmelanoma lines, 2 of the 3 antisera required further absorption with cells of the KB oral carcinoma line. The absorbed antisera reacted with 9 of 10 long-term estab lished lines, 3 of 3 short-term cultures of human malignant melanoma, and i of 10 nonmelanoma epithelial and fibro blastic cell lines. Absorption experiments using a variety of cultured cells and fresh tissue homogenates of adult human melanoma and nonmelanoma sources further substantiated the results obtained with the direct tests. While each mela noma line showed a differing degree and pattern of neactiv ity, the antisera were most reactive against their respective immunizing lines as revealed by both the direct tests and absorption analysis. By quantitative absorption analysis, no evidence for individually specific melanoma antigens was obtained. The only positive absorption with nonmelanoma adult tissues was obtained with a retinoblastoma cell line, indicating the presence of antigens shared by tumors of common neunoectodermal origin. Extensive absorption with two xenogeneic malignant melanoma (munine and porcine) homogenates, normal human adult skin, and spleen tissues, or Bacillus Calmette-Guéninfailed to reduce the reactivity against melanoma-associated antigens. Fur then absorption with human fetal tissues of 8 to 20 weeks of gestation removed part but not all of the reactivity. These studies with xenogeneic monkey antisera provide evidence for the existence of common melanoma-associated anti gens as well as distinct but shared human fetal antigens on human melanoma cells. , This is Paper 8 of the series, â€oeCharacterization of Human Malignant MelanomaCell Lines.― The work was supportedby the Medical Research Council of Canada and the Ontario Cancer Treatment and Research Foun dation.
Human melanoma-specific oncofetal antigen defined by a mouse monoclonal …
… Journal of Cancer
140.240, an lgC2a mouse monoclonal antibody raised against a cultured human melanoma cell line, was highly specific for melanoma cells as determined by direct and absorption analyses in a mixed hemadsorption assay. Supematants of doubly cloned hybridomas producing antibody 140.240 reacted with all cultured and fresh melanomas tested but failed to react with a variety of carcinomas, sarcomas, lymphomas, leukemias and other tumors of neuroectoderrnal origin. This antibody did not react with B-lymphoid cell lines, ruling out HLA-DR specificity. Non-reactivity of antibody 140.240 with peripheral blood lymphocytes obtained from the donor of the immunizing melanoma line excluded the possibility of detecting histocompatibility antigens. Nevus cells were also non-reactive. However, antibody 140.240 did identify an antigenic determinant on tissue homogenates prepared from fetuses of 10-14 weeks' gestation. The antigen involved was shed by cultured melanoma lines and, by immunoprecipitation analysis of radiolabelled lysates, had a molecular weight of approximately 87kdal. Thus, the structure identified by monoclonal antibody 140.240 is a melanoma-specific oncofetal antigen.
Common human melanoma-associated antigen(s) detected by monoclonal antibodies
Cancer research, 1980
Hybridoma cells have been derived from a fusion between mouse myeloma cells (P3-NSI/1Ag4) and spleen cells from a mouse immunized with membrane-enriched fractions from the human melanoma cell line Me-43. Of the 26 hybrids obtained, seven secreted antibodies which reacted with the melanoma cell line used for immunoassay. The specificity of the antibodies produced by the seven positive hybrids was further investigated on 16 melanoma cell lines, 15 other tumors, and 14 lymphoblastoid cell lines. The antibodies from four positive hybrids showed a broad reactivity, whereas those from three hybrids reacted exclusively with melanoma cells. The antibodies from two of these three hybrids, alpha-Mel/5 and alpha-Mel/14, seem to be directed against common melanoma antigen(s) since they reacted with all (with one exception) of the 16 melanoma cell lines tested only with five of the 16 melanoma lines. Reciprocal binding inhibition tests using [3H]leeucine-labeled antibodies showed that alpha-Mel/...
Cell surface antigens of human melanoma identified by monoclonal antibody
Proceedings of the National Academy of Sciences, 1979
Mouse NS-1 myeloma cells were fused with spleen cells from mice that had been immunized with cells from a human melanoma, M1804. Hybrid cells were grown in selective medium and tested for production of antibody to surface antigens of M1804 cells. Three hybrids that produced antibodies that bound to the melanoma cells but not to autologous skin fibroblasts were cloned. Antibodies produced by two of the clones were cytotoxic to M1804 cells in the presence of rabbit complement. Extensive specificity tests showed that the antibodies produced by the clones bound strongly only to M1804 cells; significant, although weaker, binding occurred with 2 of 11 allogeneic melanomas. Apart from weak binding of the antibody produced by one of the clones to a breast carcinoma, binding assays of five carcinomas, one sarcoma, and fibroblasts from 17 individuals were negative, as were cytotoxic tests of 10
British Journal of Cancer, 1993
The melanosome is a secretory organelle unique to the melanocyte and its neoplastic counterpart, malignant melanoma. The synthesis and assembly of these intracytoplasmic organelles is not yet fully understood. We have developed a murine monoclonal antibody (MoAb) against melanosomes isolated from human melanocytes (newborn foreskin) cultured in the presence of 12-0 tetradecanoyl phorbol-13-acetate (TPA). This MoAb, designated HMSA-5 (Human Melanosome-Specific Antigen-5) (IgG1), recognised a cytoplasmic antigen in both normal human melanocytes and neoplastic cells, such as common and dysplastic melanocytic nevi, and malignant melanoma. None of the carcinoma or sarcoma specimens tested showed positive reactivity with MoAb HMSA-5. Under immunoelectron microscopy, immuno-gold deposition was seen on microvesicles associated with melanosomes, and a portion of the ER-Golgi complexes. Radioimmunoprecipitation analysis showed that the HMSA-5 reactive antigen was a glycoprotein of M, 69 to 73 kDa. A pulse-chase time course study showed that the amount of antigen detected by MoAb HMSA-5 decreased over a 24 h period without significant expression on the cell surface, or corresponding appearance of the antigen in the culture supernatant. This glycoprotein appears to play a role in the early stages of melanosomal development, and the HMSA-5 reactive epitope may be lost during subsequent maturation processes. Importantly, HMSA-5 can be identified in all forms of human melanocytes, hence it can be considered a new common melanocytic marker even on routine paraffin sections. dose of geneticin, 100ligml-' was added to the medium (usually at 3-4 weeks).
Cancer research, 1985
Mouse monoclonal antibodies (mAb) detecting 13 distinct systems of surface antigens on cultured melanocytes and melanomas were tested for reactivity with panels of (a) normal and malignant cultured cells; (b) normal adult and fetal tissues; and (c) specimens of metastatic melanoma and other tumor types. The objectives of this study were to compare antigen expression in cultured versus noncultured cells, to develop a panel of mAbs that identify subsets of melanomas, and to provide requisite information about antibody specificity in preparation for the use of antibodies in the diagnosis, imaging, and therapy of melanoma. Five of the melanoma surface antigens have been well characterized biochemically [GD3, chondroitin sulfate proteoglycan, HLA Class II antigens, glycoprotein of molecular weight 130,000 (gp130), and glycoprotein Mr 95,000/protein Mr 97,000 (gp95/p97)]. Three antigens have been related to melanocyte differentiation (HLA Class II, M111/M231, and M144), and six provide ad...
Cancer research, 1988
We have characterized properties of a melanoma antigen with a mouse-specific melanoma epitope expressed on B16 melanoma by using syngeneic monoclonal antibodies with antimetastatic ability. The molecule recognized by the antibody is a membrane glycoprotein with a molecular weight of 80,000. Studies on tunicamycin treatment indicated that the core size of the molecule appeared to have a molecular weight of 69,000 and also suggested that the carbohydrate moiety was greatly responsible for the conformation of the mouse melanoma epitope. The antigen was released or shed into the culture medium from the cell surface, and the turnover rate of the antigen was within 1.5 h.
Analysis of two human monoclonal antibodies against melanoma
International Journal of Cancer, 1992
B cells derived from peripheral-blood lymphocytes (PBL) and tumor-infiltrating lymphocytes (TlL) from a patient with a high serum antibody titer to autologous melanoma were transformed with Epstein-Barr virus (EBV) and evaluated for reactivity against autologous tumor. B cells producing antibody reactive with autologous tumor and unreactive with normal fibroblasts were detected both in TIL and in PBL. One cell line derived from PBL and another derived from TIL sustained production of tumor-reactive antibody for I 0 weeks and over I 5 months respectively. The cell line derived from PBL, 2DI I, produced an antibody reactive with a tryprin-resistant antigen expressed on the cell membrane of autologous and allogeneic melanoma cell lines. The cell line derived from TIL, IF6, produced an antibody reactive with a cell-surface glycoprotein expressed by 5 autologous melanoma cell lines derived from 5 different metastases and I6/ I 9 allogeneic melanoma cell lines. IF6 also showed reactivity with cell lines derived from a blue nevus, a congenital nevus, an astrocytoma, and I /4 renal-cell carcinomas; but it was not reactive with 5 foreskin melanocyte cell lines, 2 normal fibroblast lines, 5 leukemia/lymphoma lines, 8 lung-cancer lines, 8 glioblastoma lines, or lines derived from I ovarian carcinoma, I colon carcinoma, I vulvar carcinoma, I fibrosarcoma, I murine melanoma, or 4 murine leukemia/ lymphomas. We describe here an antibody that detects a new melanoma specificity obtained by EBV transformation of tumorinfiltrating B cells.