Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0025202 (melanoma)
 69,561 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have previously reported the isolation of a 66 kDa melanoma-associated antigen, identified by autologous antibody, in serum and unfractionated spent tissue culture media by Western blot analysis. The antigen, detected by autologous serum S150, was found to be broadly represented on melanoma, glioma, renal cell carcinoma, neuroblastoma and head and neck carcinoma cell lines. S150 did not react with bladder or colon carcinoma, fetal fibroblasts, pooled platelets, lymphocytes and red blood cells, autologous cultured lymphocytes or fetal calf serum. To further characterize the antigen, spent tissue culture media, obtained from autologous melanoma cell line, Y-Mel 84:420, was separated by an isoelectric focusing column. Unabsorbed control serum S150 was noted to have a maximum titer of 1:2040 against autologous melanoma cells as measured by protein A hemadsorption. Following isoelectric focusing the greatest decrease in autologous antibody titer (30-fold) occurred with fractions having a pI between 2 and 3. Further resolution of the antigen was accomplished with high-pressure ion-exchange chromatography. One of these fractions showed a significantly higher concentration of antigen and was distinctly resolved from bulk serum albumin. Subsequent Western blot analysis, with autologous antibody, of the isolated antigen-containing fraction, confirmed the presence of a single 66 kDa band. Exposure of the antigen, purified by high-pressure ion-exchange chromatography, to neuraminidase ablated recognition by autologous antibody and suggests that sialic acid is present on the protein and may be part of the antigenic epitope. Binding of antigen, obtained following DEAE anion exchange chromatography, was noted to lectins derived from Triticum vulgaris, Dolichos biflorus and Lycopersicon esculentum. Preparative purification of the antigen was accomplished by anion exchange followed by lectin affinity chromatography with a Dolichos biflorus column. Antigen obtained following lectin affinity chromatography subjected to SDS-PAGE and silver stain revealed a single band at 66 kDa. We conclude that a melanoma-associated antigen detected by autologous antibody in spent tissue culture media is an unusually acidic glycoprotein (pI 2-3).
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PMID:Purification and partial characterization of a shed 66 kDa melanoma-associated antigen identified by autologous antibody. 193 77

Brefeldin A has dramatic, well-documented, effects on the structural and functional organization of the Golgi complex. We have examined the effects of brefeldin A (BFA) on the Golgi-localized synthesis and addition of chondroitin sulfate glycosaminoglycan carbohydrate side chains. BFA caused a dose-dependent inhibition of chondroitin sulfate glycosaminoglycan elongation and sulfation onto the core proteins of the melanoma-associated proteoglycan and the major histocompatibility complex class II-associated invariant chain. In the presence of BFA, the melanoma proteoglycan core protein was retained in the ER but still acquired complex, sialylated, N-linked oligosaccharides, as measured by digestion with endoglycosidase H and neuraminidase. The initiation of glycosaminoglycan synthesis was not affected by BFA, as shown by the incorporation of [6-3H]galactose into a protein-carbohydrate linkage region that was sensitive to beta-elimination. The ability of cells to use an exogenous acceptor, p-nitrophenyl-beta-D-xyloside, to elongate and sulfate core protein-free glycosaminoglycans, was completely inhibited by BFA. The effects of BFA were completely reversible in the absence of new protein synthesis. These experiments indicate that BFA effectively uncouples chondroitin sulfate glycosaminoglycan synthesis by segregating initiation reactions from elongation and sulfation events. Our findings support the proposal that glycosaminoglycan elongation and sulfation reactions are associated with the trans-Golgi network, a BFA-resistant, Golgi subcompartment.
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PMID:Uncoupling of chondroitin sulfate glycosaminoglycan synthesis by brefeldin A. 195 86

GM2 ganglioside is a common cell surface constituent of human melanoma and other tumors of neuroectodermal origin, and vaccination with GM2 ganglioside results in high levels of anti-GM2 antibodies in patients with melanoma. Lymphocytes from a GM2-vaccinated patient (VS) were transformed by Epstein-Barr virus and tested for production of antibodies with reactivity for GM2-positive tumor cells. A high percentage of antibody-producing B cells was detected, but antibody reactivity was generally lost during culture expansion. Two cultures, however, remained stable for antibody productivity and one was used to develop a stable hybrid line with mouse myeloma. The monoclonal antibody (designated 3-207) derived from patient VS has dual specificity for GM2 and GD2, despite the fact that only GM2 antibody could be detected in the patient's serum. Monoclonal antibody 3-207 shows high-titered reactivity with a range of melanoma, astrocytoma, neuroblastoma, and leukemia cell lines, cells with prominent cell surface expression of GM2 and GD2. The cell surface reactivity of monoclonal antibody 3-207 was not abolished by treatment of target cells with neuraminidase, as the enzyme converted GD2 to GM2, which was still detected by monoclonal antibody 3-207.
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PMID:Human monoclonal antibody with dual GM2/GD2 specificity derived from an immunized melanoma patient. 215 45

Spleen cells from inbred Biozzi mice, immunized against the human breast cancer cell line T47D, were fused with murine myeloma SP2O cells to generate monoclonal antibodies. One of these, 1BE12, of IgM isotype, reacted with five of six human breast tumor cell lines, while no binding was detectable with normal lymphocytes, RBC, or fibroblasts. The antigen recognized by monoclonal antibody 1BE12 was localized on the surface of T47D and MCF7 cells and was detected in cell-free supernatants of cultures. The antigen was found also on the surface of milk secretory cells. Immunohistochemical staining of frozen and paraffin-embedded sections of human tissues showed apical polarized reactivity in normal breast glands, while in all breast cancers staining was either cytoplasmic or membranous and heterogeneously distributed. Immunostaining was also observed in some other normal epithelia, including salivary gland, gastroduodenal mucosa, exocrine pancreas, and cervix. The antigen was not detectable in secretory endometrium, whereas proliferative endometrium was strongly stained. Colon carcinoma, and cancers of the bladder and endometrium were strongly reactive. No staining was detected in melanoma, lymphoma, mesothelioma, non-small cell lung carcinoma, and thyroid, renal, and ovarian carcinomas. Lectin absorption of MCF7 membrane extracts reduced 1BE12 binding. A large reduction in 1BE12 reactivity was observed after digestion of T47D and MCF7 membrane extracts with proteases. Treatment with sodium periodate resulted in complete loss of antigenicity, while neuraminidase treatment did not affect 1BE12 binding. These findings suggest that the 1BE12 epitope is expressed on the carbohydrate moiety of a glycoprotein and does not contain sialic acid. Immunoblotting of the perchloric acid-soluble fraction of MCF7 membrane extracts after electrophoresis in 1% agarose detected the antigen as a high molecular weight species (Mr greater than 900,000). The antigen was purified by perchloric acid extraction of MCF7 membrane preparations followed by affinity chromatography on 1BE12 antibody coupled to Sepharose-4B and gel exclusion fast protein liquid chromatography. No reactivity of the purified material was found with monoclonal antibodies directed against human milk fat globule membrane-associated mucins HMFG1 and DF3.
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PMID:Characterization and distribution in human tissues of a glycoproteic antigen defined by monoclonal antibody 1BE12 raised against the human breast cancer cell line T47D. 222 61

Two autologous human melanoma cell lines were studied to determine their capacities to bind wheat germ agglutinin (WGA). Both cell lines were derived from the same patient, the first, IGR 39, originated from the primary tumor, the second, IGR 37, was established from a metastatic lymph node. WGA binding sites on the surface of these cell lines were compared before and after sialidase and/or tunicamycin treatments. IGR 39 cells exhibited two classes of WGA binding sites with high and low affinities, whereas IGR 37 cells had only one class of high affinity binding sites. After tunicamycin treatment, the capacity of IGR 39 cells to bind WGA was markedly altered, since only one class of WGA binding sites with high affinity was observed under these conditions, whereas tunicamycin did not induce significant changes in the lectin binding of IGR 37 cells. The low affinity WGA binding sites, which were only found on IGR 39 cells, corresponded to sialyl residues present in N-linked glycoproteins. The high affinity binding sites present on both cell lines probably involved sialyl and N-acetyl-glucosaminyl residues associated with O-linked glycoproteins and/or glycolipids. No direct correlation could be drawn between the number of WGA binding sites and the overall sialic acid levels exposed to sialidase treatment. The 3-fold increase in the amount of cell surface glycopeptides obtained after pronase digestion and specifically binding to WGA-Sepharose was in good agreement with the overall higher number of WGA binding sites on IGR 39 compared to IGR 37 cells. Thus, subtle carbohydrate changes of cell surface glycoconjugates might account for the differences between the biological properties of human melanoma cell lines of low and high tumorigenicity.
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PMID:WGA binding to the surface of two autologous human melanoma cell lines: different expression of sialyl and N-acetylglucosaminyl residues. 234 28

Neuroglandular antigen (NGA) was identified as a human melanoma-associated antigen by a panel of murine monoclonal antibodies of both IgG2a (LS62, LS76, LS159) and IgG1 (LS113, LS140, LS152) subclasses, developed in this laboratory (L. Sikora, A. Pinto, D. Demetrick, W. Dixon, S. Urbanski, and L. M. Jerry, Int. J. Cancer, 39: 138-145, 1987). Monoclonal antibody LS62 was used to immunoprecipitate NGA from radiolabeled cultured melanoma cells, and it behaved as a heterogeneous glycoprotein "smear" on sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis (Mr 29,000-70,000). Radioactive pulse-chase time course experiments using human melanoma cells cultured in the presence or absence of inhibitors of protein glycosylation showed that the antigen consisted of a core protein with a molecular weight of 22,000 on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. This molecule was modified by the addition of at least three N-linked oligosaccharide side chains (as revealed by limited N-glycanase digestion) to give a precursor form with a molecular weight of approximately 34,000. Subsequent processing steps yielded a heterogeneous family of glycoproteins with varying amounts of covalently attached carbohydrate. Much of this heterogeneity in both molecular weight and pI (as revealed by two-dimensional electrophoresis) could be removed by treatment of the antigen with neuraminidase, suggesting heavy sialylation of the glycoprotein. NGA could be detected on the surface of melanoma cells by fluorescence-activated cell sorter analysis, surface radioiodination, and, as previously shown, immunoperoxidase staining. However, there was a larger intracellular pool of the molecule and the antigen was rapidly released into the culture supernatant. The function of NGA remains unknown but its elevated expression in transformed melanocytes have prompted this characterization to understand its biochemical nature and relation to other melanoma-associated antigens.
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PMID:Biosynthesis, glycosylation and intracellular processing of the neuroglandular antigen, a human melanoma-associated antigen. 236 31

Human anomalous killer (AK) cells lyse freshly isolated human melanoma cells which are insensitive to human natural killer cell-mediated lysis. Monoclonal antibody Leo Mel 3, an IgM (k), produced by a hybridoma obtained from a mouse immunized with human melanoma cells, binds to melanoma cells and inhibits their conjugate formation with AK cells as well as their AK cell-mediated lysis. Other IgM antibodies from the same fusion that bind melanoma cells do not inhibit (Werkmeister, J. A., Triglia, T., Andrews, P., and Burns, G. F. (1985) J. Immunol. 135, 689-695). Leo Mel 3 binds several different gangliosides from melanoma cells, as determined by immunostaining thin layer chromatograms. Binding is abolished by treatment of the gangliosides with neuraminidase. In solid-phase radioimmunoassay, Leo Mel 3 binds strongly to ganglioside GD2 and less strongly to gangliosides GT3, GD3, and GQ1b. It does not bind to other gangliosides including GM1, GM2, GM3, GD1a, GD1b, and GT1b. Thus, the epitope recognized by antibody Leo Mel 3 is found in the sugar sequence of ganglioside GD2, GalNAc beta 1-4[NeuAc alpha 2-8NeuAc alpha 2-3]Gal beta 1-4Glc beta 1 .... This sequence may contain a target in melanoma cells recognized by AK cells.
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PMID:Monoclonal antibody Leo Mel 3, which inhibits killing of human melanoma cells by anomalous killer cells, binds to a sugar sequence in GD2 (II3(NeuAc)2-GgOse3Cer) and several other gangliosides. 243 16

The binding sites for human interferon-alpha (IFN-alpha) have been characterized on human lymphoblastoid, melanoma, rhabdomyosarcoma, and cervical carcinoma cells. Crosslinking of iodinated-recombinant DNA-derived IFN-alpha-Con1, an analog of the known IFN-alpha subtypes, to the cell surface with disuccinimidyl suberate yielded four IFN-receptor complexes of 118, 138, 159, and 260 kD on all cell lines that specifically bind IFN-alpha. Since IFN-alpha exists in solution as monomers, dimers, and trimers, and the three lower molecular weight IFN-alpha-receptor complexes differ by the molecular weight of IFN-alpha (20 kD), this suggests that the human IFN-alpha receptor of 100 kD binds more than one molecule of IFN-alpha. The higher molecular weight complex of 260 kD may result from dimerization of the receptor. None of these complexes was observed in a rhabdomyosarcoma subclone that does not specifically bind IFN-alpha. Pretreatment of cells with trypsin abolished the formation of these complexes. Pretreatment of cells with neuraminidase did not reduce IFN-alpha binding, but increased the electrophoretic mobility of all four IFN-alpha-receptor complexes. Other glycosidases (i.e., mannosidase, beta-galactosidase, and endoglycosidase F) had no effects on IFN-alpha binding or mobility of complexes. Thus, although the IFN-alpha receptor is a glycoprotein, the glycosylated portion is apparently not part of the IFN-alpha-binding domain. The formation of IFN-alpha-receptor complexes is independent of the duration of incubation with IFN (from 5 min to 1 h at 15 degrees C).(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Characterization of interferon-alpha binding sites on human cell lines. 246 92

GD3 is a major ganglioside of human melanoma and was shown to be an effective target for passive immunotherapy with murine monoclonal antibodies. It was noted earlier that GD3 neither purified nor in melanoma cell vaccine (MCV), could elicit an antibody response in melanoma patients. In this study, we demonstrate that melanoma patients who received MCV had autoantibodies against a derivative of GD3, O-acetylated GD3 (O-AcGD3), a minor ganglioside expressed on human melanoma cells, and that the antibodies cross-reacted with GD3. Thin layer chromatographic immunostaining revealed that all of the sera containing antibodies against O-AcGD3 also reacted to GD3. None of the other sera responded only to GD3, although the MCV contained 7- to 12-fold higher GD3 than O-AcGD3. Furthermore, the antibody activity was completely abolished by absorption with animal erythrocytes expressing either O-acetyl disialogangliosides or GD3, indicating that the antibodies recognize an epitope commonly shared by GD3 and O-AcGD3. The antibodies bound only to the sialyloligosaccharide moiety but not to the ceramide portion of GD3 after endoglycosylceramidase treatment. The antibodies failed to bind to GD3 after neuraminidase treatment. These results indicate that the sialyloligosaccharides of the gangliosides are important components of the epitope. Periodate oxidation abolished reactivity of the antibodies to GD3 but not that to O-AcGD3, revealing that the glycerol side chain of the sialic acids in both GD3s was an important structure of the epitope. The binding of the antibodies to melanoma cell surface gangliosides was confirmed by an absorption with a GD3- and O-AcGD3-positive melanoma cell line. These results in the light of previous reports on the inability of GD3 to elicit immune response in humans suggest that anti-GD3 antibodies found in the melanoma patients were induced by immunization with O-AcGD3 and O-AcGD3 present in the MCV would serve as an antigen source for GD3-targeted active specific immunotherapy of melanoma.
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PMID:An epitope common to gangliosides O-acetyl-GD3 and GD3 recognized by antibodies in melanoma patients after active specific immunotherapy. 247 99

The melanotropin (MSH) receptor of mouse B16-F1 melanoma cells was characterized by photoaffinity cross-linking, using a potent alpha-MSH photolabel, [norleucine4, D-phenylalanine7, 1'-(2-nitro-4-azidophenylsulfenyl)-tryptophan9]-alpha-melanotropin (Naps-MSH). Its monoiodinated form, 125I-Naps-MSH, displayed a approximately 6.5-fold higher biological activity than alpha-MSH. Scatchard analysis of the saturation curves with 125I-Naps-MSH revealed approximately 20,000 receptors/B16-F1 cell and an apparent KD of approximately 0.3 nM. Analysis of the cross-linked MSH receptor by sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that a photolabeled band of approximately 45 kDa occurs in B16-F1, B16-F10, and Cloudman S91 mouse melanoma, as well as in human D10 and 205 melanoma but not in non-melanoma cells. The labeled 45-kDa protein had an isoelectric point of 4.5-4.9 as determined by two-dimensional gel electrophoresis. Treatment of the labeled 45-kDa protein of B16-F1 cell membranes by neuraminidase shifted the band to approximately 42 kDa. A similar band of about 42 kDa was also observed after receptor labeling of B16-W4 cells, a cell line with a decreased number of terminal N-linked neuraminyl residues. These results indicate that the labeled 45-kDa glycoprotein contains terminal sialic acid residues, explaining the low pI of this protein, and that it is characteristic for melanoma cells and hence part of the MSH receptor.
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PMID:The receptor for alpha-melanotropin of mouse and human melanoma cells. Application of a potent alpha-melanotropin photoaffinity label. 254 92


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