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R A Reisfeld

Publications and source records attributed to R A Reisfeld.

At least 145 records · Page 8Linked to original sources
PubMed1986

Lymphokine-activated killer cells targeted by monoclonal antibodies to the disialogangliosides GD2 and GD3 specifically lyse human tumor cells of neuroectodermal origin.

Monoclonal antibodies 14.18 (IgG3) and 11C64 (IgG3) directed against disialogangliosides GD2 and GD3, respectively, when used in conjunction with human peripheral blood mononuclear cells (PBMCs) stimulated with human recombinant interleukin (rIL-2) lyse both human melanoma and neuroblastoma cells by antibody-dependent cellular cytotoxicity. Such monoclonal antibody-"armed" effector cells are specifically directed to targets expressing the given disialoganglioside without detectable cross-reactivity. In addition, antibody-dependent cellular cytotoxicity as well as the natural killing ability of human PBMCs is augmented by a brief coincubation with rIL-2. PBMCs augmented by rIL-2 and armed with monoclonal antibodies significantly suppressed tumor growth in the xenotransplant nude mouse model. Our results suggest that once a threshold level of activation of PBMCs is achieved, additional rIL-2 (over three orders of magnitude of concentration) does not significantly enhance cytolytic augmentation. Furthermore, anti-GD3 monoclonal antibody 11C64 together with rIL-2-stimulated PBMCs from melanoma patients with widely differing tumor burdens effectively lyse melanoma tumor targets in antibody-dependent cellular cytotoxicity. Our results also suggest that GD2 and GD3 represent distinct and relevant immunotherapeutic target structures on melanoma whereas GD2 does the same for neuroblastoma tumors. Our data suggest that targeting of activated human effector cells may provide a new and effective cancer immunotherapy protocol.

Animals↗
PubMed1986

Myelin-associated glycoprotein shares an antigenic determinant with a glycoprotein of human melanoma cells.

A sulfated 100K-dalton glycoprotein has been shown to be released into the culture medium of melanoma cells. Monoclonal antibodies 10C5 and 11B5, which were raised to human melanoma cells, as well as HNK-1 bind to this glycoprotein. It is shown here that mouse anti-myelin-associated glycoprotein (MAG) carbohydrate antibodies raised to human MAG and a human IgM paraprotein associated with neuropathy also bind to the same 100K molecule. However, anti-MAG antibodies recognizing peptide epitopes do not appear to react with this glycoprotein of melanoma cells, a result suggesting that its similarity to MAG is restricted to shared carbohydrate moieties. The anti-melanoma antibodies (10C5 and 11B5) resemble HNK-1 in binding to MAG and to some 19-28K-dalton glycoproteins and sulfated, glucuronic acid-containing sphingoglycolipids of the peripheral nervous system (PNS). In addition, the anti-melanoma antibodies cross-react with neural cell adhesion molecule (N-CAM), an observation emphasizing the shared antigenicity between MAG and other adhesion molecules. The results demonstrate that the anti-melanoma antibodies fall into a class of monoclonal antibodies (including HNK-1, human IgM paraproteins associated with neuropathy, anti-human MAG antibodies, and L2 antibodies) that are characterized by reactivity against related carbohydrate determinants shared by human MAG, N-CAM, and several protein and lipid glycoconjugates of the PNS.

Antibodies, Neoplasm↗
PubMed1986

Antigens associated with human squamous cell lung carcinoma defined by murine monoclonal antibodies.

A panel of 12 monoclonal antibodies that preferentially react with human squamous lung carcinoma cells has been produced. All are reactive with fresh frozen sections of squamous cell lung carcinoma tissues in immunoperoxidase assays and are unreactive with lymphoblastoid cells, red blood cells, and fibroblasts in enzyme-linked immunosorbent assay. At least eight of these antibodies interact with cell surface components. These reagents can be subdivided into four groups based upon their reactivities. Groups 1 to 3 are unreactive with normal liver, lung, kidney, colon, spleen, and pancreas in immunoperoxidase assays. Group 1 antibodies (PF1/A, PF1/B, PF1/C, PF1/D, and PF1/E) are all of IgG3 subclass and immunoprecipitate nonsulfated glycoprotein components with molecular weights of 80,000 and 180,000 and a nonglycosylated polypeptide with a molecular weight of 38,000. Group 1 antibodies are also reactive with some lung adenocarcinomas and, with the exception of PF1/E, stain certain differentiated strata within normal adult plantar and fetal epidermis. Group 2 antibodies (PF2/A and PF2/B) react also with breast, gastric, and colonic adenocarcinomas and some tumors of neuroectodermal origin. Group 2 antibodies, which are both of IgG3 subclass immunoprecipitate a nonglycosylated Mr 24,000 polypeptide. Group 3 antibodies (PF3/A, an IgG1; PF3/B, an IgGM; and PF3/C, an IgG2a) react additionally with certain other tumors, as well as with normal adult and fetal epidermis. Group 4 antibodies (PF4/A, an IgG2a; and PF4/B, an IgG1) are less specific than those of the preceding groups, as they react with some normal tissues, including pancreatic islets and pneumocytes, as well as with a variety of adenocarcinomas and tumors of neuroectodermal origin. PF4/A and PF4/B immunoprecipitate Mr 100,000 and 95,000 glycoproteins, respectively.

Animals↗
PubMed1986

Biosynthesis and expression of the disialoganglioside GD2, a relevant target antigen on small cell lung carcinoma for monoclonal antibody-mediated cytolysis.

Monoclonal antibodies (MAbs) 126 (immunoglobulin M) and 14.18 (immunoglobulin G3) react strongly with the cell surface of small cell carcinoma of the lungs (SCCL) and are unreactive with most normal tissues and other neoplasms with the notable exception of tumors derived from cells of neural crest origin. These MAbs react specifically with the oligosaccharide portion of the disialoganglioside GD2. Analysis of total gangliosides from cultured cell lines derived from SCCL indicates that GD2 is a predominant ganglioside. A comparison of the reactivities of MAbs against GD2 with those directed against gangliosides GM2 and GD3, each differing from GD2 by a single sugar residue, clearly indicates that GD2 is preferentially expressed by cultured cells derived from SCCL. Membranes isolated from these cells exhibit GD2 synthetase activity which specifically converts the precursor GD3 to GD2 in the presence of uridine diphosphate-N-acetyl galactosamine as the glycosyl donor. We present evidence that in SCCL, GD2 serves as a relevant target antigen for monoclonal antibody-mediated cytolysis. Specifically, we demonstrate that MAb 14.18 (immunoglobulin G3), can lyse small cell carcinoma of the lung targets by either complement- or antibody-dependent cellular cytotoxicity.

Antibodies, Monoclonal↗
PubMed1985

Eradication of established human melanoma tumors in nude mice by antibody-directed effector cells.

The simultaneous injection of monoclonal antibody 9.2.27, directed against a chondroitin sulfate proteoglycan preferentially expressed on human melanoma cells, and 2 X 10(7) mononuclear splenocytes, eradicated established, progressively growing human melanoma tumors in nude mice. Neither splenocytes nor antibody alone achieved significant tumor regression. The cells responsible for tumor elimination are most likely natural killer (NK) cells: they are present in splenocytes of T cell-deficient nude mice, and cloned cells with NK activity are able to suppress tumor growth. Moreover, splenocytes treated with anti-asialo GM1 and complement or harvested from NK-deficient C57BL/6 beige mice did not cause tumor rejection. Furthermore, treatment of BALB/c nude mice just before injection with anti-asialo GM1 antiserum, which is known to eliminate NK activity in vivo, resulted in better tumor growth. In addition, evidence is presented that cells with NK activity are probably the effectors responsible for melanoma target cell lysis in vitro: Antibody-dependent and -independent cell-mediated lysis of M21 melanoma cells was suppressed when splenocytes were preincubated with complement and antibodies specific for cell surface antigens of NK cells, i.e., anti-asialo GM1, anti-Qa5, and anti-NK1.1. Moreover, splenocytes of C57BL/6 beige mice were not able to lyse M21 cells in vitro. These results strongly support the conclusion that cells with NK activity are indeed responsible for the antibody-dependent destruction of M21 melanoma cells in vivo and in vitro.

Animals↗
PubMed1985

Disialoganglioside GD3 on human melanoma serves as a relevant target antigen for monoclonal antibody-mediated tumor cytolysis.

Monoclonal antibody MB3.6 (IgG3) recognizes disialoganglioside GD3, which represents a major surface marker on most human melanoma cells. We demonstrate that this antibody effectively lyses four human melanoma cell lines expressing significant levels of GD3 on their surface by either of two mechanisms: antibody-dependent cellular cytotoxicity (ADCC) or complement-mediated cytotoxicity. However, a melanoma cell line that expresses minimal levels of GD3 on 13% of the cells shows insignificant lysis by MB3.6 by either of these two mechanisms, suggesting that a threshold level of antigen expression may be required for effective in vitro cytolysis. In addition, monoclonal antibody (MAb) MB3.6 effectively inhibits establishment and growth of human melanoma tumors in the nude mouse when injected 24 hr after subcutaneous inoculation of tumor cells. Furthermore, MB3.6 produces specific regression of established melanoma tumors when injected 7 days after the subcutaneous inoculation of tumor cells. In contrast, tumor growth in animals injected with the melanoma cell line expressing minimal levels of GD3 was not affected by MAb MB3.6. These data indicate that once appropriate levels of the GD3 ganglioside are expressed on human melanoma cells, MAb MB3.6 can mediate tumor cell killing in vitro and in vivo and, thus, may prove useful for effective immunotherapy of human malignant melanoma.

Animals↗
PubMed1985

Human major histocompatibility complex class I antigens: residues 61-83 of the HLA-B7 heavy chain specify an alloreactive site.

A chemically synthesized peptide (sequence in text) homologous to residues 61-83 of the HLA-B7 heavy chain, induced antibodies that specifically recognized the HLA heavy chain-beta 2-microglobulin complex and the free heavy chain of the HLA-B7 antigen. These antibodies specifically immunoprecipitated the HLA-B7 beta 2-microglobulin complex solubilized from human lymphoblastoid cells by nonionic detergents and reacted with free HLA-B7 heavy chains in blots on nitrocellulose. These observations suggest that the antigenic conformation of this region of the HLA-B7 molecule is independent of the presence of beta 2-microglobulin and that amino acid residues 61-83 mimic an alloreactive site expressed by the HLA-B7 antigen.

Amino Acid Sequence↗
PubMed1984

Biosynthetic studies of proteoglycans in human melanoma cells with a monoclonal antibody to a core glycoprotein of chondroitin sulfate proteoglycans.

Monoclonal antibody (Mab) 9.2.27 was utilized in a combination of biosynthetic and biochemical investigations as an immunological probe for the study of chondroitin sulfate proteoglycans (CSP) in human melanoma cells. Pulse-chase and long-term intrinsic labeling immunoprecipitation experiments combined with the biosynthetic inhibitors monensin, cycloheximide, and paranitrophenol-beta-D-xyloside all suggest that Mab 9.2.27 recognizes a set of glycoprotein molecules ranging to a 250-kDa glycoprotein which serves as the core glycoprotein for CSP in human melanoma cells. Peptide maps comparing the 250-kDa and CSP molecule verify that the 250-kDa glycoprotein is the CSP core protein in human melanoma cells. Further studies document that the CSP released by melanoma cells and recognized by Mab 9.2.27 contains (2-acetamido-2-deoxy-3-O-(beta-D-gluco-4-enepyranosyluronic acid)-beta-4-O-sulfo-D-galactose and 2-acetamido-2-deoxy-3-O-(beta-D-gluco-4-enepyranosyluronic acid)-beta-6-O-sulfo-D-galactose saccharides and this CSP can interact with hyaluronic acid-Sepharose. Topographical studies indicate that this CSP has pericellular punctuated distribution on the melanoma cell surface and may play a role in cell-substrate interactions in the biology of metastatic human melanoma.

Aggrecans↗
PubMed1984

O-acetylation of disialoganglioside GD3 by human melanoma cells creates a unique antigenic determinant.

Monoclonal antibody Mab D1.1 recognizes on human melanoma cells a ganglioside antigen characterized by an alkali-labile O-acetylated sialic acid residue. Immunochemical analysis showed that this molecule is an O-acetylated product of the neuroectoderm-associated disialoganglioside GD3. Controlled chemical O-acetylation of purified GD3 resulted in the generation of this same epitope. Lysates of human melanoma cells were found to contain O-acetyltransferase activity capable of generating the antigenic epitope recognized by Mab D1.1. Thus, the addition of a single O-acetyl group to a common cell surface-associated ganglioside can create a potentially tumor-specific antigen.

Acetylation↗
PubMed1984

A monoclonal antibody recognizes an O-acylated sialic acid in a human melanoma-associated ganglioside.

Monoclonal antibody D1.1 originally prepared against the B49 cell line derived from a rat brain tumor was shown to react with a ganglioside present in fetal rat brain. We have found that this antigen is also present in human malignant melanoma tumors as well as many melanoma cell lines. The ganglioside from human melanoma cell lines migrates between GM1 and GM2 on one-dimensional thin layer chromatography. Analysis by two-dimensional thin layer chromatography with intermediate ammonia treatment suggests that the ganglioside contains one or more base-labile O-acyl esters. Mild base hydrolysis under conditions known to remove O-acyl esters results in complete loss of antigenic reactivity. Thus, the alkali-labile moiety is a critical component of the epitope recognized by the antibody. Analysis of the sialic acids of total gangliosides from [6-3H]glucosamine-labeled melanoma cells showed that approximately 10% of these molecules are O-acylated. Similar analysis of the purified ganglioside showed that greater than 30% of the sialic acids comigrated with authentic 9-O-acetyl-N-acetylneuraminic acid. The antibody did not cross-react with normal human skin melanocytes nor with any of a large number of normal human adult and fetal tissues. The antibody also did not react with numerous other malignant cell lines studied. These findings suggest that the antigenic epitope defined by antibody D1.1 contains an O-acylated sialic acid and may arise from aberrant O-acetylation occurring in human malignant melanoma cells.

Animals↗
PubMed1984

Antigenic expression of human melanoma cells in serum-free medium.

A human melanoma cell line, M14 , adapted to grow in serum free synthetic media was examined for its expression and secretion of several serologically defined melanoma associated antigens (MAA) previously described in this laboratory. Melanoma associated antigen expression and secretion was identical to that of M14 cells grown in parallel in serum supplemented medium. Spent synthetic media was found to be an enriched serum free source for the initial isolation of 100 kilodalton secreted glycoprotein MAA. M14 melanoma cells grown in synthetic media were also shown to be adaptable to the double agar clonogenic assay facilitating the examination of clonal heterogeneity in functional studies of MAA in melanoma tumor biology. Recent investigations from this laboratory have focused on characterizing human melanoma associated antigens (MAA) found either as secreted or cell surface associated glycoproteins in human melanoma cell lines. In these studies, monoclonal and polyclonal antiserums to melanoma cell components have been developed to specifically identify these MAAs immunochemically and provide a means to study the structural biochemistry of these determinants. At this time we have identified two antigens on which our research efforts are targeted: 1) a 100,000 dalton secreted glycoprotein (100K) common to melanoma, sarcoma and neuroblastoma tumor cell lines, and 2) a 250,000 dalton-high molecular weight component glycoprotein-proteoglycan complex which is thus far restricted to melanoma cells. The ultimate goal of our efforts is two-fold. Initially, we hope to develop schemes to isolate these melanoma associated antigens in sufficient quantities to obtain detailed structural information on these molecules, and secondly, we wish to implicate these glycoproteins in functional aspects of the biology of metastatic human melanoma in vitro.(ABSTRACT TRUNCATED AT 400 WORDS)

Antigens, Neoplasm↗
PubMed1984

Localization of the gangliosides GD2 and GD3 in adhesion plaques and on the surface of human melanoma cells.

The predominant gangliosides produced by two cultured human melanoma cell lines are GD3 and/or GD2. These gangliosides were found to be cell associated and present in substratum-attached material after cell removal by EDTA. Monoclonal antibodies directed to GD2 and GD3 specified the cell-surface distribution of these gangliosides and localized them in focal adhesion plaques at the interface of cells and their substratum. These attachment sites did not represent indiscriminant membrane fragments remaining after removal of cells with EDTA, because neither melanoma-associated proteoglycan nor class I histocompatibility antigens were detected by their respective antibodies. Our data suggest that the disialogangliosides GD2 and GD3 may be involved in the interaction between human melanoma cells and solid substrata.

Cell Adhesion↗
PubMed1984

Characterization of monoclonal antibody 155.8 and partial characterization of its proteoglycan antigen on human melanoma cells.

Immunization of mice with a plasma membrane-enriched fraction from human malignant melanoma cells and subsequent generation of hybridomas resulted in the isolation of an IgG1 monoclonal antibody, 155.8, that recognizes chondroitin sulfate proteoglycans. By cell binding analysis, 155.8 was shown to react with seven of eight cultured melanoma cell lines, but not with a variety of lymphoblastoid cell lines or cultured tumor cells derived from other solid tumor types. Indirect immunoprecipitation of the 155.8 antigen from intrinsically labeled melanoma cells revealed a glycoprotein of Mr = 250,000 and a sulfated molecule of Mr greater than 400,000. The antigen was identified as a chondroitin sulfate type A/C proteoglycan synthesized by melanoma cells on the basis of its sensitivity to chondroitinase ABC digestion and the identification of sulfated glycosaminoglycans released from the antigen immunoprecipitated by 155.8. The determinants recognized by antibodies 155.8 and 9.2.27, another anti-chondroitin sulfate proteoglycan, immunoprecipitate only a proteoglycan from high density cesium chloride gradient fractions, (1.487 g/liter); however, they immunoprecipitate a free glycoprotein of Mr = 250,000 from low density fractions (1.317 g/liter). This demonstrated that the 155.8 and 9.2.27 determinants, both of which reside on the glycoprotein of Mr = 250,000, are also present in the proteoglycan, suggesting that this glycoprotein is the proteoglycan core protein. Monoclonal antibody 155.8 reacts with a determinant on the core protein distinct from that recognized by 9.2.27. Proteoglycans bearing 155.8 determinants are distributed on the surface of cultured melanoma cells in a punctated fashion that apparently resolves to short, filamentous structures at high magnification. Immunohistochemical analysis demonstrated that 155.8-defined proteoglycans are found in freshly biopsied melanoma tissue, suggesting that these antigens are also synthesized in vivo by melanoma cells.

Animals↗
PubMed1984

Antigens associated with a human lung adenocarcinoma defined by monoclonal antibodies.

Monoclonal antibodies KS1/4, KS1/9, and KS1/17 were developed in this laboratory from a fusion of the murine myeloma cell line P3X63Ag8 with spleens of BALB/c mice previously primed with UCLA P3 cells derived from a human adenocarcinoma of the lung. Monoclonal antibodies KS1/4 and KS1/17 seemed to recognize similar glycoprotein antigens on the lung carcinoma cells by indirect immunoprecipitation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis. However, mapping of [3H]lysine- and [3H]arginine-labeled tryptic peptides of antigens in specific immunoprecipitates of lung carcinoma cells by high-pressure liquid chromatography revealed a one peptide difference. Antibody KS1/9 did not immunoprecipitate any identifiable protein from detergent extracts of the immunizing cell line by routine methods and appears to detect a glycolipid antigen. Immunocytochemical analysis of tissue sections showed this monoclonal antibody to be reactive with adenocarcinomas of the lung and not with the other histological types of lung carcinoma or normal tissue. Monoclonal antibodies KS1/4 and KS1/17, however, reacted with 3 major histological types of lung cancer and minimally with the proximal tubules of normal kidney and the epithelium of bronchioles.

Adenocarcinoma↗
PubMed1984

Human tumor-associated antigens defined by monoclonal antibodies.

Melanoma-associated antigens extracted from cultured human melanoma cells or isolated from their spent culture medium react specifically with monoclonal antibody and are represented by glycoproteins with molecular weights of 260,000 (260K), 250,000 (250K), and 100,000 (100K). The 260 and 250K antigens are present only in spent culture medium or detergent extracts of melanoma cells, respectively. The 100K antigen is an oncofetal marker present on tumor cells of different histological type. Indirect immunofluorescence data indicate that the 250K antigen is present on the surface of melanoma cells from excised melanoma tissues when the 100K antigen appears to be present in the cytosol. Subunit structure determination indicates that the 100K antigen is a single polypeptide chain whereas the 250K antigen extracted from melanoma cells is associated with a proteoglycan fragment which is important for its cell surface expression.

Antibodies, Monoclonal↗