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PubMed1976

Independent control of the expression of several membrane-associated antigens of murine leukemia cells: metabolism and response to specific antiserum.

The effect of specific antisera on the metabolic rates of thymus-leukemia (TL), H-2a, and tumor-associated antigens of ASL-1 and RADA-1 cells, two murine leukemia cell lines, were determined. The metabolic half-life of each antigen was distinct from the others examined. The half-life of TL antigens was 18 hr; in cells exposed to TL antiserum, it changed to 9 hr. In RADA-1 cells, the half-life of TL antigens was 16 hr; in the presence of TL antiserum it became 4 hr. The half-lives of H-2a antigens and the tumor-associated antigen were unaffected by specific antiserum. Somatic hybrids of ASL-1 or RADA-1 cells were formed with LM(TK)-cells, a thymidine kinase deficient mutant of mouse L cells. Hybrid cells formed TL antigens, but unlike their parental cells their half-life of expression, approximately 30 hr, was unaffected by TL antiserum. Hybrid cells failed to undergo antigenic modulation under conditions more stringent than required to stimulate the modulation of ASL-1 or RADA-1 cells. The hybrids formed the tumor-associated antigen of their murine leukemia parental cells; however, the metabolic rate of the tumor antigen in hybrid cells was defferent than the metabolic rate of the analogous antigen in parental cells. Hybrid cells formed H-2 antigens of both parental sources, and possessed a hybrid karyotype. The half-life of H-2 antigens, approximately 26 hr, was the same both in parental and hybrid cells. H-2a antiserum had no detectable effect upon the metabolism of TL antigens in hybrid or parental cells; nor did TL antiserum have an effect upon the metabolism of H-2a antigens.

Animals↗
PubMed1976

An indirect radioimmunoassay for thymus leukemia (TL) antigens.

An indirect radioimmunassay for thymus leukemia (TL)2 antigens has been developed and its specificity documented. The assay makes use of anti-TL antibodies produced in congenic mice (A-Tla(b)) and radioiodinated purified rabbit anti-mouse IgG. Using this assay, differences can be detected in the amounts of antigen expressed on thymocytes of the three known phenotypes (TL.1,2,3;TL.2;TL-) of inbred mouse strains. Significant differences are also detected in comparison of the thymocytes from homozygous TL.1,2,3 mice (A-Tla(a)) and heterozyhotes from Tla(a) and Tla(b) parents. Optimum conditions for the assay have been established. Its ability to detect antigens on glutaraldehyde-fixed cells and the binding of noncytolytic antibodies on both viable and fixed cells are documented. The assay has also been used to quantitate the changes in TL antigen expression on cells incubated in anti-TL antisera under conditions of antigenic modulation.

Animals↗
PubMed1999

Cancer vaccines with emphasis on a viral oncolysate melanoma vaccine.

Biotherapy of malignant diseases has become the fourth treatment modality besides surgery, chemo- and radiotherapy. Whole cell melanoma vaccines with or without BCG and other adjuvants, purified ganglioside and shed antigens, recombinant viruses carrying tumor antigens, dendritic cells pulsed with antigenic peptides etc. are in clinical trials. Efficacious viral oncolysate vaccines induce the host to mount tumor-specific cytotoxic T-cell response and prevention of relapses is supported by clinical trials. The use of "polyvalent" whole cell vaccines vs. purified or genetically engineered single antigen vaccines is justified as i. only very few single tumor antigens are present in all tumors of a given histological type; and ii. antigen modulation occurs in tumors rendering them resistant to immune attack generated by vaccine against a single antigen. Thus polyvalent vaccines immunize against several antigens vs. against a selected antigen.

Adjuvants, Immunologic↗
PubMed1979

Antibody-induced movement of common melanoma membrane antigens on the surface of unfixed human melanoma cells.

Antisera to common human melanoma antigens were obtained from melanoma patients receiving autologous immunization with their own irradiated cultured melanoma cells and Bacillus Calmette-Guérin. The antibody thus derived was used to detect common antigens on the plasma membrane of three different human melanoma cell lines by membrane immunofluorescence. The antigen-antibody complexes on the surface of melanoma cells would move to a pole (capping) and would subsequently be extruded into the extracellular milieu at room temperature. Approximately 25 to 30% of viable cells were positive by immunofluorescence. However, when the cells were fixed with methanol, 60 to 70% of cells demonstrated membrane binding. Capping was inhibited at 0 degrees or when the cells were pretreated with vinblastine sulfate. It can be concluded that common tumor antigens exist on the surface of viable human melanoma cells and that the redistribution of antigen-antibody complexes is an active process. The extrusion of antigen-antibody complexes in vitro may represent a mechanism of antigenic modulation in vivo and could indicate a basic method of tumor survival since presumably the antigen-denuded cell is viable and capable of replication but not of recognition by subsequent effector immune events.

Antibodies, Neoplasm↗
PubMed1996

Cyclosporin depresses pancreatic islet expression of antigens for islet cell autoantibodies in non obese diabetic mice.

An unexpected observation led us to investigate whether a short course (7 days) of oral cyclosporin (CsA) at different doses (5, 10, 20 or 40 mg/kg/day) in nonobese diabetic (NOD) mice could modify the expression of islet antigens related to the autoimmune process. Analysis was performed on the last day of CsA administration, and then up to 60 days after CsA withdrawal. Antigen modulation was analysed by indirect immunofluorescence using islet-cell antibody (ICA)-positive human sera for ICA antigens, and by immunoperoxidase for glutamic acid decarboxylase 67 Kd (GAD67). Concomitantly, beta-cell function was evaluated by in vivo glucose tolerance and insulin response from isolated islets. The severity of insulitis and histological damage to islets was quantified. We measured splenocyte and thymocyte subsets by cytofluorometry (CD4+ CD8-, CD4- CD8+, and double-positive thymocytes; Thy1-2+, CD3+, CD4+, and CD8+ spleen cells) and determined splenocyte mitogenesis in response to concanavalin A. Even when the lowest dose (5 mg/kg) was used, CsA concentrated in the islets. A graded reduction of ICA antigens was detected, showing no effect for 5 mg/kg/day but a significant dose-dependent reduction (P < 0.01) with 10, 20 and 40 mg/kg/day. GAD67 expression was also reduced (P < 0.03) in a CsA dose-dependent manner. On the other hand, only treatment with the highest CsA dose (40 mg/kg/day) induced glucose intolerance in vivo (P < 0.02), decreased insulin sensitivity to glucose from isolated islets (P < 0.03), reduced insulitis (P < 0.03), and altered thymocyte and splenocyte phenotypes and mitogenesis (P < 0.02). Moreover, the reversibility of the different effects was different: islet antigens were not completely recovered 2 months after CsA withdrawal, whereas other immunologic and metabolic effects obtained with the highest CsA dose were reversed within 15 days. Thus, a short course of low CsA doses in NOD mice produced a pancreatic concentration of the drug which reduced the expression of certain islet antigens for several weeks, whereas major effects on immunological parameters and islet insulin release occurred only with higher CsA doses and improved more rapidly.

Animals↗
PubMed1998

Changing patterns and re-distribution of antigen in poorly differentiated carcinomas: its implications in tumour diagnosis.

The variability of expression of tumour-associated antigens via either antigenic heterogeneity or antigenic modulation presents a basic problem in immunohistochemical diagnosis of poorly/undifferentiated tumours. This work was designed to study antigenic expression on human resected epithelial tumours by a panel of most widely used antibodies (EMA, CEA, AUAI & Cytokeratin) in relation to tumour differentiation and polarization. It was observed that poorly differentiated carcinoma with loss of polarity show homogeneous membrane staining (with antibodies against EMA, CEA & AUAI) in contrast to either apical (luminal) or basolateral membrane staining in well differentiated counterparts. Biochemical studies have shown that apical and basolateral epithelial cell membrane domains have a characteristic set of glycoproteins. Tight junctions are essential for maintaining this functional polarization. It was concluded that structural and functional abnormalities of tight junctions in poorly differentiated carcinomas results in loss of polarity with progressive invasion of the cell surface by antigenic glycoprotein and resultant homogeneous individual cell antigenic expression in poorly differentiated carcinomas. This study demonstrates that antigenic expression on tumour cells is not static, but dynamic and heterogeneity of antigenic expression may well be due to biological factors such as spatial configuration of the lesion.

Adenocarcinoma↗
PubMed1997

Superficial angiomyxoma (cutaneous myxoma): a clinicopathologic study of 17 cases arising in the genital region.

Seventeen cases of superficial angiomyxoma (cutaneous myxoma) of the genital region are reported. Thirteen patients were female (age range: 15-33 years; mean: 21 years) and four were male (age range: 18-55 years; mean: 39 years). The sites of involvement in females were the labium majus or labium, not otherwise specified (n = 6), vulva (n = 4), groin (n = 2), and mons pubis (n = 1). All lesions in male patients involved the scrotum. The tumors were present from 2 months to 4 years before resection and ranged from 0.9 to 6 centimeters in maximal dimension; 10 tumors were 3 centimeters or less in size. The predominant reason for seeking medical attention was a slow growing painless mass. All lesions were locally excised. Follow-up was obtained for 9 patients with a mean and median follow-up interval of 135 and 95 months, respectively. A recurrence developed in three patients at 8 months, 7 years 11 months, and 20 years. No patient has been shown to have Carney's complex. The tumors were immunoreactive for vimentin (11/11), CD34 (11/11), muscle-specific actin (8/12), smooth muscle actin (9/11), S100 protein (5/13), and Factor XIIIa (5/9). No immunoreactivity was present for desmin (DE-R- 11), glial fibrillary acidic protein, estrogen receptor or progesterone receptor. Superficial angiomyxomas are probably derived from fibroblast-like cells capable of antigen modulation.

Adolescent↗
PubMed1982

Participation of follicle associated epithelium (fae), macrophages, and plasma cells in the function of appendix.

Rabbit appendix was studied by correlated scanning and transmission electron microscopy and light microscopy, taking advantage of luminal bacteria as naturally occurring markers for following antigen uptake. The three-dimensional relationship of lymphoid follicles to other structures was revealed clearly. The smooth luminal surface of the appendix is interrupted by stomata each leading to a recessed interval between the dome epithelium covering the apical pole of each follicle and the goblet cell-rich epithelium forming a cupola on the opposing surface. Bacteria abound in this recess. They are taken up by follicle associated epithelial (FAE) cells which comprise part of the dome epithelium. Intraepithelial macrophages phagocytize the bacteria. Bacteria-laden macrophages are numerous in the dome area of the follicle and in the germinal center. Large lymphatic sinuses surrounding the follicles are well visualized following stimulation of fluid movement. Plasma cells are located within the dome epithelial layer and immediately beneath it. Plasma cells may be concentrated along the base of epithelial cells, with the absence of a basal lamina indicating an absence of the molecular selection which it would normally provide. The appendix, like Peyer's patches, takes up antigens, processes them, and gives evidence of reaction to them locally, with the secretions from plasma cells possibly modulating antigen entry.

Animals↗
PubMed1984

Monoclonal antibodies to breast cancer-associated antigens as potential reagents in the management of breast cancer.

Monoclonal antibodies reactive with the surface of human breast carcinoma cells have been generated and characterized. The immunogens used were membrane-enriched fractions of metastatic carcinoma lesions. The various monoclonals were shown to react with previously known as well as with novel tumor-associated antigens (TAAs). The most specific of the latter group is monoclonal B72.3, which is reactive with a 220,000 to 400,000 high-molecular-weight glycoprotein complex found in 50% of human mammary carcinomas and 80% of human colon carcinomas. Monoclonal antibody B6.2, which recognizes a 90,000-d glycoprotein, was radiolabeled and shown to efficiently localize human carcinoma transplants in athymic mice via gamma imaging without the use of second antibody or background subtraction manipulations. F(ab')2 and Fab' fragments were shown to be more efficient for tumor localization than intact immunoglobulin. Whereas the phenomenon of antigenic heterogeneity of tumor cell populations has long been known to exist, this phenomenon was also shown to manifest itself as antigenic modulation, in which specific TAAs can modulate their expression on the cell surface concurrent with different phases of the cell cycle. A phenomenon known as antigen evolution, in which a specific cloned tumor cell population can gradually drift in antigenic phenotype, has also been demonstrated. Recombinant interferon has been employed to (1) enhance the expression of specific TAAs on the surface of tumor cells already expressing the antigen; and (2) induce the expression of specific TAAs on the surface of carcinoma cells not previously expressing the antigen. The clinical implications of such phenomena in gamma scanning for the detection of tumor masses and for tumor immunotherapy are discussed. Methods for circumvention of problems inherent in the clinical use of monoclonal antibodies are also addressed.

Animals↗
PubMed1984

Interaction of monoclonal antibodies with cell surface antigens of human ovarian carcinomas.

Two monoclonal antibodies, OC 125 and OC 133, bind to distinct determinants on the surface of human epithelial ovarian carcinoma cell lines. OC 125 and OC 133 recognize determinants on molecules with molecular weights greater than 200,000 and 80,000, respectively. When binding to four different cell lines was compared, apparent affinity constants for OC 125 ranged from 3.1 X 10(9) to 6.0 X 10(7) M-1, whereas those for OC 133 ranged from 1.6 X 10(9) to 8.5 X 10(8) M-1. An estimate of the number of antigenic determinants per cell ranged from 1.0 X 10(7) to 2.8 X 10(5) for OC 125 and from 4.0 X 10(5) to 3.4 X 10(4) for OC 133. Antigenic determinants recognized by OC 125 and OC 133 could be detected in spent culture medium. When radiolabeled OC 125 was incubated with each of four ovarian tumor cell lines, approximately 90% of the antibody remained bound to the tumor cell surfaces for more than 20 hr. Similar binding of OC 133 was observed with three of the four ovarian tumor cell lines. By contrast, greater than 70% of OC 133 antibody was either shed or endocytosed after binding to OVCA 433 cells over the same period. Antigenic modulation was not induced by either antibody interacting with any of the four cell lines. These data suggest that antigen may be lost from the surface of human ovarian carcinoma cells by several different mechanisms and that antigen release is not inconsistent with binding of radiolabeled antibody to the tumor cell surface for prolonged periods.

Antibodies, Monoclonal↗
PubMed1983

Modulation of Bordetella pertussis by nicotinic acid.

Growth of Bordetella pertussis in a high concentration of nicotinic acid (NA) had a modulating effect on several properties and activities of the bacteria. Compared with normally grown cells, those grown in a high concentration of NA had reduced capacity for taking up both NA and nicotinamide (ND); they had reduced adenylate cyclase activity and showed loss of agglutinogen factors 2 and 3, but an increase in factor 1. By contrast, cells grown in a high concentration of ND showed only a slightly decreased capacity for uptake of ND and none of the other changes. Modulation of B. pertussis by NA varied with the strain and culture conditions and appeared to be distinct from the antigenic modulation induced by high Mg2+ in the culture medium. Evidence is presented for the association of a small proportion of the extracytoplasmic adenylate cyclase with the outer membrane of B. pertussis.

Adenylyl Cyclases↗
PubMed1995

Detection of apoptosis and expression of apoptosis-related proteins during human intrahepatic bile duct development.

We investigated apoptosis by nick end labeling and the expression of apoptosis-related proteins by immunohistochemistry in fetal development of human intrahepatic bile ducts and hepatocytes. During intrahepatic bile duct development, apoptosis was present at all stages, and its positive ratio was high in the remodeling ductal plate, moderate in the ductal plate, and relatively low in remodeled ducts. The cell proliferative activity as determined by proliferating cell nuclear antigen was also high in the remodeling ductal plate, and relatively low in the ductal plate and remodeled ducts. fas antigen and c-myc protein were constantly positive in the ductal plate, remodeling ductal plate and remodeled ducts. Bcl-2 protein was negative or faintly positive in the ductal plate and remodeling ductal plate, but was apparently positive in remodeled ducts. Lewisy as detected by the BM-1 antibody was present in the ductal plate, remodeling ductal plate, and remodeled ducts. p53 protein was not found in any cell types in the liver development. During hepatocyte development, many apoptotic and proliferating cell nuclear antigen-positive hepatocytes were noted. The developing hepatocytes expressed c-myc protein and fas antigen. Bcl-2 protein and Lewisy antigen were also weakly positive in the developing hepatocytes. These findings showed that balanced cell proliferation and apoptosis are involved in the normal development of intrahepatic bile ducts and hepatocytes, and suggest that c-myc protein, fas antigen, Bcl-2 protein, and Lewisy antigen modulate apoptosis of fetal intrahepatic biliary cells and hepatocytes, probably by stimulative (c-myc protein and fas and Lewisy antigens) or inhibitory (Bcl-2 protein) effects.

Antigens, Surface↗
PubMed2004

Modulation of peripheral blood T-lymphocytes in kidney transplant recipients treated with low dose OKT3 therapy.

The immunosuppressive effect of OKT3 depends upon both T cell depletion and antigenic modulation of CD3 complex. To establish the effect of low doses of OKT3 on peripheral T lymphocytes, we analyzed 47 kidney transplant recipients receiving OKT3 for the first time. OKT3 was used as rescue therapy in 39 patients and as part of induction protocols in 8. The mean age of patients was 39+/-10 years, 30 were females and 9 were re-transplants. Half of them (51.1%) received kidney from cadaver donors. Among those receiving OKT3 as rescue therapy, 82% recovered graft function, including patients with severe BANFF-graded rejections. After the first dose of OKT3, it a pronounced T cell depletion was observed followed by an increase in CD4 and CD8 expression in CD3 negative T cells, supporting the idea that T cell modulation was present. In conclusion, low dose OKT3 was effective in treating severe allograft rejection by inducing a sustained TCR/CD3 down modulation without long-lasting T cell depletion.

Adult↗
PubMed1986

Human T-cell leukemia-associated cell surface glycoprotein GP37: studies with three monoclonal antibodies and a rabbit antiserum.

Three monoclonal antibodies (mAbs), termed SN2, SN2a and SN2b, were used in the present work to study a human T-cell leukemia-associated cell surface glycoprotein, GP37. Strong specificity of mAbs SN2, SN2a and SN2b for T leukemia cells was demonstrated by radioimmunoassay and fluorescence-activated cell sorter (FACS) analysis. GP37 was not detected on normal human peripheral blood lymphocytes, purified normal T-cells, normal thymocytes nor normal bone marrow cells. Furthermore, GP37 was barely detectable on phytohemagglutinin (PHA)- and Concanavalin A (Con A)-activated T-cells. The results indicate clinical utility of these mAbs. Competitive binding experiments show that the epitopes recognized by SN2 and SN2a are sufficiently close to each other to allow complete reciprocal inhibition of binding whereas the epitopes recognized by SN2 and SN2b are less close to allow only partial reciprocal binding inhibition. The biochemical nature of antigenic determinants defined by these mAbs was studied by treating T leukemia cells with trypsin, chymotrypsin, thermolysin, neuraminidase and mixed glycosidases. The results suggest that the antigenic determinants defined by these mAbs all consist of the protein moiety of the glycoprotein GP37. No significant antigenic modulation was observed when T leukemia cells were reacted with SN2. In a sequential immunoprecipitation experiment, a 125I-labeled leukemia antigen preparation was first treated with a rabbit anti-T leukemia antiserum. The latter had been prepared by immunizing a rabbit with a partially purified human T leukemia antigen preparation and showed a good specificity for T leukemia cells. Subsequent treatment of the labeled antigen preparation with SN2 showed that SN2 antigen had been precleared. Thus, both mouse mAb SN2 and the rabbit anti-T leukemia antiserum react with the same GP37 molecule.

Animals↗
PubMed1981

Escape of metastasizing clonal tumor cell variants from tumor-specific cytolytic T lymphocytes.

A metastasizing variant of a chemically induced lymphoma from a DBA/2 mouse is shown to carry a distinct tumor-associated transplantation antigen (TATA), which can be recognized by syngeneic secondary anti-tumor cytolytic T lymphocytes (CTL). During metastasis of twice-cloned cell lines of this tumor, variants develop that are specifically immunoresistant to lysis by anti-tumor CTL. The variants are detected in the spleen of normal syngeneic mice. They remain stable over long-term passage in tissue culture. The high frequency with which these immunoresistant metastatic variants develop was found to explain the relative ineffectiveness of specific immunization against this metastatic tumor. Compared with organ-selective metastatic variants, the immunoresistant tumor variants seem to arise with a much higher frequency. The change in TATA expression described here differs from antibody-induced antigenic modulation in that it is more stable and genetically transmitted.

Antigens, Neoplasm↗
PubMed2005

Monoclonal antibodies that identify the CD3 molecules expressed specifically at the surface of porcine gammadelta-T cells.

The CD3 antigen is a surface structure associated with the T-cell receptor (TCR) to form a complex involved in antigen recognition and signal transduction. Reports on the structures of the CD3 molecules associated with alphabeta- and gammadelta-TCR have been contradictory. To investigate this issue, we raised a panel of monoclonal antibodies (mAb) against purified porcine CD3 molecules. Unlike the conventional anti-CD3, these mAb reacted specifically with peripheral gammadelta-T cells, but not with alphabeta-T cells. Immunoprecipitation showed that the antibody recognized a subset of CD3 molecules that were associated with gammadelta-TCR. Also unlike the conventional anti-CD3, these mAb, though directed at two different epitope groups, failed to induce antigenic modulation, T-cell proliferation and CD3-redirected cytotoxicity. Taken together, these results suggest that there are differences in the antigenicity, signal transduction potentials and probably structural differences between the CD3 molecules expressed at the surface of alphabeta- and gammadelta-T cells.

Animals↗
PubMed1990

Evidence that modulation requires sequences downstream of the promoters of two vir-repressed genes of Bordetella pertussis.

Gene expression in Bordetella pertussis is altered by environmental signals in a process called antigenic modulation. In the presence of modulating signals, expression of several known virulence factors and outer membrane proteins is coordinately reduced. From a bank of TnphoA fusions, we have identified five genes whose expression profiles are reciprocal of those of the major virulence determinants; that is, alkaline phosphatase activity is maximal during growth in the presence of the modulators nicotinic acid and MgSO4 (S. Knapp and J. J. Mekalanos, J. Bacteriol. 170:5059-5066, 1988). We have called these loci vir-repressed genes (vrg). Two of these gene fusions (vrg-6 and vrg-18) have been cloned in Escherichia coli, returned on low-copy-number plasmids to several strains of B. pertussis, and found to be regulated similarly to the fusions harbored on the chromosome. Deletions of the two vrg promoters were constructed and returned to B. pertussis. Regulation was maintained even when all but 24 nucleotides upstream of the vrg-18 initiation codon and 60 nucleotides upstream of the vrg-6 initiation codon were deleted, suggesting that cis-acting regulatory elements of these genes lie very near or within the coding region. We observed a 21-base palindromic sequence overlapping an 8-base direct repeat within the signal sequence coding region of vrg-6; insertion of a 6-bp linker in this region abolished regulation. These repetitive sequences are also at the site of greatest primary sequence identify between vrg-6 and vrg-18 and correspond to the signal sequence coding region. We propose models that involve recognition of this region by a vir-regulated gene product.

Alkaline Phosphatase↗
PubMedDate not supplied

Accessibility of antigenic sites recognized by AUA1, HMFG1 and HMFG2 monoclonal antibodies: its influence on antibody binding of live cells.

We assessed the immunoreactivity of live and alcohol-fixed monolayers of HRA-19, a rectal adenocarcinoma cell line, to the monoclonal antibodies AUA1, HMFG1 and HMFG2. Differences in staining patterns between live and alcohol-fixed colonies were found. The well-polarized cells forming the centers of the monolayer colonies showed strong membrane staining when the cells were alcohol-fixed prior to AUA1 incubation, but showed no staining when the cells were alive during the incubation. When AUA1 incubation was done both before and after alcohol fixation, membrane staining was again seen, ruling out the possibility of antigenic modulation. Incubation of live cells with AUA1 together with EDTA showed strong staining of dissociating cells. It is concluded that AUA1 antigenic sites, which on polarized cells are basolateral in location, are inaccessible to the antibody-containing culture fluid, which bathes the apical aspects of the cells, but they become accessible after alcohol fixation, or treatment with EDTA. HMFG1 antigenic sites are located on the apical cell membrane, and accordingly, no differences were seen between incubation of live and alcohol-fixed cells when incubated with HMFG1. The antigenic sites of HMFG2 are partly intracellular, and in our monolayer model, the staining of live cells was weaker and more scarce than on alcohol-fixed cells. It is concluded that immunostaining of cytological and histological material of tumours may not adequately predict antibody binding on live cells, and thus, these findings are of importance in the context of selection of monoclonal antibodies for clinical radio-immunotargeting.

Adenocarcinoma↗