Replication of the F'lac sex factor in the cell cycle of Escherichia coli.
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Age Factors↗
Biomedical subjects
J Zeuthen
Publications and source records attributed to J Zeuthen.
Characterization of monoclonal antibodies against bovine insulin.
Six different monoclonal antibodies (IgG1 and IgG2a) were obtained after fusions of X63-Ag8-6.5.3 myeloma cells with spleen cells from BALB/c mice immunized with bovine insulin. Definition of binding determinants was attempted by competitive binding studies with insulins, proinsulins and modified insulins from various species. The monoclonal antibodies OXI-001 and OXI-004 were inferred to react with a region including residue A10, OXI-002 with an antigenic determinant in the B26-30 region, OXI-005 with a region including B30 and OXI-006 with a tertiary structure near the N-terminus of the B chain, possibly including B3 and A10. The equilibrium binding constants for these antibodies were calculated by three different methods (Scatchard, Langmuir and non-linear regression) and were found to be in the range of 2 X 10(7)-8 X 10(9), with good agreement between the different methods of calculation. As expected for a given monoclonal antibody, the heterogeneity index was close to 1.0, as calculated from Sip's logarithmic transformation of the binding equation. These parameters were compared to those of a mixture of the six different monoclonal antibodies and those of a conventional hyperimmune anti-insulin serum (guinea-pig). The half-dissociation times (t1/2) of complexes of antibody and bovine insulin ranged from 35 min to 38 h.
Amino Acid Sequence↗
Amino acid substitutions in the melanoma antigen recognized by T cell 1 peptide modulate cytokine responses in melanoma-specific T cells.
Single amino acid substitutions in melanoma-associated peptides dramatically enhance T-cell cytotoxicity against target cells presenting the modified peptides (often referred to as heteroclitic peptides). The authors tried to determine whether peptide modifications influence other aspects of T-cell immunity toward malignant melanoma. A heteroclitic peptide, E26F, with an E to F substitution in melanoma antigen recognized by T cell 1 (MART-1)26-35, triggers an enhanced tyrosine phosphorylation response when compared with the native- and other-modified MART-1 peptides. Similarly, the E26F peptide enhances the production of mRNA for interleukin (IL)-5, IL-10, IL-13, IL-15, and interferon-gamma and significantly enhances release of IL-13 and IL-10 from anti-MART-1 cytotoxic T cells. Another heteroclitic peptide, 1L, with an A to L substitution in MART-1(27-35), also enhances the tyrosine phosphorylation response in anti-MART-1 cytotoxic CD8+ T cells. Yet, 1L does not enhance the production of T helper cell type 2-like cytokines (IL-10 and IL-13). Together these data show that minor amino acid modifications of immunodominant melanoma peptides profoundly influence the cytokine response in melanoma-specific T cells.
Amino Acid Substitution↗
Immunisation of colorectal cancer patients with autologous tumour cells.
Patients with colorectal cancer were entered into a clinical phase I trial of immunotherapy with an autologous tumour cell/bacillus Calmette-Guerin (BCG) vaccine. We attempted to describe the possible effects and side effects of the immunisation, and further to investigate whether expression of immune-response-related surface molecules on the tumour cells in the vaccine correlated with survival. The first and second vaccine comprised of 107 irradiated tumour cells mixed with BCG, the third of irradiated tumour cells only. Thirty-nine patients were considered, but only 6 patients fulfilled the criteria for inclusion. No serious side effects were observed. With three years of observation time, two patients are healthy, while the rest have had recurrence, and two of them have died. In all vaccines, all tumour cells expressed HLA class I, some expressed HLA class II and none expressed CD80. There was an inverse relation between survival and HLA class II expression. This highlights an essential problem, in the absence of CD80 expression the expression of HLA class II may induce anergy. In future attempts to develop improved vaccines this problem should be addressed.
Adenocarcinoma↗
LAK-cell-mediated cytotoxicity against tumor cell targets used to monitor the stimulatory effect of interleukin-2: cytotoxicity, target recognition and phenotype of effector cells lysing the Daudi, T24 and K562 tumor cell lines.
The T24 transitional bladder carcinoma cell line, the Daudi Burkitt lymphoma cell line and the K562 erythroleukemia cell line have all been used as target cells in 51Cr release assays to measure the in vivo induced lymphokine-activated killer (LAK) cell cytotoxicity during interleukin-2 (IL-2) therapy of cancer patients. However, different relationships between the clinical response to IL-2 treatment and the LAK cytotoxicity have been reported using these three different target cells. The purpose of the present study was to evaluate whether the LAK cytotoxicities measured against these target cells represent similar effector-to-target-cell interactions, so similar conclusions may be drawn of 51Cr release assay results in which the cell lines are used as target cells. The cytotoxicity of peripheral blood mononuclear cells (PBMC) and PBMC depleted of different natural killer and T cell subsets was measured against the three targets. LAK cell recognition of targets was evaluated by cold target inhibition experiments, and the development of LAK-cell-mediated lysis with time was evaluated in 51Cr release assays of varying duration. This study shows that LAK-mediated lysis of T24 and Daudi cells was closely related and LAK cytotoxicity measured in 51Cr release assays against these two target cells may be measurement of similar effector-to-target cell interactions.
Antigens, CD↗
Activation and phenotyping of LAK generated by exposure to product of cells transformed by interleukin 2 cDNA.
It has been previously found that local administration of X63-m-IL-2 cells transformed by interleukin 2 (IL-2) cDNA and constitutively producing large quantities of IL-2 mediated regressions of murine plasmacytomas and 3-methyl-cholanthrene-induced sarcomas transplanted in syngeneic mice. Here we show that killer cells generated by cultivation of spleen cells in supernatants from X63-m-IL-2 cultures (LAK) or by co-cultivation of murine splenocytes with X63-m-IL-2 cells were cytolytic for natural killer (NK)-sensitive as well as NK-resistant target cells, including the IL-2-producing X63-m-IL-2 cells. Spleen cells cultured in X63-m-IL-2 supernatants or co-cultivated with X63-m-IL-2 cells yielded predominantly Thy 1.2+, CD3+, LFA-1+ lymphocytes. The in vitro results suggest that the LAK cells generated due to the IL-2 production by genetically engineered cells probably help to terminate the treatment by killing the IL-2 producers.
Animals↗
Gene therapy of cancer: use of IL-2 gene transfer and kinetics of local T and NK cell subsets.
Experiments were designed to compare the efficacy of recombinant IL-2 immunotherapy and IL-2 gene therapy of i.p. growing murine plasmacytoma X63-Ag8.653. The kinetics of peritoneal exudate mononuclear cells were monitored during the progression and gene therapy of the plasmacytoma, using cytofluorometric analysis and monoclonal antibodies against T and NK cell subsets. It has been found that the percentage of mice protected against plasmacytoma transplants was higher in mice treated by transfer of genetically manipulated IL-2-producing plasmacytoma cells as compared to the mice repeatedly injected with recombinant IL-2. Intraperitoneal inoculation of the X63-Ag8.653 plasmacytoma led in most of the inoculated mice to an increased percentage of NK+, ASGM1+, Thy 1.2+, CD3+ and TCR alpha beta+ cells in the peritoneal fluid. The presence of macroscopically detectable i.p. tumours was accompanied by a higher increase in the percentage of NK+ and TCR gamma delta+ cells. Local IL-2 gene therapy of the plasmacytoma either prevented or diminished an increase in the percentage of CD3+, Thy 1.2+ and TCR alpha beta+ lymphocytes.
Animals↗
The immunogenic properties of human melanomas and melanoma-associated antigens recognized by cytotoxic T lymphocytes.
During the last years significant progress has been achieved in the identification of melanoma-associated antigens (MAA) recognized by cytotoxic T lymphocytes (CTL). These antigens belong to three main groups: tumor-associated testis-specific antigens (MAGE, BAGE, GAGE and PRAME), melanocyte differentiation antigens (tyrosinase, Melan-A/MART-1, gp100, TRP-1 and TRP-2) and mutated or aberrantly expressed antigens (MUM-1, CDK4, beta-catenin, gp100-in4, p15 and N-acetylglucosaminyltransferase V). For the identification of these antigens, CTL cultures from mainly only 4 different melanoma patients have been used. These patients developed a strong anti-melanoma response resulting in long-lasting disease-free periods, pointing to the importance of the identification of highly immunogenic melanomas. In each of these patients, the immune response was observed against a unique set of 4 to 6 individual antigenic epitopes, on one hand suggesting the low immunogenicity of the individual antigens, and on the other pointing to the importance of the identification of additional highly immunogenic melanomas for the discovery of new MAA. The analysis of the available data on the immunogenic and protective properties of individual MAA confirms their low immunogenicity. In our study, we focused on the identification of especially highly immunogenic melanomas among a panel of 40 newly established melanoma cell lines. So far, only two such melanoma cell lines, FM3 and FM57 have been identified in this panel. The immunogenic properties of uncloned FM3 cells and several FM3 clones have been further investigated. It was found that the immunogenic properties of melanoma cells are mainly determined by the expression of progression-associated antigens as well as by ecto-ATPase, a molecule which is able to modulate cell adhesion. Cloning the cultures of PBL, stimulated with uncloned FM3 or with the highly immunogenic FM3 clone, FM3.29, has permitted us to identify the immune response against eight different MAA, five of these probably representing not previously described antigens. (Tab. 2, Fig. 2, Ref. 68.)
Antigens, Neoplasm↗