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  • 1
    Electronic Resource
    Electronic Resource
    Oxford, UK : Blackwell Publishing Ltd
    Clinical and experimental pharmacology and physiology 21 (1994), S. 0 
    ISSN: 1440-1681
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Medicine
    Notes: Upon interaction with target cells, cytotoxic T lymphocytes and natural killer cells vectorially secrete highly specialized cytoplasmic granules containing perforin and a family of serine proteases (granzymes). This granule exocytosis mechanism of cytolysis is of patho-physiological importance, and usually results in target cell DNA fragmentation. Neither perforin nor granzymes possess inherent nuclease activity, but in combination they can induce target cell apoptosis. Perforin forms transmembrane pores in the target cell, thereby enabling granzymes to access target cell substrates. The target cell substrates of granzymes are unknown, but granzyme A binding and cleavage of the nuclear shuttle protein nucleolin in target cells demonstrates that granzymes may act on nuclear substrates. Furthermore, the presence of granzyme B and other granzyme activities in the nucleus of cytotoxic lymphocytes indicates that granzymes can be transported from the cytoplasm to the nucleus.It is hypothesized that perforin enables effector granzymes to enter the target cell cytoplasm and following their transport into the nucleus, granzymes cleave specific target cell nuclear proteins to activate autolytic endonucleases that fragment DNA. In cytotoxic effectors, these nuclear substrates are normally protected from granzymes by endogenous inhibitors.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1432-0851
    Keywords: Key words Monoclonal antibody ; CD19 ; Immunoconjugate ; Chimeric antibody ; Lymphoma ; Idarubicin ; Immunochemotherapy ; Anti-CD19
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract  Mouse monoclonal antibodies to CD19 detect an antigenic determinant expressed exclusively on the surface of B lymphocytes, and have previously been shown to be potentially useful therapeutic reagents for human B cell lymphoma. We report the production and characterization of a mouse/human chimeric antibody, cCD19, with potent in vivo antitumour activity. The genes encoding the variable domains for heavy (VH) and light (VL) chains were subcloned into eukaryotic expression vectors containing human constant region genes (IgG1 and κ), and co-transfected into non-secreting Sp2/0 mouse myeloma cells. Intraperitoneal administration of cCD19 produced inhibition of growth of subcutaneous CD19+ Sultan human B lymphoma tumours in scid/scid mice. When the antibody was administered 18 and 20 days after subcutaneous tumour inoculation, an approximately 30% reduction in tumour size was noted by day 29. cCD19 faithfully mimicked the in vitro binding characteristics of mCD19 as (a) the chimeric antibody was shown by flow cytometry to bind exclusively to cell lines that expressed CD19, (b) cCD19 was able to inhibit the binding of mCD19 on CD19+ cells completely and (c) the affinity of binding of the two antibodies was not significantly different [K a=(2.03±1.5)×108]. In bio-distribution studies, up to 14.8% of the total injected antibody dose per gram of tissue was localized in CD19+ Sultan tumours at 24 h approximately, 14.4% was present in the tumors at 48 h, and about 13.7% at 72 h. These levels were comparable to mCD19 administered in the same fashion. cCD19 conjugated to idarubicin was specifically and strongly cytotoxic to CD19+ cells cultured in vitro, and demonstrated an IC50 of 0.17 μM, similar to that of mCD19 (0.32 μM) and approximately 14-fold greater than the IC50 of free idarubicin. The specific cytotoxic capacity of cCD19 and its likely reduced immunogenicity suggest that it may potentially be of use in the treatment of refractory B cell lymphoma in humans.
    Type of Medium: Electronic Resource
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  • 3
    ISSN: 1432-0851
    Keywords: Monoclonal antibody ; CD19 ; Immunoconjugate ; Chimeric antibody ; Lymphoma ; Idarubicin ; Immunochemotherapy ; Anti-CD19
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Mouse monoclonal antibodies to CD19 detect an antigenic determinant expressed exclusively on the surface of B lymphocytes, and have previously been shown to be potentially useful therapeutic reagents for human B cell lymphoma. We report the production and characterization of a mouse/human chimeric antibody, cCD19, with potent in vivo antitumour activity. The genes encoding the variable domains for heavy (VH) and light (VL) chains were subcloned into eukaryotic expression vectors containing human constant region genes (IgG1 and κ), and co-transfected into non-secreting Sp2/0 mouse myeloma cells. Intraperitoneal administration of cCD19 produced inhibition of growth of subcutaneous CD19+ Sultan human B lymphoma tumours inscid/scid mice. When the antibody was administered 18 and 20 days after subcutaneous tumour inoculation, an approximately 30% reduction in tumour size was noted by day 29. cCD19 faithfully mimicked the in vitro binding characteristics of mCD19 as (a) the chimeric antibody was shown by flow cytometry to bind exclusively to cell lines that expressed CD19, (b) cCD19 was able to inhibit the binding of mCD19 on CD19+ cells completely and (c) the affinity of binding of the two antibodies was not significantly different [K a=(2.03±1.5)×108]. In biodistribution studies, up to 14.8% of the total injected antibody dose per gram of tissue was localized in CD19+ Sultan tumours at 24 h approximately, 14.4% was present in the tumors at 48 h and about 13.7% at 72 h. These levels were comparable to mCD19 administered in the same fashion. cCD19 conjugated to idarubicin was specifically and strongly cytotoxic to CD19+ cells cultured in vitro, and demonstrated an IC50 of 0.17 μM, similar to that of mCD19 (0.32 μM) and approximately 14-fold greater than the IC50 of free idarubicin. The specific cytotoxic capacity of cCD19 and its likely reduced immunogenicity suggest that it may potentially be of use in the treatment of refractory B cell lymphoma in humans.
    Type of Medium: Electronic Resource
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  • 4
    Electronic Resource
    Electronic Resource
    Springer
    Springer seminars in immunopathology 19 (1998), S. 323-343 
    ISSN: 1432-2196
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Type of Medium: Electronic Resource
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  • 5
    ISSN: 1432-0851
    Keywords: Key words Single-chain antibody ; Redirected cytotoxicity ; Fas ligand ; Breast carcinoma ; Immunotherapy
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract A chimeric receptor, consisting of the single-chain variable (scFv) domains of an anti-erbB-2 mAb linked via a CD8 membrane-proximal hinge to the Fc receptor γ chain, was expressed in the mouse cytotoxic T lymphocyte (CTL) hybridoma cell line, MD45. This cell line was grafted with the additional specificity to recognise and bind erbB-2-expressing breast carcinoma target cells T47D, MCF-7 and BT-20 in a non-MHC-restricted manner. Tumour cell lysis was antigen-specific since erbB-2-negative tumours were insensitive to lysis by MD45-scFv-anti-erbB-2-γ clones, and lysis of erbB-2+ tumour targets was inhibited in the presence of an anti-erbB-2 mAb. Furthermore, target cell death correlated with the level of chimeric receptor expression on the effector MD45 subclones. Redirected MD45 CTL utilised Fas ligand to induce target cell death since soluble Fas-Fc fusion protein completely inhibited cytolysis. The sensitivity of tumour target cells to Fas ligand was further enhanced by treating them with interferon-γ, a regulator of Fas and downstream signalling components of the Fas pathway. Overall, this study has demonstrated the requirement for successful activation of Fas ligand function in conjunction with cytokine treatment for effective lysis of breast carcinoma target cells mediated by redirected CTL.
    Type of Medium: Electronic Resource
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  • 6
    ISSN: 1432-1211
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Type of Medium: Electronic Resource
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  • 7
    ISSN: 1432-1211
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract It has previously been shown that a mouse monoclonal antibody, designated 4E, reacts with an epitope common to all HLA-B and -C antigens and those of the HLA-Aw19 cross-reactive group, namely, HLA-A29,-A30, -A31, -A32, -Aw33, and -Aw74. In order to pinpoint the amino acid residues which comprise the public specificity recognized by 4E, an HLA-A29 cDNA clone was isolated and its predicted amino acid sequence compared with those of other clonedHLA class I genes. The isolated HLA-A29 cDNA corresponded to the rarer of the twoA29 variant alleles,A29.1. Two amino acid residues of HLA-A29.1, gln-144 and arg-151, were found in all 24HLA-B andHLA-C alleles examined but were present in only one of 15HLA-A alleles for which sequence data are available. Importantly, this exceptional allele wasHLA-A32, another member of the HLA-Aw19 cross-reactive group. Gln-144 and arg-151 should be capable of jointly contributing to the binding site for 4E, as they are situated in successive alpha-helical subregions and are predicted to be juxtaposed in the three-dimensional HLA molecule. Four other residues in the first or second external domains of HLA-A29.1 (thr-9, leu-62, gln-63, and his-102) were unique among theHLA-A alleles, but none of these was found in corresponding positions ofHLA-B or-C alleles and thus failed to correlate with presence or absence of the 4E determinant. These observations are consistent with the notion that gln-144 and arg-151 define a determinant common to HLA-B, HLA-C, and the HLA-Awl9 cross-reactive group and the binding site of the monoclonal antibody 4E.
    Type of Medium: Electronic Resource
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  • 8
    ISSN: 1432-1211
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The human IFI16 gene is a member of an interferon-inducible family of mouse and human genes closely linked on syntenic regions of chromosome 1. Expression of these genes is largely restricted to hemopoietic cells, and is associated with the differentiation of cells of the myeloid lineages. As a prelude to defining the mechanisms governing IFI16 expression, we have deduced its genomic organization using a combination of genomic cloning and polymerase chain reaction amplification of genomic DNA. IFI16 consists of ten exons and nine intervening introns spanning at least 28 kilobases (kb) of DNA. The reiterated domain structure of IFI16 protein is closely reflected in its intron/exon boundaries, and may represent the evolutionary fusion of several independent functional domains. Thus, exon 1 consists of 5' untranslated (UT) sequences and contains sequence motifs that may confer interferon-inducibility, and exon 2 encodes the lysine-rich amino-terminal (“K”) region, which possesses DNA-binding activity. Exon 3 codes for a domain which is poorly conserved between family members, except for a strongly retained basic motif likely to provide nuclear localization. The first of two 200 amino acid repeat domains that are the hallmark of this family (domain A) is represented jointly on exons 4 and 5, which are reiterated as exons 8 and 9, respectively, to encode the second 200 amino acid domain (B). Two intervening serine-threonine-rich domains (C and C'), unique to IFI16, are each encoded by single exons of identical length (exons 5 and 6). These domains are predicted to encode semi-rigid “spacer” domains between the 200 amino acid repeats. The reiterated nature of exons 4 to 6 and the insertion of introns into a single reading frame strongly suggest that IFI16 and related genes arose by a series of exon duplications, some of which antedated speciation into mouse and humans. Several alternative mRNA cap sites downstream of a TATA consensus sequence were defined, using primer extension analysis of mRNA. Sequencing of ~1.7 kb of DNA upstream of this region revealed no recognizable consensus elements for induction by interferon-α (interferon-α/ß-stimulated response elements), but two motifs resembling interferon-γ activation sites were located. IFNs α and γ both induce IFI16 mRNA expression in myeloid cells. Interferon-α inducibility of IFI16 may be regulated by an interferon-α/ß-stimulated response consensus element in the 5' UT exon, as a similar motif is conserved in the corresponding position in the related myeloid cell nuclear differentiation antigen gene. An interferon-γ-activation site consensus was also located in this region of IFI16.
    Type of Medium: Electronic Resource
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  • 9
    ISSN: 1432-1211
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Type of Medium: Electronic Resource
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  • 10
    ISSN: 1432-1211
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract The HLA-C encoded gene products display several characteristics which distinguish them from HLA-A and -B. The HLA-C antigens are poorly expressed on the cell surface, they display multiple proteins with different isoelectric points, and alloimmunization to HLA-C antigens is less common. To investigate whether the multiple products result from differential splicing of HLA-C gene transcripts, we have isolated a full-length cDNA clone encoding the Cw6 antigen. Class I antigens produced by the cDNA clone in transfected cells were of the same relative mass as those observed in the parental cells when analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Isoelectric focusing (IEF) gel analysis of the cDNA translated products in transfectants revealed multiple IEF bands. All IEF bands detected in the transfectants were also found in the parental cells, indicating that the multiplicity of the C-locus products was not due to differential splicing of HLA-C gene transcripts, but was probably due to post-translational modification. Comparison of the sequences of C-locus alleles with those of A and B alleles did not show any apparent sequences which would generate multiple IEF bands. Comparison of the coding regions for seven HLA-C alleles and one HLA-C-related class I gene with available data for 15 HLA-A and 20 HLA-B alleles demonstrated several unique features for the HLA-C locus. Six sites in the extra cellular domains, three in al and three in a3, were unique. While the cytoplasmic (CP) domain of HLA-A and -B are almost identical, the CP of HLA-C alleles is unique. Similar unique features of HLA-C are also observed in the transmembrane domain, resulting in locus-specific residues between positions 295 and 300. The present study has ruled out differential mRNA splicing as a mechanism for the multiplicity of Cw6 antigens and demonstrated unique HLA-C locus sequences.
    Type of Medium: Electronic Resource
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