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1.
J Gen Virol ; 88(Pt 3): 792-802, 2007 Mar.
Article in English | MEDLINE | ID: mdl-17325351

ABSTRACT

Human papillomavirus type 16 (HPV-16) is a major cause of human cancer. Effective prophylactic vaccines are based on type-specific neutralizing antibodies. A major neutralizing epitope has been defined by the monoclonal antibody H16.V5. To investigate the importance of this epitope for overall immunogenicity of HPV-16, HPV-16 virus-like particles devoid of the H16.V5 epitope were engineered by site-directed mutagenesis of ten non-conserved, surface-exposed residues. Removal of the H16.V5-defined epitope had only a marginal effect on antigenic reactivity with antibodies in sera from infected subjects, but affected immunogenicity in experimental immunization of mice, with reduced induction of both antibody responses and CTL responses.


Subject(s)
Antigens, Viral/immunology , Capsid Proteins/immunology , Epitopes/immunology , Human papillomavirus 16/immunology , Oncogene Proteins, Viral/immunology , Animals , Antibodies, Monoclonal/immunology , Antibodies, Viral/blood , Antibodies, Viral/immunology , Antigens, Viral/chemistry , Antigens, Viral/genetics , Blotting, Western , Capsid Proteins/chemistry , Capsid Proteins/genetics , Cytotoxicity Tests, Immunologic , Enzyme-Linked Immunosorbent Assay , Epitopes/genetics , Human papillomavirus 16/genetics , Humans , Immunization , Mice , Mice, Inbred C57BL , Models, Molecular , Mutagenesis, Site-Directed , Neutralization Tests , Oncogene Proteins, Viral/chemistry , Oncogene Proteins, Viral/genetics , T-Lymphocytes, Cytotoxic/immunology
2.
Int Immunol ; 9(4): 599-606, 1997 Apr.
Article in English | MEDLINE | ID: mdl-9138021

ABSTRACT

A murine mAb (BAL-1) was previously shown to induce apoptosis when cross-linked on the cell surface of different B acute lymphocytic leukemia (ALL) and pro-myelocytic cell lines. The present study shows that BAL-1 specifically recognizes the MHC class I (MHC-I). The apoptotic response was not dependent on the epitope specificity, since other anti-MHC-I antibodies, reacting with different monomorphic determinants of the alpha chain or beta 2-microglobulin, also induced apoptosis in these cells. However, external cross-linking of antibodies was strictly required for the apoptotic effect. Among cells originating from mature peripheral blood B cells, anti-CD40-stimulated cells were susceptible to anti-MHC-I-induced apoptosis, whereas B cells activated with Staphylococcus aureus Cowan I (SAC) or with the superantigen staphylococcal enterotoxin A (SEA) were non-responsive. Mature SEA-activated T cells were also resistant to MHC-I-induced apoptosis. In situ terminal deoxynucleotidyl transferase staining of apoptotic cells at various stages during MHC-I-induced cell death revealed that apoptosis occurred predominantly in the G2/M phase of the cell cycle, with the first apoptotic cells appearing after approximately 12 h of incubation. These results suggest a role for MHC-I-mediated apoptosis during differentiation and activation of certain hematopoietic cells.


Subject(s)
Apoptosis/immunology , B-Lymphocytes/cytology , B-Lymphocytes/immunology , CD40 Antigens/physiology , Hematopoietic Stem Cells/immunology , Histocompatibility Antigens Class I/metabolism , Lymphocyte Activation , Animals , Antibodies, Monoclonal/chemistry , Antibodies, Monoclonal/pharmacology , Cell Division/immunology , Cell Line , DNA Fragmentation/immunology , HL-60 Cells , Histocompatibility Antigens Class I/immunology , Humans , Interphase/immunology , Mice , Mitosis/immunology , Precursor Cell Lymphoblastic Leukemia-Lymphoma/immunology , Tumor Cells, Cultured
3.
Exp Cell Res ; 231(1): 190-7, 1997 Feb 25.
Article in English | MEDLINE | ID: mdl-9056426

ABSTRACT

The major histocompatibility complex class I (MHC-I) has recently been shown not only to present antigens to the immune system but also to mediate transmembrane signaling, resulting in activation, inactivation, or apoptosis. Such signaling has been observed in both normal and malignant cells of the B and T cell lineage. Cross-linking of MHC-I on the pre-B-acute-lymphocytic cell line KM-3 induces an apoptotic process, which becomes evident after approximately 12 h. In order to better understand the mechanisms regulating this apoptotic process, we have investigated both gene expression and the effect of cross-linking on certain intracellular events. Differential display PCR was used to isolate two gene fragments whose level of expression was associated with the induction of apoptosis as they were downregulated in KM-3 cells following MHC-I cross-linking. These genes encode novel molecules whose function remains to be elucidated. It was further demonstrated that the apoptotic process was not accompanied by changes in [Ca2+]i, the level of activation of NF-kappaB, or changes in protein kinase C activity and that the initiation of apoptosis could be prevented by phorbol ester treatment. It is thus suggested that multiple, fine-tuned molecular events determine the outcome of cross-linking of MHC-I in this pre-B-lymphocytic cell line.


Subject(s)
Apoptosis , Gene Expression Regulation, Leukemic , Histocompatibility Antigens Class I/metabolism , Precursor B-Cell Lymphoblastic Leukemia-Lymphoma/genetics , Amino Acid Sequence , Base Sequence , Calcium/metabolism , DNA Fragmentation , Humans , Molecular Sequence Data , NF-kappa B/metabolism , Polymerase Chain Reaction , Precursor B-Cell Lymphoblastic Leukemia-Lymphoma/metabolism , Precursor B-Cell Lymphoblastic Leukemia-Lymphoma/pathology , Protein Kinase C/metabolism , RNA Probes , Tetradecanoylphorbol Acetate/pharmacology , Tumor Cells, Cultured
4.
Int J Cancer ; 57(4): 544-52, 1994 May 15.
Article in English | MEDLINE | ID: mdl-8181858

ABSTRACT

The lack of apoptosis or programmed cell death in human tumor cells has been suggested to be one factor allowing uncontrolled growth of neoplasms. We have developed a mouse monoclonal antibody (MAb) that induces programmed cell death in a human acute leukemia cell line (KM-3) of the pre B-cell type. Stable, antibody-producing hybridomas were produced by fusing mouse myeloma cells to spleen cells from mice immunized with viable KM-3 cells. Incubation of KM-3 cells with the MAb (designated anti-BAL) resulted in growth inhibition and subsequent cell death within 2-3 days. Anti-BAL required cross-linking with a rabbit anti-mouse antibody to induce DNA fragmentation typical of apoptosis. Immunoblotting experiments with anti-BAL identified a 37-kDa protein, apparently different from any previously described apoptosis-related surface antigen. Strongest expression of the antigen was generally found on cells of lymphoid or myeloid origin. However, several other cell types such as fibroblasts and endothelial cells were also stained by anti-BAL in flow cytometry but less intensively. Despite the apparent presence of this cell surface-bound 37-kDa antigen on several normal and malignant cell types, anti-BAL induced cell death only in human malignant cell lines expressing a more immature phenotype.


Subject(s)
Antibodies, Monoclonal/pharmacology , Antigens, Neoplasm/analysis , Antigens, Surface/analysis , Apoptosis/drug effects , Precursor B-Cell Lymphoblastic Leukemia-Lymphoma/pathology , Precursor Cell Lymphoblastic Leukemia-Lymphoma/pathology , Animals , Antibodies, Monoclonal/therapeutic use , Antigens, Neoplasm/immunology , Antigens, Neoplasm/metabolism , Antigens, Surface/immunology , Antigens, Surface/metabolism , Cell Division/drug effects , Child , Chromatin/drug effects , Chromatin/metabolism , Cycloheximide/pharmacology , DNA Damage , DNA, Neoplasm/drug effects , DNA, Neoplasm/metabolism , Humans , Mice , Mice, Inbred BALB C , Precursor B-Cell Lymphoblastic Leukemia-Lymphoma/immunology , Precursor B-Cell Lymphoblastic Leukemia-Lymphoma/therapy , Precursor Cell Lymphoblastic Leukemia-Lymphoma/immunology , Precursor Cell Lymphoblastic Leukemia-Lymphoma/therapy , Tumor Cells, Cultured/drug effects
5.
Cancer Lett ; 75(2): 103-9, 1993 Dec 10.
Article in English | MEDLINE | ID: mdl-8293420

ABSTRACT

The mouse monoclonal anti-BAL antibody induced apoptosis in a pre-B acute lymphocytic leukemia cell line within 2 days of incubation, after being crosslinked by a secondary antibody. The antibody specifically recognized a 37 kDa membrane protein that was expressed on a wide spectrum of normal and malignant cells, but induced programmed cell death in only very few of these cells. In this study, we have followed the initial kinetics of the antibody-induced cell death in the human acute lymphocytic leukemia cell line KM-3, by microcalorimetric measurements in conjunction with determination of the cellular proliferation rate and DNA fragmentation. An increase in metabolic activity was observed already after incubating the cells for 20 min with crosslinked anti-BAL antibody, which was several hours before significant growth inhibition and DNA fragmentation were detected. These data show for the first time that the initiation phase of antibody-induced apoptosis is an active, energy-dependent process and not merely an effect of receptor blocking.


Subject(s)
Antibodies, Monoclonal/immunology , Apoptosis/immunology , Burkitt Lymphoma/metabolism , DNA, Neoplasm/metabolism , Precursor B-Cell Lymphoblastic Leukemia-Lymphoma/metabolism , Antibodies, Neoplasm/immunology , Antigens, Neoplasm/immunology , Burkitt Lymphoma/immunology , Calorimetry , Cross-Linking Reagents , DNA Damage , Energy Metabolism , Hot Temperature , Humans , Kinetics , Lactates/metabolism , Oxygen Consumption , Precursor B-Cell Lymphoblastic Leukemia-Lymphoma/immunology , Tumor Cells, Cultured/metabolism , Up-Regulation
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