The Czech Republic, a part of the former Czechoslovakia, has been at the forefront of several research directions in virology, genetics and physiology [...].

• MeSH
• Virology * MeSH
• Publication type
• Research Support, Non-U.S. Gov't MeSH
• Editorial MeSH
• Geographicals
• Czech Republic MeSH

Downregulation of MHC class I expression on tumour cells, a common mechanism by which tumour cells can escape from specific immune responses, can be associated with coordinated silencing of antigen-presenting machinery genes. The expression of these genes can be restored by IFNγ. In this study we documented association of DNA demethylation of selected antigen-presenting machinery genes located in the MHC genomic locus (TAP-1, TAP-2, LMP-2, LMP-7) upon IFNγ treatment with MHC class I upregulation on tumour cells in several MHC class I-deficient murine tumour cell lines (TC-1/A9, TRAMP-C2, MK16 and MC15). Our data also documented higher methylation levels in these genes in TC-1/A9 cells, as compared to their parental MHC class I-positive TC-1 cells. IFNγ-mediated DNA demethylation was relatively fast in comparison with demethylation induced by DNA methyltransferase inhibitor 5-azacytidine, and associated with increased histone H3 acetylation in the promoter regions of APM genes. Comparative transcriptome analysis in distinct MHC class I-deficient cell lines upon their treatment with either IFNγ or epigenetic agents revealed that a set of genes, significantly enriched for the antigen presentation pathway, was regulated in the same manner. Our data demonstrate that IFNγ acts as an epigenetic modifier when upregulating the expression of antigen-presenting machinery genes.

• MeSH
• Down-Regulation MeSH
• Epigenesis, Genetic MeSH
• Fibrosarcoma genetics   immunology   metabolism   MeSH
• Genes, MHC Class I * MeSH
• Interferon-gamma genetics   immunology   metabolism   MeSH
• DNA Methylation * MeSH
• Mice, Inbred C57BL MeSH
• Mice MeSH
• Tumor Cells, Cultured MeSH
• Antigen Presentation genetics   MeSH
• Gene Expression Regulation, Neoplastic MeSH
• Signal Transduction MeSH
• Transfection MeSH
• Up-Regulation MeSH
• Animals MeSH
• Check Tag
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Interferon-gamma MeSH

We constructed recombinant vaccinia viruses (VACVs) coexpressing the insulin-like growth factor-binding protein-3 (IGFBP-3) gene and the fusion gene encoding the SigE7Lamp antigen. The expression of the IGFBP-3 transgene was regulated either by the early H5 promoter or by the synthetic early/late (E/L) promoter. We have shown that IGFBP-3 expression regulated by the H5 promoter yielded higher amount of IGFBP-3 protein when compared with the E/L promoter. The immunization with P13-SigE7Lamp-H5-IGFBP-3 virus was more effective in inhibiting the growth of TC-1 tumors in mice and elicited higher T-cell response against VACV-encoded antigen than the P13-SigE7Lamp-TK(-) control virus. We found that high-level production of IGFBP-3 enhanced virus replication both in vitro and in vivo, resulting in more profound antigen stimulation. Production of IGFBP-3 was associated with a higher adsorption rate of P13-SigE7Lamp-H5-IGFBP-3 to CV-1 cells when compared with P13-SigE7Lamp-TK(-). Intracellular mature virions (IMVs) of the IGFBP-3-expressing virus P13-SigE7Lamp-H5-IGFBP-3 have two structural differences: they incorporate the IGFBP-3 protein and they have elevated phosphatidylserine (PS) exposure on outer membrane that could result in increased uptake of IMVs by macropinocytosis. The IMV PS content was measured by flow cytometry using microbeads covered with immobilized purified VACV virions.

• MeSH
• Antigens, Viral immunology   MeSH
• Insulin-Like Growth Factor Binding Protein 3 genetics   immunology   MeSH
• Immunization methods   MeSH
• Human papillomavirus 16 genetics   immunology   MeSH
• Mice, Inbred C57BL MeSH
• Mice MeSH
• Papillomavirus E7 Proteins genetics   immunology   MeSH
• Promoter Regions, Genetic MeSH
• Virus Replication immunology   MeSH
• T-Lymphocytes immunology   MeSH
• Antibody Formation MeSH
• Vaccination methods   MeSH
• Viral Vaccines immunology   pharmacology   MeSH
• Vaccinia virus genetics   immunology   MeSH
• Animals MeSH
• Check Tag
• Mice MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Antigens, Viral MeSH
• Insulin-Like Growth Factor Binding Protein 3 MeSH
• oncogene protein E7, Human papillomavirus type 16 MeSH Browser
• Papillomavirus E7 Proteins MeSH
• Viral Vaccines MeSH

Epigenetic events play an important role in tumour progression and also contribute to escape of the tumour from immune surveillance. In this study, we investigated the up-regulation of major histocompatibility complex (MHC) class I surface expression on tumour cells by epigenetic mechanisms using a murine tumour cell line expressing human E6 and E7 human papilloma virus 16 (HPV16) oncogenes and deficient in MHC class I expression, as a result of impaired antigen-presenting machinery (APM). Treatment of the cells with the histone deacetylase inhibitor Trichostatin A, either alone or in combination with the DNA demethylating agent 5-azacytidine, induced surface re-expression of MHC class I molecules. Consequently, the treated cells became susceptible to lysis by specific cytotoxic T lymphocytes. Further analysis revealed that epigenetic induction of MHC class I surface expression was associated with the up-regulation of APM genes [transporter associated with antigen processing 1 (TAP-1), TAP-2, low-molecular-mass protein 2 (LMP-2) and LMP-7]. The results demonstrate that expression of the genes involved in APM are modulated by epigenetic mechanisms and suggest that agents modifying DNA methylation and/or histone acetylation have the potential to change the effectiveness of antitumour immune responses and therapeutically may have an impact on immunological output.

• MeSH
• Apoptosis drug effects   MeSH
• Azacitidine pharmacology   MeSH
• Epigenesis, Genetic immunology   MeSH
• Neoplasms, Experimental genetics   immunology   virology   MeSH
• Genes, MHC Class I * MeSH
• Histones metabolism   MeSH
• Papillomavirus Infections complications   MeSH
• Enzyme Inhibitors pharmacology   MeSH
• Hydroxamic Acids pharmacology   MeSH
• Humans MeSH
• Human papillomavirus 16 * MeSH
• Mice, Inbred C57BL MeSH
• Mice MeSH
• Tumor Cells, Cultured MeSH
• Reverse Transcriptase Polymerase Chain Reaction methods   MeSH
• Antigen Presentation genetics   immunology   MeSH
• Up-Regulation drug effects   genetics   immunology   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Azacitidine MeSH
• Histones MeSH
• Enzyme Inhibitors MeSH
• Hydroxamic Acids MeSH
• trichostatin A MeSH Browser

Experiments were designed to examine whether local cytokine therapy of subcutaneous (s.c.) tumours results in inhibition of their lung metastases. Moderately immunogenic, major histocompatibility complex (MHC) class I and II negative. B7 negative, metastasizing murine carcinoma MK16 transplantable in syngeneic mice was obtained by co-transfection of human papilloma virus type 16 (HPV 16) E6/E7 and activated H-ras oncogene plasmid DNA into C57BL/6 kidney cells. After s.c. transplantation of the malignantly converted MK16 cells, the majority of the transplanted mice developed lung metastases; the number and size of the lung metastases increased with the increasing size of the s.c. tumour. Therapy of 5-day MK16 tumours by peritumoral administration of recombinant interleukin-2 (IL-2) and recombinant interleukin-12 (IL-12) inhibited growth of the s.c. MK16 tumour transplants and reduced the number of MK16 lung metastases. To investigate the antimetastatic effect of IL-2 and IL- 12 in a clinically more relevant setting, surgical minimal residual tumour disease was utilized. Subcutaneously growing MK16 carcinomas, 8-12 mm in diameter, were removed on day 30 and the operated mice were injected with IL-2 or IL- 12 on days 35-39 and 42-46 at the site of the operation. Treatment with IL-2 significantly reduced the percentage of MK16 tumour recurrences as well as the number of lung metastases, whereas the effect of IL- 12 was substantially weaker and statistically insignificant.

• MeSH
• B7-1 Antigen metabolism   MeSH
• B7-2 Antigen MeSH
• Cell Division drug effects   MeSH
• Antigens, CD metabolism   MeSH
• Histocompatibility Antigens metabolism   MeSH
• Tumor Virus Infections drug therapy   pathology   MeSH
• Papillomavirus Infections drug therapy   pathology   MeSH
• Interleukin-12 therapeutic use   MeSH
• Interleukin-2 therapeutic use   MeSH
• Carcinoma drug therapy   secondary   virology   MeSH
• Membrane Glycoproteins metabolism   MeSH
• Mice, Inbred C57BL MeSH
• Mice MeSH
• Lung Neoplasms drug therapy   secondary   MeSH
• Antineoplastic Agents therapeutic use   MeSH
• Cell Line, Transformed MeSH
• Neoplasm Transplantation MeSH
• Animals MeSH
• Check Tag
• Mice MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• B7-1 Antigen MeSH
• B7-2 Antigen MeSH
• Antigens, CD MeSH
• Cd86 protein, mouse MeSH Browser
• Histocompatibility Antigens MeSH
• Interleukin-12 MeSH
• Interleukin-2 MeSH
• Membrane Glycoproteins MeSH
• Antineoplastic Agents MeSH