RESUMO
The host-immune response against adenoviruses forms a major obstacle for their use as gene therapy vectors for treatment of genetic defects. None the less, they are the preferred vectors for in vivo gene transfer in experimental gene therapy protocols for cancer. In this article we demonstrate the antitumor efficacy of adenovirus-mediated transfer of human interleukin-2 cDNA in the rat-CC531 model for hepatic metastases of colorectal cancer: intratumoral administration of 10 plaque-forming units of the hlL-2-expressing adenoviral vector, AdCAIL-2, resulted in a cessation of tumor growth in 80% of the injected tumors. In control groups receiving AdCnull, a vector with the same viral backbone, but lacking transgene expression, none of the tumors responded. However, intratumoral treatment with this vector significantly enhanced tumor regression induced by systemic IL-2 protein treatment, which was used as a positive control. In addition we show, by performing delayed-type of hypersensitivity assays, that AdCnull when injected intratumorally enhances recognition of tumor antigens by T lymphocytes to the same extent as intratumoral treatment with the IL-2-expressing vector. The replication-deficient adenoviruses appear to have a therapeutic advantage in cytokine-mediated immunotherapy: even adenovirus vectors that do not express a transgene, show adjuvant activity and stimulate an antitumor immune response.
Assuntos
Adenoviridae/genética , Terapia Genética/métodos , Vetores Genéticos/imunologia , Neoplasias Hepáticas Experimentais/secundário , Neoplasias Hepáticas Experimentais/terapia , Linfócitos T/imunologia , Animais , Neoplasias Colorretais/imunologia , Citometria de Fluxo , Antígenos de Histocompatibilidade Classe I/análise , Antígenos de Histocompatibilidade Classe II/análise , Humanos , Molécula 1 de Adesão Intercelular/análise , Interleucina-2/genética , Neoplasias Hepáticas Experimentais/imunologia , Ativação Linfocitária , Ratos , Ratos Endogâmicos , Células Tumorais CultivadasRESUMO
The myelin-associated protein, alphaB-crystallin, is considered a candidate autoantigen in multiple sclerosis (MS). In the present study, we examined the potential of alphaB-crystallin to induce experimental autoimmune encephalomyelitis (EAE) in Lewis rats. Attempts to induce EAE with either bovine, rat or murine alphaB-crystallin or alphaB-crystallin peptides consistently failed. Immunization with either autologous rat or murine alphaB-crystallin did not trigger any antigen-specific T cell response. Immunization with bovine alphaB-crystallin or a synthetic peptide representing the cryptic epitope 49-64 did trigger T cell responses but these failed to crossreact with autologous rat alphaB-crystallin. Examination of lymphoid tissues of the Lewis rat revealed constitutive expression of alphaB-crystallin in thymus, spleen, and peripheral lymphocytes. Our data show that in Lewis rats, constitutive lymphoid expression of alphaB-crystallin is associated with a state of nonresponsiveness to autologous alphaB-crystallin that effectively controls the development of EAE in response to this myelin antigen.
Assuntos
Cristalinas/imunologia , Encefalomielite Autoimune Experimental/imunologia , Animais , Autoantígenos/imunologia , Bovinos , Cristalinas/biossíntese , Cristalinas/genética , Cristalinas/metabolismo , Relação Dose-Resposta Imunológica , Hipersensibilidade Tardia/imunologia , Tolerância Imunológica/imunologia , Imunidade Celular/imunologia , Epitopos Imunodominantes/imunologia , Tecido Linfoide/imunologia , Tecido Linfoide/metabolismo , Masculino , Camundongos , Especificidade de Órgãos , Fragmentos de Peptídeos/imunologia , Fosforilação , RNA Mensageiro/metabolismo , Ratos , Ratos Endogâmicos Lew , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Especificidade da Espécie , Linfócitos T/imunologiaRESUMO
Several findings indicate that infectious events play a role in the pathogenesis of multiple sclerosis (MS). At the same time, T-cell autoimmunity to myelin antigens is widely believed to be crucial to the development of MS lesions. Several mechanisms have been put forward to explain the presumed link between microbial infections and myelin-directed autoimmunity. These include molecular mimicry, bystander activation including epitope spreading and superantigenic activation of T cells. Evidence that either one of these mechanisms actually occurs in MS patients, however, is still weak. Also, none of the above mechanisms explain why MS is unique to humans. We propose an alternative link between microbial infection and myelin autoimmunity, which we refer to as 'mistaken self'. In this mechanism, peripheral microbial infections of lymphoid cells prime the human T-cell repertoire not only to microbial antigens but also to the stress protein alpha B-crystallin that is expressed de novo in infected lymphoid cells. Subsequently, stress-induced accumulation of this self antigen in oligodendocytes/myelin can provoke pro-inflammatory responses as the recruited memory T-cell repertoire then mistakes the self protein for a microbial antigen. In this paper we review the currently available evidence that 'mistaken self' centering on alpha B-crystallin represents a powerful source of anti-myelin autoimmunity in a way that is unique to humans.
Assuntos
Autoimunidade , Infecções Bacterianas/complicações , Cristalinas/imunologia , Esclerose Múltipla/etiologia , Bainha de Mielina/imunologia , Linfócitos T/imunologia , Cristalinas/fisiologia , Humanos , Memória Imunológica , Esclerose Múltipla/imunologia , Especificidade de ÓrgãosRESUMO
In the development of multiple sclerosis (MS), (re)activation of infiltrating T cells by myelin-derived Ags is considered to be a crucial step. Previously, alpha B-crystallin has been shown to be an important myelin Ag to human T cells. Since alpha B-crystallin is an intracellular heat shock protein, the question arises at what stage, if any, during lesional development in MS this Ag becomes available for CD4+ T cells. In 3 of 10 active MS lesions, alpha B-crystallin could be detected inside phagocytic vesicles of perivascular macrophages, colocalizing with myelin basic protein and myelin oligodendrocyte glycoprotein (MOG). Although the detectability of MOG in phagosomes is considered as a marker for very recent demyelination, MOG was detected in more macrophages and in more lesions than alpha B-crystallin. The disappearance of alpha B-crystallin from macrophages even before MOG was confirmed by in vitro studies; within 6 h after myelin-uptake alpha B-crystallin disappears from the phagosomes. Alpha B-crystallin-containing macrophages colocalized with infiltrating T cells and they were characterized by expression of MHC class II, CD40, and CD80. To examine functional presentation of myelin Ags to T cells, purified macrophages were pulsed in vitro with whole myelin membranes. These macrophages activated both myelin-primed and alpha B-crystallin-primed T cells in terms of proliferation and IFN-gamma secretion. In addition, alpha B-crystallin-pulsed macrophages activated myelin-primed T cells to the same extent as myelin-pulsed macrophages, whereas myelin basic protein-pulsed macrophages triggered no response at all. These data indicate that, in active MS lesions, alpha B-crystallin is available for functional presentation to T cells early during inflammatory demyelination.
Assuntos
Apresentação de Antígeno/imunologia , Cristalinas/imunologia , Cristalinas/metabolismo , Doenças Desmielinizantes/imunologia , Doenças Desmielinizantes/patologia , Esclerose Múltipla/imunologia , Esclerose Múltipla/patologia , Linfócitos T/imunologia , Autoantígenos/imunologia , Autoantígenos/metabolismo , Antígeno B7-1/metabolismo , Antígenos CD40/metabolismo , Catepsinas/metabolismo , Movimento Celular/imunologia , Doenças Desmielinizantes/metabolismo , Antígenos de Histocompatibilidade Classe II/metabolismo , Humanos , Hidrólise , Inflamação/imunologia , Inflamação/metabolismo , Inflamação/patologia , Macrófagos/enzimologia , Macrófagos/imunologia , Macrófagos/metabolismo , Macrófagos/patologia , Esclerose Múltipla/metabolismo , Proteínas da Mielina/metabolismo , Bainha de Mielina/imunologia , Bainha de Mielina/metabolismo , Fagocitose , Linfócitos T/metabolismo , Linfócitos T/patologiaRESUMO
N-ras and c-myc oncogenes were found to be activated in melanoma. High c-myc expression renders melanoma cell lines sensitive to lysis by natural killer (NK) cells. This effect is mediated by locus-specific downmodulation of HLA-B expression by c-myc. Cell lines with a mutation in the N-ras gene were relatively sensitive to NK cells irrespective of HLA class I expression. These findings indicate that NK cells can kill tumor cells with activated myc or ras oncogenes in various ways, thus providing potential mechanisms to eliminate cancer cells with an activation of these oncogenes.
Assuntos
Citotoxicidade Imunológica , Antígenos de Histocompatibilidade Classe I/metabolismo , Células Matadoras Naturais/imunologia , Melanoma/imunologia , Oncogenes/fisiologia , Northern Blotting , Linhagem Celular , Regulação para Baixo , Regulação Neoplásica da Expressão Gênica , Genes ras/fisiologia , Antígenos HLA-A/metabolismo , Antígenos HLA-B/metabolismo , Antígenos HLA-C/metabolismo , Humanos , Proteínas Proto-Oncogênicas c-myc/fisiologia , RNA Mensageiro/análise , Microglobulina beta-2/biossínteseRESUMO
NK cells kill a wide variety of tumor cells, but usually leave normal cells intact. It was earlier reported that low class I HLA expression can be one of the factors that render target cells relatively susceptible to NK lysis. In this contribution, we show that in human melanomas the class I HLA expression is down-modulated by high expression of transfected c-myc oncogenes. The extent of down-modulation depended on the level of c-myc expression in a dose-dependent way. Taken together, these data suggested to us that one of the results of high c-myc expression could be the induction of a NK susceptible phenotype in melanoma cells. Therefore, we analyzed the effect of c-myc on NK susceptibility. We have found that high expression of transfected c-myc genes indeed converts the melanoma cell lines from poor into good targets for NK cells. IFN-gamma was used to restore the class I HLA expression of the c-myc transfectants, and the resulting cells showed a decreased NK susceptibility. These results suggest the possibility that the c-myc-induced NK susceptibility is mediated by the reduction of class I HLA expression.
Assuntos
Citotoxicidade Imunológica/genética , Regulação Neoplásica da Expressão Gênica , Células Matadoras Naturais/imunologia , Melanoma/genética , Oncogenes , Linhagem Celular , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Antígenos de Histocompatibilidade Classe I/imunologia , Humanos , Imunidade Inata , Interferon gama/farmacologia , Melanoma/imunologia , Oncogenes/efeitos dos fármacos , Transfecção , Células Tumorais CultivadasRESUMO
The c-myc oncogene downregulates class I HLA expression in human melanoma. The major class I HLA antigens are encoded by three loci, A, B, and C, and we investigated whether these loci are suppressed equally by c-myc. In three melanoma cell lines with high c-myc expression, we analyzed mRNA, protein, and cell surface expression of the class I HLA antigens. Whereas the HLA-B locus expression was found to be strongly reduced, the HLA-A locus was expressed normally. Analysis of c-myc-transfected clones of two melanoma cell lines confirmed that c-myc preferentially suppresses the class I HLA-B locus. Immunohistochemical analysis of fresh melanoma lesions also showed that in the tumors the HLA-A loci are expressed normally, while on the majority of tumor cells no HLA-B antigen expression was found. This downregulation may have consequences for the recognition of malignant cells by tumor-infiltrating lymphocytes. Our results predict that HLA-B-restricted cytotoxic T cells will be unable to kill high c-myc-expressing melanoma cells.