RESUMO
In preparation of the therapeutic genetic constructs aimed to the gene-programmed enzymatic transformation of the non-toxic prodrug into toxin within cancer cells the right choice of regulatory elements (promoters and enhancers) is essential. This is widely accepted that the efficiency of the gene therapy constructions is dependent, in particular, on the strength of promoters driving the expression of the therapeutic genes. In this work we demonstrated, using the melanoma-specific promoters and enhancers of human melanoma inhibitory activity and mouse tyrosinase gene, that for the development of cytotoxic effect the promoter strength is not of primary importance. In the case of HSVtk, coding for the herpes simplex virus thymidine kinase, and FCU1, coding for cytosine deaminase/uracil phosphoribosyltransferase hybrid protein genes, their cytotoxic activity was determined by the quantity of the added prodrug.
Assuntos
Citosina Desaminase/genética , Terapia Genética , Melanoma Experimental/genética , Pentosiltransferases/genética , Animais , Citosina Desaminase/uso terapêutico , Genes Transgênicos Suicidas , Humanos , Melanoma Experimental/patologia , Melanoma Experimental/terapia , Camundongos , Pentosiltransferases/uso terapêutico , Pró-Fármacos/uso terapêutico , Regiões Promotoras Genéticas , Simplexvirus/enzimologia , Simplexvirus/genéticaRESUMO
Melanoma is one of the most malignant tumors, aggressively metastasizing by lymphatic and hematogenous routes. Due to the resistance of melanoma cells to many types of chemotherapy, this disease causes high mortality rate. High hopes are pinned on gene therapeutic approaches to melanoma treatment. At present, one of the main problems of the efficient use of the post-genomic generation therapeutic means is the lack of optimal techniques of delivery of foreign genetic material to the patient's target cells. Surface specific markers of melanoma cells can be considered as promising therapeutic targets. This review describes currently known melanoma specific receptors and its stem cells, as well as contains data on melanoma antigens presented on the cell surface by major histocompatibility complex proteins. The ability of surface proteins to internalize might be successfully used for the development of methods of targeted delivery of gene therapeutic constructs. In conclusion, a concept of multilevel gene therapy and the possible role therein of surface determinants as targets of gene systems delivery to the tumor are discussed.
Assuntos
Moléculas de Adesão Celular/metabolismo , Terapia Genética/métodos , Melanoma/terapia , Células-Tronco Neoplásicas/metabolismo , Receptores de Superfície Celular/metabolismo , Neoplasias Cutâneas/terapia , Apresentação de Antígeno , Antígenos de Neoplasias/imunologia , Antígenos de Neoplasias/metabolismo , Humanos , Complexo Principal de Histocompatibilidade , Melanócitos/imunologia , Melanócitos/metabolismo , Melanoma/imunologia , Melanoma/metabolismo , Proteínas Proto-Oncogênicas c-met/metabolismo , Receptor Tipo 1 de Melanocortina/metabolismo , Receptores de Fatores de Crescimento/metabolismo , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/metabolismo , Linfócitos T Citotóxicos/imunologiaRESUMO
Polyfunctional protein PML contributes significantly in vital activity of cell. 11 isoforms of PML differ from one another by C-terminal domain. In spite of intensive research into the protein, the role of the isoforms in cellular processes remains obscure. In addition, the literature contains various names of the isoforms. The goal of this work was to review the structure of the PML gene, variants of alternative splicing of mPNA, and domain organization of corresponding protein forms. The PML isoforms were classified and functional specificity of each PML isoform was characterized: contribution to gene transcription, contribution to cell apoptosis, cell growth, immune response, formation of nuclear bodies.
Assuntos
Processamento Alternativo , Regulação Leucêmica da Expressão Gênica , Leucemia Promielocítica Aguda/genética , Proteínas Nucleares/classificação , Proteínas Nucleares/genética , Fatores de Transcrição/classificação , Fatores de Transcrição/genética , Proteínas Supressoras de Tumor/classificação , Proteínas Supressoras de Tumor/genética , Dedos de Zinco , Animais , Apoptose/genética , Processos de Crescimento Celular/genética , Núcleo Celular/metabolismo , Humanos , Proteínas Nucleares/metabolismo , Proteína da Leucemia Promielocítica , Isoformas de Proteínas/classificação , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Estrutura Terciária de Proteína/fisiologia , Fatores de Transcrição/metabolismo , Proteínas Supressoras de Tumor/metabolismoRESUMO
The level of ferritin in serum is known to be increased frequently in most human cancers. Ferritin consists of the heavy and light chains, encoded by FTL and FTH genes. The analysis of the EST database showed that the level of FTL and FTH mRNA is decreased in lung squamous cell carcinomas as compared to the normal tissues, no change in the mRNA level was observed in clear cell renal cell carcinoma. Using real-time PCR we estimated the mRNA level of these genes in primary tumors. It was shown significant and frequent decrease of FTL and FTH mRNA level in lung squamous cell carcinoma: on the average by 11 and 9 times in 83% (33/40) and 73% (11/15) of cases, respectively. In clear cell renal cell carcinoma the changes were not so marked both with respect to the level of decrease (on the average 6 and 3 times) and to its frequency (58 and 27%). In the present work it has been shown for the first time that the FTL mRNA is frequently down-regulated even at the early stages of lung squamous cell carcinoma in all studied samples. This fact permits to consider this gene as potential oncomarker of early diagnosis. The FTL mRNA content may be quantified by non-concurrent hybridization on expression DNA microarrays. The possible causes of a serum ferritin increase in lung cancer and renal cancer are discussed.
Assuntos
Apoferritinas/biossíntese , Biomarcadores Tumorais/biossíntese , Carcinoma de Células Renais/metabolismo , Carcinoma de Células Escamosas/metabolismo , Ferritinas/biossíntese , Regulação Neoplásica da Expressão Gênica , Neoplasias Renais/metabolismo , Neoplasias Pulmonares/metabolismo , Proteínas de Neoplasias/biossíntese , Adulto , Idoso , Apoferritinas/genética , Biomarcadores Tumorais/genética , Carcinoma de Células Renais/diagnóstico , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/patologia , Carcinoma de Células Escamosas/diagnóstico , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patologia , Bases de Dados Genéticas , Feminino , Ferritinas/genética , Perfilação da Expressão Gênica , Genes Neoplásicos , Humanos , Neoplasias Renais/diagnóstico , Neoplasias Renais/genética , Neoplasias Renais/patologia , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Masculino , Pessoa de Meia-Idade , Proteínas de Neoplasias/genética , Análise de Sequência com Séries de Oligonucleotídeos , OxirredutasesRESUMO
Survivin (BIRC5) is one of the members of IAP-family apoptosis inhibitors. The BIRCS gene is expressed in most human embryonic tissues and malignant tumors but not in normal differentiated tissues of adult human. It was suggested that BIRC5 proteins inhibit apoptosis and play an essential role in tumorigenesis, makings surviving an attractive target for anticancer therapy. The mechanisms regulating level of survivin are not completely understood. It was supposed that natural inhibitors of survivin, namely SMAC and PML, play an important role in these processes. Using RT-PCR and immunoblotting we analyzed the transcription level of BIRC5, SMAC and PML genes and content of corresponding proteins in normal and tumor human tissues in non-small cell lung cancer and esophageal squamous cell carcinoma. It was demonstrated that BIRC5 is transcribed only in tumor tissues, whereas expression levels of SMAC and PML are the same in normal and tumor tissues. The contents of proteins correspond to levels of mRNA of the respective genes. Thus the increase of level of survivin in tumor tissues is not the result of decrease in content of its inhibitors SMAC and PML, as their content in tumor and normal cells is the same.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/metabolismo , Neoplasias Esofágicas/metabolismo , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/metabolismo , Proteínas Associadas aos Microtúbulos/biossíntese , Proteínas Mitocondriais/biossíntese , Proteínas de Neoplasias/biossíntese , Neoplasias de Células Escamosas/metabolismo , Proteínas Nucleares/biossíntese , Fatores de Transcrição/biossíntese , Proteínas Supressoras de Tumor/biossíntese , Adulto , Idoso , Apoptose , Proteínas Reguladoras de Apoptose , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/patologia , Feminino , Humanos , Proteínas Inibidoras de Apoptose , Peptídeos e Proteínas de Sinalização Intracelular/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Masculino , Proteínas Associadas aos Microtúbulos/antagonistas & inibidores , Proteínas Associadas aos Microtúbulos/genética , Pessoa de Meia-Idade , Proteínas Mitocondriais/genética , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/genética , Neoplasias de Células Escamosas/genética , Neoplasias de Células Escamosas/patologia , Proteínas Nucleares/genética , Proteína da Leucemia Promielocítica , Survivina , Fatores de Transcrição/genética , Proteínas Supressoras de Tumor/genéticaRESUMO
Chromosomal and genome abnormalities of 3p are frequent events in many epithelial tumours, including lung cancer. Several critical regions with high frequency of hemi--and homozygous deletions in tumours were detected on 3p and more then 20 different cancer-related genes were identified in 3p21.3 locus. Real-time PCR was used to measure mRNA level of tumour-suppressor genes and candidates in 3p21.3 (RBSP3/CTDSPL, NPRL2/G21, RASSF1A, ITGA9, HYAL1 and HYAL2 in basic types of non-small cell lung cancer (NSCLC)--squamous cell lung cancer (SCC) and lung adenocarcinoma (AC). Significant (from 2 to 100 times) and frequent (from 44 to 100%) mRNA level decrease was shown in NSCLC. Level and frequency of mRNA decrease for all genes depended on histological type of NSCLC. Down-regulation of RASSF1A and ITGA9 was associated significantly with AC progression, the same tendency was found for genes RBSP3/CTDSPL, NPRL2/G21, HYAL1 and HYAL2. On the contrary, down-regulation of all genes in SCC was not associated with clinical stages, tumor cells differentiation and metastases in lymph nodes. Significant decrease of RBSP3/CTDSPL, NPRL2/G21, ITGA9, HYAL1 and HYAL2 mRNA levels (on average, 5-13 times) with high frequency (83-100%) was already shown at the first stage of SCC. Simultaneous decrease of all six genes mRNA level was found in the same tumor samples and was not depended on their localization on 3p21.3 and functions of the proteins. Spearman's correlation coefficient r(s) was from 0.63 to 0.91, P < 0.001. Co-regulation of gene pairs ITGA9 and HYAL2, HYAL1 and HYAL2, which mediate cell-cell adhesion and cell-matrix interaction, was suggested based on the obtained data. It was shown that genetic and epigenetic mechanisms were important for down-regulation of RBSP3/CTDSPL and ITGA9 genes. These results supported the hypothesis on simultaneous inactivation of cluster cancer-related genes in extended 3p21.3 locus during development and progression of lung cancer and other epithelial tumors. Significant and frequent decrease of mRNA level of six genes in SCC could be important for development of specific biomarker sets for early SCC diagnosis and new therapeutic approaches/strategies for NSCLC.
Assuntos
Biomarcadores Tumorais/biossíntese , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Moléculas de Adesão Celular/biossíntese , Regulação Neoplásica da Expressão Gênica , Hialuronoglucosaminidase/biossíntese , Neoplasias Pulmonares/metabolismo , Proteínas Supressoras de Tumor/biossíntese , Biomarcadores Tumorais/genética , Carcinoma Pulmonar de Células não Pequenas/diagnóstico , Carcinoma Pulmonar de Células não Pequenas/genética , Moléculas de Adesão Celular/genética , Cromossomos Humanos Par 3/genética , Cromossomos Humanos Par 3/metabolismo , Regulação para Baixo , Epigênese Genética/genética , Feminino , Proteínas Ligadas por GPI , Humanos , Hialuronoglucosaminidase/genética , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/genética , Masculino , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , RNA Neoplásico/biossíntese , RNA Neoplásico/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteínas Supressoras de Tumor/genéticaRESUMO
The KLRB1 gene encodes the CD161 receptor of natural killer cells (NK-cells). The gene is also expressed in the NKT-cells. The two cell types are cytotoxic and capable of recognizing and eliminating various cell species, e.g. tumorous, virus-infected, and allotransplants. The biological function of human CD161 is still insufficiently understood; probably it is involved in regulation of the cytotoxic functions of the cells and in regulation of cytokine production. Because immune system, in particular, the activity of cytotoxic cells, is suppressed in most cancer patients, it was suggested that the KLRB1 expression might be suppressed in cancerous cells. The results of this work demonstrated that the transcription of the KLRB1 was suppressed in tumor tissues in 68% patients with nonsmall-lung-cancer (p < 0.0001) and 57% patients with esophageal squamous-cell carcinoma (p = 0.0003). High frequency of the KLRB1 transcription suppression upon cancers makes it possible to use this parameter as a highly informative marker of lung and esophageal cancers.
Assuntos
Antígenos de Superfície/genética , Biomarcadores Tumorais/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Neoplasias Esofágicas/genética , Regulação Neoplásica da Expressão Gênica , Lectinas Tipo C/genética , Neoplasias Pulmonares/genética , Regulação para Baixo , Feminino , Humanos , Masculino , Subfamília B de Receptores Semelhantes a Lectina de Células NK , Transcrição Gênica , Células Tumorais CultivadasRESUMO
Secretory polymeric immunoglobulins (IgA dimers and IgM pentamers) are unique in that, apart from L- and H-chains, they contain J-chains responsible for their oligomerization. These antibodies are part of the local adaptive immune system acting on mucosa membranes of the respiratory and digestive systems as the first protection barrier to potential infectious agents. Secretory polymeric immunoglobulins are produced by highly specific B-cells and actively transported to the surface of mucosa membrane through epithelium cells. Therefore, their synthesis and J-chain content are dependent upon epithelium translocation function and condition that are markedly affected by tumorous transformation. Here, we used RT-PCR and immunoblotting to study of the J-chain content and its mRNA expression level in normal and tumorous tissues in lung squamous cell cancer and adenocarcinoma at various stages of disease progression.
Assuntos
Adenocarcinoma/imunologia , Carcinoma de Células Escamosas/imunologia , Cadeias J de Imunoglobulina/metabolismo , Neoplasias Pulmonares/imunologia , Adenocarcinoma/química , Carcinoma de Células Escamosas/química , Regulação para Baixo , Expressão Gênica , Humanos , Immunoblotting , Cadeias J de Imunoglobulina/análise , Cadeias J de Imunoglobulina/genética , Neoplasias Pulmonares/química , RNA Mensageiro/análise , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Changes in WIFI expression, an extracellular inhibitor of Wnt pathway, in non-small cell lung carcinomas were analyzed. Frequent (67% cases) suppression of WIFI transcript in non-small cell lung carcinomas were found. Our results, together with previously published data, suggest that inhibition of WIFI expression often occurs in squamous cell carcinomas and is less typical of adenocarcinomas. It was also found that a decrease in the WIFI transcript in tumors is parallel to concomitant suppression of the WIFI protein level. Our results provide further evidence that the WIFI suppression is a frequent event in the lung carcinogenesis, which might lead to disregulation of Wnt signaling pathway and contribute to tumor progression.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/biossíntese , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/metabolismo , Proteínas de Neoplasias/biossíntese , RNA Mensageiro/biossíntese , RNA Neoplásico/biossíntese , Proteínas Repressoras/biossíntese , Proteínas Adaptadoras de Transdução de Sinal/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Regulação para Baixo , Feminino , Humanos , Neoplasias Pulmonares/genética , Masculino , Proteínas de Neoplasias/genética , RNA Mensageiro/genética , RNA Neoplásico/genética , Proteínas Repressoras/genética , Transdução de Sinais , Proteínas Wnt/metabolismoRESUMO
Alterations in the c-Met/HGF system are frequently observed in various types of tumors and can directly influence tumor progression. In particular, the c-Met/HGF system can influence progression of lung tumors. The altered c-Met and HGF expression occurs in non-small-cell lung carcinomas. In this work we analyzed changes in c-Met and HGF expression in non-small-cell lung carcinomas by comparing expression levels with those in adjacent non-malignant tissues using semi-quantitative PCR. The c-Met transcript was detected in 50% of squamous cell carcinoma samples with an elevated level observed in 2 out of 25 cases (8%). HGF expression was detected only in two squamous cell carcinomas. At the same time, the c-Met transcript was observed in all 5 studied adenocarcinoma samples with an increased level compared to adjacent non- malignant tissue in 3 cases. HGF transcript was found in 2 adenocarcinoma samples. Therefore, c-Met rather than HGF transcript is frequently observed in non-small-cell lung carcinomas, especially in adenocarcinomas. According to the results of other studies, the c-Met transcript can serve as an indicator of the aggressive behavior and progression of non-small-cell lung carcinomas.
Assuntos
Biomarcadores Tumorais/biossíntese , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Regulação Neoplásica da Expressão Gênica , Fator de Crescimento de Hepatócito/biossíntese , Neoplasias Pulmonares/metabolismo , Proteínas Proto-Oncogênicas c-met/biossíntese , Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Biomarcadores Tumorais/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma de Células Escamosas , Fator de Crescimento de Hepatócito/genética , Humanos , Neoplasias Pulmonares/genética , Proteínas Proto-Oncogênicas c-met/genética , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , RNA Neoplásico/biossíntese , RNA Neoplásico/genética , Regulação para CimaRESUMO
Lung cancer is one of the most frequent neoplasia in the Russia, the United States and Europe. This cancer is associated with functional activity changes of many genes. In the present study TIMP3, DAPK1 and AKR1B10 genes transcription analysis of squamous cell lung cancer specimens was carried out using reverse transcription-PCR. Substantial increasing of AKR1B10 transcription level is revealed in 80% tumor samples. TIMP3 and DAPK1 transcription level is considerably decreased in 76 and 72% tumor specimens, accordingly. These results may point out that all three genes are important for squamous cell lung cancer tumorogenesis while AKR1B10 is potential oncogene whereas TIMP3 and DAPK1 are potential tumor suppressor genes. We suggest that revealed substantial transcription level-changes of investigated genes may be used for oncodiagnostics.
Assuntos
Aldeído Redutase/genética , Proteínas Reguladoras de Apoptose/genética , Proteínas Quinases Dependentes de Cálcio-Calmodulina/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Neoplasias Pulmonares/genética , Inibidor Tecidual de Metaloproteinase-3/genética , Adulto , Idoso , Aldeído Redutase/biossíntese , Aldo-Ceto Redutases , Proteínas Reguladoras de Apoptose/biossíntese , Proteínas Quinases Dependentes de Cálcio-Calmodulina/biossíntese , Carcinoma Pulmonar de Células não Pequenas/enzimologia , Proteínas Quinases Associadas com Morte Celular , Indução Enzimática/genética , Repressão Enzimática/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Pulmonares/enzimologia , Masculino , Pessoa de Meia-Idade , Inibidor Tecidual de Metaloproteinase-3/biossíntese , Transcrição GênicaAssuntos
Neovascularização Fisiológica/fisiologia , Fator A de Crescimento do Endotélio Vascular/genética , Idoso , Processamento Alternativo , Animais , Doenças Cardiovasculares/terapia , Amplificação de Genes , Humanos , Mamíferos , Reação em Cadeia da Polimerase , Isoformas de Proteínas/genética , Reação em Cadeia da Polimerase Via Transcriptase ReversaAssuntos
Proteínas de Ligação a DNA , Expressão Gênica , Hematopoese/genética , Proteínas de Homeodomínio , Animais , Sequência de Bases , Proteína Quinase CDC2/genética , Diferenciação Celular/genética , Divisão Celular/genética , Células Cultivadas , Clonagem Molecular , Primers do DNA , DNA Complementar , Hematopoese/fisiologia , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Proteínas/genética , Proteínas Proto-Oncogênicas/genética , Receptores Proteína Tirosina Quinases/genética , Tirosina Quinase 3 Semelhante a fmsRESUMO
In order to elucidate the nature of endogenous proton conductance of rat liver inner mitochondrial membrane, the dependence of the rate of Ca2+ transport on pH was studied. It was found that the inhibiting effect of H+ is independent of protonation of functional groups of hypothetical Ca2+ carrier, but results from electrogenic transfer of H+ across the membrane, which is highly permeable for the proton. The adsorption of H+ by mitochondria is inhibited by ruthenium red and other specific inhibitors of Ca2+ transport. It is concluded that endogenous proton conductance of the inner mitochondrial membrane depends on the functioning of the same transport system essential for membrane permeability for Ca2+ and other bivalent cations. The correlation observed between the rates of H+ and Ca2+ transport in mitochondria and the ratio of cation mobilities in aqueous solutions is in favour of a "porous" mechanism of cation transport across the mitochondrial membrane.
