Assuntos
Catecolaminas , Doenças em Gêmeos/diagnóstico , Ganglioneuroma/diagnóstico , Neuroblastoma/diagnóstico , Neoplasias Retroperitoneais/diagnóstico , Gêmeos Monozigóticos , Catecolaminas/metabolismo , Pré-Escolar , Doenças em Gêmeos/genética , Feminino , Ganglioneuroma/genética , Ganglioneuroma/metabolismo , Humanos , Lactente , Masculino , Pessoa de Meia-Idade , Neuroblastoma/genética , História Reprodutiva , Neoplasias Retroperitoneais/genética , Neoplasias Retroperitoneais/metabolismo , Gêmeos Monozigóticos/genéticaRESUMO
PURPOSE: Decision making regarding the urethra before and after radical cystectomy due to urothelial carcinoma has always been controversial. To determine whether anterior urethra sparing cystoprostatectomy for bladder cancer is an oncologically-safe procedure, we evaluated the long-term oncologic clinical outcome. PATIENTS AND METHODS: A total of 51 male patients with cTa-4N0-2M0 bladder cancer were treated with anterior urethra sparing cystoprostatectomy and simultaneous urinary diversion between 2000 and 2013, and underwent follow up for 4 months or more. We assessed differences in the perioperative outcomes, oncologic outcomes and recurrence rates according to the urinary diversion. RESULTS: The median patient age and follow-up period were 66 years and 35 months, respectively. The 5- and 10-year recurrence free survival (RFS) rates in ileal conduit (IC) group vs. orthotopic neobladder reconstruction (NB) group were 45.0 and 20.3% vs. 39.3 and 19.6%, respectively. Likewise, the 5- and 10-year disease specific survival (DSS) were 52.7 and 32.1% vs. 39.3 and 29.5%, respectively. Multivariate analysis revealed two independent prognostic factors for RFS and DSS, including age at surgery and lymph node status. Local recurrence in the remnant anterior urethra occurred in only 1 patient (2.0%) at 57 months after surgery. CONCLUSIONS: Our long-term data show that anterior urethra sparing cystoprostatectomy is an oncologically-safe procedure regardless of the type of urinary diversion in a subset of carefully selected patients with bladder cancer without evidence of urothelial carcinoma in the urethra/bladder neck and urethral surgical margin.
RESUMO
BACKGROUND: The sarcomatoid variant of metastatic renal cell carcinoma (RCC) has often an aggressive course and a poor prognosis, particularly when accompanied with brain metastasis. CASE REPORT: We describe the case of a patient with sarcomatoid variant RCC in whom brain metastasis was observed as a new lesion during treatment with temsirolimus, despite other extracerebral metastatic lesions being well-controlled and progression-free. RESULTS: This discrepancy between the effectiveness of temsirolimus for extracerebral metastases and the simultaneous progression of brain metastases of RCC raises a concern that while vascular endothelial growth factor (VEGF)-targeted therapy may have clinical efficacy, it may also carry a risk for new brain metastases due to weakening of the structure of the blood brain barrier. CONCLUSION: This case indicates that computed tomography monitoring of the brain should be regularly performed during VEGF-targeted therapy in patients with sarcomatoid variant RCC, even if brain metastases are absent and extracerebral metastatic lesions are well controlled.
Assuntos
Antineoplásicos/uso terapêutico , Neoplasias Encefálicas/secundário , Carcinoma de Células Renais/tratamento farmacológico , Neoplasias Renais/tratamento farmacológico , Metástase Neoplásica/prevenção & controle , Sirolimo/análogos & derivados , Carcinoma de Células Renais/patologia , Humanos , Neoplasias Renais/patologia , Sirolimo/uso terapêuticoRESUMO
The functional significance of Wnt antagonist DKK1 has not been investigated in renal cell carcinoma (RCC). Therefore, we hypothesized that DKK1 may be a tumor suppressor gene and is epigenetically silenced, thus decreased DKK1 may cause progression of RCC. To assess the function of DKK1, we established stable DKK1 transfected cells and monitored them regarding cell viability, colony formation, apoptosis, cell cycle, and invasive capability. RCC cell lines had decreased levels of DKK1, which were increased after treatment with 5-Aza-2'-deoxycytidine and trichostatin A. In chromatin immunoprecipitation assay, the level of dimethyl H3K9 and trimethyl H3K27 was decreased after 5-Aza-2'-deoxycytidine/trichostatin A treatment in RCC cell lines. Increased methylation was also associated with higher pathological stages in primary RCC tissues. T-cell factor/lymphoid enhancer factor activity and nuclear beta-catenin expression were not changed in DKK1 transfectants. Also the expression of cyclinD1 and c-Myc was not changed in DKK1 transfectants. These results suggest that DKK1 may not be involved in the beta-catenin dependent pathway. We also evaluated the expression of various related genes. Cleaved caspase3, p53, p21 and puma expression were significantly upregulated in the DKK1 transfected cells. The population of apoptotic cells was increased in stable DKK1 cells and tumor growth suppression was also observed in nude mice with DKK1 transfected cells. In conclusion, this is the first report to show that DKK1 expression is epigenetically silenced in kidney cancer and its reexpression induces apoptosis and cell cycle arrest in RCC.
Assuntos
Apoptose , Carcinoma de Células Renais/patologia , Proliferação de Células , Genes Supressores de Tumor , Peptídeos e Proteínas de Sinalização Intercelular/fisiologia , Neoplasias Renais/patologia , Proteínas Wnt/antagonistas & inibidores , Antimetabólitos Antineoplásicos/farmacologia , Proteínas Reguladoras de Apoptose/genética , Proteínas Reguladoras de Apoptose/metabolismo , Azacitidina/análogos & derivados , Azacitidina/farmacologia , Western Blotting , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/metabolismo , Caspase 3/genética , Caspase 3/metabolismo , Imunoprecipitação da Cromatina , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Metilação de DNA , Decitabina , Epigênese Genética , Feminino , Humanos , Ácidos Hidroxâmicos/farmacologia , Técnicas Imunoenzimáticas , Neoplasias Renais/genética , Neoplasias Renais/metabolismo , Masculino , Regiões Promotoras Genéticas/genética , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , RNA Mensageiro/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células Tumorais Cultivadas , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismoRESUMO
Wnt inhibitory factor-1 (WIF-1) has been identified as one of the secreted antagonists that bind Wnt protein. WIF-1 has been described as a tumor suppressor in various types of cancer. However, the molecular function of WIF-1 gene has never been examined in human renal cell carcinoma (RCC). Therefore, we hypothesized that WIF-1 functions as a tumor suppressor gene and overexpression of this gene may induce apoptosis and inhibit tumor growth in RCC cells. Immunohistochemistry and real-time reverse transcription-PCR revealed that WIF-1 was significantly downregulated in RCC samples and RCC cell lines, respectively. Bisulfite sequencing of the WIF-1 promoter region in RCC cell lines showed it to be densely methylated, whereas there was no methylation of WIF-1 promoter in normal kidney. Significant inhibition of cell growth and colony formation in WIF-1-transfected cells compared with controls were observed. WIF-1 transfection significantly induced apoptosis and suppressed in vivo tumor growth. Also, Wnt signaling activity and beta-catenin expression were reduced by WIF-1 transfection. In conclusion, this is the first report documenting that the WIF-1 is downregulated by promoter methylation and functions as a tumor suppressor gene by inducing apoptosis in RCC cells.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Carcinoma de Células Renais/genética , Genes Supressores de Tumor , Neoplasias Renais/genética , Proteínas Repressoras/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Apoptose/fisiologia , Carcinoma de Células Renais/metabolismo , Metilação de DNA/genética , Regulação para Baixo , Feminino , Citometria de Fluxo , Expressão Gênica , Humanos , Imuno-Histoquímica , Neoplasias Renais/metabolismo , Camundongos , Camundongos Nus , Regiões Promotoras Genéticas/genética , Proteínas Repressoras/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/fisiologia , Análise Serial de Tecidos , Transfecção , Proteínas Wnt/fisiologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
PURPOSE: Wnt/beta-catenin signaling is involved in renal cancer. DKK2, a Wnt antagonist, is silenced in some cancers, although its function has not been investigated. We hypothesized that DKK2 may be epigenetically silenced and inhibits progression of renal cell carcinoma (RCC). EXPERIMENTAL DESIGN: RCC cell lines and a normal kidney cell line were used for methylation and chromatin immunoprecipitation assays. To assess various functions of DKK2, we established stable DKK2-transfected cells and examined them with regard to cell viability, colony formation, apoptosis, cell cycle, and invasive capability. A total of 52 patients with confirmed conventional RCC were enrolled in this study. RESULTS: RCC cell lines had decreased levels of DKK2, which were significantly increased after treatment with 5-Aza-2'-deoxycytidine alone or 5-Aza-2'-deoxycytidine and trichostatin A. In chromatin immunoprecipitation assay, the levels of acetyl H3, acetyl H4, and dimethylated H3K4 were decreased, whereas the level of dimethylated H3K9 was increased in RCC cell lines compared with HK2 cells. Increased methylation in RCC tissues was associated with higher grades, pathologic stages, and pathologic tumor in RCC. Functional analysis showed that the numbers of viable A498 cells were significantly decreased in DKK2-transfected cells compared with mock cells. The number of apoptotic cells and S/G(2)-M phase cells was significantly increased and decreased after DKK2 transfection, respectively. Corresponding to these results, Bcl2 and cyclin D1 expression were also decreased in DKK2-overexpressing cells. CONCLUSION: DKK2 is epigenetically silenced by methylation in higher grades and stages of RCC. These results suggest that DKK2 inhibits renal cancer progression through apoptotic and cell cycle pathways.
Assuntos
Apoptose , Ciclo Celular , Progressão da Doença , Inativação Gênica , Peptídeos e Proteínas de Sinalização Intercelular/genética , Neoplasias Renais/genética , Neoplasias Renais/patologia , Adulto , Idoso , Azacitidina/análogos & derivados , Azacitidina/farmacologia , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/metabolismo , Carcinoma de Células Renais/patologia , Imunoprecipitação da Cromatina , Decitabina , Feminino , Humanos , Ácidos Hidroxâmicos/farmacologia , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Neoplasias Renais/metabolismo , Masculino , Pessoa de Meia-Idade , Proteínas Wnt/antagonistas & inibidoresRESUMO
PURPOSE: A single nucleotide polymorphism (-938C/A, rs2279115) was found in the bcl2 gene, whose -938A allele is significantly associated with increased Bcl2 expression compared with that of the C allele. Bcl2 up-regulation was reported to be associated with longer survival in patients with renal cancer. However, to our knowledge there is currently no information on the role of the bcl2-938C/A single nucleotide polymorphism in renal cell carcinoma cases. Therefore, we investigated the polymorphism at the bcl2 -938C/A site and its effects on clinical characteristics in patients with renal cell carcinoma. MATERIALS AND METHODS: We genotyped the bcl2-938C/A single nucleotide polymorphism in 216 patients with renal cancer, and in 209 healthy age and gender matched controls. We also investigated the relationship between the bcl2 -938C/A polymorphism, Bcl2 expression, proliferation and apoptosis status in renal cell carcinoma tissues using immunohistochemistry and TUNEL assay. The association of the bcl2 -938C/A single nucleotide polymorphism with survival in patients with renal cell carcinoma was also analyzed by Kaplan-Meier curves. RESULTS: Survival in Bcl2 positive cases was significantly longer than in negative cases. On univariate and multivariate analyses the bcl2 -938CC genotype was independently associated with poor prognosis. Kaplan-Meier analysis showed that survival in patients with CC genotypes was significantly worse than in those with CA+AA genotypes. CC genotype carriers had significantly lower Bcl2 expression and higher proliferative activity in renal cancer tissues than CA+AA genotype carriers. CONCLUSIONS: To our knowledge this is the first report to show that the bcl2 -938C/C genotype has worse prognosis and lower survival in patients with renal cell carcinoma. In addition, the bcl2 -938C/A single nucleotide polymorphism was shown to be an independent adverse prognostic factor for renal cell carcinoma.
Assuntos
Carcinoma de Células Renais/genética , Carcinoma de Células Renais/mortalidade , Proteínas de Transporte/genética , Neoplasias Renais/genética , Neoplasias Renais/mortalidade , Polimorfismo de Nucleotídeo Único , Proteínas Reguladoras de Apoptose , Feminino , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Prognóstico , Taxa de SobrevidaRESUMO
BACKGROUND: Epigenetic silencing of several wingless-type mouse mammary tumor virus integration site (Wnt) pathway-related genes has been reported in renal cancer. Except for the T-cell factor 4 gene TCF4, there are no reports regarding Wnt pathway gene polymorphisms in renal cancer. Therefore, the authors of this report hypothesized that the polymorphisms in Wnt signaling genes may be risk factors for renal cancer. METHODS: In total, 210 patients (145 men and 65 women) with pathologically confirmed renal cell carcinoma (RCC) and 200 age-matched and sex-matched control individuals were enrolled in this study. We genotyped 14 single nucleotide polymorphisms (SNPs) in 6 genes. including Dickkopf 2 (DKK2) (reference SNP identification number 17037102 [rs17037102], rs419558, and rs447372), DKK3 (rs3206824, rs11022095, rs1472189, rs7396187, and rs2291599), DKK4 (rs2073664), secreted frizzled-related protein 4 (sFRP4) (rs1802073 and rs1802074), mothers against decapentaplegic homolog (SMAD) family member 7 or SMAD7 (rs12953717), and disheveled associated activator of morphogenesis 2 or DAAM2 (rs6937133 and rs2504106) using polymerase chain reaction-restriction fragment length polymorphism analysis and direct sequencing in the patients with RCC and in the healthy, age-matched control group. The relations also were tested between these polymorphisms and clinicopathologic data, including sex, tumor grade, tumor stage, lymph node involvement, distant metastasis, and overall survival. RESULTS: A significant decrease in the frequency of the guanine/adenine (G/A) + A/A genotypes in the DKK3 codon 335 rs3206824 was observed in the patients with RCC compared with the control group. The frequency of the rs3206824 (G/A) A-rs7396187 (guanine/cytosine [G/C]) C haplotype was significantly lower in patients with RCC compared with other haplotypes. In addition, DKK3 rs1472189 cytosine/thymine (C/T) was associated with distant metastasis, and, DKK2 rs17037102 G-homozygous patients had a decreased risk for death in multivariate Cox regression analysis. CONCLUSIONS: To the authors' knowledge, this is the first report documenting that DKK3 polymorphisms are associated with RCC and that the DKK2 rs17037102 polymorphism may be a predictor for survival in patients with RCC after radical nephrectomy.
Assuntos
Carcinoma de Células Renais/genética , Peptídeos e Proteínas de Sinalização Intercelular/genética , Neoplasias Renais/genética , Polimorfismo de Nucleotídeo Único , Proteínas Wnt/antagonistas & inibidores , Proteínas Adaptadoras de Transdução de Sinal , Animais , Carcinoma de Células Renais/mortalidade , Estudos de Casos e Controles , Quimiocinas , Feminino , Genótipo , Humanos , Neoplasias Renais/mortalidade , Masculino , Camundongos , Pessoa de Meia-Idade , Proteínas Wnt/genética , beta Catenina/metabolismoRESUMO
PURPOSE: Approximately 10% to 26% of patients show biochemical failure after radical prostatectomy or radiation therapy. The importance of cell cycle and apoptosis pathways in prostate cancer has been reported. However, to our knowledge there is currently no information on the role of apoptosis and cell cycle related gene polymorphisms in prostate cancer cases. We investigated several polymorphisms related to the cell cycle and apoptosis, and their role in biochemical failure after radical prostatectomy. MATERIALS AND METHODS: We genotyped 6 single nucleotide polymorphisms in 6 genes, including p53 (rs1042522), p21 (rs1801270), MDM2 (rs2279744), PTEN (rs701848), GNAS1 (rs7121) and bcl2 (rs2279115), using polymerase chain reaction-restriction fragment length polymorphism and direct DNA sequencing in 140 patients with prostate cancer and 167 age matched controls. The association of polymorphic variants with prostate specific antigen failure in patients with prostate cancer was analyzed by Kaplan-Meier curves. RESULTS: A significant increase in the frequency of the C/C genotype of GNAS1 rs7121 was observed in patients compared with controls. Interestingly we found a significant difference in biochemical recurrence-free time between the bcl2 C/C and C/A+A/A genotypes. It was also noted that the bcl2 C/C genotype was an independent risk factor for biochemical recurrence after radical prostatectomy on multivariate analysis. There was no statistical difference in the genotype distributions of the other genes between patients and controls. CONCLUSIONS: To our knowledge this is the first report documenting that bcl2 promoter region -938 C/C genotype carriers more frequently show biochemical recurrence than -938C/A+A/A carriers.
Assuntos
Recidiva Local de Neoplasia/genética , Polimorfismo Genético , Prostatectomia , Neoplasias da Próstata/genética , Neoplasias da Próstata/cirurgia , Proteínas Proto-Oncogênicas c-bcl-2/genética , Idoso , Humanos , Masculino , Recidiva Local de Neoplasia/sangue , Antígeno Prostático Específico/sangue , Neoplasias da Próstata/sangue , Fatores de RiscoRESUMO
OBJECTIVE: To determine the combined effects of nerve growth factor (NGF) and vascular endothelial growth factor (VEGF) on regeneration of the bladder acellular matrix graft (BAMG) in spinal cord injury (SCI)-mediated neurogenic bladder in rats. MATERIALS AND METHODS: In all, 40 female Sprague-Dawley rats were used. At 8 weeks after spinalization surgery (neurogenic bladder), they were divided into five groups consisting of untreated controls and those whose bladders were injected with either no growth factor, NGF (2 microg/rat), VEGF (2 microg/rat) or both at partial BAMG replacement surgery. After 8 weeks, bladder function was assessed by urodynamic studies and the bladders were harvested for histological examination. Smooth muscle induction, collagen and nerve fibre regeneration were assessed immunohistochemically using antibodies to smooth muscle actin (alpha-actin), Masson's trichrome and protein gene product 9.5, respectively. RESULTS: Bladder capacity and compliance were significantly increased in all BAMG groups 8 weeks after surgery compared with that before bladder replacement surgery. Bladder capacity and compliance were much higher in the VEGF and NGF combined group than in the control, or NGF and VEGF alone groups. There was no significant difference in the residual volume ratio among all groups. CONCLUSIONS: This is the first report showing that NGF has a significant synergistic effect on the development, differentiation and functional restoration of the BAMG when administered with VEGF in neurogenic bladder. Our results indicate that NGF may be a useful cytokine for enhancing the regeneration of a functional bladder following acellular matrix grafting in a neurogenic rat model.
Assuntos
Fator de Crescimento Neural/farmacologia , Regeneração/efeitos dos fármacos , Traumatismos da Medula Espinal/complicações , Bexiga Urinaria Neurogênica/fisiopatologia , Bexiga Urinária/fisiologia , Fator A de Crescimento do Endotélio Vascular/farmacologia , Animais , Matriz Extracelular/efeitos dos fármacos , Matriz Extracelular/transplante , Feminino , Sobrevivência de Enxerto , Ratos , Ratos Sprague-Dawley , Recuperação de Função Fisiológica/fisiologia , Engenharia Tecidual , Bexiga Urinaria Neurogênica/tratamento farmacológicoRESUMO
Genistein is a phytoestrogen that has been reported to suppress the AKT signaling pathway in several malignancies. However, the molecular mechanism of genistein action is not known. We tested the hypothesis that genistein activates expression of several aberrantly silenced tumor suppressor genes (TSGs) that have unmethylated promoters such as PTEN, CYLD, p53 and FOXO3a. We report here that genistein activates TSGs through remodeling of the heterochromatic domains at promoters in prostate cancer cells by modulating histone H3-Lysine 9 (H3-K9) methylation and deacetylation. Genistein activation involved demethylation and acetylation of H3-K9 at the PTEN and the CYLD promoter, while acetylation of H3-K9 at the p53 and the FOXO3a promoter occurred through reduction of endogenous SIRT1 activity. There was a decrease of SIRT1 expression and accumulation of SIRT1 in the cytoplasm from the nucleus. Increased expression of these TSGs was also reciprocally related to attenuation of phosphorylated-AKT and NF-kappaB binding activity in prostate cancer cells. This is the first report describing a novel epigenetic pathway that activates TSGs by modulating either histone H3-Lysine 9 (H3-K9) methylation or deacetylation at gene promoters leading to inhibition of the AKT signaling pathway. These findings strengthen the understanding of how genistein may be chemoprotective in prostate cancer.
Assuntos
Anticarcinógenos/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Genes Supressores de Tumor/efeitos dos fármacos , Genisteína/farmacologia , Histonas/metabolismo , PTEN Fosfo-Hidrolase/metabolismo , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Acetilação/efeitos dos fármacos , Antimetabólitos Antineoplásicos/farmacologia , Azacitidina/análogos & derivados , Azacitidina/farmacologia , Western Blotting , Linhagem Celular Tumoral , Imunoprecipitação da Cromatina , Cromonas/farmacologia , Ilhas de CpG/efeitos dos fármacos , Decitabina , Enzima Desubiquitinante CYLD , Regulação para Baixo/efeitos dos fármacos , Ensaio de Desvio de Mobilidade Eletroforética , Inibidores Enzimáticos/farmacologia , Proteína Forkhead Box O3 , Fatores de Transcrição Forkhead/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Genes p53/efeitos dos fármacos , Histonas/efeitos dos fármacos , Humanos , Ácidos Hidroxâmicos/farmacologia , Masculino , Metilação/efeitos dos fármacos , Morfolinas/farmacologia , NF-kappa B/efeitos dos fármacos , NF-kappa B/metabolismo , PTEN Fosfo-Hidrolase/efeitos dos fármacos , PTEN Fosfo-Hidrolase/genética , Fosfatidilinositol 3-Quinases/metabolismo , Inibidores de Fosfoinositídeo-3 Quinase , Fitoestrógenos/farmacologia , Neoplasias da Próstata/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sirtuína 1 , Sirtuínas/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Regulação para Cima/efeitos dos fármacosRESUMO
Genistein (4',5,7-trihydroxyisoflavone) is the most abundant isoflavone found in the soybean. The effects of genistein on various cancer cell lines have been extensively studied but the precise molecular mechanisms are not known. We report here the epigenetic mechanism of the action of genistein on androgen-sensitive (LNCaP) and androgen-insensitive (DuPro) human prostate cancer cell lines. Genistein induced the expression of tumor suppressor genes p21 (WAF1/CIP1/KIP1) and p16 (INK4a) with a concomitant decrease in cyclins. There was a G(0)-G(1) cell cycle arrest in LNCaP cells and a G(2)-M arrest in DuPro cells after genistein treatment. Genistein also induced apoptosis in DuPro cells. DNA methylation analysis revealed the absence of p21 promoter methylation in both cell lines. The effect of genistein on chromatin remodeling has not been previously reported. We found that genistein increased acetylated histones 3, 4, and H3/K4 at the p21 and p16 transcription start sites. Furthermore, we found that genistein treatment also increased the expression of histone acetyl transferases that function in transcriptional activation. This is the first report on epigenetic regulation of various genes by genistein through chromatin remodeling in prostate cancer. Altogether, our data provide new insights into the epigenetic mechanism of the action of genistein that may contribute to the chemopreventive activity of this dietary isoflavone and have important implications for epigenetic therapy.
Assuntos
Cromatina/genética , Inibidor p16 de Quinase Dependente de Ciclina/genética , Inibidor de Quinase Dependente de Ciclina p21/genética , Genisteína/farmacologia , Neoplasias da Próstata/genética , Acetilação , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Cromatina/efeitos dos fármacos , DNA de Neoplasias/efeitos dos fármacos , DNA de Neoplasias/genética , DNA de Neoplasias/isolamento & purificação , Perfilação da Expressão Gênica , Genes Supressores/efeitos dos fármacos , Genes Supressores de Tumor/efeitos dos fármacos , Histonas/efeitos dos fármacos , Histonas/metabolismo , Humanos , Masculino , Reação em Cadeia da Polimerase , Neoplasias da Próstata/patologiaRESUMO
Secreted frizzled-related protein 2 (sFRP2) is a negative modulator of the Wingless-type (Wnt) signaling pathway, and shown to be inactivated in renal cell carcinoma (RCC). However, the molecular mechanism of silencing of sFRP2 is not fully understood. Our study was designed to elucidate the silencing mechanism of sFRP2 in RCC. Expression of sFRP2 was examined in 20 pairs of primary cancers by immunohistochemistry. Kidney cell lines (HK-2, Caki-1, Caki-2, A-498 and ACHN) were analyzed for sFRP2 expression using real-time RT-PCR and Western blotting. The methylation status at 46 CpG sites of the 2 CpG islands in the sFRP2 promoter was characterized by bisulfite DNA sequencing. Histone modifications were assessed by chromatin immunoprecipitation (ChIP) assay using antibodies against AcH3, AcH4, H3K4 and H3K9. sFRP2 was frequently repressed in primary cancers and in RCC cells. The majority of sFRP2 negative cells had a methylated promoter. Meanwhile, sFRP2 expression was repressed by a hypomethylated promoter in Caki-1 cells, and these cells had a repressive histone modification at the promoter. In Caki-1 cells, sFRP2 was reactivated by trichostatin A (TSA). Repressive histone modifications were also observed in RCC cells with hypermethylated promoters, but sFRP2 was reactivated only by 5-aza-2'-deoxycytidine (DAC) and not by TSA. However, the activation of the silenced sFRP2 gene could be achieved in all cells using a combination of DAC and TSA. This is the first report indicating that aberrant DNA methylation and histone modifications work together to silence the sFRP2 gene in RCC cells.
Assuntos
Carcinoma de Células Renais/metabolismo , Metilação de DNA , Inativação Gênica , Histonas/metabolismo , Neoplasias Renais/metabolismo , Proteínas de Membrana/genética , Proteínas Wnt/metabolismo , Antimetabólitos Antineoplásicos/farmacologia , Apoptose/genética , Azacitidina/análogos & derivados , Azacitidina/farmacologia , Western Blotting , Carcinoma de Células Renais/genética , Linhagem Celular Tumoral , Ilhas de CpG/genética , Metilases de Modificação do DNA/antagonistas & inibidores , Decitabina , Inibidores Enzimáticos/farmacologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Inativação Gênica/efeitos dos fármacos , Inibidores de Histona Desacetilases , Humanos , Ácidos Hidroxâmicos/farmacologia , Imuno-Histoquímica/métodos , Imunoprecipitação , Neoplasias Renais/genética , Regiões Promotoras Genéticas , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/efeitos dos fármacosRESUMO
PURPOSE: The mismatch repair system is a DNA repair mechanism that corrects mispaired bases during DNA replication errors. Cancer cells deficient in MMR proteins have a 10(2) to 10(3)-fold increase in the mutation rate. Single nucleotide polymorphisms of mismatch repair genes have been shown to cause a decrease in DNA repair activity. We hypothesized that mismatch repair gene polymorphism could be a risk factor for prostate cancer and p53 Pro/Pro genotype carriers could influence MSH3 and MSH6 polymorphisms. MATERIALS AND METHODS: DNA samples from 110 patients with prostate cancer and 110 healthy controls were analyzed by single strand conformational polymorphism and polymerase chain reaction-restriction fragment length polymorphism to determine the genotypic frequency of 5 polymorphic loci on 2 MMR genes (MSH3 and MSH6) and p53 codon72. The chi-square test was applied to compare genotype frequency between patients and controls. RESULTS: A significant increase in the G/A+A/A genotype of MSH3 Pro222Pro was observed in patients compared to controls (OR 1.87, 95% CI 1.0-3.5). The frequency of A/G + G/G genotypes of MSH3 exon23 Thr1036Ala also tended to increase in patients (OR 1.57, 95% CI 0.92-2.72). In p53 codon72 Arg/Pro + Pro/Pro carriers the frequency of the AG + GG genotype of MSH3 exon23 was significantly increased in patients compared to controls (OR 2.1, 95% CI 1.05-4.34). CONCLUSIONS: To our knowledge this is the first report of the association of MSH3 gene polymorphisms in prostate cancer. These results suggest that the MSH3 polymorphism may be a risk factor for prostate cancer.
Assuntos
Pareamento Incorreto de Bases , Reparo de Erro de Pareamento de DNA , Proteínas de Ligação a DNA/genética , Predisposição Genética para Doença , Polimorfismo Genético , Neoplasias da Próstata/genética , Idoso , Genes p53/genética , Marcadores Genéticos , Genótipo , Humanos , Masculino , Proteína 3 Homóloga a MutS , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Polimorfismo Conformacional de Fita SimplesRESUMO
In estrogen metabolic pathways, the COMT enzyme is related to detoxification. COMT gene polymorphisms have been shown to effect enzyme function. We hypothesized that these polymorphisms may be risk factors for endometrial cancer (EC). DNA samples from 150 cases of EC and healthy controls (n = 165) were analyzed by PCR-RFLP to determine the genotypic frequency of four different polymorphic loci on COMT [codon 62 (rs4633), 102 (rs5031015), 136 (rs4818), 158 (rs4680)]. Genotyping was confirmed by direct DNA sequencing. We also conducted haplotype analysis of COMT and investigated the relationship between COMT expression and COMT SNPs in EC tissues by immunohistochemistry. A significant increase in the T/T genotype of codon 62 (C/T) was observed in patients compared to controls (OR = 2.39, 95% CI: 1.31-4.37, P = 0.004). The frequency of the C-G haplotype of codon 62 C/T and codon 158 G/A was significantly higher in controls (P < 0.0001) than in patients. The expression level of COMT protein in EC tissues was significantly lower in COMT codon 62 variant TT and codon 158 variant AA genotype carriers. Therefore, the EC samples with polymorphic variants of COMT lead to lower expression of COMT protein whereas EC samples with wild-type codon 62 C/C and codon 158 G/G have higher expression of COMT protein. This is the first study demonstrating that polymorphisms in COMT codon 62 and codon 158 altered protein expression levels in EC, suggesting that they may be risk factors for EC in Caucasians.
Assuntos
Catecol O-Metiltransferase/genética , Neoplasias do Endométrio/enzimologia , Polimorfismo Genético , Adulto , Idoso , Idoso de 80 Anos ou mais , Sequência de Bases , Estudos de Casos e Controles , Primers do DNA , Neoplasias do Endométrio/genética , Feminino , Haplótipos , Humanos , Imuno-Histoquímica , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Fatores de RiscoRESUMO
BACKGROUND: In estrogen biosynthetic pathways, many enzymes are important for metabolism, detoxification, and bioavailability. Polymorphisms in these genes may have an effect on the enzymes' function. For example, higher expression and activation of biosynthetic enzymes and lower expression and activation of conjugation enzymes may lead to high toxicity or carcinogenesis. The authors hypothesized that single nucleotide polymorphisms (single nucleotide polymorphisms) of CYP1A1, CYP1A2, CYP1B1, CYP17, SULT1A1, SULT1E1, and SHBG genes may be risk factors for endometrial cancer. METHODS: DNA samples from 150 cases of endometrial cancer and healthy controls (n = 165) were analyzed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) to determine the genotypic frequency of 13 different polymorphic loci on the CYP1A1 (m1, m2, m3, m4), CYP1A2 1F, CYP1B1 codon432, COMT codon158, CYP17, SULT1A1 (Arg213His, 14A/G, 85C/T in the 3' flanking region), SULT1E1-64G/A promoter region, and SHBG genes. Genotyping was validated by direct DNA sequencing. The authors also investigated the relation between expression of CYP1A1 in endometrial cancer tissues and genotypes of CYP1A1 m1. RESULTS: A decreased frequency of TC + CC genotype of the CYP1A1 m1 (T/C) polymorphism was observed in endometrial cancer patients compared with controls (OR = 0.42; 95% CI, 0.27-0.69). The T-A haplotype of CYP1A1 m1 and m2 was increased in endometrial cancer patients (P = .017). The frequency of CYP1A1 m1 T/C + C/C was higher in a high CYP1A1 expression group (P = .009). The authors also found that individuals carrying the variants of SULT1A1 codon213 and 2 single nucleotide polymorphisms in the 3' flanking region (14A/G and 85C/T) had an increased risk for endometrial cancer. The frequencies of G-A-C and A-G-T haplotypes of these 3 variants were higher in endometrial cancer patients (P < .0001; P = .0002). In addition, the frequency of combined genotypes (SULT1A1 213 GA + AA and CYP1A1 m1 TT) was higher in endometrial cancer patients. (OR, 4.58; 95% CI, 2.35-8.93). CONCLUSIONS: This is the first report on the combined association of CYP1A1 and SULT gene polymorphisms in endometrial cancer that suggests a decreased single nucleotide polymorphism of CYP1A1 and an increased single nucleotide polymorphism for SULT1A1 and SULT1E1 genes may be risk factors for endometrial cancer in Caucasians.
Assuntos
Arilsulfotransferase/genética , Citocromo P-450 CYP1A1/genética , Neoplasias do Endométrio/genética , Predisposição Genética para Doença , Polimorfismo de Nucleotídeo Único , Sulfotransferases/genética , Adulto , Idoso , Feminino , Genótipo , Haplótipos , Humanos , Pessoa de Meia-Idade , Fatores de RiscoRESUMO
PURPOSE: The chemokine CXCL12 and its receptor CXCR4 have been found to be associated with cancer metastasis. A single nucleotide polymorphism of CXCL12 G801A has been described and is regarded as a target for cis-acting factor that has the ability to up-regulate CXCL12 expression. Currently, there are no reports investigating the role of CXCL12 G801A polymorphism in prostate cancer (PC). EXPERIMENTAL DESIGN: We genotyped CXCL12 G801A and p53Arg72Pro in 167 PC patients and 167 age-matched healthy subjects. Genotyping was done with PCR-RFLP and confirmed by direct DNA sequencing. To investigate the effect of the CXCL12 G801A polymorphism on CXCL12 and CXCR4 expression, immunohistochemistry was done in genotyped PC tissues. RESULTS: A significant increase in the GA + AA genotype of the CXCL12 G801A polymorphism was observed in PC patients compared with healthy controls. The frequency of CXCL12 AA genotype was significantly higher in a group of patients with lymph node metastasis (23%) compared with those without metastasis (7%). The frequency of CXCL12 expression in AA + GA genotype carriers was significantly higher than that in GG genotype carriers. Among the carriers with CXCL12 GA + AA genotypes, CXCR4 expression was also significantly higher compared with those with the GG genotype. Moreover, among the groups with both CXCL12- and CXCR4-positive staining, the frequency of the CXCL12 GA + AA genotype was high. Although we did not find a significant relationship between the frequency of the Arg/Pro + Pro/Pro genotype of p53 Arg72Pro and susceptibility in PC, there was a combined effect of CXCL12 GA + AA genotype and the p53 72Arg/Pro + Pro/Pro genotype on the frequency of PC. These results indicate that the p53 codon 72 polymorphism may interact with CXCL12 G801A. CONCLUSIONS: This is the first report showing that CXCL12 G801A polymorphism may be a risk factor for PC. Moreover, this study suggests that this polymorphism can be an important marker for detecting microinvasion and PC metastasis.
Assuntos
Quimiocinas CXC/genética , Predisposição Genética para Doença , Polimorfismo Genético , Neoplasias da Próstata/genética , Idoso , Quimiocina CXCL12 , Códon , Genes p53 , Genótipo , Humanos , Metástase Linfática , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único , Neoplasias da Próstata/etiologia , Neoplasias da Próstata/patologia , Fatores de RiscoRESUMO
PURPOSE: MDM2 is a major negative regulator of p53, and a single nucleotide polymorphism in the MDM2 promoter region SNP309 (rs2279744) has been shown to increase the affinity of the transcriptional activator Sp1, resulting in elevated MDM2 transcription and expression in some cancers. There is currently no information about the role of MDM2 polymorphism in renal cell carcinoma (RCC). We investigated polymorphisms in p53-related genes, including MDM2, and their interactions in renal cancer. EXPERIMENTAL DESIGN: We genotyped three single nucleotide polymorphisms of three genes (p53 Arg(72)Pro, p21 Ser(31)Arg, and MDM2 SNP309) in 200 patients with renal cancer and 200 age- and gender-matched healthy subjects. Genotyping was confirmed by direct DNA sequencing. Samples that showed significant polymorphic variants were analyzed for MDM2 expression by immunohistochemistry. Association of polymorphic variants on survival of RCC patients was analyzed by Kaplan-Meier curves. RESULTS: A significant increase in the GG genotype of the MDM2 SNP309 was observed in RCC patients compared with healthy controls (odds ratio, 1.80; 95% confidence interval, 1.14-2.84). To investigate the effect of the MDM2 SNP309 polymorphism on MDM2 expression, immunohistochemistry was done in genotyped RCC tissues. Positive staining for MDM2 was detected in 2 of 15 (13%) TT genotype, 4 of 15 (26%) TG genotype, and 5 of 10 (50%) GG genotype carriers. The frequency of MDM2 expression in GG genotype carriers was significantly higher than that in TT genotype carriers. Polymorphisms of p53 Arg(72)Pro and p21 Ser(31)Arg did not show significant association with RCC. In univariate and multivariate analysis, MDM2 SNP309 GG genotype was independently associated with poor prognosis. Kaplan-Meier curve analysis showed that survival of patients with GG carriers was significantly worse than that of carriers with TG + TT genotypes. CONCLUSIONS: This is the first report to show a significant association between functional polymorphisms in MDM2 and increased risk of developing renal cancer. In addition, the MDM2 polymorphism was shown to be an independent adverse prognostic factor for RCC. Patients with MDM2 309GG genotype showed worse prognosis and low survival.
Assuntos
Carcinoma de Células Renais/genética , Neoplasias Renais/genética , Polimorfismo de Nucleotídeo Único , Proteínas Proto-Oncogênicas c-mdm2/genética , Idoso , Carcinoma de Células Renais/mortalidade , Carcinoma de Células Renais/patologia , Feminino , Regulação Neoplásica da Expressão Gênica , Predisposição Genética para Doença , Genótipo , Humanos , Neoplasias Renais/mortalidade , Neoplasias Renais/patologia , Masculino , Pessoa de Meia-Idade , Prognóstico , Fatores de Risco , Análise de Sobrevida , Transcrição Gênica , Proteína Supressora de Tumor p53/genéticaRESUMO
PURPOSE: The RAS-association domain family 1, isoform A (RASSF1A) gene is shown to be inactivated in prostate cancers. However, the molecular mechanism of silencing of the RASSFIA gene is not fully understood. The present study was designed to investigate the mechanisms of inactivation of the RASSF1A gene through the analysis of CpG methylation and histone acetylation and H3 methylation associated with the RASSF1A promoter region. EXPERIMENTAL DESIGN: Methylation status of the RASSF1A gene was analyzed in 131 samples of prostate cancer, 65 samples of benign prostate hypertrophy (BPH), and human prostate cell lines using methylation-specific PCR. Histone acetylation (acetyl-H3, acetyl-H4) and H3 methylation (dimethyl-H3-K4, dimethyl-H3-K9) status associated with the promoter region in prostate cells were analyzed by chromatin immunoprecipitation (ChIP) assay. RESULTS: Aberrant methylation was detected in 97 (74.0%) prostate cancer samples and 12 (18.5%) BPH samples. The methylation frequency of RASSF1A showed a significant increase with high Gleason sum and high stage. The ChIP assays showed enhancement of histone acetylation and dimethyl-H3-K4 methylation on the unmethylated RASSF1A promoter. TSA alone was unable to alter key components of the histone code. However, after 5-aza-2'-deoxy-cytidine treatment, there was a complete reversal of the histone components in the hypermethylated promoter. Levels of acetyl-H3, acetyl-H4, and dimethyl-H3-K4 became more enriched, whereas H3K9me2 levels were severely depleted. CONCLUSIONS: This is the first report suggesting that reduced histone acetylation or H3K4me2 methylation and increased dimethyl-H3-K9 methylation play a critical role in the maintenance of promoter DNA methylation-associated RASSF1A gene silencing in prostate cancer.