RESUMO
OBJECTIVE: The aim of this work was to evaluate a pediatric ependymoma protein expression that may be useful as a molecular biomarker candidate for prognosis, correlated with clinical features such as age, gender, histopathological grade, ependymal tumor recurrence and patient survival. PATIENTS AND METHODS: Immunohistochemistry assays were performed for GNAO1, ASAH1, IMMT, IPO7, Cyclin D1, P53 and Ki-67 proteins. Kaplan-Meier and Cox analysis were performed for age, gender, histopathological grade, relapse and survival correlation. RESULTS: We found that three proteins correlate with histopathological grade and relapse; two proteins correlate with survival; one protein does not correlate with any clinical feature. CONCLUSION: Our results suggest that, out of the proteins analyzed, five may be considered suitable prognostic biomarkers and one may be considered a predictive biomarker for response to treatment of pediatric ependymoma.
Assuntos
Ceramidase Ácida/biossíntese , Neoplasias Encefálicas/metabolismo , Ependimoma/metabolismo , Subunidades alfa Gi-Go de Proteínas de Ligação ao GTP/biossíntese , Carioferinas/biossíntese , Proteínas Mitocondriais/biossíntese , Proteínas Musculares/biossíntese , Receptores Citoplasmáticos e Nucleares/biossíntese , Ceramidase Ácida/genética , Adolescente , Biomarcadores Tumorais/biossíntese , Biomarcadores Tumorais/genética , Neoplasias Encefálicas/diagnóstico , Neoplasias Encefálicas/genética , Criança , Pré-Escolar , Estudos de Coortes , Ependimoma/diagnóstico , Ependimoma/genética , Feminino , Subunidades alfa Gi-Go de Proteínas de Ligação ao GTP/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Lactente , Recém-Nascido , Carioferinas/genética , Masculino , Proteínas Mitocondriais/genética , Proteínas Musculares/genética , Prognóstico , Receptores Citoplasmáticos e Nucleares/genética , Fatores de TempoRESUMO
Bioactive sphingolipids are modulators of immune processes and their metabolism is often dysregulated in ulcerative colitis, a major category of inflammatory bowel disease (IBD). While multiple axes of sphingolipid metabolism have been investigated to delineate mechanisms regulating ulcerative colitis, the role of acid ceramidase (AC) in intestinal inflammation is yet to be characterized. Here we demonstrate that AC expression is elevated selectively in the inflammatory infiltrate in human and murine colitis. To probe for mechanistic insight into how AC up-regulation can impact intestinal inflammation, we investigated the selective loss of AC expression in the myeloid population. Using a model of intestinal epithelial injury, we demonstrate that myeloid AC conditional knockout mice exhibit impairment of neutrophil recruitment to the colon mucosa as a result of defective cytokine and chemokine production. Furthermore, the loss of myeloid AC protects from tumor incidence in colitis-associated cancer (CAC) and inhibits the expansion of neutrophils and granulocytic myeloid-derived suppressor cells in the tumor microenvironment. Collectively, our results demonstrate a tissue-specific role for AC in regulating neutrophilic inflammation and cytokine production. We demonstrate novel mechanisms of how granulocytes are recruited to the colon that may have therapeutic potential in intestinal inflammation, IBD, and CAC.-Espaillat, M. P., Snider, A. J., Qiu, Z., Channer, B., Coant, N., Schuchman, E. H., Kew, R. R., Sheridan, B. S., Hannun, Y. A., Obeid, L. M. Loss of acid ceramidase in myeloid cells suppresses intestinal neutrophil recruitment.
Assuntos
Ceramidase Ácida/biossíntese , Colite Ulcerativa/enzimologia , Colo/enzimologia , Regulação Enzimológica da Expressão Gênica , Mucosa Intestinal/enzimologia , Neutrófilos/enzimologia , Regulação para Cima , Ceramidase Ácida/genética , Animais , Quimiocinas/biossíntese , Quimiocinas/genética , Colite Ulcerativa/genética , Colite Ulcerativa/patologia , Colo/patologia , Neoplasias do Colo/enzimologia , Neoplasias do Colo/genética , Neoplasias do Colo/patologia , Feminino , Humanos , Mucosa Intestinal/patologia , Masculino , Camundongos , Camundongos Knockout , Células Supressoras Mieloides/enzimologia , Células Supressoras Mieloides/patologia , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Neutrófilos/patologia , Microambiente Tumoral/genéticaRESUMO
Glioblastoma multiforme (GBM) is the most common primary, intracranial malignancy of the central nervous system. The standard treatment protocol, which involves surgical resection, and concurrent radiation with adjuvant temozolomide (TMZ), still imparts a grim prognosis. Ultimately, all GBMs exhibit recurrence or progression, developing resistance to standard treatment. This study demonstrates that GBMs acquire resistance to radiation via upregulation of acid ceramidase (ASAH1) and sphingosine1-phosphate (Sph-1P). Moreover, inhibition of ASAH1 and Sph-1P, either with humanized monoclonal antibodies, small molecule drugs (i.e. carmofur), or a combination of both, led to suppression of GBM cell growth. These results suggest that ASAH1 and Sph-1P may be excellent targets for the treatment of new GBMs and recurrent GBMs, especially since the latter overexpresses ASAH1.
Assuntos
Ceramidase Ácida/metabolismo , Neoplasias Encefálicas/enzimologia , Neoplasias Encefálicas/radioterapia , Glioblastoma/enzimologia , Glioblastoma/radioterapia , Ceramidase Ácida/biossíntese , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Glioblastoma/metabolismo , Glioblastoma/patologia , Humanos , Imuno-Histoquímica , Lisofosfolipídeos/metabolismo , Recidiva Local de Neoplasia/enzimologia , Recidiva Local de Neoplasia/patologia , Tolerância a Radiação , Esfingosina/análogos & derivados , Esfingosina/metabolismo , Regulação para CimaRESUMO
Acid ceramidase (ASAH1) a key enzyme of sphingolipid metabolism converting pro-apoptotic ceramide to sphingosine has been shown to be overexpressed in various cancers. We previously demonstrated higher expression of ASAH1 in ER positive compared to ER negative breast cancer. In the current study we performed subtype specific analyses of ASAH1 gene expression in invasive and non invasive breast cancer. We show that expression of ASAH1 is mainly associated with luminal A - like cancers which are known to have the best prognosis of all breast cancer subtypes. Moreover tumors with high ASAH1 expression among the other subtypes are also characterized by an improved prognosis. The good prognosis of tumors with high ASAH1 is independent of the type of adjuvant treatment in breast cancer and is also detected in non small cell lung cancer patients. Moreover, even in pre-invasive DCIS of the breast ASAH1 is associated with a luminal phenotype and a reduced frequency of recurrences. Thus, high ASAH1 expression is generally associated with an improved prognosis in invasive breast cancer independent of adjuvant treatment and could also be valuable as prognostic factor for pre-invasive DCIS.
Assuntos
Ceramidase Ácida/biossíntese , Neoplasias da Mama/enzimologia , Neoplasias da Mama/mortalidade , Carcinoma Intraductal não Infiltrante/enzimologia , Carcinoma Intraductal não Infiltrante/mortalidade , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Neoplasias da Mama/tratamento farmacológico , Carcinoma Intraductal não Infiltrante/tratamento farmacológico , Quimioterapia Adjuvante , Intervalo Livre de Doença , Feminino , Humanos , Pessoa de Meia-Idade , Invasividade Neoplásica , Taxa de SobrevidaRESUMO
Acid ceramidase (AC), a key enzyme of sphingolipid metabolism, seems to play an important role in cancer progression. The objective of this study was to explore the expression of AC in ovarian cancer and its impact on prognosis. Expression analysis of AC in n=112 ovarian cancer patients was performed by immunohistochemical analysis of primary paraffin-embedded tumor samples. The results were scored on the basis of the staining intensity and percentage of positive tumor cells, resulting in an immunoreactive score from 0 to 12. These results were correlated to clinical and pathologic characteristics and survival. AC expression correlated significantly only with FIGO stage (0.047). In serous carcinoma, low level of AC was independently associated with reduced progression-free survival and overall survival of 12.0 mo [95% confidence interval (CI), 5.78-18.23] versus 18.1 mo (95% CI, 11.61-24.59; P=0.008) and 35.7 mo (95% CI, 22.24-47.16) versus 58.7 mo (95% CI, 36.48-80.91; P=0.032), respectively. In multivariate analysis, AC presents as an independent prognostic factor for progression-free survival (hazard ratio 1.88; 95% CI, 1.13-3.11; P=0.015). AC is a prognostic factor in epithelial ovarian cancer. Low AC expression can be associated with tumor progression in carcinoma of the ovaries. These results are in contrast to the concept of AC as a promoter for cancer progression. Nevertheless, they are supported by the lately discovered tumor-suppressing function of sphingosine, the enzymatic product of AC.
Assuntos
Ceramidase Ácida/biossíntese , Biomarcadores Tumorais/análise , Neoplasias Epiteliais e Glandulares/enzimologia , Neoplasias Epiteliais e Glandulares/mortalidade , Neoplasias Ovarianas/enzimologia , Neoplasias Ovarianas/mortalidade , Ceramidase Ácida/análise , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma Epitelial do Ovário , Intervalo Livre de Doença , Feminino , Humanos , Imuno-Histoquímica , Estimativa de Kaplan-Meier , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Prognóstico , Modelos de Riscos Proporcionais , Esfingolipídeos/metabolismo , Adulto JovemRESUMO
Dacarbazine (DTIC) is the treatment of choice for metastatic melanoma, but its response in patients remains very poor. Ceramide has been shown to be a death effector and to play an important role in regulating cancer cell growth upon chemotherapy. Among ceramidases, the enzymes that catabolize ceramide, acid ceramidase (aCDase) has been implicated in cancer progression. Here we show that DTIC elicits a time- and dose-dependent decrease of aCDase activity and an increase of intracellular ceramide levels in human A375 melanoma cells. The loss of enzyme activity occurred as a consequence of reactive oxygen species-dependent activation of cathepsin B-mediated degradation of aCDase. These events preceded autophagic features and loss of cell viability. Down-regulation of acid but not neutral or alkaline ceramidase 2 resulted in elevated levels of ceramide and sensitization to the toxic effects of DTIC. Conversely, inducible overexpression of acid but not neutral ceramidase reduced ceramide levels and conferred resistance to DTIC. In conclusion, we report that increased levels of ceramide, due to enhanced degradation of aCDase, are in part responsible for the cell death effects of DTIC. These results suggest that down-regulation of aCDase alone or in combination with DTIC may represent a useful tool in the treatment of metastatic melanoma.
Assuntos
Ceramidase Ácida/biossíntese , Ceramidase Ácida/genética , Antineoplásicos Alquilantes/farmacologia , Dacarbazina/farmacologia , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Melanoma/tratamento farmacológico , Melanoma/enzimologia , Autofagia/efeitos dos fármacos , Autofagia/genética , Relação Dose-Resposta a Droga , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Regulação Enzimológica da Expressão Gênica/genética , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Melanoma/genética , Melanoma/patologia , Metástase NeoplásicaRESUMO
Sphingolipid metabolites, such as ceramide (Cer), sphingosine (SPH), and sphingosine 1-phosphate (S1P), contribute to multiple aspects of carcinogenesis including cell proliferation, migration, angiogenesis, and tumor resistance. The cellular balance between Cer and S1P levels, for example, is an important determinant of cell fate, with the former inducing apoptosis and the later mitogenesis. Acid ceramidase (ASAH1) plays a pivotal role in regulating the intracellular concentration of these two metabolites by hydrolyzing Cer into SPH, which is rapidly phosphorylated to form S1P. Genistein is a phytoestrogen isoflavone that exerts agonist and antagonist effects on the proliferation of estrogen-dependent MCF-7 cells in a dose-dependent manner, primarily as a ligand for estrogen receptors. Genistein can also activate signaling through GPR30, a G-protein-coupled cell surface receptor. Based on the relationship between bioactive sphingolipids and tumorigenesis, we sought to determine the effect of genistein on ASAH1 transcription in MCF-7 breast cancer cells. We show herein that nanomolar concentrations of genistein induce ASAH1 transcription through a GPR30-dependent, pertussis toxin-sensitive pathway that requires the activation of c-Src and extracellular signal regulated kinase 1/2 (ERK1/2). Activation of this pathway promotes histone acetylation and recruitment of phospho-estrogen receptor α and specificity protein-1 to the ASAH1 promoter, ultimately culminating in increased ceramidase activity. Finally, we show that genistein stimulates cyclin B2 expression and cell proliferation in an ASAH1-dependent manner. Collectively, these data identify a mechanism through which genistein promotes sphingolipid metabolism and support a role for ASAH1 in breast cancer cell growth.
Assuntos
Ceramidase Ácida/biossíntese , Anticarcinógenos/farmacologia , Neoplasias da Mama/enzimologia , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Genisteína/farmacologia , Acetilação/efeitos dos fármacos , Ceramidase Ácida/genética , Neoplasias da Mama/genética , Proteína Tirosina Quinase CSK , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Feminino , Regulação Enzimológica da Expressão Gênica/genética , Regulação Neoplásica da Expressão Gênica/genética , Histonas/genética , Histonas/metabolismo , Humanos , Proteína Quinase 1 Ativada por Mitógeno/genética , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/genética , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Proteínas Tirosina Quinases/genética , Proteínas Tirosina Quinases/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Receptores de Estrogênio , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Esfingolipídeos/genética , Esfingolipídeos/metabolismo , Transcrição Gênica/efeitos dos fármacos , Transcrição Gênica/genética , Quinases da Família srcRESUMO
IFN regulatory factor 8 (IRF8) is a key transcription factor for myeloid cell differentiation and its expression is frequently lost in hematopoietic cells of human myeloid leukemia patients. IRF8-deficient mice exhibit uncontrolled clonal expansion of undifferentiated myeloid cells that can progress to a fatal blast crisis, thereby resembling human chronic myelogeneous leukemia (CML). Therefore, IRF8 is a myeloid leukemia suppressor. Whereas the understanding of IRF8 function in CML has recently improved, the molecular mechanisms underlying IRF8 function in CML are still largely unknown. In this study, we identified acid ceramidase (A-CDase) as a general transcription target of IRF8. We demonstrated that IRF8 expression is regulated by IRF8 promoter DNA methylation in myeloid leukemia cells. Restoration of IRF8 expression repressed A-CDase expression, resulting in C16 ceramide accumulation and increased sensitivity of CML cells to FasL-induced apoptosis. In myeloid cells derived from IRF8-deficient mice, A-CDase protein level was dramatically increased. Furthermore, we demonstrated that IRF8 directly binds to the A-CDase promoter. At the functional level, inhibition of A-CDase activity, silencing A-CDase expression, or application of exogenous C16 ceramide sensitized CML cells to FasL-induced apoptosis, whereas overexpression of A-CDase decreased CML cells' sensitivity to FasL-induced apoptosis. Consequently, restoration of IRF8 expression suppressed CML development in vivo at least partially through a Fas-dependent mechanism. In summary, our findings determine the mechanism of IRF8 downregulation in CML cells and they determine a primary pathway of resistance to Fas-mediated apoptosis and disease progression.
Assuntos
Ceramidase Ácida/biossíntese , Apoptose/fisiologia , Fatores Reguladores de Interferon/metabolismo , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Animais , Linhagem Celular Tumoral , Ceramidas/metabolismo , Metilação de DNA , Proteína Ligante Fas/imunologia , Proteína Ligante Fas/farmacologia , Células HT29 , Humanos , Fatores Reguladores de Interferon/biossíntese , Fatores Reguladores de Interferon/genética , Células K562 , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/imunologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Células Mieloides/enzimologia , Células Mieloides/metabolismo , Regiões Promotoras Genéticas , Transcrição GênicaRESUMO
CONCLUSION: Fas ligand (FasL) gene therapy may provide a new efficient therapeutic model for head and neck squamous cell cancer (HNSCC). Acid ceramidase (AC) may not play an important role in the sensitivity of HNSCC cell lines to Fas-induced apoptosis. OBJECTIVES: The aims of this study were to investigate the efficacy of FasL gene therapy for HNSCC in vitro and to determine whether the expression of AC in different kinds of HNSCC cell lines is related to the sensitivity of HNSCC cell lines to Fas-mediated apoptotic induction. METHODS: Three HNSCC cell lines (Hep-2, MMSI, and SCCVII) were transfected with pEGFP-FasL, a plasmid containing a modified human FasL gene fused to enhanced green fluorescent protein (GFP). pEGFP-C1, a plasmid containing the GFP gene alone, was used as a control. Cell death was observed by fluorescence imaging and quantified using a tetrazolium-based (MTS) assay. SCCVII cells were analyzed by flow cytometry to determine the presence of apoptotic induction. Hep-2 and MMSI cells were evaluated by quantitative real-time PCR to evaluate the expression of AC. RESULTS: Transfection of pEGFP-FasL plasmid was shown to be able to induce cell death, the sensitivity of Fas-mediated apoptosis in HNSCC was different, and the level of AC did not correlate with the sensitivity of HNSCC cells to Fas-induced apoptosis.
Assuntos
Apoptose/genética , Proteína Ligante Fas/genética , Técnicas de Transferência de Genes , Terapia Genética/métodos , Neoplasias de Cabeça e Pescoço/terapia , Ceramidase Ácida/biossíntese , Ceramidase Ácida/genética , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Neoplasias de Cabeça e Pescoço/genética , Neoplasias de Cabeça e Pescoço/patologia , Humanos , Microscopia de Fluorescência , RNA Neoplásico/genética , Reação em Cadeia da Polimerase em Tempo Real , Sensibilidade e EspecificidadeRESUMO
Bioactive sphingolipids, such as ceramide, sphingosine and sphingosine-1-phosphate are known bio-effector molecules which play important roles in various aspects of cancer biology including cell proliferation, growth arrest, apoptosis, metastasis, senescence and inflammation. Therefore, enzymes involved in ceramide metabolism are gaining recognition as being critical regulators of cancer cell growth and/or survival. We previously observed that the ceramide metabolizing enzyme, acid ceramidase (AC) is upregulated in tumor tissues. Studies have now concluded that this creates a dysfunctional ceramide pathway, which is responsible for tumor progression and resistance to chemotherapy and radiation. This suggests that development of small-molecule drugs that inhibit AC enzyme activity is a promising approach for improving standard cancer therapy and patient's clinical outcomes.