RESUMEN
Cardiomyopathy is the predominant defect in Barth syndrome (BTHS) and is caused by a mutation of the X-linked Tafazzin (TAZ) gene, which encodes an enzyme responsible for remodeling mitochondrial cardiolipin. Despite the known importance of mitochondrial dysfunction in BTHS, how specific TAZ mutations cause diverse BTHS heart phenotypes remains poorly understood. We generated a patient-tailored CRISPR/Cas9 knock-in mouse allele (TazPM) that phenocopies BTHS clinical traits. As TazPM males express a stable mutant protein, we assessed cardiac metabolic dysfunction and mitochondrial changes and identified temporally altered cardioprotective signaling effectors. Specifically, juvenile TazPM males exhibit mild left ventricular dilation in systole but have unaltered fatty acid/amino acid metabolism and normal adenosine triphosphate (ATP). This occurs in concert with a hyperactive p53 pathway, elevation of cardioprotective antioxidant pathways, and induced autophagy-mediated early senescence in juvenile TazPM hearts. However, adult TazPM males exhibit chronic heart failure with reduced growth and ejection fraction, cardiac fibrosis, reduced ATP, and suppressed fatty acid/amino acid metabolism. This biphasic changeover from a mild-to-severe heart phenotype coincides with p53 suppression, downregulation of cardioprotective antioxidant pathways, and the onset of terminal senescence in adult TazPM hearts. Herein, we report a BTHS genotype/phenotype correlation and reveal that absent Taz acyltransferase function is sufficient to drive progressive cardiomyopathy.
Asunto(s)
Aciltransferasas , Síndrome de Barth , Cardiomiopatías , Síndrome de Barth/genética , Síndrome de Barth/metabolismo , Síndrome de Barth/patología , Animales , Ratones , Aciltransferasas/genética , Cardiomiopatías/genética , Cardiomiopatías/metabolismo , Cardiomiopatías/patología , Masculino , Humanos , Mutación Puntual , Modelos Animales de Enfermedad , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , FenotipoRESUMEN
BACKGROUND: Gestational trophoblastic disease (GTD) is a heterogeneous group of diseases developed from trophoblasts. ASPP (Ankyrin-repeat, SH3-domain and proline-rich region containing protein) family proteins, ASPP1 and ASPP2, have been reported to be dysregulated in GTD. They modulate p53 activities and are responsible for multiple cellular processes. Nevertheless, the functional role of the ASPP family inhibitory member, iASPP, is not well characterized in GTD. METHODS: To study the functional role of iASPP in GTD, trophoblastic tissues from normal placentas, hydatidiform mole (HM) and choriocarcinoma were used for immunohistochemistry, whereas siRNAs were used to manipulate iASPP expression in choriocarcinoma cell lines and study the subsequent molecular changes. RESULTS: We demonstrated that iASPP was overexpressed in both HM and choriocarcinoma when compared to normal placenta. Progressive increase in iASPP expression from HM to choriocarcinoma suggests that iASPP may be related to the development of trophoblastic malignancy. High iASPP expression in HM was also significantly associated with a high expression of autophagy-related protein LC3. Interestingly, iASPP silencing retarded the growth of choriocarcinoma through senescence instead of induction of apoptosis. LC3 expression decreased once iASPP was knocked down, suggesting a downregulation on autophagy. This may be due to iASPP downregulation rendered decrease in Atg5 expression and concomitantly hindered autophagy in choriocarcinoma cells. Autophagy inhibition per se had no effect on the growth of choriocarcinoma cells but increased the susceptibility of choriocarcinoma cells to oxidative stress, implying a protective role of iASPP against oxidative stress through autophagy in choriocarcinoma. CONCLUSIONS: iASPP regulates growth and the cellular responses towards oxidative stress in choriocarcinoma cells. Its overexpression is advantageous to the pathogenesis of GTD. (266 words).
Asunto(s)
Autofagia , Coriocarcinoma/metabolismo , Mola Hidatiforme/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Estrés Oxidativo , Proteínas Represoras/metabolismo , Adolescente , Adulto , Proteína 5 Relacionada con la Autofagia/metabolismo , Línea Celular Tumoral , Coriocarcinoma/patología , Femenino , Estudios de Seguimiento , Técnicas de Silenciamiento del Gen , Humanos , Mola Hidatiforme/patología , Péptidos y Proteínas de Señalización Intracelular/clasificación , Péptidos y Proteínas de Señalización Intracelular/genética , Proteínas Asociadas a Microtúbulos/metabolismo , Persona de Mediana Edad , Placenta/citología , Placenta/metabolismo , Embarazo , Proteínas Represoras/clasificación , Proteínas Represoras/genética , Transfección , Proteína p53 Supresora de Tumor/metabolismo , Adulto JovenRESUMEN
Ovarian clear cell carcinoma (OCCC) is a type of epithelial ovarian cancer that is strongly associated with endometriosis, resistance against conventional chemotherapy and thus poorer prognosis. The expression of inhibitory member of the ASPP family proteins (iASPP) and Polo-like kinase (PLK)1 were significantly higher in OCCC compared to benign cystadenomas and endometriosis. Both protein expressions were found to correlate with chemoresistance in patients with OCCC while high iASPP expression alone was significantly associated with a poor patient survival. The growth of OCCC cell lines, OVTOKO and KK, were inhibited after iASPP silencing. Such effect was related to senescence triggering as evidenced by increased SA-ß-Gal staining and p21WAF1/Cip1 expression. Moreover, knockdown of iASPP induced PLK1 downregulation, whereas either genes' silencing sensitized the cells in response to cisplatin treatment. More prominent apoptosis was induced by cisplatin in OCCC cells after the knockdown of either iASPP or PLK1 as evidenced by the formation of more cleaved caspase 3. Heightened chemosensitivity to cisplatin after iASPP knockdown was further demonstrated in in vivo xenograft model. Additionally, both iASPP and PLK1 were shown to regulate autophagic flux as the induction of LC3B-II and LC3 puncta were much less in OCCC cells with either knockdown. Importantly, inhibition of autophagy also enhanced chemosensitivity to cisplatin in OCCC cells. These findings strongly imply that iASPP and PLK1 affect the chemoresistance of OCCC via the regulation of autophagy and apoptosis. Both iASPP and PLK1 can be potential therapeutic targets for treating OCCC in combination with conventional chemotherapy.
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Adenocarcinoma de Células Claras/patología , Autofagia , Proteínas de Ciclo Celular/metabolismo , Cisplatino/farmacología , Resistencia a Antineoplásicos , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Neoplasias Ováricas/patología , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Represoras/metabolismo , Adenocarcinoma de Células Claras/tratamiento farmacológico , Adenocarcinoma de Células Claras/metabolismo , Animales , Antineoplásicos/farmacología , Apoptosis , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Proteínas de Ciclo Celular/genética , Proliferación Celular , Femenino , Estudios de Seguimiento , Regulación Neoplásica de la Expresión Génica , Humanos , Péptidos y Proteínas de Señalización Intracelular/genética , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/metabolismo , Pronóstico , Proteínas Serina-Treonina Quinasas/genética , Proteínas Proto-Oncogénicas/genética , Proteínas Represoras/genética , Transducción de Señal , Tasa de Supervivencia , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de Xenoinjerto , Quinasa Tipo Polo 1RESUMEN
Human placental trophoblasts can be considered pseudomalignant, with tightly controlled proliferation, apoptosis, and invasiveness. Gestational trophoblastic disease (GTD) represents a family of heterogeneous trophoblastic lesions with aberrant apoptotic and proliferative activities and dysregulation of cell signaling pathways. We characterize the oncogenic effects of factor that binds to the inducer of short transcripts of HIV-1 [FBI-1, alias POZ and Krüppel erythroid myeloid ontogenic factor (POKEMON)/ZBTB7A] in GTD and its role in promoting cell aggressiveness in vitro and tumor growth in vivo. IHC studies showed increased nuclear expression of FBI-1, including hydatidiform moles, choriocarcinoma (CCA), and placental site trophoblastic tumor, in GTD. In JAR and JEG-3 CCA cells, ectopic FBI-1 expression opposed apoptosis through repression of proapoptotic genes (eg, BAK1, FAS, and CASP8). FBI-1 overexpression also promoted Akt activation, as indicated by Akt-pS473 phosphorylation. FBI-1 overexpression promoted mobility and invasiveness of JEG-3 and JAR, but not in the presence of the phosphoinositide 3-kinase inhibitor LY294002. These findings suggest that FBI-1 could promote cell migration and invasion via phosphoinositide 3-kinase/Akt signaling. In vivo, nude mice injected with CCA cells with stable FBI-1 knockdown demonstrated reduced tumor growth compared with that in control groups. These findings suggest that FBI-1 is clinically associated with the progression of, and may be a therapeutic target in, GTD, owing to its diverse oncogenic effects on dysregulated trophoblasts.
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Coriocarcinoma/patología , Proteínas de Unión al ADN/genética , Enfermedad Trofoblástica Gestacional/patología , Proteína Oncogénica v-akt/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Transducción de Señal , Factores de Transcripción/genética , Animales , Anticuerpos , Apoptosis , Pruebas de Carcinogenicidad , Movimiento Celular , Coriocarcinoma/genética , Coriocarcinoma/metabolismo , Proteínas de Unión al ADN/metabolismo , Femenino , Regulación Neoplásica de la Expresión Génica , Enfermedad Trofoblástica Gestacional/genética , Enfermedad Trofoblástica Gestacional/metabolismo , Humanos , Mola Hidatiforme/genética , Mola Hidatiforme/metabolismo , Mola Hidatiforme/patología , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Proteína Oncogénica v-akt/genética , Fosfatidilinositol 3-Quinasas/genética , Placenta/metabolismo , Embarazo , Conejos , Factores de Transcripción/metabolismo , Trofoblastos/metabolismoRESUMEN
Emerging evidence has indicated a role of the bone morphogenetic proteins (BMP) in the pathogenesis of certain cancers. The signaling of BMP family members is tightly regulated by their antagonists, including noggin and SOST, which are, in turn, positively regulated by BMP, thereby forming a negative feedback loop. Consequently, the expression of these antagonists should be taken into account in studies on the prognostic significance of BMP. In the present paper, we correlated protein and mRNA expression levels of BMP6, noggin and SOST, alone or in combination, with patient survival in various types of cancer. We found that BMP6 alone was not significantly correlated with esophageal squamous cell carcinoma patient survival. Instead, a high level of inhibitor of differentiation 1, a downstream factor of BMP6, was associated with shorter survival in patients whose tumors stained strongly for BMP6. Knockdown of noggin in esophageal cancer cell line EC109, which expresses BMP6 strongly and SOST weakly, enhanced the non-adherent growth of the cells. Noggin and SOST expression levels, when analyzed alone, were not significantly correlated with patient survival. However, high BMP6 activity, defined by strong BMP6 expression coupled with weak noggin or SOST expression, was significantly associated with shorter survival in esophageal squamous cell carcinoma patients. We further confirmed that BMP6 activity could be used as a prognostic indicator in prostate, bladder and colorectal cancers, using publicly available data on BMP6, noggin and SOST mRNA expression and patient survival. Our results strongly suggest that BMP6, noggin and SOST could be used in combination as a prognostic indicator in cancer progression.
Asunto(s)
Proteína Morfogenética Ósea 6/metabolismo , Proteínas Morfogenéticas Óseas/metabolismo , Proteínas Portadoras/metabolismo , Neoplasias Esofágicas/genética , Proteínas Adaptadoras Transductoras de Señales , Adulto , Anciano , Anciano de 80 o más Años , Proteína Morfogenética Ósea 6/genética , Proteínas Morfogenéticas Óseas/genética , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/mortalidad , Proteínas Portadoras/genética , Línea Celular Tumoral , Neoplasias Colorrectales/metabolismo , Progresión de la Enfermedad , Neoplasias Esofágicas/mortalidad , Neoplasias Esofágicas/patología , Femenino , Marcadores Genéticos/genética , Humanos , Proteína 1 Inhibidora de la Diferenciación/metabolismo , Masculino , Persona de Mediana Edad , Neoplasias de la Próstata/metabolismo , Interferencia de ARN , ARN Mensajero/genética , ARN Mensajero/metabolismo , ARN Interferente Pequeño , Transducción de Señal , Neoplasias de la Vejiga Urinaria/metabolismoRESUMEN
The transcription factors Pea3, Erm, and Er81 can promote cancer initiation and progression in various types of solid tumors. However, their role in esophageal squamous cell carcinoma (ESCC) has not been elucidated. In this study, we found that the expression levels of Pea3 and Erm, but not that of Er81, were significantly higher in ESCC compared with nontumor esophageal epithelium. A high level of Pea3 expression was significantly correlated with a shorter overall survival in a cohort of 81 patients with ESCC and the subgroup with N1 stage tumor (Wilcoxon-Gehan test, P = 0.016 and P = 0.001, respectively). Pea3 was overexpressed in seven ESCC cell lines compared with two immortalized esophageal cell lines. Pea3 knockdown reduced cell proliferation and suppressed nonadherent growth, migration, and invasion in ESCC cells in vitro. In addition, Pea3 knockdown in ESCC cells resulted in a down-regulation of phospho-Akt and matrix metalloproteinase 13, whereas a significant positive correlation in the expression levels was observed between Pea3 and phospho-Akt (r = 0.281, P < 0.013) and between Pea3 and matrix metalloproteinase 13 in the human specimens (r = 0.462, P < 0.001). Moreover, Pea3 modulated the sensitivity of EC109 cells to doxorubicin, probably via reduced activity of the phosphatidylinositol 3-kinase-Akt-mammalian target of Rapamycin complex 1 pathway on Pea3 knockdown. In conclusion, our results suggest that Pea3 plays an important role in the progression of ESCC.
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Carcinoma de Células Escamosas/metabolismo , Neoplasias Esofágicas/metabolismo , Regulación Neoplásica de la Expresión Génica , Factores de Transcripción/fisiología , Adulto , Anciano , Anciano de 80 o más Años , Línea Celular Tumoral , Movimiento Celular , Proteínas de Unión al ADN/metabolismo , Femenino , Humanos , Masculino , Persona de Mediana Edad , Invasividad Neoplásica , Pronóstico , Factores de Transcripción/metabolismoRESUMEN
Using inverse polymerase chain reaction, we identified CD44, located on chromosome 11p13, as a novel translocation partner of IGH in 9 of 114 cases of gastric, nongastric extranodal, follicular, and nodal diffuse large B-cell lymphoma (DLBCL). Notably, these translocations involving IGHSmu were detected in follicular lymphomas and exclusively in germinal center B cell-ike (GCB)-DLBCLs. CD44 is not expressed in reactive GC B cells. The IGHSmu/CD44 translocations substitute Smu for the CD44 promoter and remove exon 1 of CD44, resulting in the overexpression of Imu-CD44 hybrid mRNA transcripts activated from derivative 11 that encode a new CD44 variant lacking the leader peptide and with a unique C-terminus (CD44DeltaEx1). When overexpressed in vitro in the CD44(-) GCB-DLBCL cell line BJAB, CD44DeltaEx1-green fluorescent protein localized to the cytoplasm and nucleus, whereas CD44s-green fluorescent protein (standard form) localized to the plasma membrane. The ectopic expression of CD44DeltaEx1 in BJAB cells enhanced their proliferation rate and clonogenic ability, indicating a possible pathogenic role of the translocation.
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Receptores de Hialuranos/genética , Cadenas Pesadas de Inmunoglobulina/genética , Linfoma Folicular/genética , Linfoma de Células B Grandes Difuso/genética , Neoplasias Gástricas/genética , Translocación Genética , Línea Celular Tumoral , Puntos de Rotura del Cromosoma , Cromosomas Humanos Par 11 , Cromosomas Humanos Par 14 , Regulación Neoplásica de la Expresión Génica , Proteínas Fluorescentes Verdes/genética , Humanos , Receptores de Hialuranos/metabolismo , Linfoma Folicular/patología , Linfoma de Células B Grandes Difuso/patología , Reacción en Cadena de la Polimerasa , Neoplasias Gástricas/patologíaRESUMEN
The Fanconi anemia (FA) pathway safeguards genomic stability through cell cycle regulation and DNA damage repair. The canonical tumor suppressive role of FA proteins in the repair of DNA damage during interphase is well established, but their function in mitosis is incompletely understood. Here, we performed a kinome-wide synthetic lethality screen in FANCA-/- fibroblasts, which revealed multiple mitotic kinases as necessary for survival of FANCA-deficient cells. Among these kinases, we identified the depletion of the centrosome kinase SIK2 as synthetic lethal upon loss of FANCA. We found that FANCA colocalizes with SIK2 at multiple mitotic structures and regulates the activity of SIK2 at centrosomes. Furthermore, we found that loss of FANCA exacerbates cell cycle defects induced by pharmacological inhibition of SIK2, including impaired G2-M transition, delayed mitotic progression, and cytokinesis failure. In addition, we showed that inhibition of SIK2 abrogates nocodazole-induced prometaphase arrest, suggesting a novel role for SIK2 in the spindle assembly checkpoint. Together, these findings demonstrate that FANCA-deficient cells are dependent upon SIK2 for survival, supporting a preclinical rationale for targeting of SIK2 in FA-disrupted cancers.
Asunto(s)
Anemia de Fanconi , Ciclo Celular , Anemia de Fanconi/genética , Anemia de Fanconi/metabolismo , Anemia de Fanconi/patología , Proteína del Grupo de Complementación A de la Anemia de Fanconi/genética , Proteína del Grupo de Complementación A de la Anemia de Fanconi/metabolismo , Proteínas del Grupo de Complementación de la Anemia de Fanconi/genética , Humanos , Mitosis/genética , Proteínas Serina-Treonina Quinasas , Mutaciones Letales SintéticasRESUMEN
Deregulation of nuclear factor (NF)-kappaB signalling is common in cancers and is essential for tumourigenesis. Constitutive NF-kappaB activation in extranodal natural killer (NK)-cell lymphoma, nasal type (ENKL) is known to be associated with aberrant nuclear translocation of BCL10. Here we investigated the mechanisms leading to NF-kappaB activation and BCL10 nuclear localization in ENKLs. Given that ENKLs are dependent on T-cell-derived interleukin-2 (IL2) for cytotoxicity and proliferation, we investigated whether IL2 modulates NF-kappaB activation and BCL10 subcellular localization in ENKLs. In the present study, IL2-activated NK lymphoma cells were found to induce NF-kappaB activation via the PI3K/Akt pathway, leading to an increase in the entry of G(2)/M phase and concomitant transcription of NF-kappaB-responsive genes. We also found that BCL10, a key mediator of NF-kappaB signalling, participates in the cytokine receptor-induced activation of NF-kappaB. Knockdown of BCL10 expression resulted in deficient NF-kappaB signalling, whereas Akt activation was unaffected. Our results suggest that BCL10 plays a role downstream of Akt in the IL2-triggered NF-kappaB signalling pathway. Moreover, the addition of IL2 to NK cells led to aberrant nuclear translocation of BCL10, which is a pathological feature of ENKLs. We further show that BCL10 can bind to BCL3, a transcriptional co-activator and nuclear protein. Up-regulation of BCL3 expression was observed in response to IL2. Similar to BCL10, the expression and nuclear translocation of BCL3 were induced by IL2 in an Akt-dependent manner. The nuclear translocation of BCL10 was also dependent on BCL3 because silencing BCL3 by RNA interference abrogated this translocation. We identified a critical role for BCL10 in the cytokine receptor-induced NF-kappaB signalling pathway, which is essential for NK cell activation. We also revealed the underlying mechanism that controls BCL10 nuclear translocation in NK cells. Our findings provide insight into a molecular network within the NF-kappaB signalling pathway that promotes the pathogenesis of NK cell lymphomas.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Núcleo Celular/metabolismo , Interleucina-2/fisiología , Linfoma Extranodal de Células NK-T/metabolismo , FN-kappa B/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Proteína 10 de la LLC-Linfoma de Células B , Proteínas del Linfoma 3 de Células B , Línea Celular , Regulación de la Expresión Génica/genética , Humanos , Interleucina-2/farmacología , FN-kappa B/genética , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Transducción de Señal/genética , Factores de Transcripción/metabolismoRESUMEN
Fanconi anemia (FA) is a disease of genomic instability and cancer. In addition to DNA damage repair, FA pathway proteins are now known to be critical for maintaining faithful chromosome segregation during mitosis. While impaired DNA damage repair has been studied extensively in FA-associated carcinogenesis in vivo, the oncogenic contribution of mitotic abnormalities secondary to FA pathway deficiency remains incompletely understood. To examine the role of mitotic dysregulation in FA pathway deficient malignancies, we genetically exacerbated the baseline mitotic defect in Fancc-/- mice by introducing heterozygosity of the key spindle assembly checkpoint regulator Mad2. Fancc-/-;Mad2+/- mice were viable, but died from acute myeloid leukemia (AML), thus recapitulating the high risk of myeloid malignancies in FA patients better than Fancc-/-mice. We utilized hematopoietic stem cell transplantation to propagate Fancc-/-; Mad2+/- AML in irradiated healthy mice to model FANCC-deficient AMLs arising in the non-FA population. Compared to cells from Fancc-/- mice, those from Fancc-/-;Mad2+/- mice demonstrated an increase in mitotic errors but equivalent DNA cross-linker hypersensitivity, indicating that the cancer phenotype of Fancc-/-;Mad2+/- mice results from error-prone cell division and not exacerbation of the DNA damage repair defect. We found that FANCC enhances targeting of endogenous MAD2 to prometaphase kinetochores, suggesting a mechanism for how FANCC-dependent regulation of the spindle assembly checkpoint prevents chromosome mis-segregation. Whole-exome sequencing revealed similarities between human FA-associated myelodysplastic syndrome (MDS)/AML and the AML that developed in Fancc-/-; Mad2+/- mice. Together, these data illuminate the role of mitotic dysregulation in FA-pathway deficient malignancies in vivo, show how FANCC adjusts the spindle assembly checkpoint rheostat by regulating MAD2 kinetochore targeting in cell cycle-dependent manner, and establish two new mouse models for preclinical studies of AML.
RESUMEN
BACKGROUND AND AIM: There is currently no safe and effective treatment for liver fibrosis. We have previously shown that Stephania tetrandra (ST) and Salvia miltiorrhiza (SM) suppress cell proliferation and enhance apoptosis of hepatic stellate cell (HSC) in vitro. In this study, we aimed to investigate the anti-fibrotic effect of these two herbs in vivo. METHODS: Liver fibrosis was induced by carbon tetrachloride (CCl(4)) injection in rats for 5 weeks. SM, ST or SM + ST was gavaged on day 1 of CCl(4) administration to study the preventive effects of herbs on hepatic fibrosis. In a separate study designed to assess possible fibrosis regression, rats were randomly allocated to be treated with SM, ST or SM + ST when fibrosis was established. Liver injury and collagen content were assessed. HSC activation and apoptosis were determined. RESULTS: As compared with the CCL(4)-only rats, serum ALT was significantly lower in CCl(4)-treated rats that received either SM (P < 0.01) or ST (P < 0.01). Administration of ST significantly prevented (P < 0.01) or reversed the hepatic fibrosis (P < 0.01) induced by CCL(4). Moreover, rats treated with ST had reduced protein expression of alpha-SMA both in prevention (P < 0.05) and in regression (P < 0.01) experiments. The double-color staining of alpha-SMA and TUNEL showed that ST increased HSC apoptosis. However, co-treatment of SM + ST did not increase the antifibrotic effect of ST. CONCLUSIONS: Stephania tetrandra safely and effectively prevents and reverses hepatic fibrosis through activating HSC apoptosis in rats.
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Células Estrelladas Hepáticas/efectos de los fármacos , Cirrosis Hepática/tratamiento farmacológico , Cirrosis Hepática/prevención & control , Hígado/efectos de los fármacos , Extractos Vegetales/farmacología , Sustancias Protectoras/farmacología , Salvia miltiorrhiza , Stephania tetrandra , Alanina Transaminasa/sangre , Animales , Apoptosis/efectos de los fármacos , Biomarcadores/sangre , Tetracloruro de Carbono , Colágeno/metabolismo , Citoprotección , Células Estrelladas Hepáticas/metabolismo , Células Estrelladas Hepáticas/patología , Hígado/metabolismo , Hígado/patología , Cirrosis Hepática/inducido químicamente , Cirrosis Hepática/metabolismo , Cirrosis Hepática/patología , Masculino , Ratas , Ratas Sprague-DawleyRESUMEN
TWIST, a helix-loop-helix transcription factor, is highly expressed in many types of human cancer. We have previously found that TWIST confers prostate cancer cells with an enhanced metastatic potential through promoting epithelial-mesenchymal transition (EMT) and a high TWIST expression in human prostate cancer is associated with an increased metastatic potential. The predilection of prostate cancer cells to metastasize to bone may be due to two interplaying mechanisms (i) by increasing the rate of bone remodeling and (ii) by undergoing osteomimicry. We further studied the role of TWIST in promoting prostate cancer to bone metastasis. TWIST expression in PC3, a metastatic prostate cancer cell line, was silenced by small interfering RNA and we found that conditioned medium from PC3 with lower TWIST expression had a lower activity on stimulating osteoclast differentiation and higher activity on stimulating osteoblast mineralization. In addition, we found that these effects were, at least partly, associated with TWIST-induced expression of dickkopf homolog 1 (DKK-1), a factor that promotes osteolytic metastasis. We also examined TWIST and RUNX2 expressions during osteogenic induction of an organ-confined prostate cancer cell, 22Rv1. We observed increased TWIST and RUNX2 expressions upon osteogenic induction and downregulation of TWIST through short hairpin RNA reduced the induction level of RUNX2. In summary, our results suggest that, in addition to EMT, TWIST may also promote prostate cancer to bone metastasis by modulating prostate cancer cell-mediated bone remodeling via regulating the expression of a secretory factor, DKK-1, and enhancing osteomimicry of prostate cancer cells, probably, via RUNX2.
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Neoplasias Óseas/secundario , Huesos/fisiopatología , Proteínas Nucleares/genética , Osteogénesis/fisiología , Neoplasias de la Próstata/patología , Proteína 1 Relacionada con Twist/genética , Células 3T3 , Fosfatasa Alcalina/metabolismo , Animales , Calcificación Fisiológica , Línea Celular , Línea Celular Tumoral , Subunidad alfa 1 del Factor de Unión al Sitio Principal/genética , Cartilla de ADN , Genes Reporteros , Humanos , Masculino , Ratones , Osteoblastos/citología , Osteoblastos/fisiología , Plásmidos , ARN Interferente Pequeño/genética , Regulación hacia ArribaRESUMEN
Hydatidiform mole (HM), an unusual pregnancy with pure or predominant paternal genetic contribution, is the most common form of gestational trophoblastic disease. Most HM regress after uterine evacuation but some will develop into persistent disease or even frank malignancy. Although p53 is highly expressed in HM, TP53 mutations have rarely been detected in previous studies. Here we screened for specific missense mutations on several TP53 hotspots in 49 HMs using a highly sensitive pyrosequencing approach and revealed the significant existence of such mutations in HM tissues. A particularly high frequency (â¼59% of the cases) of p53 inactivating mutation on exon 7 has been detected. Our identification of hitherto unreported TP53 mutations in HM suggests the presence of p53 mutants and reflects the advantages of using pyrosequencing for point mutation detection in clinical samples. Traditional sequencing method may have overlooked such mutations that only occur in a small population of trophoblasts.
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Mola Hidatiforme/genética , Mutación Puntual , Proteína p53 Supresora de Tumor/genética , Neoplasias Uterinas/genética , Análisis Mutacional de ADN , Femenino , Humanos , Mola Hidatiforme/metabolismo , Mola Hidatiforme/patología , Embarazo , Estudios Retrospectivos , Trofoblastos/metabolismo , Trofoblastos/patología , Proteína p53 Supresora de Tumor/metabolismo , Neoplasias Uterinas/metabolismo , Neoplasias Uterinas/patologíaRESUMEN
BACKGROUND: The antiapoptotic and epithelial-mesenchymal transition activities of Twist have been implicated in the neoplastic transformation and the development of metastasis, respectively. Upregulation of Twist, described in several types of human cancer, also acts as a prognostic marker of poor outcome. AIM: To investigate Twist expression in oesophageal squamous cell carcinoma (SCC) and its prognostic value in a Chinese cohort of patients with oesophageal SCC. METHODS: Twist expression in primary oesophageal SCC of 87 Chinese patients was investigated by immunohistochemical staining. Twist protein level in one immortalised normal oesophageal epithelial cell line and six oesophageal SCC cell lines was measured by western blot analysis. Twist mRNA level in 30 pairs of frozen specimens of primary oesophageal SCC and non-neoplastic oesophageal epithelium from the upper resection margin of corresponding oesophagectomy specimen was also determined by semiquantitative reverse transcription-PCR. RESULTS: It was found that Twist was upregulated in oesophageal SCC cell lines, and its mRNA and protein levels were both increased in oesophageal SCC and the non-neoplastic oesophageal epithelium (p<0.001). In addition, a high level of Twist expression in oesophageal SCC was significantly associated with a greater risk for the patient of developing distant metastasis within 1 year of oesophagectomy (OR 3.462, 95% CI 1.201 to 9.978; p=0.022). CONCLUSIONS: Our results suggest that upregulation of Twist plays a role in the neoplastic transformation to oesophageal SCC and subsequent development of distant metastasis. Twist may serve as a useful prognostic marker for predicting the development of distant metastasis in oesophageal SCC.
Asunto(s)
Biomarcadores de Tumor/biosíntesis , Carcinoma de Células Escamosas/metabolismo , Neoplasias Esofágicas/metabolismo , Proteínas Nucleares/biosíntesis , Proteína 1 Relacionada con Twist/biosíntesis , Regulación hacia Arriba , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores de Tumor/genética , Carcinoma de Células Escamosas/secundario , Carcinoma de Células Escamosas/cirugía , Transformación Celular Neoplásica/metabolismo , Neoplasias Esofágicas/patología , Neoplasias Esofágicas/cirugía , Esofagectomía , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Persona de Mediana Edad , Proteínas de Neoplasias/biosíntesis , Proteínas de Neoplasias/genética , Estadificación de Neoplasias , Proteínas Nucleares/genética , Pronóstico , ARN Mensajero/genética , ARN Neoplásico/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Células Tumorales Cultivadas , Proteína 1 Relacionada con Twist/genéticaRESUMEN
Breast cancer is a leading cause of cancer-related deaths. Anemia is common in breast cancer patients and can be treated with blood transfusions or with recombinant erythropoietin (EPO) to stimulate red blood cell production. Clinical studies have indicated decreased survival in some groups of cancer patients treated with EPO. Numerous tumor cells express the EPO receptor (EPOR), posing a risk that EPO treatment would enhance tumor growth, but the mechanisms involved in breast tumor progression are poorly understood.Here, we have examined the functional role of the EPO-EPOR axis in pre-clinical models of breast cancer. EPO induced the activation of PI3K/AKT and MAPK pathways in human breast cancer cell lines. EPOR knockdown abrogated human tumor cell growth, induced apoptosis through Bim, reduced invasiveness, and caused downregulation of MYC expression. EPO-induced MYC expression is mediated through the PI3K/AKT and MAPK pathways, and overexpression of MYC partially rescued loss of cell proliferation caused by EPOR downregulation. In a xenotransplantation model, designed to simulate recombinant EPO therapy in breast cancer patients, knockdown of EPOR markedly reduced tumor growth.Thus, our experiments in vitro and in vivo demonstrate that functional EPOR signaling is essential for the tumor-promoting effects of EPO and underline the importance of the EPO-EPOR axis in breast tumor progression.
Asunto(s)
Neoplasias de la Mama/patología , Eritropoyetina/farmacología , Receptores de Eritropoyetina/metabolismo , Animales , Apoptosis/efectos de los fármacos , Apoptosis/fisiología , Neoplasias de la Mama/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Proliferación Celular/fisiología , Progresión de la Enfermedad , Eritropoyetina/metabolismo , Femenino , Xenoinjertos , Humanos , Ratones , Ratones Desnudos , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiologíaRESUMEN
It has been shown previously that cancer cells with an activated oncogenic pathway, including Met activation, require Ran for growth and survival.Here, we show that knockdown of Ran leads to a reduction of Met receptor expression in several breast and lung cancer cell lines. This, in turn suppressed HGF expression and the Met-mediated activation of the Akt pathway, as well as cell adhesion, migration, and invasion. In a cell line model where Met amplification has previously been shown to contribute to gefitinib resistance, Ran knockdown sensitized cells to gefitinib-mediated inhibition of Akt and ERK1/2 phosphorylation and consequently reduced cell proliferation. We further demonstrate that Met reduction-mediated by knockdown of Ran, occurs at the post-transcriptional level, probably via a matrix metalloproteinase. Moreover, the level of immunoreactive Ran and Met are positively associated in human breast cancer specimens, suggesting that a high level of Ran may be a pre-requisite for Met overexpression. Interestingly, a high level of immunoreactive Ran dictates the prognostic significance of Met, indicating that the co-overexpression of Met and Ran may be associated with cancer progression and could be used in combination as a prognostic indicator.
Asunto(s)
Neoplasias/metabolismo , Neoplasias/patología , Proteínas Proto-Oncogénicas c-met/metabolismo , Transducción de Señal , Proteína de Unión al GTP ran/metabolismo , Adhesión Celular/efectos de los fármacos , Adhesión Celular/genética , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Movimiento Celular/genética , Proliferación Celular , Progresión de la Enfermedad , Resistencia a Antineoplásicos/genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Técnicas de Silenciamiento del Gen , Factor de Crecimiento de Hepatocito/metabolismo , Humanos , Neoplasias/genética , Fosforilación , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas c-akt/genética , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas Proto-Oncogénicas c-met/genética , Transducción de Señal/efectos de los fármacos , Proteína de Unión al GTP ran/genéticaRESUMEN
We characterized the effects of selective Cox-2 inhibition on the angiogenesis gastric cancer cell line SGC7901 by stable transfection of antisense Cox-2 cDNA (Cox-2-AS) or pharmacological inhibition by selective cox-2 inhibitor (NS398). The conditioned media obtained from SGC7901 treated with Cox-2-AS or NS398 suppressed the proliferation, migration and tube formation of human umbilical vein endothelial cells. Moreover, both treatments inhibited in vivo angiogenesis. These inhibitions could be partially reversed by the addition of PGE2. Our findings support that selective inhibition of Cox-2 alone plays an instrumental role on gastric cancer associated angiogenesis.
Asunto(s)
ADN sin Sentido , Neovascularización Patológica , Prostaglandina-Endoperóxido Sintasas/genética , Prostaglandina-Endoperóxido Sintasas/metabolismo , Neoplasias Gástricas/patología , Ciclooxigenasa 2 , Inhibidores de la Ciclooxigenasa 2 , Inhibidores de la Ciclooxigenasa/farmacología , ADN Complementario , Humanos , Proteínas de la Membrana , Nitrobencenos/farmacología , Sulfonamidas/farmacología , Transfección , Células Tumorales CultivadasRESUMEN
Breast cancer is one of the most prevalent malignancies worldwide. It consists of a group of tumor cells that have the ability to grow uncontrollably, overcome replicative senescence (tumor progression) and metastasize within the body. Metastases are processes that consist of an array of complex gene dysregulation events. Although these processes are still not fully understood, the dysregulation of a number of key proteins must take place if the tumor cells are to disseminate and metastasize. It is now widely accepted that future effective and innovative treatments of cancer metastasis will have to encompass all the major components of malignant transformation. For this reason, much research is now being carried out into the mechanisms that govern the malignant transformation processes. Recent research has identified key genes involved in the development of metastases, as well as their mechanisms of action. A detailed understanding of the encoded proteins and their interrelationship generates the possibility of developing novel therapeutic approaches. This review will focus on a select group of proteins, often deregulated in breast cancer metastasis, which have shown therapeutic promise, notably, EMT, E-cadherin, Osteopontin, PEA3, Transforming Growth Factor Beta (TGF-ß) and Ran.
Asunto(s)
Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Proteínas de Neoplasias/metabolismo , Animales , Femenino , Humanos , Metástasis de la NeoplasiaRESUMEN
Although gastric cancer with cyclooxygenase (COX)-2 overexpression is associated with poor prognosis, the mechanistic pathway remains unknown. We examined the associations between expressions of COX-2 and vascular endothelial growth factor (VEGF) in both gastric cancer cells and in human gastric cancer. The gastric cell line, Kato III, was transiently transfected with cox-2 expressing vector. The levels of COX-2, prostaglandin (PG) E2 and VEGF expression were measured post-transfection. Additionally, expressions of COX-2 and VEGF in human gastric cancer were determined by immunohistochemistry in archive gastrectomy specimens. Tumor angiogenesis was assessed by the microvessel density (MVD), which was determined by anti-CD34 immunostaining. Transient transfection of Kato III with cox-2 was associated with increased COX-2 expression, higher PGE2 production and upregulated VEGF expressions. Treatment with NS398, a specific COX-2 inhibitor, reduced VEGF expression in COX-2 expressing Kato III cells by 25%. Among the 67 gastric cancers examined, COX-2 overexpression was found in 45 (67%) cases whereas increased VEGF expression was detected in 46 (69%) cases. There was a significant association between COX-2 and VEGF expressions in gastric cancer (r=0.25, p=0.041). Additionally, tumor MVD was associated with both COX-2 (r=0.32, p=0.008) and VEGF (r=0.39, p=0.001) expressions. Our results showed that overexpression of COX-2 in both gastric cells and primary gastric cancer is associated with upregulation of VEGF and angiogenesis. Future studies should evaluate the potential anti-angiogenic effect of COX-2 inhibitors on human gastric cancer.
Asunto(s)
Carcinoma/enzimología , Isoenzimas/biosíntesis , Neovascularización Patológica , Prostaglandina-Endoperóxido Sintasas/biosíntesis , Neoplasias Gástricas/enzimología , Regulación hacia Arriba , Factor A de Crecimiento Endotelial Vascular/biosíntesis , Anciano , Antígenos CD34/biosíntesis , Western Blotting , Línea Celular Tumoral , Ciclooxigenasa 2 , Dinoprostona/metabolismo , Femenino , Vectores Genéticos , Humanos , Immunoblotting , Inmunohistoquímica , Isoenzimas/genética , Masculino , Proteínas de la Membrana , Microcirculación , Persona de Mediana Edad , Plásmidos/metabolismo , Prostaglandina-Endoperóxido Sintasas/genética , TransfecciónRESUMEN
BACKGROUND: Ras-related nuclear protein (Ran) is required for cancer cell survival in vitro and human cancer progression, but the molecular mechanisms are largely unknown. METHODS: We investigated the effect of the v-myc myelocytomatosis viral oncogene homolog (Myc) on Ran expression by Western blot, chromatin immunoprecipitation, and luciferase reporter assays and the effects of Myc and Ran expression in cancer cells by soft-agar, cell adhesion, and invasion assays. The correlation between Myc and Ran and the association with patient survival were investigated in 14 independent patient cohorts (n = 2430) and analyzed with Spearman's rank correlation and Kaplan-Meier plots coupled with Wilcoxon-Gehan tests, respectively. All statistical tests were two-sided. RESULTS: Myc binds to the upstream sequence of Ran and transactivates Ran promoter activity. Overexpression of Myc upregulates Ran expression, whereas knockdown of Myc downregulates Ran expression. Myc or Ran overexpression in breast cancer cells is associated with cancer progression and metastasis. Knockdown of Ran reverses the effect induced by Myc overexpression in breast cancer cells. In clinical data, a positive association between Myc and Ran expression was revealed in 288 breast cancer and 102 lung cancer specimens. Moreover, Ran expression levels differentiate better or poorer survival in Myc overexpressing breast (χ2 = 24.1; relative risk [RR] = 9.1, 95% confidence interval [CI] = 3.3 to 24.7, P < .001) and lung (χ2 = 6.04; RR = 2.8, 95% CI = 1.2 to 6.3; P = .01) cancer cohorts. CONCLUSIONS: Our results suggest that Ran is required for and is a potential therapeutic target of Myc-driven cancer progression in both breast and lung cancers.