RESUMEN
The prognosis of patients with Borrmann type IV gastric cancer (Type IV) is extremely poor. Thus, there is an urgent need to elucidate the molecular mechanisms underlying the oncogenesis of Type IV and to identify new therapeutic targets. Although previous studies using whole-exome and whole-genome sequencing have elucidated genomic alterations in gastric cancer, none has focused on comprehensive genetic analysis of Type IV. To discover cancer-relevant genes in Type IV, we performed whole-exome sequencing and genome-wide copy number analysis on 13 patients with Type IV. Exome sequencing identified 178 somatic mutations in protein-coding sequences or at splice sites. Among the mutations, we found a mutation in muscle RAS oncogene homolog (MRAS), which is predicted to cause molecular dysfunction. MRAS belongs to the Ras subgroup of small G proteins, which includes the prototypic RAS oncogenes. We analyzed an additional 46 Type IV samples to investigate the frequency of MRAS mutation. There were eight nonsynonymous mutations (mutation frequency, 17%), showing that MRAS is recurrently mutated in Type IV. Copy number analysis identified six focal amplifications and one homozygous deletion, including insulin-like growth factor 1 receptor (IGF1R) amplification. The samples with IGF1R amplification had remarkably higher IGF1R mRNA and protein expression levels compared with the other samples. This is the first report of MRAS recurrent mutation in human tumor samples. Our results suggest that MRAS mutation and IGF1R amplification could drive tumorigenesis of Type IV and could be new therapeutic targets.
Asunto(s)
Mutación , Receptores de Somatomedina/genética , Receptores de Somatomedina/metabolismo , Análisis de Secuencia de ADN/métodos , Neoplasias Gástricas/patología , Proteínas ras/genética , Adulto , Anciano , Anciano de 80 o más Años , Exoma , Femenino , Amplificación de Genes , Regulación Neoplásica de la Expresión Génica , Estudios de Asociación Genética/métodos , Humanos , Masculino , Persona de Mediana Edad , Tasa de Mutación , Receptor IGF Tipo 1 , Eliminación de Secuencia , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismoRESUMEN
BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive neoplastic diseases, associated with a remarkably poor prognosis. However, the molecular mechanisms underlying the development of PDAC remain elusive. The aim of this study was to identify genes whose expressions are correlated with a poor prognosis in PDAC patients, and to unravel the mechanisms underlying the involvement of these genes in the development of the cancer. METHODS: Global gene expression profiling was conducted in 39 specimens obtained from Japanese patients with PDAC to identify genes whose expressions were correlated with a shorter overall survival. The effect of gene silencing or overexpression of ARHGEF15 in pancreatic cancer cell lines was examined by introducing siRNAs of ARHGEF15 or the ARHGEF15 expression vector. After assessing the effect of ARHGEF15 deregulation on the Rho-family proteins by pull-down assay, wound healing, transwell and cell viability assays were carried out to investigate the cellular phenotypes caused by the perturbation. RESULTS: The global mRNA expression profiling revealed that overexpression of ARHGEF15, a Rho-specific GEF, was significantly associated with a poor prognosis in patients with PDAC. We also found that the depletion of ARHGEF15 by RNA interference in pancreatic cancer cell lines downregulated the activities of molecules of the Rho signaling pathway, including RhoA, Cdc42 and Rac1. Then, we also showed that ARHGEF15 silencing significantly reduced the motility and viability of the cells, while its overexpression resulted in the development of the opposite phenotype in multiple pancreatic cancer cell lines. CONCLUSION: These data suggest that upregulation of ARHGEF15 contributes to the development of aggressive PDAC by increasing the growth and motility of the pancreatic cancer cells, thereby worsening the prognosis of these patients. Therefore, ARHGEF15 could serve as a novel therapeutic target in patients with PDAC.
Asunto(s)
Adenocarcinoma/genética , Adenocarcinoma/mortalidad , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/mortalidad , Expresión Génica , Factores de Intercambio de Guanina Nucleótido/genética , Adenocarcinoma/patología , Adulto , Anciano , Biomarcadores de Tumor , Carcinoma Ductal Pancreático/patología , Línea Celular Tumoral , Movimiento Celular/genética , Proliferación Celular , Análisis por Conglomerados , Femenino , Perfilación de la Expresión Génica , Silenciador del Gen , Humanos , Masculino , Persona de Mediana Edad , Clasificación del Tumor , Pronóstico , ARN Interferente Pequeño/genéticaRESUMEN
BACKGROUND: We previously reported that 3'-ethynylcytidine (ECyd, TAS-106), an RNA polymerases inhibitor, enhances the anti-tumor efficacy of platinum in several tumor types in both in vitro and in vivo tumor models. However, the molecular mechanisms underlying the ECyd-induced enhancement remain elusive. METHODS: Cisplatin (CDDP)-resistant head and neck cancer KB cells were established by stepwise dose escalation with CDDP. The combination effect of ECyd and CDDP were assessed using isobologram analysis. The transcriptional and post-translational statuses of several molecules were detected using real-time PCR, immunoblot analysis and immunocytochemistry. Xenograft assays were used to confirm the mechanisms underlying the ECyd induced enhancement of CDDP anti-tumor efficacy in vivo. RESULTS: ECyd sensitized KB to CDDP by inhibiting the drug transporter Vault complex (Vaults). First, we showed that Vaults were overexpressed in CDDP-resistant KB cells. The suppression of major vault protein (MVP) by RNA interference restored the sensitivity to CDDP. Next, we showed that ECyd significantly sensitized the resistant cells to CDDP, compared with the parental paired cell line. A molecular analysis revealed that ECyd inhibited the synthesis of vRNAs as well as the induction of MVP, both of which are critical components of Vaults as a drug transporter. Furthermore, we found that the synergistic effect of ECyd and CDDP was correlated with the MVP expression level when the effect was analyzed in additional cancer cell lines. Finally, we demonstrated that ECyd decreased the vRNAs expression level in xenograft tumor. CONCLUSIONS: Our data indicated the ability of ECyd to cancel the resistance of cancer cells to CDDP by inhibiting the Vaults function and the decrease of Vaults expression itself, and the ability of the combination therapy with CDDP and ECyd to offer a new strategy for overcoming platinum resistance. Moreover, the study results suggest that Vaults could be a biomarker for stratifying patients who may benefit from the combination therapy with ECyd and platinum.
Asunto(s)
Antineoplásicos/farmacología , Cisplatino/farmacología , Citidina/análogos & derivados , Neoplasias Nasofaríngeas/tratamiento farmacológico , Partículas Ribonucleoproteicas en Bóveda/genética , Animales , Carcinoma , Línea Celular Tumoral , Citidina/farmacología , ARN Polimerasas Dirigidas por ADN/antagonistas & inhibidores , Resistencia a Antineoplásicos/efectos de los fármacos , Sinergismo Farmacológico , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Masculino , Ratones , Carcinoma Nasofaríngeo , Neoplasias Nasofaríngeas/patología , Trasplante de Neoplasias , Partículas Ribonucleoproteicas en Bóveda/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
In this study, we investigated the synergistic mechanism of anti-androgen and 5-fluorouracil (5-FU) combination therapy against castration-resistant prostate cancer (CRPC). Four prostate cancer cell lines, LNCaP, 22Rv1, DU145 and PC3, were examined for their growth dependency on androgens and the insulin-like growth factor 1 (IGF1). We assessed the expression changes of certain growth factor receptors and regulating proteins when treated with 5-FU, and found that 5-FU increased the expression of the IGF-binding protein 3 (IGFBP3). Furthermore, 5-FU inhibited the phosphorylation of Akt and p70 S6K, while the knockdown of IGFBP3 reduced the levels of poly (ADP-ribose) polymerase cleaved by 5-FU in PC3 cells. Therefore, the up-regulation of IGFBP3 by 5-FU not only inhibits cell growth by reducing the IGF1 signal but also induces apoptosis in PC3 cells. The synergistic effect of bicalutamide and 5-FU on 22Rv1 cells was reduced by IGFBP3 gene silencing using small-interfering RNA. These results suggest that the up-regulation of IGFBP3 induced by 5-FU plays an important role in the potent anti-tumor effect of 5-FU combined with anti-androgens on CRPC. Androgen-deprivation therapy combined with 5-FU could therefore be an appropriate therapy for CRPC patients.
Asunto(s)
Antagonistas de Andrógenos/farmacología , Antineoplásicos/farmacología , Proliferación Celular/efectos de los fármacos , Fluorouracilo/farmacología , Proteína 3 de Unión a Factor de Crecimiento Similar a la Insulina/metabolismo , Regulación hacia Arriba/efectos de los fármacos , Andrógenos/farmacología , Anilidas/farmacología , Apoptosis/efectos de los fármacos , Apoptosis/genética , Línea Celular Tumoral , Sinergismo Farmacológico , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/genética , Humanos , Proteína 3 de Unión a Factor de Crecimiento Similar a la Insulina/genética , Factor I del Crecimiento Similar a la Insulina/farmacología , Masculino , Nitrilos/farmacología , Neoplasias de la Próstata , Interferencia de ARN , Receptores de Factores de Crecimiento/genética , Receptores de Factores de Crecimiento/metabolismo , Transducción de Señal/efectos de los fármacos , Compuestos de Tosilo/farmacologíaRESUMEN
We investigated the antitumor effects of combination therapy with anti-androgens and 5-fluorouracil (5-FU), and examined the underlying mechanism of the treatment. Initially, we established the bicalutamide-resistant subline CDX25R from the androgen receptor (AR)-positive human prostate cancer cell line LNCaP through continuous exposure to bicalutamide. CDX25R cells lost the ability to respond to androgens, but still expressed AR. They showed significant resistance to bicalutamide, but had high sensitivity to hydroxyflutamide (OH-flutamide) compared with LNCaP cells. The CDX25R subline was thus considered to be a suitable model for prostate cancer that has developed resistance to first-line hormonal therapy but shows sensitivity to an alternative approach. Combined treatment with 5-FU and OH-flutamide had a synergistic effect on CDX25R cells. OH-flutamide decreased expression of the transcription factor E2F1, and subsequently of thymidylate synthase (TS), in CDX25R cells but not in AR-negative DU145 cells. This suggested that OH-flutamide enhanced the growth-inhibitory activity of 5-FU in CDX25R cells by reducing TS expression through the AR pathway. Combined therapy with 5-FU and OH-flutamide may, therefore, be appropriate for patients with prostate cancer that has acquired resistance to initial hormone therapy including bicalutamide.
Asunto(s)
Carcinoma/tratamiento farmacológico , Fluorouracilo/farmacología , Flutamida/análogos & derivados , Neoplasias de la Próstata/tratamiento farmacológico , Timidilato Sintasa/genética , Antagonistas de Andrógenos/administración & dosificación , Antagonistas de Andrógenos/farmacología , Anilidas/uso terapéutico , Antimetabolitos Antineoplásicos/administración & dosificación , Antimetabolitos Antineoplásicos/farmacología , Carcinoma/genética , Carcinoma/patología , Línea Celular Tumoral , Evaluación Preclínica de Medicamentos , Resistencia a Antineoplásicos/efectos de los fármacos , Resistencia a Antineoplásicos/genética , Sinergismo Farmacológico , Fluorouracilo/administración & dosificación , Flutamida/administración & dosificación , Flutamida/farmacología , Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Masculino , Modelos Biológicos , Neoplasias Hormono-Dependientes/tratamiento farmacológico , Neoplasias Hormono-Dependientes/genética , Nitrilos/uso terapéutico , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/patología , Timidilato Sintasa/metabolismo , Compuestos de Tosilo/uso terapéuticoRESUMEN
PURPOSE: Of the various microtubule-associated molecules, beta-tubulin III has been reported to be closely associated with the therapeutic efficacy of taxane-based chemotherapy against ovarian cancer. Stathmin and microtubule-associated protein 4 (MAP4) have been reported to play an important role in microtubule stabilization. In this study, we investigated whether expression of these microtubule-associated factors affects the therapeutic efficacy of taxane-based chemotherapy in ovarian clear cell adenocarcinoma. EXPERIMENTAL DESIGN: Drug sensitivity of paclitaxel or cisplatin was assessed in ovarian cancer cell lines treated with small interfering RNA of tubulin isoforms, MAP4, and stathmin. We examined 94 surgically resected ovarian clear cell adenocarcinoma specimens from patients treated with taxane-containing regimens (n=44) and with taxane-free regimens (n=50), using immunohistochemistry to detect expression of beta-tubulin III, stathmin, and MAP4. RESULTS: Knockdown of beta-tubulin III and IV specifically conferred drug resistance to paclitaxel in one ovarian cancer cell line, but not to other molecules. Estimated overall survival revealed a significant synergistic effect between taxane and beta-tubulin III in patients with ovarian clear cell adenocarcinoma. Of three microtubule-related molecules, among the taxane-based chemotherapy group, cases with higher beta-tubulin III expression were associated with a significantly more favorable prognosis compared with those having lower beta-tubulin III expression. By contrast, there was no statistical significance in the synergistic relationships between stathmin and taxane or between MAP4 and taxane. CONCLUSIONS: Taxane-based chemotherapy was effective for patients with ovarian clear cell adenocarcinomas who were positive for beta-tubulin III but not for those who were negative for these proteins.
Asunto(s)
Adenocarcinoma de Células Claras/tratamiento farmacológico , Antineoplásicos Fitogénicos/uso terapéutico , Neoplasias Ováricas/tratamiento farmacológico , Paclitaxel/uso terapéutico , Tubulina (Proteína)/fisiología , Adenocarcinoma de Células Claras/química , Adenocarcinoma de Células Claras/mortalidad , Adulto , Anciano , Línea Celular Tumoral , Cisplatino/uso terapéutico , Femenino , Humanos , Inmunohistoquímica , Proteínas Asociadas a Microtúbulos/análisis , Persona de Mediana Edad , Neoplasias Ováricas/química , Neoplasias Ováricas/mortalidad , Estatmina/análisis , Tubulina (Proteína)/análisisRESUMEN
Although 5-fluorouracil (5-FU) plus leucovorin (LV) is a standard chemotherapy regimen for colorectal cancer, the factors that determine the LV-mediated enhancement of the antitumor activity of 5-FU have remained unknown. We investigated the roles of folylpolyglutamate synthase (FPGS) and gamma-glutamyl hydrolase (GGH), which are the main enzymes involved in folate metabolism, in the effect of LV. LV enhanced the anticancer activity of 5-FU and the level of reduced folate in human colon cancer cells. Small-interfering RNA (siRNA) transfected into DLD-1 cells to downregulate FPGS reduced the basal level of reduced folate, the folate level after LV treatment, and the enhancement of 5-fluoro-2'-deoxyuridine (FdUrd)-induced cytotoxicity elicited by LV. By contrast, the downregulation of GGH by siRNA increased cellular sensitivity to FdUrd combined with LV. These results suggest that FPGS and GGH expression levels in tumors are determinants of the efficacy of LV in enhancing the antitumor activity of 5-FU.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/administración & dosificación , Neoplasias del Colon/tratamiento farmacológico , Neoplasias del Colon/enzimología , Fluorouracilo/administración & dosificación , Leucovorina/administración & dosificación , Péptido Sintasas/metabolismo , gamma-Glutamil Hidrolasa/metabolismo , Animales , Antineoplásicos/administración & dosificación , Neoplasias del Colon/patología , Relación Dosis-Respuesta a Droga , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Resultado del TratamientoRESUMEN
Although the intratumoral expression levels of thymidylate synthase (TS) and dihydropyrimidine dehydrogenase (DPD) are known to affect the antitumor activity of 5-fluorouracil (5-FU), the importance of orotate phosphoribosyltransferase (OPRT) has remained unclear. This study investigated the relationship between intratumoral OPRT expression and the antitumor activity of 5-FU using human NCI60 cell lines with similar levels of TS and DPD messenger RNAs, as well as 31 tumor xenografts. The OPRT mRNA level was positively correlated with the 5-FU efficacy in these cell lines. In vitro, the 50% growth-inhibitory concentrations of 5-FU were closely correlated with the OPRT mRNA levels in cancer cell lines with similar levels of TS mRNAs when combined with a DPD inhibitor. Moreover, downregulation of OPRT with small-interfering RNA decreased the sensitivities of the cultured tumor cells to 5-FU. These results suggest that the OPRT expression level in tumors is an additional determinant of the efficacy of 5-FU.
Asunto(s)
Biomarcadores de Tumor/metabolismo , Supervivencia Celular/efectos de los fármacos , Fluorouracilo/administración & dosificación , Proteínas de Neoplasias/metabolismo , Neoplasias/enzimología , Neoplasias/patología , Orotato Fosforribosiltransferasa/metabolismo , Antineoplásicos/administración & dosificación , Línea Celular Tumoral , Regulación Enzimológica de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , HumanosRESUMEN
Dihydropyrimidine dehydrogenase (DPD) is the rate-limiting enzyme in the catabolism of 5-fluorouracil (5-FU) and its activity is closely associated with cellular sensitivity to 5-FU. This study examines the role of DPD in the antiproliferative effects of 5-FU combined with IFN-alpha on hepatocellular carcinoma (HCC) cells in culture and asks whether IFN-alpha could affect DPD expression. The combined action of IFN-alpha and 5-FU on three HCC lines was quantified by a combination index method. Coadministration of IFN-alpha and 5-FU showed synergistic effects against HAK-1A and KYN-2 but antagonistic effects against KYN-3. The cellular expression levels of DPD mRNA and protein were markedly up-regulated in KYN-3 cells by IFN-alpha but were down-regulated in HAK-1A and KYN-2. The expression of thymidylate synthase mRNA and protein was down-regulated by IFN-alpha in all three cell lines. Coadministration of a selective DPD inhibitor, 5-chloro-2,4-dihydroxypyridine (CDHP), enhanced the antiproliferative effect of 5-FU and IFN-alpha on KYN-3 approximately 4-fold. However, the synergistic effects of 5-FU and IFN-alpha on HAK-1A and KYN-2 were not affected by CDHP. The antiproliferative effect of 5-FU could thus be modulated by IFN-alpha, possibly through DPD expression, in HCC cells. Inhibition of DPD activity by CDHP may enhance the efficacy of IFN-alpha and 5-FU combination therapy in patients with HCC showing resistance to this therapy.
Asunto(s)
Carcinoma Hepatocelular/enzimología , Carcinoma Hepatocelular/patología , Dihidrouracilo Deshidrogenasa (NADP)/genética , Dihidrouracilo Deshidrogenasa (NADP)/metabolismo , Fluorouracilo/farmacología , Interferón-alfa/farmacología , Carcinoma Hepatocelular/genética , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Dihidrouracilo Deshidrogenasa (NADP)/antagonistas & inhibidores , Sinergismo Farmacológico , Inhibidores Enzimáticos/farmacología , Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Timidilato Sintasa/genética , Timidilato Sintasa/metabolismoRESUMEN
We investigated the effects of pegylated (PEG)-IFN-alpha2b on alpha-fetoprotein (AFP) expression as demonstrated by protein and mRNA levels in six human hepatocellular carcinoma (HCC) cell lines. The number of KIM-1 cells in culture with PEG-IFN-alpha2b decreased between 24 amd 240 h, whereas the levels of intracellular and secreted AFP per cellular protein increased (except at 192 h), with levels 1.9-fold and 2.9-fold higher at maximum, respectively, than cells without PEG-IFN-alpha2b (control). The mRNA level increased between 72 and 192 h, when the level was 3-fold higher than that of the control. In the 72-h culture with 40-5000 IU/mL PEG-IFN-alpha2b, there were dose-dependent increases in AFP protein and mRNA expression and dose-dependent decrease in cell number resulting from apoptosis and blockage of the cell cycle at the S-phase. The rate of fucosylated AFP in the cell lysate decreased in a dose-dependent and time-dependent manner. In the PEG-IFN-alpha2b culture of the other five HCC cell lines, cell proliferation was suppressed, but the expressions of AFP protein and mRNA increased in only two cell lines, and suppression of cell proliferation was not related to the increase in AFP expressions. Our findings demonstrated that PEG-IFN-alpha2b induces an increase in AFP expression at both the protein and mRNA levels.
Asunto(s)
Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Carcinoma Hepatocelular/metabolismo , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Interferón-alfa/farmacología , alfa-Fetoproteínas/biosíntesis , Línea Celular Tumoral , Relación Dosis-Respuesta a Droga , Humanos , Interferón alfa-2 , ARN Mensajero/biosíntesis , Proteínas Recombinantes , Fase S/efectos de los fármacos , Factores de TiempoRESUMEN
Combination therapy with interferon (IFN)-alpha and 5-fluorouracil (5-FU) has been reported to show an improved therapeutic efficacy in patients with advanced hepatocellular carcinoma (HCC) but the mechanism behind this has not been completely elucidated. We examined the molecular events underlying the antiproliferative effects of IFN-alpha and 5-FU in combination using six human HCC cell lines. When the antiproliferative effects of administering IFN-alpha and 5-FU together were analyzed using isobolograms, we found that the cell lines could be divided into two groups: the S-group containing three cell lines, which showed synergistic effects, and the A-group, containing the remaining three cell lines, which showed additive effects. Real-time RT-PCR and Western blot analyses revealed that the expression levels of type I IFN receptor subunits, IFNAR1 and IFNAR2, were specifically up-regulated by 5-FU in all three cell lines of the S-group with the exception of IFNAR2 in one cell line, but not in those of the A-group. IFN-alpha modulated the protein expression levels of six enzymes regulating sensitivity to 5-FU, but none of them were down- or up-regulated in the same way in all members of the S- or A-group. In conclusion, the 5-FU-induced modulation of IFN receptor expression could play a pivotal role in the therapeutic efficacy of IFN-alpha combined with 5-FU. Measuring the expression levels of IFN receptors, and their ability to be up-regulated, may be a promising method for selecting HCC patients for this type of combination therapy.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Carcinoma Hepatocelular/tratamiento farmacológico , Carcinoma Hepatocelular/genética , Neoplasias Hepáticas/tratamiento farmacológico , Neoplasias Hepáticas/genética , Receptor de Interferón alfa y beta/biosíntesis , Carcinoma Hepatocelular/metabolismo , Procesos de Crecimiento Celular/efectos de los fármacos , Línea Celular Tumoral , Sinergismo Farmacológico , Fluorouracilo/administración & dosificación , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Interferón-alfa/administración & dosificación , Neoplasias Hepáticas/metabolismo , ARN Interferente Pequeño/genética , Receptor de Interferón alfa y beta/antagonistas & inhibidores , Receptor de Interferón alfa y beta/genética , Regulación hacia Arriba/efectos de los fármacosRESUMEN
Insulin-like growth factor binding protein-3 (IGFBP-3) modulates cell proliferation of various cancer cell types. However, it remains unclear how IGF-IGFBP-3-signaling is involved in growth and progression of hepatocellular carcinoma (HCC). The aim of the present study was to evaluate the role of IGFBP-3 in HCC. Type 1 receptor for IGF (IGF-1R) was expressed at various levels in the seven lines examined, but IGF-2R was not expressed. Of the seven lines, the growth of HAK-1B, KIM-1, KYN-2 and HepG2 cells was stimulated in a dose-dependent manner by the exogenous addition of IGF-I or IGF-II, but the HAK-1A, KYN-1 and KYN-3 cell lines showed no growth. Exogenous addition of IGFBP-3 markedly blocked IGF-I and IGF-II-stimulated cell growth of KYN-2 and HepG2 cells, and moderately stimulated that of KIM-1 and HAK-1B cells, but no growth of the KYN-1, KYN-3 and HAK-1A cell lines was observed. IGF-I enhanced the phosphorylation of IGF-1R, Akt and Erk1/2 in KYN-2 cells, and coadministration of IGFBP-3 blocked all types of activation by IGF-I investigated here. In contrast, no such activation by IGF-I was detected in KYN-3 cells. IGFBP-3 also suppressed IGF-I-induced cell invasion by KYN-2 cells. Moreover, we were able to observe the apparent expression of IGFBP-3 in KYN-3 cells, but not in the other six cell lines. Furthermore reduced expression of IGFBP-3, but not that of IGF-1R, was significantly correlated with tumor size, histological differentiation, capsular invasion and portal venous invasion. Low expression of IGFBP-3 was independently associated with poor survival. IGFBP-3 could be a molecular target of intrinsic importance for further development of novel therapeutic strategy against HCC.
Asunto(s)
Carcinoma Hepatocelular/metabolismo , Proteína 3 de Unión a Factor de Crecimiento Similar a la Insulina/metabolismo , Neoplasias Hepáticas/metabolismo , Invasividad Neoplásica/patología , Vena Porta/patología , Anciano , Anciano de 80 o más Años , Carcinoma Hepatocelular/patología , Femenino , Humanos , Técnicas para Inmunoenzimas , Proteína 3 de Unión a Factor de Crecimiento Similar a la Insulina/antagonistas & inhibidores , Proteína 3 de Unión a Factor de Crecimiento Similar a la Insulina/genética , Factor I del Crecimiento Similar a la Insulina/metabolismo , Neoplasias Hepáticas/patología , Masculino , Persona de Mediana Edad , Vena Porta/metabolismo , Pronóstico , Proteínas Proto-Oncogénicas c-akt/metabolismo , ARN Interferente Pequeño/farmacología , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/antagonistas & inhibidores , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/genética , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/metabolismo , Tasa de Supervivencia , Células Tumorales CultivadasRESUMEN
PURPOSE: We investigated the effects of pegylated IFN-alpha2b (PEG-IFN-alpha2b) on the growth of human liver cancer cells. METHODS: The effect of PEG-IFN-alpha2b on the proliferation of 13 liver cancer cell lines was investigated in vitro. Chronological changes in growth and IFN-alpha receptor-2 (IFNAR-2) expression were monitored in hepatocellular carcinoma (HCC) cells (HAK-1B) cultured with PEG-IFN-alpha2b. After HAK-1B cells were transplanted into nude mice, various doses of PEG-IFN-alpha2b or IFN-alpha2b were administered, and tumor volume, weight, histology, and IFNAR-2 expression were examined. RESULTS: PEG-IFN-alpha2b inhibited the growth of nine cell lines with apoptosis in a dose- and time-dependent manner. Continuous contact with PEG-IFN-alpha2b induced time-dependent growth inhibition and down-regulation of IFNAR-2 expression. PEG-IFN-alpha2b induced a dose-dependent decrease in tumor volume and weight, a significant increase of apoptotic cells, and a decrease in IFNAR-2 expression in the tumor. The clinical dose for chronic hepatitis C was also effective. The antitumor effect of PEG-IFN-alpha2b was significantly stronger than that of non-PEG-IFN-alpha2b in vivo. CONCLUSIONS: Continuous contact with PEG-IFN-alpha2b induces strong antitumor effects and the down-regulation of IFNAR-2 in HCC cells. The data suggest potential clinical application of PEG-IFN-alpha2b for the prevention and treatment of HCC.
Asunto(s)
Carcinoma Hepatocelular/tratamiento farmacológico , Carcinoma Hepatocelular/patología , Interferón-alfa/farmacología , Neoplasias Hepáticas/tratamiento farmacológico , Neoplasias Hepáticas/patología , Animales , Carcinoma Hepatocelular/metabolismo , Procesos de Crecimiento Celular/efectos de los fármacos , Línea Celular Tumoral , Femenino , Humanos , Interferón alfa-2 , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas Experimentales/tratamiento farmacológico , Neoplasias Hepáticas Experimentales/patología , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Polietilenglicoles , Receptor de Interferón alfa y beta/biosíntesis , Receptores de Interferón/inmunología , Proteínas RecombinantesRESUMEN
PURPOSE: Cap43 is known as a nickel- and calcium-inducible gene. In the present study, we examined whether 17beta-estradiol (E2) could affect the expression of Cap43 in breast cancer. EXPERIMENTAL DESIGN: Real-time PCR, immunoblotting, and immunocytochemistry were used to examine the expression of Cap43 and estrogen receptor-alpha (ER-alpha) in breast cancer cell lines. MDA-MB-231 and SK-BR-3 cell lines were transfected with ER-alpha cDNA to establish cells overexpressing ER-alpha. Immunohistochemistry was used to evaluate the expression of the Cap43 protein in breast cancer patients (n = 96), and the relationship between Cap43 expression and clinicopathologic findings was examined. RESULTS: Of the eight cell lines, four expressed higher levels of Cap43 with very low levels of ER-alpha, whereas the other four expressed lower levels of Cap43 with high ER-alpha levels. Treatment with E2 decreased the expression of Cap43 dose-dependently in ER-alpha-positive cell lines but not in ER-alpha-negative lines. Administration of antiestrogens, tamoxifen and ICI 182780, abrogated the E2-induced down-regulation of Cap43. Overexpression of ER-alpha in both ER-alpha-negative cell lines, SK-BR-3 and MDA-MB-231, resulted in down-regulation of Cap43. Immunostaining studies showed a significant correlation between Cap43 expression and the histologic grade of tumors (P = 0.0387). Furthermore, Cap43 expression was inversely correlated with the expression of ER-alpha (P = 0.0374). CONCLUSIONS: E2-induced down-regulation of Cap43 seems to be mediated through ER-alpha-dependent pathways in breast cancer cells both in culture and in patients. Cap43 has potential as a molecular marker to determine the therapeutic efficacy of antiestrogenic anticancer agents in breast cancer.
Asunto(s)
Estradiol/fisiología , Neoplasias/genética , Proteínas/análisis , Proteínas de Ciclo Celular , Proliferación Celular , Regulación hacia Abajo , Antagonistas de Estrógenos/farmacología , Receptor alfa de Estrógeno/análisis , Femenino , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Inmunohistoquímica , Péptidos y Proteínas de Señalización Intracelular , Neoplasias/patología , Reacción en Cadena de la Polimerasa , Proteínas/efectos de los fármacos , Tamoxifeno/farmacología , Células Tumorales CultivadasRESUMEN
A cell line, designated as RMG-V, was established from a patient with clear cell adenocarcinoma of the ovary. The cell line has grown without interruption and has been propagated continuously by serial passaging (more than 36 times) over 5 years. The cells are spindle-shaped, display neoplastic and pleomorphic features, and grow in a jigsaw puzzle-like arrangement while forming monolayers without contact inhibition. These cells proliferate rapidly, and the population doubling time is about 15.5 hours. The number of chromosomes ranges between 77 and 85, with a modal number of 83.
Asunto(s)
Adenocarcinoma de Células Claras/patología , Proliferación Celular , Neoplasias Ováricas/patología , Adenocarcinoma de Células Claras/genética , Línea Celular Tumoral , Cromosomas Humanos/genética , Femenino , Humanos , Cariotipificación , Persona de Mediana Edad , Neoplasias Ováricas/genética , Factores de TiempoRESUMEN
Among various types of surface epithelial ovarian carcinoma, clear cell adenocarcinoma often has a particularly poor prognosis even when diagnosed in stage I. It is resistant to existing anticancer drugs and appears to have different biological properties to other histological types of ovarian cancer. The present study was conducted using cell lines derived from ovarian clear cell adenocarcinoma in order to identify genes associated with the acquisition of malignant potential by this type of cancer. Two cell lines derived from ovarian clear cell adenocarcinoma (RMG-I and RMG-V), with different levels of invasive potential in an invasion assay, were used. DNA fragments were extracted from the band showing differences in the level of expression by RT-PCR with fluorescent differential display. A total of 56 different DNA base sequences were determined by direct sequencing. Primers were established using these base sequences and the level of expression in each cancer cell line was determined by real-time PCR. Integrin a3, the gene of which is present on chromosome 17q, was identified. It was also detected by a microarray analysis as one of the molecules showing a different level of expression between the two cell lines. Then the pattern of integrin a3 expression was investigated using immunocytochemical staining, and was found to differ between the two cell lines. The findings obtained using these cell lines derived from ovarian clear cell adenocarcinoma indicate that integrin alpha3 may associated with the acquisition of malignant potential by clear cell adenocarcinoma.
Asunto(s)
Adenocarcinoma de Células Claras/genética , Adenocarcinoma de Células Claras/patología , Regulación Neoplásica de la Expresión Génica , Integrina alfa3/genética , Neoplasias Ováricas/genética , Neoplasias Ováricas/patología , Línea Celular Tumoral , Femenino , Humanos , Inmunohistoquímica , Integrina alfa3/metabolismo , Invasividad Neoplásica , Análisis de Secuencia por Matrices de Oligonucleótidos , Reacción en Cadena de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
OBJECTIVES: A novel retinobenzoic acid derivative, 4-[3,5-bis (trimethylsilyl) benzamido] benzoic acid (TAC-101), was reported to suppress the growth and invasion of human gastric cancer or hepatocellular carcinoma by induction of apoptosis. We examined the antitumor activity of TAC-101 against human ovarian carcinoma cell lines. METHODS: Apoptosis of human epithelial ovarian carcinoma-derived cell lines (RMG-I, RMG-II, RTSG, RMUG-S, RMUG-L, and KF) was investigated by detecting DNA laddering and was quantified by an enzyme-linked immunosorbent assay. Inhibition of apoptosis was also examined using a caspase inhibitor. Furthermore, TAC-101 (8 mg kg(-1) day(-1) orally for 30 days) was investigated in nude mice with subcutaneous RMG-II tumors. A prominent apoptotic response to TAC-101 was observed. The antitumor effects of cisplatin (7 mg/kg intravenously on day 1) and paclitaxel (36 mg/kg intravenously on days 1 and 5) were also assessed for comparison. RESULTS: Apoptosis occurred in all of the cell lines (except KF) in a concentration-dependent manner after exposure to TAC-101 and was markedly induced in RMG-I and RMG-II cells (derived from ovarian clear cell adenocarcinomas). A caspase inhibitor blocked the induction of apoptosis by TAC-101. The maximum inhibition of RMG-II tumor growth in nude mice by TAC-101, cisplatin, and paclitaxel was 45%, 34%, and 47%, respectively. CONCLUSION: Oral TAC-101 shows potential as a novel antitumor agent for ovarian carcinoma, especially ovarian clear cell adenocarcinoma.