RESUMEN
Ornithine decarboxylase 1 (ODC1), a metabolic enzyme critically involved in the polyamine biosynthesis, is commonly upregulated in hepatocellular carcinoma (HCC). Despite its altered expression in human HCC tissues, the molecular mechanism by which ODC1 alters the course of HCC progression and functions in HCC cell survival is unknown. Here we identified that silencing of ODC1 expression with small interfering (si) RNA causes inhibition of HCC cell growth through blockade of cell cycle progression and induction of apoptosis. Next, to obtain insights into the molecular changes in response to ODC1 knockdown, global changes in gene expression were examined using RNA sequencing. It revealed that 119 genes show same directional regulation (76 up- and 43 down-regulated) in both Huh1 and Huh7 cells and were considered as a common ODC1 knockdown signature. Particularly, we found through a network analysis that KLF2, which is known to inhibit PPARγ expression and adipogenesis, was commonly up-regulated. Subsequent Western blotting affirmed that the downregulation of ODC1 was accompanied by a decrease in the levels of PPARγ as well as of PARP-1, cyclin E1 and pro-caspase 9 delaying cell cycle progression and accelerating apoptotic signaling. Following the down-regulation of PPARγ expression, ODC1 silencing resulted in a strong inhibition in the expression of important regulators of glucose transport and lipid biogenesis, and caused a marked decrease in lipid droplet accumulation. In addition, ODC1 silencing significantly inhibited the growth of human HCC xenografts in nude mice. These findings indicate that the function of ODC1 is correlated with HCC lipogenesis and suggest that targeting ODC1 could be an attractive option for molecular therapy of HCC.
Asunto(s)
Carcinoma Hepatocelular/genética , Proliferación Celular/genética , Metabolismo de los Lípidos/genética , Neoplasias Hepáticas/genética , Ornitina Descarboxilasa/genética , Interferencia de ARN , Animales , Apoptosis/genética , Western Blotting , Carcinoma Hepatocelular/enzimología , Carcinoma Hepatocelular/patología , Caspasa 9/genética , Caspasa 9/metabolismo , Ciclo Celular/genética , Línea Celular Tumoral , Ciclina E/genética , Ciclina E/metabolismo , Regulación Neoplásica de la Expresión Génica , Redes Reguladoras de Genes , Humanos , Factores de Transcripción de Tipo Kruppel/genética , Factores de Transcripción de Tipo Kruppel/metabolismo , Neoplasias Hepáticas/enzimología , Neoplasias Hepáticas/patología , Masculino , Ratones Endogámicos BALB C , Ratones Desnudos , Proteínas Oncogénicas/genética , Proteínas Oncogénicas/metabolismo , Ornitina Descarboxilasa/metabolismo , PPAR gamma/genética , PPAR gamma/metabolismo , Poli(ADP-Ribosa) Polimerasa-1/genética , Poli(ADP-Ribosa) Polimerasa-1/metabolismo , Tratamiento con ARN de Interferencia/métodos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Ensayos Antitumor por Modelo de Xenoinjerto/métodosRESUMEN
Histone deacetylase 2 (HDAC2) is a chromatin modifier involved in epigenetic regulation of cell cycle, apoptosis, and differentiation that is upregulated commonly in human hepatocellular carcinoma (HCC). In this study, we show that specific targeting of this HDAC isoform is sufficient to inhibit HCC progression. siRNA-mediated silencing of HDAC inhibited HCC cell growth by blocking cell-cycle progression and inducing apoptosis. These effects were associated with deregulation of HDAC-regulated genes that control cell cycle, apoptosis, and lipid metabolism, specifically, by upregulation of p27 and acetylated p53 and by downregulation of CDK6 and BCL2. We found that HDAC2 silencing in HCC cells also strongly inhibited PPARγ signaling and other regulators of glycolysis (ChREBPα and GLUT4) and lipogenesis (SREBP1C and FAS), eliciting a marked decrease in fat accumulation. Notably, systemic delivery of HDAC2 siRNA encapsulated in lipid nanoparticles was sufficient to blunt the growth of human HCC in a murine xenograft model. Our findings offer preclinical proof-of-concept for HDAC2 blockade as a systemic therapy for liver cancer.
Asunto(s)
Carcinoma Hepatocelular/genética , Histona Desacetilasa 2/genética , Neoplasias Hepáticas/genética , Isoformas de Proteínas/genética , Animales , Apoptosis/genética , Carcinoma Hepatocelular/patología , Ciclo Celular/genética , Línea Celular Tumoral , Quinasa 6 Dependiente de la Ciclina/genética , Progresión de la Enfermedad , Regulación hacia Abajo/genética , Regulación Neoplásica de la Expresión Génica/genética , Glucólisis/genética , Células Hep G2 , Humanos , Metabolismo de los Lípidos/genética , Lipogénesis/genética , Neoplasias Hepáticas/patología , Masculino , Ratones , Ratones SCID , PPAR gamma/genética , Antígeno Nuclear de Célula en Proliferación/genética , Transducción de Señal/genética , Proteína p53 Supresora de Tumor/genética , Regulación hacia Arriba/genética , Proteína X Asociada a bcl-2/genéticaRESUMEN
PURPOSE: Acute hemorrhagic conjunctivitis (AHC), a highly contagious eye disease, is caused primarily by either enterovirus 70 (EV70) or coxsackievirus A24 (CVA24) infection. Yet methods to prevent or cure AHC are not available. Recent evidence has shown that small-interfering RNAs (siRNAs), mediators of posttranscriptional gene knockdown, can act as effective antiviral agents. Thus, the authors attempted to develop a novel siRNA-based anti-AHC agent effective against both EV70 and CVA24. METHODS: Concurrent screening of the entire viral genome sequences of EV70 and CVA24 using the CAPSID program identified five different siRNA candidates complementary to genome regions of both viruses. The antiviral potentials of these siRNAs were assessed by treating MRC5 and primary human conjunctival cells with the siRNAs and following this with viral challenge. RESULTS: Among the five siRNAs, AHCe-3D-3 siRNA showed excellent cytoprotective effects and dramatic decreases in virus replication and virus protein synthesis. This siRNA, targeting the virus polymerase 3D gene, also induced similar antiviral effects in primary human conjunctival cells. CONCLUSIONS: These findings strongly suggest that the AHCe-3D-3 siRNA, homologous to two different AHC-associated enteroviruses, can provide equivalent antiviral activities against both AHC-causing enteroviruses. Such an siRNA may be developed as a clinically valuable AHC control agent.