RESUMEN
Dysregulation of cellular energy metabolism is closely linked to cancer development and progression. Calorie or glucose restriction (CR or GR) inhibits energy-dependent pathways, including IGF-1/PI3K/Akt/mTOR, in cancer cells. However, alterations in proton dynamics and reversal of the pH gradient across the cell membrane, which results in intracellular alkalinization and extracellular acidification in cancer tissues, have emerged as important etiopathogenic factors. We measured glucose, lactate, and ATP production after GR, plant-derived CR-mimetic curcumin treatment, and curcumin plus GR in human hepatoma cells. Intracellular pH regulatory effects, in particular, protein-protein interactions within mTOR complex-1 and its structural change, were investigated. Curcumin treatment or GR mildly inhibited Na+/H+ exchanger-1 (NHE1). vATPase, monocarboxylate transporter (MCT)-1, and MCT4 level. Combination treatment with curcumin and GR further enhanced the inhibitory effects on these transporters and proton-extruding enzymes, with intracellular pH reduction. ATP and lactate production decreased according to pH change. Modeling of mTOR protein revealed structural changes upon treatments, and curcumin plus GR decreased binding of Raptor and GßL to mTOR, as well as of Rag A and Rag B to Raptor. Consequently, 4EBP1 phosphorylation was decreased and cell migration and proliferation were inhibited in a pH-dependent manner. Autophagy was increased by curcumin plus GR. In conclusion, curcumin treatment combined with GR may be a useful supportive approach for preventing intracellular alkalinization and cancer progression.
Asunto(s)
Antineoplásicos/farmacología , Carcinoma Hepatocelular/metabolismo , Curcumina/farmacología , Glucosa/deficiencia , Neoplasias Hepáticas/metabolismo , Álcalis/metabolismo , Línea Celular , Proliferación Celular/efectos de los fármacos , Glucosa/metabolismo , Células Hep G2 , Humanos , Transportadores de Ácidos Monocarboxílicos/metabolismo , Proteína Reguladora Asociada a mTOR/metabolismo , ATPasas de Translocación de Protón Vacuolares/metabolismoRESUMEN
Calorie restriction or a low-carbohydrate diet (LCD) can increase life span in normal cells while inhibiting carcinogenesis. Various phytochemicals also have calorie restriction-mimetic anticancer properties. We investigated whether an isocaloric carbohydrate-restriction diet and AMP-activated protein kinase (AMPK)-activating phytochemicals induce synergic tumor suppression. We used a mixture of AMPK-activating phytochemical extracts including curcumin, quercetin, catechins, and resveratrol. Survival analysis was carried out in a B16F10 melanoma model fed a control diet (62.14% kcal carbohydrate, 24.65% kcal protein and 13.2% kcal fat), a control diet with multiple phytochemicals (MP), LCD (16.5, 55.2, and 28.3% kcal, respectively), LCD with multiple phytochemicals (LCDmp), a moderate-carbohydrate diet (MCD, 31.9, 62.4, and 5.7% kcal, respectively), or MCD with phytochemicals (MCDmp). Compared with the control group, MP, LCD, or MCD intervention did not produce survival benefit, but LCDmp (22.80±1.58 vs. 28.00±1.64 days, P=0.040) and MCDmp (23.80±1.08 vs. 30.13±2.29 days, P=0.008) increased the median survival time significantly. Suppression of the IGF-1R/PI3K/Akt/mTOR signaling, activation of the AMPK/SIRT1/LKB1pathway, and NF-κB suppression were the critical tumor-suppression mechanisms. In addition, SIRT1 suppressed proliferation of the B16F10 and A375SM cells under a low-glucose condition. Alterations in histone methylation within Pten and FoxO3a were observed after the MCDmp intervention. In the transgenic liver cancer model developed by hydrodynamic transfection of the HrasG12V and shp53, MCDmp and LCDmp interventions induced significant cancer-prevention effects. Microarray analysis showed that PPARα increased with decreased IL-6 and NF-κB within the hepatocytes after an MCDmp intervention. In conclusion, an isocaloric carbohydrate-restriction diet and natural AMPK-activating agents induce synergistic anticancer effects. SIRT1 acts as a tumor suppressor under a low-glucose condition.
Asunto(s)
Proteínas Quinasas Activadas por AMP/metabolismo , Carbohidratos de la Dieta/administración & dosificación , Suplementos Dietéticos , Neoplasias Hepáticas Experimentales/prevención & control , Melanoma Experimental/prevención & control , Fitoquímicos/administración & dosificación , Sirtuina 1/metabolismo , Animales , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Proliferación Celular/efectos de los fármacos , Inmunoprecipitación de Cromatina , Carbohidratos de la Dieta/farmacología , Sinergismo Farmacológico , Neoplasias Hepáticas Experimentales/metabolismo , Neoplasias Hepáticas Experimentales/mortalidad , Neoplasias Hepáticas Experimentales/patología , Masculino , Melanoma Experimental/metabolismo , Melanoma Experimental/mortalidad , Melanoma Experimental/patología , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Análisis de Secuencia por Matrices de Oligonucleótidos , Fosforilación , Fitoquímicos/farmacocinética , Transducción de Señal , Sirtuina 1/genética , Tasa de Supervivencia , Células Tumorales CultivadasRESUMEN
PURPOSE: Perifosine has shown antitumor activity via inhibition of Akt phosphorylation in many advanced solid tumors. This study investigated the efficacy of perifosine alone and in combination with sorafenib in a transgenic mouse model of HCC. METHODS: The mouse model of HCC was generated by hydrodynamic injection of transposons encoding HrasG12V and short-hairpin RNA downregulating p53. The transgenic mice were treated with perifosine alone and in combination with sorafenib to evaluate efficacy of drugs on tumor growth and survival. RESULTS: Treatment with perifosine for 5 weeks, alone and in combination with sorafenib, strongly inhibited tumor growth and increased survival. Perifosine inhibited HCC cell proliferation, induced apoptosis, and decreased tumor angiogenesis. Furthermore, its combination with sorafenib enhanced these effects. In addition, Akt phosphorylation was decreased by perifosine and further decreased by combination treatment. Although perifosine alone did not appear to activate the caspase pathway, combination treatment increased the cleavage of caspase-3, caspase-9, and poly (ADP-ribose) polymerase. CONCLUSIONS: The preclinical effect that current study showed represents a strong rationale for clinical trials using perifosine alone and in combination with sorafenib in the treatment of HCC patients.
Asunto(s)
Antineoplásicos/uso terapéutico , Carcinoma Hepatocelular/tratamiento farmacológico , Neoplasias Hepáticas Experimentales/tratamiento farmacológico , Niacinamida/análogos & derivados , Compuestos de Fenilurea/uso terapéutico , Fosforilcolina/análogos & derivados , Proteínas Proto-Oncogénicas p21(ras)/genética , ARN Interferente Pequeño/genética , Proteína p53 Supresora de Tumor/genética , Animales , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Apoptosis/efectos de los fármacos , Carcinoma Hepatocelular/irrigación sanguínea , Carcinoma Hepatocelular/patología , Caspasa 3/metabolismo , Caspasa 9/metabolismo , Proliferación Celular/efectos de los fármacos , Elementos Transponibles de ADN , Sinergismo Farmacológico , Neoplasias Hepáticas Experimentales/irrigación sanguínea , Neoplasias Hepáticas Experimentales/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Neovascularización Patológica/tratamiento farmacológico , Niacinamida/uso terapéutico , Fosforilcolina/uso terapéutico , Poli(ADP-Ribosa) Polimerasas/metabolismo , SorafenibRESUMEN
The purpose of this study was to analyze the combined effects of regular exercise and ginseng supplementation on peritoneal exudate ROS (reactive oxygen species), lymphocyte proliferation by splenocytes, and DNA damage following exhaustive exercise stress. Thirty-six female BALB/c mice were randomly divided into control (UT, n = 12), trained (TR, n = 12), and ginseng supplemented and trained (GT, n = 12) groups. Each group was divided into two equal subgroups where mice were studied at rest (UTre, TRre, and GTre) or immediately after exhaustive exercise stress (UTex, TRex, and GTex). Animals were bred in the animal facility, where they were housed at 22-24 degrees C and relative humidity (RH) 50-60% in a controlled environment with a 12-hour photoperiod, and provided food and water ad libitum. The trained mice underwent 10 weeks of endurance swim training (5 times/week) in water at 27-30 degrees C for 60 minutes. The analytical items examined were weight, proliferative activity, the production of ROS from peritoneal exudate cells, and DNA damage following exhaustive exercise stress (2 h exercise stress). Significant level was set at p < 0.05. The results obtained showed that the trained group had a significantly lower mean body weight than the untrained group (p < 0.05). However, there was no significant difference between UT and GT. Swim training increased swim survival time in TRex and GTex, and TRex showed the highest swim survival time. With regard to mitogenic activities of splenocytes in response to exhaustive exercise stress, all groups showed much lower lymphocyte proliferative activity when stimulated with media (Med), concanavalin A (ConA), or lipopolysaccharide (LPS) after exhaustive exercise stress. However, GTex had a higher proliferative activity than the other groups. Trained and ginseng-supplemented groups showed lower peritoneal ROS responses and lymphocyte DNA damage levels after exhaustive exercise. These findings suggest that the combined effect of swim training and ginseng supplementation sustain lymphocyte function in the presence of reduced ROS production and DNA damage following acute exercise stress.
Asunto(s)
Daño del ADN , Panax , Condicionamiento Físico Animal/fisiología , Resistencia Física/fisiología , Especies Reactivas de Oxígeno/metabolismo , Animales , Linfocitos B/inmunología , Exudados y Transudados/efectos de los fármacos , Exudados y Transudados/fisiología , Femenino , Activación de Linfocitos/efectos de los fármacos , Masculino , Ratones , Ratones Endogámicos BALB C , Resistencia Física/efectos de los fármacos , Natación/fisiología , Linfocitos T/inmunologíaRESUMEN
Protein-tyrosine phosphatases (PTPs) constitute a family of receptor-like, and cytoplasmic enzymes, which catalyze the dephosphorylation of phosphotyrosine residues in a variety of receptors and signaling molecules. Together with protein tyrosine kinases (PTKs), PTPs are critically involved in regulating many cellular signaling processes. In this study, diverse compounds were screened for PTP inhibition and selectively screened for inhibitors with the end product inhibition properties. Among phosphate analogues and their derivatives for PTP inhibition, Keggin compounds phosphomolybdate (PM) and phosphotungstate (PT) strongly inhibited both PTP-1B and SHP-1, with K(i) values of 0.06-1.2 micromM in the presence of EDTA. Unlike the vanadium compounds, inhibition potencies of PM and PT were not significantly affected by EDTA. PM and PT were potent, competitive inhibitors for PTPs, but relatively poor inhibitors of Ser/Thr phosphatase. Interestingly, PM and PT did not inhibit alkaline phosphatase at all. The crystal structure of PTP-1B in complex with PM, at 2.0 A resolution, reveals that MoO(3), derived from PM by hydrolysis, binds at the active site. The molybdenium atom of the inhibitor is coordinated with six ligands: three oxo-ligands, two apical water molecules and a S atom of the catalytic cysteine residue. In support of the crystallographic finding, we observed that molybdenium oxides (MoO(3), MoO(2), and MoO(2)Cl(2)) inhibited PTP-1B with IC(50) in the range 5-15 micromM.