RESUMEN
Feeding a high-fat diet (HFD) coupled with sugar, mimicking a Western diet, causes fatty liver disease in mice. Histamine induces biliary proliferation and fibrosis and regulates leptin signaling. Wild-type (WT) and l-histidine decarboxylase (Hdc-/-) mice were fed a control diet or an HFD coupled with a high fructose corn syrup equivalent. Hematoxylin and eosin and Oil Red O staining were performed to determine steatosis. Biliary mass and cholangiocyte proliferation were evaluated by immunohistochemistry. Senescence and fibrosis were measured by quantitative PCR and immunohistochemistry. Hepatic stellate cell activation was detected by immunofluorescence. Histamine and leptin levels were measured by enzyme immunoassay. Leptin receptor (Ob-R) was evaluated by quantitative PCR. The HDC/histamine/histamine receptor axis, ductular reaction, and biliary senescence were evaluated in patients with nonalcoholic fatty liver disease, nonalcoholic steatohepatitis, or end-stage liver disease. Hdc-/- HFD mice had increased steatosis compared with WT HFD mice. WT HFD mice had increased biliary mass, biliary proliferation, senescence, fibrosis, and hepatic stellate cell activation, which were reduced in Hdc-/- HFD mice. In Hdc-/- HFD mice, serum leptin levels increased, whereas biliary Ob-R expression decreased. Nonalcoholic steatohepatitis patients had increased HDC/histamine/histamine receptor signaling. Hdc-/- HFD mice are susceptible to obesity via dysregulated leptin/Ob-R signaling, whereas the lack of HDC protects from HFD-induced fibrosis and cholangiocyte damage. HDC/histamine/leptin signaling may be important in managing obesity-induced biliary damage.
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Dieta Alta en Grasa , Histamina/metabolismo , Histidina Descarboxilasa/metabolismo , Leptina/metabolismo , Cirrosis Hepática/metabolismo , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Adulto , Anciano , Animales , Femenino , Histidina Descarboxilasa/genética , Humanos , Cirrosis Hepática/genética , Cirrosis Hepática/patología , Masculino , Ratones , Ratones Noqueados , Persona de Mediana Edad , Enfermedad del Hígado Graso no Alcohólico/genética , Enfermedad del Hígado Graso no Alcohólico/patología , Transducción de Señal/fisiologíaRESUMEN
Effectiveness and low-toxicity to normal tissues are ideal properties for a cancer treatment, and one that numerous research programs are aiming for. Vitamin C has long been used in the field of Complementary and Alternative Medicine as a cancer treatment, with profound safety and anecdotal efficacy. Recent studies revealed the scientific basis for this use, and indicated that vitamin C, at supra-nutritional doses, holds considerable promise as an effective and low-toxic therapeutic strategy to treat cancer. Reviewed here are the early controversies surrounding vitamin C and cancer treatment, the breakthrough discoveries that led to the current advancement, and recent clinical studies, as well as research into its mechanisms of action.
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Ácido Ascórbico/farmacología , Ácido Ascórbico/uso terapéutico , Neoplasias/tratamiento farmacológico , Animales , HumanosRESUMEN
The present study investigates the anticancer effect of ascorbate in MIA-PaCa-2 human pancreatic cancer cells using both in vitro and in vivo models, with a focus on assessing the role of oxidative stress and autophagy as important mechanistic elements in its anticancer actions. We showed that ascorbate suppresses the growth of human pancreatic cancer cells via the induction of oxidative stress and caspase-independent cell death. Ascorbate induces the formation of autophagosomes and the presence of autophagy inhibitors suppresses ascorbate-induced cell death. These data suggest that the induction of autophagosome formation contributes to ascorbate-induced pancreatic cancer cell death.
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Antineoplásicos/farmacología , Proteínas Reguladoras de la Apoptosis/metabolismo , Ácido Ascórbico/farmacología , Autofagia/efectos de los fármacos , Proteínas de la Membrana/metabolismo , Neoplasias Pancreáticas/tratamiento farmacológico , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Animales , Proteínas Reguladoras de la Apoptosis/genética , Beclina-1 , Muerte Celular/efectos de los fármacos , Línea Celular Tumoral/efectos de los fármacos , Femenino , Humanos , Proteínas de la Membrana/genética , Ratones Desnudos , Proteínas Asociadas a Microtúbulos/metabolismo , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , ARN Interferente Pequeño , Especies Reactivas de Oxígeno/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The prenyl diphosphate synthase subunit 2 (PDSS2) gene has recently been proposed as a novel tumor suppressor in several types of solid tumors. However, the mechanism of its tumor-suppressing activity is not known. Our previous study found a decreased expression of PDSS2 in clinical samples of non-small-cell lung cancer, and an inverse correlation between PDSS2 levels and stages of tumor differentiation and lymph node metastasis. In this study, we further investigated the tumor-suppressing activity of PDSS2 in lung cancer cells using cellular and molecular tools. The PDSS2 gene has low levels of expression in human lung cancer cell lines. We transfected and overexpressed PDSS2 in the NCI-H1299 lung cancer cell line. The forced overexpression caused massive cell death (~70%) through apoptotic pathways and significantly inhibited colony formation. At the same time, repression of PDSS2 expression by siRNA enhanced the growth of a noncancerous lung epithelial cell line MRC-5. There was an inverse correlation (Pearson's test, r=-0.9373) between PDSS2 expression and gelsolin expression, which is known to inhibit apoptosis and enhance cell invasion and metastasis. The ability of PDSS2 to repress gelsolin might contribute to its tumor-suppressing activity. However, PDSS2 did not influence the sensitivity of the lung cancer cells to chemotherapeutic drugs. Taken together, PDSS2 has tumor-suppressing activity in human lung cancer cells by enhancing apoptosis and inhibiting tumorigenic capacity.
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Transferasas Alquil y Aril/metabolismo , Neoplasias Pulmonares/patología , Transferasas Alquil y Aril/genética , Antineoplásicos/farmacología , Apoptosis , Línea Celular Tumoral , Proliferación Celular , Gelsolina/genética , Gelsolina/metabolismo , Genes Supresores de Tumor , Humanos , Neoplasias Pulmonares/metabolismo , ARN Interferente Pequeño/genéticaRESUMEN
The pyrimidine nucleus is an important component of nucleic acids (DNA and RNA) and vitamins (B(2) and folic acid). It is evident from the literature that pyrimidine derivatives possess a wide spectrum of biological activities such as antioxidant, anticancer, antibacterial, and anti-inflammatory activities. On the basis of diverse biological activities, we attempted to synthesize a series of novel bis(2-(pyrimidin-2-yl)ethoxy)alkanes 5a-j in four steps with good yields. 2-Chloropyrimidine (1) was reacted with diethyl malonate in the presence of sodium hydride in dry dimethyl formamide to yield the intermediate diethyl 2-(pyrimidin-2-yl)malonate (2), which on further reaction with sodium chloride and dimethyl sulfoxide yielded ethyl 2-(pyrimidin-2-yl)ethanoate (3). Reduction with sodium borohydride (NaBH(4) ) resulted in the formation of 2-(pyrimidin-2-yl)ethanol (4). This was further reacted with various dibromoalkanes to obtain the title compounds 5a-j. In this current study, we evaluated the antioxidant properties of the title compounds using four in vitro test systems: the 2,2-diphenyl-2-picrylhydrazyl radical-, superoxide radical-, and hydroxyl radical-scavenging assays, and the anti-lipid peroxidation activity test. The title compounds showed promising antioxidant activity when compared to butylated hydroxytoluene. The potency of their antioxidant activity was mainly influenced by the alkyl fragment attached to 2-(pyrimidin-2-yl)ethanol. The ethyl and butyl fragments linked to oxygen led to increased antioxidant activity of the title compounds (i.e., 5b and 5d) in all our in vitro assays.
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Alcanos/síntesis química , Alcanos/farmacología , Antioxidantes/síntesis química , Antioxidantes/farmacología , Pirimidinas/síntesis química , Hidroxitolueno Butilado/farmacología , Evaluación Preclínica de Medicamentos/métodos , Técnicas In Vitro , Peroxidación de Lípido/efectos de los fármacos , Pirimidinas/farmacología , Relación Estructura-Actividad , Temperatura de TransiciónRESUMEN
Methotrexate (MTX) is widely used in the treatment of autoimmune arthritis but is limited by its unpredictable and variable response profile. Currently, no biomarkers exist to predict or monitor early therapeutic responses to MTX. Using a collagen-induced arthritis (CIA) mouse model, this study aimed to identify biochemical pathways and biomarkers associated with MTX efficacy in autoimmune arthritis. Following arthritis disease induction, DBA/1J mice were treated with subcutaneous MTX (20 mg/kg/week) and disease activity was assessed based on disease activity scores (DAS) and paw volume (PV) measurements. Red blood cell (RBC) and plasma samples were collected at the end of the study and were assessed for folate and MTX content. Plasma samples were analyzed by semitargeted global metabolomic profiling and analyzed by univariate and multivariate analysis. Treatment with MTX was associated with significant reductions in disease activity based on both DAS (p = 0.0006) and PV (p = 0.0006). MTX therapy resulted in significant reductions in 5-methyltetrahydrofolate (5mTHF) levels in plasma (p = 0.02) and RBCs (p = 0.001). Reductions in both RBC and plasma 5mTHF were associated with lower DAS (p = 0.0007, p = 0.01, respectively) and PV (p = 0.001, p = 0.005, respectively). Increases in RBC MTX were associated with lower DAS (p = 0.003) but not PV (p = 0.23). Metabolomic analysis identified N-methylisoleucine (NMI) and quinolone as metabolites significantly altered in disease mice, which were corrected towards healthy control levels in mice treated with MTX. Reductions in plasma NMI were associated with lower DAS (p = 0.0002) and PV (p = 9.5 × 10-6). Increases in plasma quinolone were associated with lower DAS (p = 0.02) and PV (p = 0.01). Receiver-operating characteristic curve analysis identified plasma NMI (AUC = 1.00, p = 2.4 × 10-8), RBC 5mTHF (AUC = 0.99, p = 2.4 × 10-5), and plasma quinolone (AUC = 0.89, p = 0.01) as top discriminating metabolites of MTX treatment. Our data support a relationship between MTX efficacy and its effect on circulating folates and identified 5mTHF, NMI, and quinolone as potential therapeutic biomarkers of disease activity and MTX response in the CIA mouse model of autoimmune arthritis.
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BACKGROUND: Inadequate systemic exposure to infliximab (IFX) is associated with treatment failure. This work evaluated factors associated with reduced IFX exposure in children with autoimmune disorders requiring IFX therapy. METHODS: In this single-center cross-sectional prospective study IFX trough concentrations and anti-drug antibodies (ADAs) were measured in serum from children diagnosed with inflammatory bowel disease (IBD) (n = 73), juvenile idiopathic arthritis (JIA) (n = 16), or uveitis (n = 8) receiving maintenance IFX infusions at an outpatient infusion clinic in a tertiary academic pediatric hospital. IFX concentrations in combination with population pharmacokinetic modeling were used to estimate IFX clearance. Patient demographic and clinical data were collected by chart review and evaluated for their relationship with IFX clearance. RESULTS: IFX trough concentrations ranged from 0 to > 40 µg/mL and were 3-fold lower in children with IBD compared to children with JIA (p = 0.0002) or uveitis (p = 0.001). Children with IBD were found to receive lower IFX doses with longer dosing intervals, resulting in dose intensities (mg/kg/day) that were 2-fold lower compared to children with JIA (p = 0.0002) or uveitis (p = 0.02). Use of population pharmacokinetic analysis to normalize for variation in dosing practices demonstrated that increased IFX clearance was associated with ADA positivity (p = 0.004), male gender (p = 0.02), elevated erythrocyte sedimentation rate (ESR) (p = 0.02), elevated c-reactive protein (CRP) (p = 0.001), reduced serum albumin concentrations (p = 0.0005), and increased disease activity in JIA (p = 0.009) and IBD (p ≤ 0.08). No significant relationship between diagnosis and underlying differences in IFX clearance was observed. Multivariable analysis by covariate population pharmacokinetic modeling confirmed increased IFX clearance to be associated with anti-IFX antibody positivity, increased ESR, and reduced serum albumin concentrations. CONCLUSIONS: Enhanced IFX clearance is associated with immunogenicity and inflammatory burden across autoimmune disorders. Higher systemic IFX exposures observed in children with rheumatologic disorders are driven primarily by provider drug dose and interval selection, rather than differences in IFX pharmacokinetics across diagnoses. Despite maintenance IFX dosing at or above the standard recommended range for IBD (i.e., 5 mg/kg every 8 weeks), the dosing intensity used in the treatment of IBD is notably lower than dosing intensities used to treat JIA and uveitis, and may place some children with IBD at risk for suboptimal maintenance IFX exposures necessary for treatment response.
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Artritis Juvenil , Enfermedades Autoinmunes , Monitoreo de Drogas , Enfermedades Inflamatorias del Intestino , Infliximab , Uveítis , Adolescente , Anticuerpos Antiidiotipos/sangre , Artritis Juvenil/sangre , Artritis Juvenil/diagnóstico , Artritis Juvenil/tratamiento farmacológico , Enfermedades Autoinmunes/tratamiento farmacológico , Enfermedades Autoinmunes/epidemiología , Estudios Transversales , Relación Dosis-Respuesta Inmunológica , Monitoreo de Drogas/métodos , Monitoreo de Drogas/normas , Monitoreo de Drogas/estadística & datos numéricos , Femenino , Humanos , Enfermedades Inflamatorias del Intestino/sangre , Enfermedades Inflamatorias del Intestino/diagnóstico , Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , Infliximab/administración & dosificación , Infliximab/inmunología , Infliximab/farmacocinética , Masculino , Tasa de Depuración Metabólica/fisiología , Pediatría/métodos , Inhibidores del Factor de Necrosis Tumoral/administración & dosificación , Inhibidores del Factor de Necrosis Tumoral/inmunología , Inhibidores del Factor de Necrosis Tumoral/farmacocinética , Estados Unidos/epidemiología , Uveítis/sangre , Uveítis/diagnóstico , Uveítis/tratamiento farmacológicoRESUMEN
Anaplastic large cell lymphoma (ALCL) is an aggressive type of non-Hodgkin lymphoma. More than three-fourths of anaplastic lymphoma kinase (ALK)-positive ALCL cases express the nucleophosmin 1 (NPM1)-ALK fusion gene as a result of t(2;5) chromosomal translocation. The homodimerization of NPM1-ALK fusion protein mediates constitutive activation of the chimeric tyrosine kinase activity and downstream signaling pathways responsible for lymphoma cell proliferation and survival. Gilteritinib is a tyrosine kinase inhibitor recently approved by the FDA for the treatment of FMS-like tyrosine kinase mutation-positive acute myeloid leukemia. In this study, we demonstrate for the first time gilteritinib-mediated growth inhibitory effects on NPM1-ALK-driven ALCL cells. We utilized a total of five ALCL model cell lines, including both human and murine. Gilteritinib treatment inhibits NPM1-ALK fusion kinase phosphorylation and downstream signaling, resulting in induced apoptosis. Gilteritinib-mediated apoptosis was associated with caspase 3/9, PARP cleavage, the increased expression of proapoptotic protein BAD, and decreased expression of antiapoptotic proteins, survivin and MCL-1. We also found downregulation of fusion kinase activity resulted in decreased c-Myc protein levels. Furthermore, cell-cycle analysis indicated gilteritinib induced G0-G1-phase cell-cycle arrest and reduced CD30 expression. In summary, our preclinical studies explored the novel therapeutic potential of gilteritinib in the treatment of ALCL cells expressing NPM1-ALK and potentially in other ALK or ALK fusion-driven hematologic or solid malignancies. IMPLICATIONS: Our preclinical results explore the use of gilteritinib for the treatment of NPM1-ALK-driven ALCL cells and pave a path for developing future clinical trials. VISUAL OVERVIEW: http://mcr.aacrjournals.org/content/molcanres/19/5/913/F1.large.jpg.
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Quinasa de Linfoma Anaplásico/metabolismo , Compuestos de Anilina/uso terapéutico , Linfoma Anaplásico de Células Grandes/tratamiento farmacológico , Pirazinas/uso terapéutico , Compuestos de Anilina/farmacología , Humanos , Linfoma Anaplásico de Células Grandes/genética , Pirazinas/farmacologíaRESUMEN
A series of 3-amino-2-hydroxybenzofused 2-phosphalactones (4a-l) has been synthesized from the Kabachnik-Fields reaction via a facile route from a one-pot three-component reaction of diphenylphosphite with various 2-hydroxybenzaldehyes and heterocyclic amines in a new way of expansion. The in vitro anti-cell proliferation studies by MTT assay have revealed them as potential Panc-1, Miapaca-2, and BxPC-3 pancreatic cell growth inhibitors, and the same is supported by molecular docking, QSAR, and ADMET studies. The MTT assay of their SAHA derivatives against the same cell lines evidenced them as potential HDAC inhibitors and identified 4a, 4b, and 4k substituted with 1,3-thiazol, 1,3,4-thiadiazol, and 5-sulfanyl-1,3,4-thiadiazol moieties on phenyl and diethylamino phenyl rings as potential ones. Additionally, the flow cytometric analyses of 4a, 4b, and 4k against BxPC-3 cells revealed compound 4k as a lead compound that arrests the S phase cell cycle growth at low micromolar concentrations. The ADMET properties have ascertained their inherent pharmacokinetic potentiality, and the wholesome results prompted us to report it as the first study on anti-pancreatic cancer activity of cyclic α-aminophosphonates. Ultimately, this study serves as a good contribution to update the existing knowledge on the anticancer organophosphorus heterocyclic compounds and elevates the scope for generation of new anticancer drugs. Further, the studies like QSAR, drug properties, toxicity risks, and bioactivity scores predicted for them have ascertained the synthesized compounds as newer and potential drug candidates. Hence, this study had augmented the array of α-aminophosphonates by adding a new collection of 3-amino-2-hydroxybenzofused 2-phosphalactones, a class of cyclic α-aminophosphonates, to it, which proved them as potential anti-pancreatic cancer agents.
RESUMEN
Pancreatic cancer has poor prognosis and treatment outcomes due to its highly metastatic nature and resistance to current treatments. The RNA-binding protein (RBP) Hu-antigen R (HuR) is a central player in posttranscriptional regulation of cancer-related gene expression, and contributes to tumorigenesis, tumor growth, metastasis, and drug resistance. HuR has been suggested to regulate pancreatic cancer epithelial-to-mesenchymal transition (EMT), but the mechanism was not well understood. Here, we further elucidated the role HuR plays in pancreatic cancer cell EMT, and developed a novel inhibitor specifically interrupting HuR-RNA binding. The data showed that HuR binds to the 3'-UTR of the mRNA of the transcription factor Snail, resulting in stabilization of Snail mRNA and enhanced Snail protein expression, thus promoted EMT, metastasis, and formation of stem-like cancer cells (CSC) in pancreatic cancer cells. siRNA silencing or CRISPR/Cas9 gene deletion of HuR inhibited pancreatic cancer cell EMT, migration, invasion, and inhibited CSCs. HuR knockout cells had dampened tumorigenicity in immunocompromised mice. A novel compound KH-3 interrupted HuR-RNA binding, and KH-3 inhibited pancreatic cancer cell viability, EMT, migration/invasion in vitro KH-3 showed HuR-dependent activity and inhibited HuR-positive tumor growth and metastasis in vivo.
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Proteína 1 Similar a ELAV/genética , Proteína 1 Similar a ELAV/metabolismo , Transición Epitelial-Mesenquimal/genética , Células Madre Neoplásicas/metabolismo , Neoplasias Pancreáticas/etiología , Neoplasias Pancreáticas/metabolismo , Animales , Antineoplásicos/farmacología , Línea Celular Tumoral , Movimiento Celular/genética , Modelos Animales de Enfermedad , Susceptibilidad a Enfermedades , Proteína 1 Similar a ELAV/antagonistas & inhibidores , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Técnicas de Silenciamiento del Gen , Genes Reporteros , Humanos , Ratones , Células Madre Neoplásicas/patología , Neoplasias Pancreáticas/patología , Estabilidad del ARN/efectos de los fármacos , Proteínas de Unión al ARN/genética , Proteínas de Unión al ARN/metabolismo , Factores de Transcripción de la Familia Snail/genética , Factores de Transcripción de la Familia Snail/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Non-small cell lung cancer remains a highly lethal malignancy. Using the tamoxifen inducible Hnf1b:CreERT2 (H) transgenic mouse crossed to the LsL-KrasG12D (K) transgenic mouse, we recently discovered that an Hnf1b positive cell type in the lung is sensitive to adenoma formation when expressing a mutant KrasG12D allele. In these mice, we observe adenoma formation over a time frame of three to six months. To study specificity of the inducible Hnf1b:CreERT2 in the lung, we employed lineage tracing using an mTmG (G) reporter allele. This technique revealed recombined, GFP+ cells were predominantly SPC+. We further employed this technique in HKG mice to determine Hnf1b+ cells give rise to adenomas that express SPC and TTF1. Review of murine lung tissue confirmed a diagnosis of adenoma and early adenocarcinoma, a pathologic subtype of non-small cell lung cancer. Our expanded mouse model revealed loss of Mst1/2 promotes aggressive lung adenocarcinoma and large-scale proteomic analysis revealed upregulation of PKM2 in the lungs of mice with genetic deletion of Mst1/2. PKM2 is a known metabolic regulator in proliferating cells and cancer. Using a human lung adenocarcinoma cell line, we show pharmacologic inhibition of Mst1/2 increases the abundance of PKM2, indicating genetic loss or pharmacologic inhibition of Mst1/2 directly modulates the abundance of PKM2. In conclusion, here we report a novel model of non-small cell lung cancer driven by a mutation in Kras and deletion of Mst1/2 kinases. Tumor development is restricted to a subset of alveolar type II cells expressing Hnf1b. Our data show loss of Mst1/2 regulates levels of a potent metabolic regulator, PKM2.
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Carcinoma de Pulmón de Células no Pequeñas/patología , Transformación Celular Neoplásica , Modelos Animales de Enfermedad , Neoplasias Pulmonares/patología , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas ras/genética , Animales , Carcinoma de Pulmón de Células no Pequeñas/genética , Proteínas Portadoras/metabolismo , Línea Celular Tumoral , Eliminación de Gen , Factor Nuclear 1-beta del Hepatocito/genética , Péptidos y Proteínas de Señalización Intracelular , Pulmón/metabolismo , Pulmón/patología , Neoplasias Pulmonares/genética , Proteínas de la Membrana/metabolismo , Ratones , Ratones Transgénicos , Proteínas Serina-Treonina Quinasas/deficiencia , Proteínas Serina-Treonina Quinasas/genética , Serina-Treonina Quinasa 3 , Hormonas Tiroideas/metabolismo , Proteínas de Unión a Hormona TiroideRESUMEN
Pancreatic cancer remains a highly lethal malignancy. We have recently shown that simultaneous expression of Kras and mutant Tp53R175H promotes invasive ductal adenocarcinoma from pancreatic ductal cells. We hypothesized specific mutations in TP53 have divergent mechanisms of transforming ductal cells. In order to understand the role of mutant TP53 in transforming pancreatic ductal cells, we used a lentiviral system to express mutant TP53R175H and TP53R273H, two of the most frequently mutated TP53 alleles in pancreatic cancer patients, in immortalized, but not transformed, pancreatic ductal epithelial cells carrying a KRAS mutation (HPNE:KRASG12D). Mutant TP53 expression enhanced colony formation and an RPPA assay results revealed TP53R175H uniquely induced HSP70 expression in HPNE:KRASG12D cells. In the context of TP53R175H expression; we observed nuclear localization of HSP70. We performed immunoprecipitation experiments to show mutant p53R175H binds to HSP70. We also provide evidence mutant p53R175H is important for HSP70 stability and, more importantly, HSP70 is required for mutant p53 stability. These data are critical in the context of events leading to cellular transformation in the pancreas.
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Proteínas HSP70 de Choque Térmico/metabolismo , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismo , Carcinogénesis , Línea Celular Tumoral , Núcleo Celular/metabolismo , Supervivencia Celular/fisiología , Proteínas HSP70 de Choque Térmico/biosíntesis , Proteínas HSP70 de Choque Térmico/genética , Humanos , Mutación , Neoplasias Pancreáticas/patología , ProteómicaRESUMEN
Pancreatic cancer is among the most lethal cancers with poorly tolerated treatments. There is increasing interest in using high-dose intravenous ascorbate (IVC) in treating this disease partially because of its low toxicity. IVC bypasses bioavailability barriers of oral ingestion, provides pharmacological concentrations in tissues, and exhibits selective cytotoxic effects in cancer cells through peroxide formation. Here, we further revealed its anti-pancreatic cancer mechanisms and conducted a phase I/IIa study to investigate pharmacokinetic interaction between IVC and gemcitabine. Pharmacological ascorbate induced cell death in pancreatic cancer cells with diverse mutational backgrounds. Pharmacological ascorbate depleted cellular NAD+ preferentially in cancer cells versus normal cells, leading to depletion of ATP and robustly increased α-tubulin acetylation in cancer cells. While ATP depletion led to cell death, over-acetylated tubulin led to inhibition of motility and mitosis. Collagen was increased, and cancer cell epithelial-mesenchymal transition (EMT) was inhibited, accompanied with inhibition in metastasis. IVC was safe in patients and showed the possibility to prolong patient survival. There was no interference to gemcitabine pharmacokinetics by IVC administration. Taken together, these data revealed a multi-targeting mechanism of pharmacological ascorbate's anti-cancer action, with minimal toxicity, and provided guidance to design larger definitive trials testing efficacy of IVC in treating advanced pancreatic cancer.
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Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Apoptosis/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Neoplasias Pancreáticas/tratamiento farmacológico , Animales , Protocolos de Quimioterapia Combinada Antineoplásica/farmacocinética , Ácido Ascórbico/administración & dosificación , Desoxicitidina/administración & dosificación , Desoxicitidina/análogos & derivados , Femenino , Estudios de Seguimiento , Humanos , Infusiones Parenterales , Ratones , Ratones Desnudos , Metástasis de la Neoplasia , Neoplasias Pancreáticas/patología , Pronóstico , Estudios Prospectivos , Distribución Tisular , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de Xenoinjerto , GemcitabinaRESUMEN
Pancreatic cancer is one of the most lethal cancers among all malignances, with a median overall survival of <1 year and a 5-year survival of ~5%. The dismal survival rate and prognosis are likely due to lack of early diagnosis, fulminant disease course, high metastasis rate, and disappointing treatment outcome. Pancreatic cancers harbor a variety of genetic alternations that render it difficult to treat even with targeted therapy. Recent studies revealed that pancreatic cancers are highly enriched with a cancer stem cell (CSC) population, which is resistant to chemotherapeutic drugs, and therefore escapes chemotherapy and promotes tumor recurrence. Cancer cell epithelial to mesenchymal transition (EMT) is highly associated with metastasis, generation of CSCs, and treatment resistance in pancreatic cancer. Reviewed here are the molecular biology of pancreatic cancer, the major signaling pathways regulating pancreatic cancer EMT and CSCs, and the advancement in current clinical and experimental treatments for pancreatic cancer.
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BACKGROUND: Pancreatic cancer has an enrichment of stem-like cancer cells (CSCs) that contribute to chemoresistant tumors prone to metastasis and recurrence. Drug screening assays based on cytotoxicity cannot identify specific CSC inhibitors, because CSCs comprise only a small portion of cancer cell population, and it is difficult to propagate stable CSC populations in vitro for high-throughput screening (HTS) assays. Based on the important role of cancer cell epithelial-to-mesenchymal transition (EMT) in promoting CSCs, we hypothesized that inhibition of EMT can be a useful strategy for inhibiting CSCs, and therefore a feasible approach for HTS can be built for identification of CSC inhibitors, based on assays detecting EMT inhibition. METHODS: An immunofluorescent assay was established and optimized for HTS to identify compounds that enhance E-cadherin expression, as a hallmark of inhibition of EMT. Four chemical libraries containing 41,472 compounds were screened in PANC-1 pancreatic cancer cell line. Positive hits were validated for EMT and CSC inhibition in vitro using sphere formation assay, western blotting, immune fluorescence, and scratch assay. RESULTS: Initial hits were refined to 73 compounds with a secondary screening, among which 17 exhibited concentration dependent induction of E-cadherin expression. Six compounds were selected for further study which belonged to 2 different chemical structural clusters. A novel compound 1-(benzylsulfonyl) indoline (BSI, Compound #38) significantly inhibited pancreatic cancer cell migration and invasion. BSI inhibited histone deacetylase, increased histone 4 acetylation preferably, resulting in E-cadherin up-regulation. BSI effectively inhibited tumor spheres formation. Six more analogues of BSI were tested for anti-migration and anti-CSC activities. CONCLUSION: This study demonstrated a feasible approach for discovery of agents targeting EMT and CSCs using HTS, and identified a class of novel chemicals that could be developed as anti-EMT and anti-CSC drug leads.
Asunto(s)
Transición Epitelial-Mesenquimal/efectos de los fármacos , Bibliotecas de Moléculas Pequeñas/toxicidad , Antígeno CD24/metabolismo , Cadherinas/genética , Cadherinas/metabolismo , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Molécula de Adhesión Celular Epitelial/metabolismo , Ensayos Analíticos de Alto Rendimiento , Humanos , Receptores de Hialuranos/metabolismo , Microscopía Fluorescente , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Bibliotecas de Moléculas Pequeñas/química , Factores de Transcripción de la Familia Snail/antagonistas & inhibidores , Factores de Transcripción de la Familia Snail/metabolismo , Proteína de la Zonula Occludens-1/genética , Proteína de la Zonula Occludens-1/metabolismoRESUMEN
Aberrant regulation of cellular extrusion can promote invasion and metastasis. Here, we identify molecular requirements for early cellular invasion using a premalignant mouse model of pancreatic cancer with conditional knockout of p120 catenin (Ctnnd1). Mice with biallelic loss of p120 catenin progressively develop high-grade pancreatic intraepithelial neoplasia (PanIN) lesions and neoplasia accompanied by prominent acute and chronic inflammatory processes, which is mediated, in part, through NF-κB signaling. Loss of p120 catenin in the context of oncogenic Kras also promotes remarkable apical and basal epithelial cell extrusion. Abundant single epithelial cells exit PanIN epithelium basally, retain epithelial morphology, survive, and display features of malignancy. Similar extrusion defects are observed following p120 catenin knockdown in vitro, and these effects are completely abrogated by the activation of S1P/S1pr2 signaling. In the context of oncogenic Kras, p120 catenin loss significantly reduces expression of genes mediating S1P/S1pr2 signaling in vivo and in vitro, and this effect is mediated at least, in part, through activation of NF-κB. These results provide insight into mechanisms controlling early events in the metastatic process and suggest that p120 catenin and S1P/S1pr2 signaling enhance cancer progression by regulating epithelial cell invasion. Cancer Res; 76(11); 3351-63. ©2016 AACR.
Asunto(s)
Carcinoma in Situ/patología , Carcinoma Ductal Pancreático/patología , Cateninas/metabolismo , Células Epiteliales/patología , Metaplasia/patología , Neoplasias Pancreáticas/patología , Animales , Apoptosis , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Western Blotting , Carcinoma in Situ/genética , Carcinoma in Situ/metabolismo , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/metabolismo , Cateninas/genética , Proliferación Celular , Células Epiteliales/metabolismo , Humanos , Metaplasia/genética , Metaplasia/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , FN-kappa B/genética , FN-kappa B/metabolismo , Invasividad Neoplásica , Estadificación de Neoplasias , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Pronóstico , ARN Mensajero/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transducción de Señal , Células Tumorales Cultivadas , Catenina deltaRESUMEN
Ascorbate (vitamin C) was an early, unorthodox therapy for cancer, with an outstanding safety profile and anecdotal clinical benefit. Because oral ascorbate was ineffective in two cancer clinical trials, ascorbate was abandoned by conventional oncology but continued to be used in complementary and alternative medicine. Recent studies provide rationale for reexamining ascorbate treatment. Because of marked pharmacokinetic differences, intravenous, but not oral, ascorbate produces millimolar concentrations both in blood and in tissues, killing cancer cells without harming normal tissues. In the interstitial fluid surrounding tumor cells, millimolar concentrations of ascorbate exert local pro-oxidant effects by mediating hydrogen peroxide (H(2)O(2)) formation, which kills cancer cells. We investigated downstream mechanisms of ascorbate-induced cell death. Data show that millimolar ascorbate, acting as a pro-oxidant, induced DNA damage and depleted cellular adenosine triphosphate (ATP), activated the ataxia telangiectasia mutated (ATM)/adenosine monophosphate-activated protein kinase (AMPK) pathway, and resulted in mammalian target of rapamycin (mTOR) inhibition and death in ovarian cancer cells. The combination of parenteral ascorbate with the conventional chemotherapeutic agents carboplatin and paclitaxel synergistically inhibited ovarian cancer in mouse models and reduced chemotherapy-associated toxicity in patients with ovarian cancer. On the basis of its potential benefit and minimal toxicity, examination of intravenous ascorbate in combination with standard chemotherapy is justified in larger clinical trials.
Asunto(s)
Antineoplásicos/efectos adversos , Antineoplásicos/uso terapéutico , Ácido Ascórbico/administración & dosificación , Ácido Ascórbico/uso terapéutico , Infusiones Parenterales , Neoplasias Ováricas/tratamiento farmacológico , Animales , Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Ácido Ascórbico/efectos adversos , Carboplatino/farmacología , Carboplatino/uso terapéutico , Muerte Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Sinergismo Farmacológico , Femenino , Ratones , Neoplasias Ováricas/patología , Paclitaxel/farmacología , Paclitaxel/uso terapéutico , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
A series of novel substituted dihydropyrimidine and 5H-thiazolo [3, 2-a] pyrimidine derivatives were designed and synthesized as a potential target to discover drugs fighting against the viral diseases. The main objective of the present work is to carry out the QSAR studies for all the series of the compounds starting from 4a to 6j to find out their molecular descriptors and predict the biological properties. All of them are showing the best QSAR descriptors, hence chosen for the prediction of anti-viral activity against Newcastle disease virus (NDV). Initially their inhibitory activity was predicted by molecular docking of these compounds against haemaglutinin-neuraminidase (HN) protein using molecular operating environment (MOE) software. Based on the best affinity and highest docking scores 4b, 5b and 6b were assayed in vivo on NDV infected chicks and it was found that there is significant improvement in the survival of the chicks with the treatment (P<0.05). 4b and 6b showed better curative effect than 5b at the dose concentration of 40 mg/kg body weight of chicks. The results from molecular docking study and biological assays can be inferred to consider these molecules as potential antiviral drugs.
Asunto(s)
Antivirales/química , Antivirales/farmacología , Virus de la Enfermedad de Newcastle/efectos de los fármacos , Pirimidinas/química , Pirimidinas/farmacología , Animales , Antivirales/administración & dosificación , Pollos , Simulación de Dinámica Molecular , Enfermedad de Newcastle/tratamiento farmacológico , Enfermedades de las Aves de Corral/tratamiento farmacológico , Pirimidinas/administración & dosificación , Relación Estructura-Actividad Cuantitativa , Análisis de SupervivenciaRESUMEN
ABCB6 is a member of the adenosine triphosphate (ATP)-binding cassette family of transporter proteins that is increasingly recognized as a relevant physiological and therapeutic target. Evaluation of modulators of ABCB6 activity would pave the way toward a more complete understanding of the significance of this transport process in tumor cell growth, proliferation and therapy-related drug resistance. In addition, this effort would improve our understanding of the function of ABCB6 in normal physiology with respect to heme biosynthesis, and cellular adaptation to metabolic demand and stress responses. To search for modulators of ABCB6, we developed a novel cell-based approach that, in combination with flow cytometric high-throughput screening (HTS), can be used to identify functional modulators of ABCB6. Accumulation of protoporphyrin, a fluorescent molecule, in wild-type ABCB6 expressing K562 cells, forms the basis of the HTS assay. Screening the Prestwick Chemical Library employing the HTS assay identified four compounds, benzethonium chloride, verteporfin, tomatine hydrochloride and piperlongumine, that reduced ABCB6 mediated cellular porphyrin levels. Validation of the identified compounds employing the hemin-agarose affinity chromatography and mitochondrial transport assays demonstrated that three out of the four compounds were capable of inhibiting ABCB6 mediated hemin transport into isolated mitochondria. However, only verteporfin and tomatine hydrochloride inhibited ABCB6's ability to compete with hemin as an ABCB6 substrate. This assay is therefore sensitive, robust, and suitable for automation in a high-throughput environment as demonstrated by our identification of selective functional modulators of ABCB6. Application of this assay to other libraries of synthetic compounds and natural products is expected to identify novel modulators of ABCB6 activity.
Asunto(s)
Transportadoras de Casetes de Unión a ATP/metabolismo , Citometría de Flujo/métodos , Ensayos Analíticos de Alto Rendimiento , Mitocondrias/efectos de los fármacos , Porfirinas/farmacología , Protoporfirinas/metabolismo , Tomatina/farmacología , Transportadoras de Casetes de Unión a ATP/antagonistas & inhibidores , Bencetonio/farmacología , Transporte Biológico/efectos de los fármacos , Cromatografía de Afinidad , Dioxolanos/farmacología , Hemina/análogos & derivados , Hemina/antagonistas & inhibidores , Hemina/metabolismo , Humanos , Células K562 , Mitocondrias/metabolismo , Modelos Moleculares , Protoporfirinas/antagonistas & inhibidores , Sefarosa/análogos & derivados , Bibliotecas de Moléculas Pequeñas , VerteporfinaRESUMEN
ABCB6 is a mitochondrial transporter that regulates porphyrin biosynthesis. ABCB6 expression is upregulated in hepatocellular carcinoma (HCC) but the significance of this upregulation to HCC is not known. In the present study, we investigated: 1) ABCB6 expression in 18 resected human hepatocellular carcinoma (HCC) tissues and 3 human hepatoma cell lines; 2) pattern of ABCB6 expression during liver disease progression; and 3) functional significance of ABCB6 expression to HCC using the hepatoma cell line Huh7. ABCB6 expression was determined by real-time quantitative reverse transcription-polymerase chain reaction and western blotting. ABCB6 expression was upregulated in all the HCC specimens and the three-hepatoma cell lines. Increased ABCB6 expression correlated with liver disease progression with the pattern of expression being HCC > cirrhosis > steatosis. Small hairpin RNA (shRNA)-mediated knockdown of ABCB6 in Huh7 cells lead to decreased cellular proliferation and colony formation. Attenuation of ABCB6 expression did not affect Huh7 apoptosis but lead to a delay in G2/M phase of the cell cycle. In contrast, ABCB6 overexpression resulted in increased growth and proliferation of Huh7 cells. Since ABCB6 expression is induced in multiple tumor types we explored the role of ABCB6 in other cancer cells. ShRNA mediated knockdown of ABCB6 in HEK293 and K562 cells reduced cellular proliferation leading to a delay in G2/M phase, while ABCB6 overexpression promoted cell growth and proliferation. Collectively, these findings, obtained by loss of function and gain of function analysis, suggest that ABCB6 plays a role in cell growth and proliferation by targeting the cell cycle.