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Estrogen receptor (ER)-negative breast cancers are known to be aggressive and unresponsive to anti-estrogen therapy, and triple negative breast cancers are associated with poor prognosis and metastasis. Thus, new targeted therapies are needed. FOXM1 is abundantly expressed in human cancers and implicated in protecting tumor cells from oxidative stress by reducing the levels of intracellular reactive oxygen species (ROS). Aspirin, a prototypical anti-cancer agent with deleterious side effects, has been modified to release nitric oxide and hydrogen sulfide, called NOSH-aspirin (NOSH-ASA), generating a 'safer' class of new anti-inflammatory agents. We evaluated NOSH-ASA against (ER)-negative breast cancer using cell lines and a xenograft mouse model. NOSH-ASA strongly inhibited growth of MDA-MB-231 and SKBR3 breast cancer cells with low IC50s of 90{plus minus}5 and 82{plus minus}5 nM, respectively, with marginal effects on a normal breast epithelial cell line. NOSH-ASA inhibited cell proliferation, caused G0/G1 phase arrest, increased apoptosis, and was associated with increases in ROS. In MDA-MB-231 cell xenografts, NOSH-ASA reduced tumor size markedly, which was associated with reduced proliferation (decreased PCNA expression), induction of apoptosis (increased TUNEL positive cells), and increased ROS, while NF-kB and FoxM1 that were high in untreated xenografts were significantly reduced. mRNA data for FoxM1, p21 and CyclinD1 corroborated with the respective protein expressions and arrest of cells. Taken together, these molecular events contribute to NOSH-ASA mediated growth inhibition and apoptotic death of (ER)-negative breast cells in vitro and in vivo. Additionally, as a ROS-inducer and FOXM1-inhibitor, NOSH-ASA has potential as a targeted therapy. Significance Statement In this investigation, we examined the cellular effects and xenograft tumor inhibitory potential of NOSH-aspirin, an NO and H2S-donating hybrid, against ER-negative breast cancer, which currently lacks effective therapeutic options. The induction of reactive oxygen species and subsequent downregulation of FOXM1 represents a plausible mechanism contributing to the observed decrease in cell proliferation and concurrent increase in apoptosis. NOSH-ASA demonstrated a remarkable reduction in tumor size by 90% without inducing any observable gross toxicity, underscoring its promising translational potential.
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Post-traumatic stress disorder (PTSD) can develop following severe trauma, but the extent to which genetic and environmental risk factors contribute to individual clinical outcomes is unknown. Here, we compared transcriptional responses to hydrocortisone exposure in human induced pluripotent stem cell (hiPSC)-derived glutamatergic neurons and peripheral blood mononuclear cells (PBMCs) from combat veterans with PTSD (n = 19 hiPSC and n = 20 PBMC donors) and controls (n = 20 hiPSC and n = 20 PBMC donors). In neurons only, we observed diagnosis-specific glucocorticoid-induced changes in gene expression corresponding with PTSD-specific transcriptomic patterns found in human postmortem brains. We observed glucocorticoid hypersensitivity in PTSD neurons, and identified genes that contribute to this PTSD-dependent glucocorticoid response. We find evidence of a coregulated network of transcription factors that mediates glucocorticoid hyper-responsivity in PTSD. These findings suggest that induced neurons represent a platform for examining the molecular mechanisms underlying PTSD, identifying biomarkers of stress response, and conducting drug screening to identify new therapeutics.
Asunto(s)
Células Madre Pluripotentes Inducidas , Trastornos por Estrés Postraumático , Humanos , Trastornos por Estrés Postraumático/genética , Glucocorticoides/farmacología , Leucocitos Mononucleares , Interacción Gen-Ambiente , Células Madre Pluripotentes Inducidas/metabolismo , Expresión Génica , Neuronas/metabolismoRESUMEN
This Article was originally published without the correct Supplemental Table file (Table S1 was missing). In total, there are seven Supplemental Tables, and six were in the original submission. Furthermore, Fig. 1 was misplaced in the main text; it was embedded in the manuscript file even before the results section. Both issues have now been fixed in the HTML and PDF versions of this Article.
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Pancreatic cancer has poor survival rates and largely ineffective therapies. Aspirin is the prototypical anti-cancer agent but its long-term use is associated with significant side effects. NOSH-aspirin belongs to a new class of anti-inflammatory agents that were designed to be safer alternatives by releasing nitric oxide and hydrogen sulfide. In this study we evaluated the effects of NOSH-aspirin against pancreatic cancer using cell lines and a xenograft mouse model. NOSH-aspirin inhibited growth of MIA PaCa-2 and BxPC-3 pancreatic cancer cells with IC50s of 47 ± 5, and 57 ± 4 nM, respectively, while it did not inhibit growth of a normal pancreatic epithelial cell line at these concentrations. NOSH-aspirin inhibited cell proliferation, caused G0/G1 phase cycle arrest, leading to increased apoptosis. Treated cells displayed increases in reactive oxygen species (ROS) and caspase-3 activity. In MIA PaCa-2 cell xenografts, NOSH-aspirin significantly reduced tumor growth and tumor mass. Growth inhibition was due to reduced proliferation (decreased PCNA expression) and induction of apoptosis (increased TUNEL positive cells). Expressions of ROS, iNOS, and mutated p53 were increased; while that of NF-κB and FoxM1 that were high in vehicle-treated xenografts were significantly inhibited by NOSH-aspirin. Taken together, these molecular events and signaling pathways contribute to NOSH-aspirin mediated growth inhibition and apoptotic death of pancreatic cancer cells in vitro and in vivo.
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Aspirina/análogos & derivados , Proliferación Celular/efectos de los fármacos , Disulfuros/farmacología , Nitratos/farmacología , Neoplasias Pancreáticas/tratamiento farmacológico , Ensayos Antitumor por Modelo de Xenoinjerto , Animales , Aspirina/química , Aspirina/farmacología , Caspasa 3/metabolismo , Línea Celular Tumoral , Disulfuros/química , Proteína Forkhead Box M1/metabolismo , Masculino , Ratones Desnudos , Ratones SCID , FN-kappa B/metabolismo , Nitratos/química , Óxido Nítrico Sintasa de Tipo II/metabolismo , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Especies Reactivas de Oxígeno/metabolismo , Carga Tumoral/efectos de los fármacosRESUMEN
Post-traumatic stress disorder (PTSD) is a condition of stress reactivity, whose clinical manifestations are evident when patients are triggered following exposure to a traumatic event. While baseline differences in gene expression of glucocorticoid signaling and inflammatory cytokines in peripheral blood mononuclear cells (PBMCs) have been associated with PTSD, these alterations do not fully recapitulate the molecular response to physiological triggers, such as stress hormones. Therefore, it is critical to develop new techniques that will capture the dynamic transcriptional response associated with stress-activated conditions relative to baseline conditions. To achieve this goal, cultured PBMCs from combat-exposed veterans with PTSD(+) (n = 10) and without PTSD(-) (n = 10) were incubated with increasing concentrations (vehicle, 2.5 nM, 5 nM, 50 nM) of dexamethasone (DEX). Across diagnosis and dosage, several genes and gene networks were reliable markers of glucocorticoid stimulation (FDR < 5%), including enhanced expression of FKPB5, VIPR1, NR1I3, and apoptosis-related pathways, and reduced expression of NR3C1, STAT1, IRF1, and related inflammatory and cellular stress-responsive pathways. Dose-dependent differential transcriptional changes in several genes were also identified between PTSD+ and PTSD-. Robust changes in expression were observed at 2.5 nM DEX in PTSD- but not PTSD+ participants; whereas, with increasing concentrations (5 nM and 50 nM), several genes were identified to be uniquely up-regulated in PTSD+ but not PTSD- participants. Collectively, these preliminary findings suggest that genome-wide gene expression profiling of DEX-stimulated PBMCs is a promising method for the exploration of the dynamic differential molecular responses to stress hormones in PTSD, and may identify novel markers of altered glucocorticoid signaling and responsivity in PTSD.
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Dexametasona/administración & dosificación , Expresión Génica/efectos de los fármacos , Redes Reguladoras de Genes/efectos de los fármacos , Glucocorticoides/administración & dosificación , Leucocitos Mononucleares/metabolismo , Trastornos por Estrés Postraumático/diagnóstico , Transcripción Genética/efectos de los fármacos , Adulto , Biomarcadores/metabolismo , Receptor de Androstano Constitutivo , Relación Dosis-Respuesta a Droga , Perfilación de la Expresión Génica , Humanos , Leucocitos Mononucleares/efectos de los fármacos , Masculino , Persona de Mediana Edad , Trastornos por Estrés Postraumático/metabolismo , Veteranos , Adulto JovenRESUMEN
Vascular endothelial cells play an important role in the regulation of vascular function in response to mechanical stimuli in both healthy and diseased states. Prostaglandin I2 (PGI2) is an important antiatherogenic prostanoid and vasodilator produced in endothelial cells through the action of the cyclooxygenase (COX) isoenzymes COX-1 and COX-2. However, the mechanisms involved in sustained, shear-induced production of COX-2 and PGI2 have not been elucidated but are determined in the present study. We used cultured endothelial cells exposed to steady fluid shear stress (FSS) of 10 dyn/cm2 for 5 h to examine shear stress-induced induction of COX-2/PGI2 Our results demonstrate the relationship between the mechanosensor platelet endothelial cell adhesion molecule-1 (PECAM-1) and the intracellular mechanoresponsive molecules phosphatidylinositol 3-kinase (PI3K), focal adhesion kinase (FAK), and mitogen-activated protein kinase p38 in the FSS induction of COX-2 expression and PGI2 release. Knockdown of PECAM-1 (small interference RNA) expression inhibited FSS-induced activation of α5ß1-integrin, upregulation of COX-2, and release of PGI2 in both bovine aortic endothelial cells (BAECs) and human umbilical vein endothelial cells (HUVECs). Furthermore, inhibition of the PI3K pathway (LY294002) substantially inhibited FSS activation of α5ß1-integrin, upregulation of COX-2 gene and protein expression, and release of PGI2 in BAECs. Inhibition of integrin-associated FAK (PF573228) and MAPK p38 (SB203580) also inhibited the shear-induced upregulation of COX-2. Finally, a PECAM-1-/- mouse model was characterized by reduced COX-2 immunostaining in the aorta and reduced plasma PGI2 levels compared with wild-type mice, as well as complete inhibition of acute flow-induced PGI2 release compared with wild-type animals.NEW & NOTEWORTHY In this study we determined the major mechanotransduction pathway by which blood flow-driven shear stress activates cyclooxygenase-2 (COX-2) and prostaglandin I2 (PGI2) release in endothelial cells. Our work has demonstrated for the first time that COX-2/PGI2 mechanotransduction is mediated by the mechanosensor platelet endothelial cell adhesion molecule-1 (PECAM-1).
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Ciclooxigenasa 2/biosíntesis , Células Endoteliales/metabolismo , Epoprostenol/biosíntesis , Estrés Mecánico , Animales , Bovinos , Línea Celular , Cilios/metabolismo , Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Humanos , Inmunohistoquímica , Integrinas/metabolismo , Péptidos/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Fosforilación , Molécula-1 de Adhesión Celular Endotelial de Plaqueta/metabolismo , Transducción de Señal/fisiología , Regulación hacia Arriba , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
Naproxen (NAP) is a potent nonsteroidal anti-inflammatory drug (NSAID) with a favorable cardiovascular profile. However, its long-term use may lead to serious gastrointestinal and renal side effects. NOSH- (nitric oxide and hydrogen sulfide) releasing naproxen (NOSH-NAP, AVT-219) belongs to a new class of anti-inflammatory agents designed to overcome these limitations. We compared the gastrointestinal safety, anti-inflammatory, analgesic, antipyretic, and antiplatelet properties of AVT-219 to that of NAP in preclinical animal models. We also evaluated its anticancer effects in 11 human cancer cell (HCC) lines of six different tissue origins and in a chemotherapeutic xenograft mouse model of colon cancer. AVT-219: (1) was orders of magnitude more potent than NAP in inhibiting the growth of cultured HCC; (2) was safe to the stomach, whereas NAP caused significant ulceration; (3) showed strong anti-inflammatory, analgesic, antipyretic, and antiplatelet properties comparable to NAP; and (4) NAP caused a significant rise in plasma tumor necrosis factor-alpha (TNFα), whereas in the AVT-219-treated rats this rise was significantly less. Mechanistically, AVT-219 was a strong antioxidant, inhibited cyclooxygenase (COX)-1 and -2, thus reducing prostaglandin (PG) E2. In xenografts, AVT-219 significantly reduced tumor growth and tumor mass with no sign of GI toxicity, whereas NAP-treated mice died due to GI bleeding. AVT-219 displayed considerable safety and potency in inhibiting HCC growth; was an effective analgesic, antipyretic, antiplatelet, and anti-inflammatory; and was significantly more efficacious than NAP in reducing the growth of established tumors in a xenograft mouse model.
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Colorectal cancer (CRC) is the second leading cause of death due to cancer and the third most common cancer in men and women in the USA. Nuclear factor kappa B (NF-κB) is known to be activated in CRC and is strongly implicated in its development and progression. Therefore, activated NF-κB constitutes a bona fide target for drug development in this type of malignancy. Many epidemiological and interventional studies have established nonsteroidal anti-inflammatory drugs (NSAIDs) as a viable chemopreventive strategy against CRC. Our previous studies have shown that several novel hydrogen sulfide-releasing NSAIDs are promising anticancer agents and are safer derivatives of NSAIDs. In this study, we examined the growth inhibitory effect of a novel H2S-releasing naproxen (HS-NAP), which has a repertoire as a cardiovascular-safe NSAID, for its effects on cell proliferation, cell cycle phase transitions, and apoptosis using HT-29 human colon cancer cells. We also investigated its effect as a chemo-preventive agent in a xenograft mouse model. HS-NAP suppressed the growth of HT-29 cells by induction of G0/G1 arrest and apoptosis and downregulated NF-κB. Tumor xenografts in mice were significantly reduced in volume. The decrease in tumor mass was associated with a reduction of cell proliferation, induction of apoptosis, and decreases in NF-κB levels in vivo. Therefore, HS-NAP demonstrates strong anticancer potential in CRC.
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Anticarcinógenos/farmacología , Proliferación Celular/efectos de los fármacos , Neoplasias Colorrectales/prevención & control , Sulfuro de Hidrógeno/farmacología , FN-kappa B/metabolismo , Naproxeno/farmacología , Transducción de Señal/efectos de los fármacos , Animales , Anticarcinógenos/metabolismo , Apoptosis/efectos de los fármacos , Ciclo Celular/efectos de los fármacos , Neoplasias Colorrectales/metabolismo , Neoplasias Colorrectales/patología , Relación Dosis-Respuesta a Droga , Células HT29 , Humanos , Sulfuro de Hidrógeno/metabolismo , Masculino , Ratones Desnudos , Naproxeno/metabolismo , Reductasa de Tiorredoxina-Disulfuro/metabolismo , Factores de Tiempo , Carga Tumoral/efectos de los fármacos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
We recently reported the synthesis of NOSH-aspirin, a novel hybrid compound capable of releasing both nitric oxide (NO) and hydrogen sulfide (H2S). In NOSH-aspirin, the two moieties that release NO and H2S are covalently linked at the 1, 2 positions of acetyl salicylic acid, i.e., ortho-NOSH-aspirin. Here we report on the synthesis of meta- and para-NOSH-aspirins. We also made a head-to-head evaluation of the effects of these three positional isomers of NOSH-aspirin on colon cancer cell kinetics and induction of reactive oxygen species, which in recent years has emerged as a key event in causing cancer cell regression. Electron donating/withdrawing groups incorporated about the benzoate moiety significantly affected the potency of these compounds with respect to colon cancer cell growth inhibition.
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Antineoplásicos/síntesis química , Antineoplásicos/farmacología , Aspirina/análogos & derivados , Disulfuros/farmacología , Sulfuro de Hidrógeno/metabolismo , Nitratos/farmacología , Óxido Nítrico/metabolismo , Antineoplásicos/química , Apoptosis/efectos de los fármacos , Aspirina/síntesis química , Aspirina/química , Aspirina/farmacología , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Disulfuros/síntesis química , Disulfuros/química , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Células HT29 , Humanos , Estructura Molecular , Nitratos/síntesis química , Nitratos/química , Especies Reactivas de Oxígeno/metabolismo , Relación Estructura-ActividadRESUMEN
Aspirin is chemopreventive; however, side effects preclude its long-term use. NOSH-aspirin (NBS-1120), a novel hybrid that releases nitric oxide and hydrogen sulfide, was designed to be a safer alternative. Here we compare the gastrointestinal safety, anti-inflammatory, analgesic, anti-pyretic, anti-platelet, and chemopreventive properties of aspirin and NBS-1120 administered orally to rats at equimolar doses. Gastrointestinal safety: 6h post-administration, the number and size of hemorrhagic lesions in stomachs were counted; tissue samples were frozen for PGE2, SOD, and MDA determination. Anti-inflammatory: 1h after drug administration, the volume of carrageenan-induced rat paw edemas was measured for 5h. Anti-pyretic: fever was induced by LPS (ip) an hour before administration of the test drugs, core body temperature was measured hourly for 5h. Analgesic: time-dependent analgesic effects were evaluated by carrageenan-induced hyperalgesia. Antiplatelet: anti-aggregatory effects were studied on collagen-induced platelet aggregation of human platelet-rich plasma. Chemoprevention: nude mice were gavaged daily for 25 days with vehicle, aspirin or NBS-1120. After one week, each mouse was inoculated subcutaneously in the right flank with HT-29 human colon cancer cells. Both agents reduced PGE2 levels in stomach tissue; however, NBS-1120 did not cause any stomach ulcers, whereas aspirin caused significant bleeding. Lipid peroxidation induced by aspirin was higher than that exerted by NBS-1120. SOD activity was significantly inhibited by aspirin but increased by NBS-1120. Both agents showed similar anti-inflammatory, analgesic, anti-pyretic, and anti-platelet activities. Aspirin increased plasma TNFα more than NBS-1120-treated animals. NBS-1120 was better than aspirin as a chemopreventive agent; it dose-dependently inhibited tumor growth and tumor mass.
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Antiinflamatorios no Esteroideos/uso terapéutico , Aspirina/análogos & derivados , Quimioprevención/métodos , Disulfuros/uso terapéutico , Mucosa Gástrica/metabolismo , Sulfuro de Hidrógeno/metabolismo , Nitratos/uso terapéutico , Óxido Nítrico/metabolismo , Animales , Antiinflamatorios no Esteroideos/farmacología , Aspirina/farmacología , Aspirina/uso terapéutico , Disulfuros/farmacología , Relación Dosis-Respuesta a Droga , Mucosa Gástrica/efectos de los fármacos , Tracto Gastrointestinal/efectos de los fármacos , Tracto Gastrointestinal/metabolismo , Células HT29 , Humanos , Masculino , Ratones , Ratones Desnudos , Nitratos/farmacología , Ratas , Ratas Wistar , Ensayos Antitumor por Modelo de Xenoinjerto/métodosRESUMEN
We recently reported the synthesis of NOSH-aspirin, a novel hybrid that releases both nitric oxide (NO) and hydrogen sulfide (H2S). In NOSH-aspirin, the two moieties that release NO and H2S are covalently linked at the 1, 2 positions of acetyl salicylic acid, i.e. ortho-NOSH-aspirin (o-NOSH-aspirin). In the present study, we compared the effects of the positional isomers of NOSH-ASA (o-NOSH-aspirin, m-NOSH-aspirin and p-NOSH-aspirin) to that of aspirin on growth of HT-29 and HCT 15 colon cancer cells, belonging to the same histological subtype, but with different expression of cyclooxygenase (COX) enzymes; HT-29 express both COX-1 and COX-2, whereas HCT 15 is COX-null. We also analyzed the effect of these compounds on proliferation and apoptosis in HT-29 cells. Since the parent compound aspirin, inhibits both COX-1 and COX-2, we also evaluated the effects of these compounds on COX-1 and COX-2 enzyme activities and also performed modeling of the interactions between the positional isomers of NOSH-aspirin and COX-1 and COX-2 enzymes. We observed that the three positional isomers of NOSH aspirin inhibited the growth of both colon cancer cell lines with IC50s in the nano-molar range. In particular in HT-29 cells the IC50s for growth inhibition were: o-NOSH-ASA, 0.04±0.011 µM; m-NOSH-ASA, 0.24±0.11 µM; p-NOSH-ASA, 0.46±0.17 µM; and in HCT 15 cells the IC50s for o-NOSH-ASA, m-NOSH-ASA, and p-NOSH-ASA were 0.062 ±0.006 µM, 0.092±0.004 µM, and 0.37±0.04 µM, respectively. The IC50 for aspirin in both cell lines was >5mM at 24h. The reduction of cell growth appeared to be mediated through inhibition of proliferation, and induction of apoptosis. All 3 positional isomers of NOSH-aspirin preferentially inhibited COX-1 over COX-2. These results suggest that the three positional isomers of NOSH-aspirin have the same biological actions, but that o-NOSH-ASA displayed the strongest anti-neoplastic potential.
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Antineoplásicos/química , Aspirina/análogos & derivados , Ciclooxigenasa 1/química , Ciclooxigenasa 2/química , Inhibidores de la Ciclooxigenasa/química , Disulfuros/química , Nitratos/química , Donantes de Óxido Nítrico/química , Animales , Antineoplásicos/síntesis química , Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Aspirina/síntesis química , Aspirina/química , Aspirina/farmacología , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Ciclooxigenasa 1/genética , Ciclooxigenasa 1/metabolismo , Ciclooxigenasa 2/genética , Ciclooxigenasa 2/metabolismo , Inhibidores de la Ciclooxigenasa/síntesis química , Inhibidores de la Ciclooxigenasa/farmacología , Disulfuros/síntesis química , Disulfuros/farmacología , Expresión Génica , Células HT29 , Humanos , Sulfuro de Hidrógeno/química , Sulfuro de Hidrógeno/metabolismo , Concentración 50 Inhibidora , Simulación del Acoplamiento Molecular , Nitratos/síntesis química , Nitratos/farmacología , Óxido Nítrico/química , Óxido Nítrico/metabolismo , Donantes de Óxido Nítrico/síntesis química , Donantes de Óxido Nítrico/farmacología , Ovinos , Estereoisomerismo , Relación Estructura-ActividadRESUMEN
Sulindac is chemopreventive and has utility in patients with familial adenomatous polyposis; however, side effects preclude its long-term use. NOSH-sulindac (AVT-18A) releases nitric oxide and hydrogen sulfide, was designed to be a safer alternative. Here we compare the gastrointestinal safety, anti-inflammatory, analgesic, anti-pyretic, anti-platelet, and anti-cancer properties of sulindac and NOSH-sulindac administered orally to rats at equimolar doses. Gastrointestinal safety: 6h post-administration, number/size of hemorrhagic lesions in stomachs were counted. Tissue samples were frozen for PGE2, SOD, and MDA determination. Anti-inflammatory: 1h after drug administration, the volume of carrageenan-induced rat paw edemas was measured for 5h. Anti-pyretic: fever was induced by LPS (ip) an hour before administration of the test drugs, core body temperature was measured hourly for 5h. Analgesic: time-dependent analgesic effects were evaluated by carrageenan-induced hyperalgesia. Antiplatelet: anti-aggregatory effects were studied on collagen-induced platelet aggregation of human platelet-rich plasma. Anti-cancer: We examined the effects of NOSH-sulindac on the growth properties of 12 human cancer cell lines of six different tissue origins. Both agents reduced PGE2 levels in stomach tissue; however, NOSH-sulindac did not cause any stomach ulcers, whereas sulindac caused significant bleeding. Lipid peroxidation induced by sulindac was higher than that from NOSH-sulindac. SOD activity was significantly lowered by sulindac but increased by NOSH-sulindac. Both agents showed similar anti-inflammatory, analgesic, anti-pyretic, and anti-platelet activities. Sulindac increased plasma TNFα whereas this rise was lower in the NOSH-sulindac-treated animals. NOSH-sulindac inhibited the growth of all cancer cell lines studied, with potencies of 1000- to 9000-fold greater than that of sulindac. NOSH-sulindac inhibited cell proliferation, induced apoptosis, and caused G2/M cell cycle block. These results demonstrate that NOSH-sulindac is gastrointestinal safe, and maintains the anti-inflammatory, analgesic, antipyretic, and antiplatelet properties of its parent compound sulinsac, with anti-growth activity against a wide variety of human cancer cells.
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Antiinflamatorios no Esteroideos/farmacología , Antineoplásicos/farmacología , Antipiréticos/farmacología , Sulfuro de Hidrógeno/farmacología , Óxido Nítrico/farmacología , Sulindac/farmacología , Administración Oral , Analgésicos/química , Analgésicos/farmacología , Animales , Antiinflamatorios no Esteroideos/química , Antineoplásicos/química , Antipiréticos/química , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Dinoprostona/metabolismo , Edema/tratamiento farmacológico , Edema/metabolismo , Edema/patología , Fiebre/tratamiento farmacológico , Fiebre/metabolismo , Fiebre/patología , Sulfuro de Hidrógeno/química , Hiperalgesia/tratamiento farmacológico , Hiperalgesia/metabolismo , Hiperalgesia/patología , Masculino , Malondialdehído/metabolismo , Óxido Nítrico/química , Donantes de Óxido Nítrico/química , Donantes de Óxido Nítrico/farmacología , Agregación Plaquetaria/efectos de los fármacos , Inhibidores de Agregación Plaquetaria/química , Inhibidores de Agregación Plaquetaria/farmacología , Ratas , Ratas Wistar , Sulindac/análogos & derivados , Superóxido Dismutasa/metabolismoRESUMEN
Estrogen receptor negative (ER(-)) breast cancer is aggressive, responds poorly to current treatments and has a poor prognosis. The NF-κB signaling pathway is implicated in ER(-) tumorigenesis. Aspirin (ASA) is chemopreventive against ER(+) but not for ER(-) breast cancers. Nitric oxide-releasing aspirin (NO-ASA) is a safer ASA where ASA is linked to an NO-releasing moiety through a spacer. In vitro, we investigated anti-proliferation effects of NO-ASA (para- and meta-isomers) against ER(-) breast cancer cells MDA-MB-231 and SK-BR-23, effects on NF-κB signaling, and reactive oxygen species by standard techniques. In vivo, effects of NO-ASA were evaluated in a mouse xenograft model using MDA-MB-231 cells. p-NO-ASA inhibited the growth of MDA-MB-231 and SK-BR-3 cells at 24 h, the respective IC50s were 13 ± 2 and 17 ± 2 µM; ASA had an IC50 of >3000 µM in both cell lines. The IC50s for m-NO-ASA in MDA-MB-231 and SK-BR-3 were 173 ± 15 and 185 ± 12 µM, respectively, therefore, implying p-NO-ASA as a stronger inhibitor of growth p-NO-ASA reduced cell growth by inhibiting proliferation, inducing apoptosis and causing G0/G1 cell cycle block. Activation of NF-κB was inhibited by both isomers as demonstrated by decreases in NF-κB-DNA binding and luciferase activity at 24 h, However, m-NO-ASA produced transient effects at 3 h such as increased NF-κB-DNA-binding, increased levels of nuclear p50, even though both isomers inhibited IκB degradation. Increase in nuclear p50 by m-NO-ASA was associated with translocation of p50 in to the nucleus as observed by immunoflouresence at 3 h. NO-ASA induced reactive oxygen species (ROS) as evidenced by overall increases in both H2DCFDA (2',7'-dichlorodihydrofluorescein) and DHE (dihydroethidium)-derived fluorescence. Inhibition of ROS by N-acetyl-cysteine reversed the m-NO-ASA-mediated translocation of p50 in to the nucleus. In xenografts, p-NO-ASA inhibited tumor growth by inhibiting proliferation (PCNA and tumor volume), inducing apoptosis (TUNEL positive cells) and reducing NF-κB expression. Both isomers inhibit cancer cells, inhibit NF-κB pathway and induce ROS, and have potential as anticancer compounds.
Asunto(s)
Antineoplásicos/farmacología , Aspirina/análogos & derivados , Neoplasias de la Mama/tratamiento farmacológico , Regulación Neoplásica de la Expresión Génica , FN-kappa B/antagonistas & inhibidores , Receptor ErbB-2/deficiencia , Receptores de Estrógenos/deficiencia , Animales , Apoptosis/efectos de los fármacos , Aspirina/farmacología , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Femenino , Genes Reporteros , Humanos , Concentración 50 Inhibidora , Inyecciones Subcutáneas , Luciferasas/genética , Luciferasas/metabolismo , Ratones , Ratones Desnudos , FN-kappa B/genética , FN-kappa B/metabolismo , Antígeno Nuclear de Célula en Proliferación/genética , Antígeno Nuclear de Célula en Proliferación/metabolismo , Especies Reactivas de Oxígeno/agonistas , Especies Reactivas de Oxígeno/metabolismo , Receptor ErbB-2/genética , Receptores de Estrógenos/genética , Transducción de Señal , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
BACKGROUND: NOSH-aspirin, a novel hybrid that releases nitric oxide (NO) and hydrogen sulfide (H2S) was designed to overcome the potential side effects of aspirin. AIM: We compared the cell growth inhibitory properties of ortho-, meta-, and para-NOSH-aspirins. Effects of electron donating/withdrawing groups on the stability and biological activity of these novel compounds were also evaluated. METHODS: Cell line: HT-29 (Cyclooxygenase, COX-1 & -2 expressing) and HCT 15 (COX null) human colon adenocarcimoa; Cell growth: MTT; Cell cycle phase distribution: Flow cytometry; Apoptosis: subdiploid (sub-G0/G1) peak in DNA content histograms; Proliferation: PCNA; ROS: measured hydrogen peroxide and super oxide by flow cytometry using DCFDA and DHE dyes. RESULTS: The IC50s for growth inhibition in µM at 24h were, HT-29: ortho-NOSH-ASA (0.04±0.011), meta-NOSH-ASA (0.24±0.11), para-NOSH-ASA (0.46±0.17); significance between the groups were: o vs m P>0.05, o vs p P<0.05, m vs p P>0.05; HCT 15: ortho-NOSH-ASA (0.062±0.006), meta-NOSH-ASA (0.092±0.004), para-NOSH-ASA (0.37±0.04); significance between the groups were: o vs m P<0.01, o vs p P<0.001, m vs p P<0.001. Electron donating/withdrawing groups significantly affected these IC50s. All positional isomers qualitatively had similar effects on proliferation, apoptosis, and caused G0/G1 cell cycle arrest in both colon cancer cell lines. The underlying mechanism for these observations appeared to be mediated through ROS, as pretreatment of the cells with N-acetylcysteine, partially blocked these effects. CONCLUSIONS: Positional isomerism affects the potency of NOSH-aspirin. The effects appear to be COX independent.
Asunto(s)
Aspirina/análogos & derivados , Neoplasias del Colon/tratamiento farmacológico , Disulfuros/química , Disulfuros/farmacología , Nitratos/química , Nitratos/farmacología , Aspirina/química , Aspirina/farmacología , Línea Celular Tumoral , Neoplasias del Colon/metabolismo , Neoplasias del Colon/patología , Ciclooxigenasa 1/biosíntesis , Ciclooxigenasa 1/genética , Ciclooxigenasa 2/biosíntesis , Ciclooxigenasa 2/genética , Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Proteínas de Neoplasias/biosíntesis , Proteínas de Neoplasias/genéticaRESUMEN
Nitric oxide- (NO) and hydrogen sulfide- (H2S) releasing naproxen (NOSH-naproxen) and NO and H2S-releasing sulindac (NOSH-sulindac) were synthesized and their cell growth inhibitory properties were evaluated in four different human cancer cell lines. These cell lines are of adenomatous (colon, pancreas), epithelial (breast), and lymphocytic (leukemia) origin. Using HT-29 human colon cancer cells, NOSH-naproxen and NOSH-sulindac increased apoptosis, and inhibited proliferation. NOSH-naproxen caused a G0/G1 whereas NOSH-sulindac caused a G2/M block in the cell cycle. Both compounds exhibited significant anti-inflammatory properties, using the carrageenan rat paw edema model. Reconstitution and structure-activity studies representing a fairly close approximation to the intact molecule showed that NOSH-naproxen was approximately 8000-fold more potent than the sum of its parts in inhibiting cell growth. Our data suggest that these compounds merit further investigation as potential anti-cancer agents.
RESUMEN
Hydrogen sulfide-releasing aspirin (HS-ASA) is a novel compound with potential against cancer. It inhibited the growth of Jurkat T-leukemia cells with an IC50 of 1.9 ± 0.2 µM whereas that of ASA was >5000 µM. It dose-dependently inhibited proliferation and induced apoptosis in these cells, causing a G0/G1 cell cycle arrest. HS-ASA down-regulated ß-catenin protein levels and reduced mRNA and protein expression of ß-catenin/TCF downstream target genes cyclinD1 and c-myc. Aspirin up to 5 mM had no effect on ß-catenin expression. HS-ASA also increased caspase-3 protein levels and dose-dependently increased its activity. These effects were substantially blocked by z-VAD-fmk, a pan-caspase inhibitor.
Asunto(s)
Aspirina/química , Aspirina/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Sulfuro de Hidrógeno/química , beta Catenina/genética , Caspasa 3/metabolismo , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Activación Enzimática/efectos de los fármacos , Humanos , Células Jurkat , beta Catenina/metabolismoRESUMEN
We compared the differential effects of positional isomers of acetylsalicylic acid (o-ASA, m-ASA, and p-ASA) on cyclooxygenase (COX) inhibition, gastric prostaglandin E2 (PGE2), malondialdehyde, tumor necrosis factor-alpha (TNF-α) levels, superoxide dismutase (SOD) activity, human adenocarcinoma colon cancer cell growth inhibition, cell proliferation, apoptosis, and cell-cycle progression. We also evaluated the gastric toxicity exerted by ASA isomers. All ASA isomers inhibit COX enzymes, but only the o-ASA exerted an irreversible inhibitory profile. We did not observe a significant difference between ASA isomers in their ability to decrease the in vivo synthesis of PGE2 and SOD activity. Furthermore, all isomers increased the levels of gastric and TNF-α when administered orally at equimolar doses. We observed a dose-dependent cell growth inhibitory effect; the order of potency was p-ASA > m-ASA ≈ o-ASA. There was a dose-dependent decrease in cell proliferation and an increase in apoptosis, with a concomitant Go/G1 arrest. The ulcerogenic profile of the three ASA isomers showed a significant difference between o-ASA (aspirin) and its two positional isomers when administered orally at equimolar doses (1 mmol/kg); the ulcer index (UI) for o-ASA indicated extensive mucosal injury (UI = 38), whereas m-ASA and p-ASA produced a significantly decreased toxic response (UI = 12 and 8, respectively) under the same experimental conditions. These results suggest that the three positional isomers of ASA exert practically the same biologic profile in vitro and in vivo but showed different safety profiles. The mechanism of gastric ulcer formation exerted by aspirin and its two isomers warrants a more detailed and thorough investigation.
Asunto(s)
Antineoplásicos/farmacología , Aspirina/farmacología , Proliferación Celular/efectos de los fármacos , Neoplasias del Colon/prevención & control , Inhibidores de la Ciclooxigenasa/farmacología , Prostaglandina-Endoperóxido Sintasas/metabolismo , Animales , Antineoplásicos/química , Antineoplásicos/toxicidad , Apoptosis/efectos de los fármacos , Aspirina/química , Aspirina/toxicidad , Ciclo Celular/efectos de los fármacos , Cromatografía Líquida de Alta Presión , Neoplasias del Colon/enzimología , Neoplasias del Colon/patología , Inhibidores de la Ciclooxigenasa/química , Inhibidores de la Ciclooxigenasa/toxicidad , Relación Dosis-Respuesta a Droga , Células HT29 , Humanos , Isomerismo , Masculino , Modelos Moleculares , Estructura Molecular , Ratas , Ratas Wistar , Úlcera Gástrica/inducido químicamente , Relación Estructura-ActividadRESUMEN
A series of new hybrids of aspirin (ASA), bearing both nitric oxide (NO) and hydrogen sulfide (H(2)S)-releasing moieties were synthesized and designated as NOSH compounds (1-4). NOSH-1 (4-(3-thioxo-3H-1,2-dithiol-5-yl) phenyl 2-((4-(nitrooxy)-butanoyl)oxy) benzoate); NOSH-2 (4-(nitrooxy)butyl (2-((4-(3-thioxo-3H-1,2-dithiol-5-yl)phenoxy)carbonyl)phenyl)); NOSH-3 (4-carbamothioylphenyl 2-((4-(nitrooxy)butanoyl)-oxy)benzoate); and NOSH-4 (4-(nitrooxy)butyl 2-(5-((R)-1,2-dithiolan-3-yl)pentanoyloxy)-benzoate). The cell growth inhibitory properties of compounds 1-4 were evaluated in eleven different human cancer cell lines of six different tissue origins. These cell lines are of adenomatous (colon, pancreatic, lung, prostate), epithelial (breast), and lymphocytic (leukemia) origin. All NOSH compounds were extremely effective in inhibiting the growth of these cell lines. NOSH-1 was the most potent, with an IC(50) of 48 ± 3 nM in HT-29 colon cancer cells. This is the first NSAID-based compound with such potency. This compound was also devoid of any cellular toxicity, as determined by LDH release. NOSH-1 was comparable to aspirin in its anti-inflammatory properties, using the carrageenan rat paw edema model.
RESUMEN
Nonsteroidal anti-inflammatory drugs (NSAIDs) are prototypical anti-cancer agents. However, their long-term use is associated with adverse gastrointestinal effects. Recognition that endogenous gaseous mediators, nitric oxide (NO) and hydrogen sulfide (H(2)S) can increase mucosal defense mechanisms has led to the development of NO- and H(2)S-releasing NSAIDs with increased safety profiles. Here we report on a new hybrid, NOSH-aspirin, which is an NO- and H(2)S-releasing agent. NOSH-aspirin inhibited HT-29 colon cancer growth with IC(50)s of 45.5 ± 2.5, 19.7 ± 3.3, and 7.7 ± 2.2 nM at 24, 48, and 72 h, respectively. This is the first NSAID based agent with such high degree of potency. NOSH-aspirin inhibited cell proliferation, induced apoptosis, and caused G(0)/G(1) cell cycle block. Reconstitution and structure-activity studies representing a fairly close approximation to the intact molecule showed that NOSH-aspirin was 9000-fold more potent than the sum of its parts towards growth inhibition. NOSH-aspirin inhibited ovine COX-1 more than ovine COX-2. NOSH-ASA treatment of mice bearing a human colon cancer xenograft caused a reduction in volume of 85%. Taken together, these results demonstrate that NOSH-aspirin has strong anti-cancer potential and merits further evaluation.
Asunto(s)
Antineoplásicos/farmacología , Aspirina/análogos & derivados , Neoplasias del Colon/metabolismo , Inhibidores de la Ciclooxigenasa/farmacología , Disulfuros/farmacología , Sulfuro de Hidrógeno/metabolismo , Nitratos/farmacología , Donantes de Óxido Nítrico/farmacología , Óxido Nítrico/metabolismo , Animales , Antineoplásicos/química , Apoptosis/efectos de los fármacos , Aspirina/química , Aspirina/farmacología , Ciclo Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Neoplasias del Colon/enzimología , Inhibidores de la Ciclooxigenasa/química , Disulfuros/química , Células HT29 , Humanos , Ratones , Ratones Desnudos , Nitratos/química , Donantes de Óxido Nítrico/química , Prostaglandina-Endoperóxido Sintasas/metabolismo , Ratas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The balance between phase-I carcinogen-activating and phase-II detoxifying xenobiotic metabolizing enzymes is critical to determining an individual's risk for cancer. We evaluated the effect of Hydrogen sulfide-releasing aspirin (HS-ASA) on xenobiotic metabolizing enzymes in HT-29 human colon and Hepa 1c1c7 mouse liver adenocarcinoma cells and in Wistar rats. HS-ASA inhibited the growth of HT-29 and Hepa 1c1c7 cells, with an IC(50) of 3.2 ± 0.3 µM and 4.2 ± 0.4 µM, respectively. The IC(50) for ASA in both cell lines was greater than 5000 µM at 24h. In these cell lines, HS-ASA caused a dose-dependent increase in activity and expression of the phase-II enzymes glutathione S-transferase (GST) and NAD(P)H:quinoneoxireductase (NQO1). It also caused an increase in UDP-glucuronosyltransferase (UGT) expression. The levels of CYP 1A1 a phase-I enzyme was increased by HS-ASA in both cell lines. Pretreatment of cells with NaF, an esterase inhibitor, abrogated the HS-ASA-mediated increases in NQO1 enzyme activity. HS-ASA increased the protein levels of the transcription factor Nrf2, which is a regulator of the phase-II enzymes. In vivo, HS-ASA at 100mg/kg/day had no effect on rat's weights; it induced a 3.4-fold and 1.4-fold increase in hepatic GST and NQO1 enzyme activities, respectively. GST and NQO1 protein levels were also increased. In contrast to that in cultured cells, CYP 1A1 protein levels were not altered in vivo. Therefore, HS-ASA induces phase-II enzymes, at least in part, through the action of H(2)S and by modulating Nrf2; these effects may be part of its mechanism of action against carcinogenesis.