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Carboxyamidotriazole (CAI) was initially considered a non-cytotoxic anticancer agent. However, recently, pronounced anti-inflammatory properties of CAI have been reported. Rheumatoid arthritis (RA) is an autoimmune inflammatory disease characterized by aberrant activation of signaling pathways. Therefore, this study explored the therapeutic effects and potential mechanism of action of CAI on RA in the adjuvant arthritis (AA) model. The results showed that CAI reduced the severity of arthritis in AA rats as demonstrated by inhibited hind paw swelling, reduced body weight, and decreased infiltration of joint pathological inflammatory cells. Importantly, pathological scoring of new blood vessels and immunohistochemical assays revealed that CAI inhibited pannus formation. CAI decreased the expression of pro-angiogenic growth factors, such as vascular epidermal growth factor, basic fibroblast growth factor, and metalloproteinases (MMPs), namely, MMP-1 and MMP-3 in the synovium of AA rats. Furthermore, CAI significantly reduced the increased levels of phosphorylated p38, c-Jun N-terminal kinase (JNK)1/2, and extracellular signal-regulated kinase (ERK)1/2 proteins in AA rats. In addition, the proliferation of fibroblast-like synoviocytes (FLS) was downregulated by CAI both in vivo and in vitro. In conclusion, this investigation illustrates the therapeutic effect of CAI on synovitis and erosion of articular cartilage in RA. Furthermore, the mechanism might involve inhibition of aberrantly activated mitogen-activated protein kinase signaling, as well as a decrease in pro-angiogenic factors, MMP expression, and FLS proliferation.
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OBJECTIVES: This study aimed to explore the changes of four major inflammasomes in adult-onset Still's disease (AOSD) and preliminarily evaluate the therapeutic effect of carboxyamidotriazole (CAI), which has previously been reported to have the significant anti-inflammatory activity. METHOD: The mRNA expressions of proinflammatory cytokines and inflammasome components in peripheral blood mononuclear cells (PBMCs) from AOSD patients and healthy controls (HC) were determined by reverse transcription-quantitative PCR. Poly(dA:dT)-induced AIM2 inflammasome and flagellin-induced NLRC4 inflammasome activation models were established in bone marrow-derived macrophages (BMDMs). The levels of cytokines in serum and culture supernatants were measured by ELISA method. RESULTS: The serum levels of IL-1ß, IL-6, and TNF-α in AOSD patients were significantly higher than those in HC. However, the mRNA expressions of IL-1ß, IL-6, IL-18, and TNF-α in PBMCs did not differ markedly in AOSD patients in comparison with HC. Significantly increased mRNA levels of AIM2, NLRC4, ASC, and caspase-1 were observed in patients with AOSD when compared with HC, while NLRP1 and NLRP3 showed no change in AOSD samples. In addition, CAI treatment could significantly reduce the levels of IL-1ß, IL-6, and TNF-α secreted by AOSD PBMCs and inhibit AIM2 and NLRC4 inflammasomes activation in BMDMs. CONCLUSIONS: Increased levels of proinflammatory cytokines in AOSD might be associated with NLRC4 and AIM2 inflammasomes activation. CAI is likely to have the therapeutic potential for AOSD by inhibiting NLRC4 and AIM2 inflammasomes activation and reducing the proinflammatory cytokines and worthy of further investigation. These results provide new ideas for elucidating the pathogenesis of AOSD and providing specific targeted therapy. Key points ⢠Significantly higher mRNA levels of AIM2 and NLRC4 inflammsome signaling were observed in AOSD patients compared with health controls, indicating that AIM2 and NLRC4 inflammsome activation might be related to the increased proinflammatory cytokines in AOSD. ⢠CAI treatment markedly reduced the secretion levels of cytokines IL-1ß, IL-6, and TNF-α in AOSD PBMCs and inhibited AIM2 and NLRC4 inflammasome activation.
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Inflamasomas , Enfermedad de Still del Adulto , Adulto , Humanos , Inflamasomas/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Interleucina-6/metabolismo , Leucocitos Mononucleares/metabolismo , Citocinas/metabolismo , Interleucina-1beta/metabolismo , ARN Mensajero/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Proteínas de Unión al Calcio , Proteínas Adaptadoras de Señalización CARD/metabolismo , Proteínas de Unión al ADNRESUMEN
Inflammation and immunity are closely related to the occurrence and development of a variety of immune diseases. Although IL-1ß has been identified as a key cytokine in many immune diseases, safe and specific small molecular IL-1ß releasement inhibitors are still scarce and urgently required in clinic. The investigation prospect of triazoleis limited by its complicated pharmacological effect which exhibited inferior effects on IL-1ß and TNF-α. Herein, 36 novel derivatives were designed and synthesized, and nearly half of the derivatives exhibited much better selectivity on IL-1ß releasement inhibition as well as keep similar inhibitory activities to lead compound. In 20 µM, compound 19 exhibited IL-1ß releasement inhibitory activity (IC50 = 5.489 µM) which closed to the original compound, and 4.5-fold superior selectivity (SI = 4.71) to the lead compound (SI = 0.82). A probable SAR model of triazole derivatives for IL-1ß releasement inhibition and selectivity was also proposed, which might promote the discovery of more effective and specific IL-1ß releasement inhibitors in the future.
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Descubrimiento de Drogas , Interleucina-1beta/antagonistas & inhibidores , Triazoles/farmacología , Relación Dosis-Respuesta a Droga , Humanos , Interleucina-1beta/inmunología , Estructura Molecular , Relación Estructura-Actividad , Triazoles/síntesis química , Triazoles/químicaRESUMEN
Carboxyamidotriazole (CAI), originally developed as a non-cytotoxic anti-cancer drug, was shown to have anti-inflammatory activity according to recent studies in a number of animal models of inflammation. However, its mechanism of action has not been characterized. Therefore, the present study was performed to identify the anti-inflammatory action of CAI in lipopolysaccharide (LPS)-induced RAW 264.7 macrophages and to identify the signal transduction pathways involved. The in vitro results revealed that CAI had no direct effect on the activity of cyclooxygenase (COX), suggesting a different anti-inflammatory mechanism compared with that of COX-inhibiting non-steroidal anti-inflammatory drugs. Further investigation in RAW264.7 macrophages revealed that CAI decreased the production of nitric oxide via decreasing the LPS-stimulated expression of inducible nitric oxide synthase, and downregulated both mRNA and protein expression levels of the cytokines tumor necrosis factor-α, interleukin (IL)-1ß, and IL-6. CAI also significantly reduced the increased DNA-binding activity of nuclear factor (NF)-κB induced by LPS stimulation. With respect to the mechanisms involved on NF-κB activity, CAI exhibited suppression of the phosphorylation and degradation of the inhibitor of nuclear factor-κBα (IκB), and decreased the phosphorylation levels of the p65 subunit and its subsequent nuclear translocation. In addition, CAI significantly decreased the phosphorylated forms of p38, JNK and ERK, which were increased following LPS stimulation, while the total expression levels of p38, JNK and ERK remained unaltered. The results in the present study indicate that CAI alleviates the inflammatory responses of RAW 264.7 macrophages in response to LPS stimulation via attenuating the activation of NF-κB and MAPK signaling pathways and decreasing the levels of pro-inflammatory mediators. This offers a novel perspective for understanding the anti-inflammatory mechanism of CAI and suggests its potential use as a therapeutic treatment in inflammatory diseases with excessive macrophage activation.
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BACKGROUND: Cancer immunotherapy has generated significant excitement, mainly as a result of the development of immune checkpoint inhibitors. The blockade of PD-1 or its ligand with antibodies has resulted in impressive clinical efficacy. However, a subset of patients does not respond to biologic therapeutics, and another subset suffers from severe immune-related adverse events in certain cases. The modulation of the immune system with small molecules might yield surprising benefits. METHODS: CD8+ cells were obtained through a magnetic cell sorting system (MACS), and their capabilities for IFN-γ release and PD-1 expression were analyzed. The in vitro effects of drugs were studied in a coculture system of tumor cells and activated CD8+ cells. We further isolated the primary tumor cells in tumor-bearing mice treated with CAI, DMF, 1-MT or a combination (CAI and DMF/CAI and 1-MT) and analyzed the percentages of CD8+ T cells and PD-1+CD8+ T cells among TILs. The selective anti-tumor immune reactions of the two drug combinations were confirmed in a coculture system consisting of B16-OVA cells and OVA-specific CTLs derived from OT-1 transgenic mice. The anti-tumor effects of the single drugs or combined therapies were assessed according to their capability to slow tumor growth and extend the life span of tumor-bearing mice, and they were compared with the effects of PD-1 antibody. RESULTS: CAI increased IFN-γ release from activated T cells, which might strengthen the anti-proliferative and anti-metastatic effects on cancer cells. However, CAI also stimulated IDO1-Kyn metabolic circuitry in the tumor microenvironment and facilitated tumor cell immune evasion. Combining CAI with 1-MT or DMF disrupted PD-1 expression and promoted IFN-γ production in CD8+ T cells, and it also increased T lymphocyte infiltration in the tumor microenvironment, inhibited tumor growth and prolonged the life spans of tumor-bearing mice. CONCLUSION: Inhibitors of the IDO1-Kyn-AhR pathway could abolish the negative effects of CAI on CD8+ T cells and result in complementary and beneficial anti-tumor immune effects. The combination of CAI with 1-MT or DMF greatly augmented the ability of CD8+ T cells to kill malignant cells and showed a strong anti-cancer capability that was superior to that of either of the single agents was is comparable with that of anti-PD-1 antibody. The combinations of small molecules utilized in this study may serve as valuable new immunotherapy strategies for cancer treatment.
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Inmunoterapia , Indolamina-Pirrol 2,3,-Dioxigenasa/antagonistas & inhibidores , Quinurenina/metabolismo , Melanoma/tratamiento farmacológico , Receptores de Hidrocarburo de Aril/antagonistas & inhibidores , Triazoles/farmacología , Microambiente Tumoral/inmunología , Animales , Antineoplásicos/farmacología , Apoptosis , Linfocitos T CD8-positivos/efectos de los fármacos , Linfocitos T CD8-positivos/inmunología , Proliferación Celular , Quimioterapia Combinada , Flavonas/farmacología , Humanos , Activación de Linfocitos/inmunología , Melanoma/inmunología , Melanoma/patología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Transgénicos , Triptófano/farmacología , Células Tumorales Cultivadas , Microambiente Tumoral/efectos de los fármacosRESUMEN
The anticancer and anti-inflammatory effects of carboxyamidotriazole (CAI) have been demonstrated in several studies, but the underlying mechanisms remain to be elucidated. This study showed that CAI caused metabolic reprogramming of pancreatic cancer cells. The inhibition of mitochondrial oxidative metabolism by CAI led to increased glutamine-dependent reductive carboxylation and enhanced glycolytic metabolism. The presence of environmental substances that affect cellular metabolism, such as glutamine and pyruvate, attenuated the anticancer efficacy of CAI. Based on the action of CAI: 1) when glutamine was removed, the NAD+/NADH ratio was decreased, the synthesis of cellular aspartate was reduced, and autophagy flux was blocked; and 2) when glycolysis was pharmacologically inhibited, the ATP level was significantly decreased, the cell viability was greatly inhibited, and the compensatory rescue effect of glutamine was eliminated. When combined with chemotherapy, cotreatment with CAI and the glycolysis inhibitor 2-deoxyglucose (2-DG) inhibited the pancreatic cancer progression after chemotherapy. As the inhibition of mitochondrial oxidative metabolism can explain several anticancer activities of CAI reported previously, including inhibition of calcium entry and induction of reactive oxygen species, we demonstrate that inhibition of mitochondrial oxidative phosphorylation may be the fundamental mechanism of CAI. The combination of CAI and 2-DG causes energy depletion in cancer cells, eliminating the rescue effect of the metabolic environment. Inhibiting pancreatic cancer progression after chemotherapy is a rational application of this metabolism-disturbing combination strategy.
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Antineoplásicos/farmacología , Progresión de la Enfermedad , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Triazoles/farmacología , Animales , Antineoplásicos/uso terapéutico , Ácido Aspártico/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Respiración de la Célula/efectos de los fármacos , Desoxiglucosa/farmacología , Interacciones Farmacológicas , Femenino , Glucólisis/efectos de los fármacos , Humanos , Espacio Intracelular/metabolismo , Ratones , Ratones Endogámicos BALB C , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Neoplasias Pancreáticas/tratamiento farmacológico , Triazoles/uso terapéutico , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
BACKGROUND: Diabetic neuropathic pain (DNP) is a common and distressing complication in patients with diabetes, and the underlying mechanism remains unclear. Tricyclic antidepressants (TCAs) and serotonin and norepinephrine reuptake inhibitors (SNRIs) are recommended as first-line drugs for DNP. Ammoxetine is a novel and potent SNRI that exhibited a strong analgesic effect on models of neuropathic pain, fibromyalgia-related pain, and inflammatory pain in our primary study. The present study was undertaken to investigate the chronic treatment properties of ammoxetine on DNP and the underlying mechanisms for its effects. METHODS: The rat model of DNP was established by a single streptozocin (STZ) injection (60 mg/kg). Two weeks after STZ injection, the DNP rats were treated with ammoxetine (2.5, 5, and 10 mg/kg/day) for 4 weeks. The mechanical allodynia and locomotor activity were assayed to evaluate the therapeutic effect of ammoxetine. In mechanism study, the activation of microglia, astrocytes, the protein levels of pro-inflammatory cytokines, the mitogen-activated protein kinases (MAPK), and NF-κB were evaluated. Also, microglia culture was used to assess the direct effects of ammoxetine on microglial activation and the signal transduction mechanism. RESULTS: Treatment with ammoxetine for 4 weeks significantly relieved the mechanical allodynia and ameliorated depressive-like behavior in DNP rats. In addition, DNP rats displayed increased activation of microglia in the spinal cord, but not astrocytes. Ammoxetine reduced the microglial activation, accumulation of pro-inflammatory cytokines, and activation of p38 and c-Jun N-terminal kinase (JNK) in the spinal cord of DNP rats. Furthermore, ammoxetine displayed anti-inflammatory effects upon challenge with LPS in BV-2 microglia cells. CONCLUSION: Our results suggest that ammoxetine may be an effective treatment for relieving DNP symptoms. Moreover, a reduction in microglial activation and pro-inflammatory release by inhibiting the p-p38 and p-JNK pathways is involved in the mechanism.
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Benzodioxoles/uso terapéutico , Neuropatías Diabéticas/complicaciones , Neuropatías Diabéticas/tratamiento farmacológico , Hipoglucemiantes/uso terapéutico , Microglía/efectos de los fármacos , Mielitis , Propilaminas/uso terapéutico , Animales , Benzodioxoles/química , Proteínas de Unión al Calcio/metabolismo , Línea Celular Transformada , Neuropatías Diabéticas/inducido químicamente , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Clorhidrato de Duloxetina/uso terapéutico , Conducta Exploratoria/efectos de los fármacos , Hiperalgesia/tratamiento farmacológico , Hiperalgesia/fisiopatología , Hipoglucemiantes/química , Lipopolisacáridos/farmacología , Locomoción/efectos de los fármacos , Proteínas de Microfilamentos/metabolismo , Mielitis/tratamiento farmacológico , Mielitis/etiología , Mielitis/patología , Propilaminas/química , Ratas , Estreptozocina/toxicidadRESUMEN
Deficits in mitochondrial function is a critical inducement in the major pathways that drive neuronal cell death in ischemic process particularly. Drugs target to improve the mitochondrial function may be a feasible therapeutic choice in treatment with ischemic diseases. In the present study, we investigated whether 5-(4-hydroxy-3-dimethoxybenzylidene)-2-thioxo-4-thiazolidinone (RD-1), a compound derived from rhodanine, could protect against ischemic neuronal damage via improving mitochondrial function. We tested the neuroprotective effect of RD-1 both in rats modeled by middle cerebral artery occlusion reperfusion in vivo and in primary cortical neurons subjected to hypoxia/reperfusion injury in vitro. Results showed that treatment with RD-1 for 14 days remarkably reduced infarct size, decreased neurological deficit score and accelerated the recovery of somatosensory function in vivo. Meanwhile, RD-1 also increased the cellular viability after 48 h treatment in vitro. In addition, RD-1 protected the primary cortical neurons against mitochondrial damage as evidenced by stabilizing the mitochondrial membrane potential and reducing the overproduction of reactive oxygen species. Furthermore, hypoxia/reperfusion injury induced damaged mitochondrial axonal transport and consequently neurotransmitter release disorder, which were ameliorated by RD-1 treatment. Besides, RD-1 inhibited the downregulation of proteins related with mitochondrial transport and neurotransmitter release induced by ischemic injury both in vivo and in vitro. The obtained data demonstrated the neuroprotective effect of RD-1 and the involved mechanisms were partially attributed to the improvement in mitochondrial function and the synaptic activity. Our study indicated that RD-1 may be a potential therapeutic drug for the ischemic stroke therapy.
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Lung cancer is currently the leading cause of cancer-related deaths worldwide. In this study, we investigated the combination of carboxyamidotriazole (CAI) and sorafenib in non-small cell lung cancer (NSCLC) in vitro and in vivo to test whether CAI enhances the antitumor effects of sorafenib and reduces its side effects. The combination index (CI) showed that coadministration of CAI and sorafenib synergistically inhibited the proliferation of NSCLC cells (Lewis lung carcinoma, A549, and NCI-H1975 cells). Cell death as a result of the combination treatment was attributed to apoptosis, which was accompanied by activation of caspase-3 and poly(ADP-ribose) polymerase. In addition, combination therapy induced the accumulation of mitochondrial-associated reactive oxygen species, as well as depolarization of mitochondrial and reduced NANOG (homeobox protein NANOG) mRNA and protein expression. Basic fibroblast growth factor, a stimulator of NANOG, was applied to identify the possible mechanism. The addition of basic fibroblast growth factor followed by combined treatment may stimulate NANOG expression and synchronously rescue the accumulation of reactive oxygen species. C57BL/6J mice bearing Lewis lung carcinoma were randomized to receive vehicle (polyethylene glycol 400), CAI (30 mg/kg), low-dose sorafenib (SFB-L; 10 mg/kg), high-dose sorafenib (SFB-H; 30 mg/kg), or a CAI and SFB-L combination. Tumor growth was significantly suppressed in the combination group, and the efficacy of combination treatment was equivalent to that of the SFB-H monotherapy group. Furthermore, the combination group had reduced side effects compared with the SFB-H group, as indicated by weight preservation in mice. Our study illustrates that CAI enhances the antitumor activity of sorafenib in NSCLC and provides a novel strategy for NSCLC treatment.
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Apoptosis/efectos de los fármacos , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Neoplasias Pulmonares/tratamiento farmacológico , Proteína Homeótica Nanog/antagonistas & inhibidores , Niacinamida/análogos & derivados , Compuestos de Fenilurea/administración & dosificación , Triazoles/administración & dosificación , Animales , Antineoplásicos/administración & dosificación , Protocolos de Quimioterapia Combinada Antineoplásica/administración & dosificación , Apoptosis/fisiología , Carcinoma Pulmonar de Lewis/tratamiento farmacológico , Carcinoma Pulmonar de Lewis/metabolismo , Carcinoma Pulmonar de Lewis/patología , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Carcinoma de Pulmón de Células no Pequeñas/patología , Relación Dosis-Respuesta a Droga , Sinergismo Farmacológico , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Proteína Homeótica Nanog/metabolismo , Niacinamida/administración & dosificación , Sorafenib , Ensayos Antitumor por Modelo de Xenoinjerto/métodosRESUMEN
Excess proinflammatory cytokines owing to the activation of NF-κB and NLRP3 inflammasome play the key role in inflammatory bowel disease (IBD). Previously, we reported the anti-inflammatory activity of carboxyamidotriazole (CAI) resulting from decreasing cytokines. Therefore, we investigated the therapeutic effects of CAI in 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced rat colitis and the involvement of CAI action with NLRP3 inflammasome and NF-κB pathway. CAI was orally administered to TNBS-induced colitis rat. The severity of colitis was assessed, and NLRP3 inflammasome, NF-κB pathway and cytokines were determined. Our results showed that CAI significantly reduced weight loss and disease activity index (DAI) scores in colitis rats and alleviated the colonic macroscopic signs and pathological damage. In addition, the intestinal inflammatory markers and permeability index were markedly ameliorated by CAI treatment. The decreased levels of tumor necrosis factor-α (TNF-α), interleukin (IL)-1ß, IL-6, IL-18 were also detected in the colon tissues of CAI-treated colitis rats. Moreover, the activation of NLRP3 inflammasome in inflamed colon was significantly suppressed by showing an obvious reduction in the NLRP3 and activated caspase-1 levels. Furthermore, CAI reduced NF-κB p65 expression and IκBα phosphorylation and degradation in colitis rats. Therefore, CAI attenuates TNBS-induced colitis, which may be attributed to its inhibition of NLRP3 inflammasome and NF-κB activation, and down-regulation of proinflammatory cytokines. These results provide further understanding of the intestinal anti-inflammatory effect of CAI and highlight it as a potential drug for the treatment of IBD.
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Colitis/tratamiento farmacológico , Colon/efectos de los fármacos , Inflamasomas/metabolismo , Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , Intestinos/efectos de los fármacos , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Triazoles/uso terapéutico , Animales , Células Cultivadas , Colitis/inducido químicamente , Colon/patología , Citocinas/metabolismo , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Intestinos/patología , FN-kappa B/metabolismo , Ratas , Ratas Sprague-Dawley , Transducción de Señal/efectos de los fármacos , Ácido TrinitrobencenosulfónicoRESUMEN
Cancer cachexia is a complex disorder characterized by inflammatory responses, and it is associated with poor performance status and high mortality rate of cancer patients. Carboxyamidotriazole (CAI), a noncytotoxic chemotherapy agent, shows anti-inflammatory features in the treatment of many diseases. Here, we investigated the preventive and therapeutic effects of CAI on muscle loss that occurred in mice with advanced Lewis lung carcinoma (LLC). The carcass weights of CAI-treated mice were significantly higher than that of mice in the vehicle group from Day 19 to the end of the study. The gastrocnemius and epididymal adipose tissue weights were also increased by CAI treatment. The protective mechanisms might be attributed to the following points: CAI treatment inhibited the proteolysis in muscles by decreasing expressions of muscle-specific FoxO3 transcription factor and ubiquitin E3 ligases (MuRF1 and atrogin1). Moreover, CAI restricted the NF-κB signaling, downregulated the level of TNF-α in muscle and both TNF-α and IL-6 levels in serum, directly stimulated SIRT1 activity in vitro, and increased SIRT1 content in muscle. These results indicate that CAI can alleviate muscle wasting and is a promising drug against lung cancer cachexia.
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Antineoplásicos , Caquexia/tratamiento farmacológico , Atrofia Muscular/tratamiento farmacológico , FN-kappa B/antagonistas & inhibidores , Sirtuina 1/metabolismo , Triazoles , Tejido Adiposo/efectos de los fármacos , Tejido Adiposo/crecimiento & desarrollo , Animales , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Caquexia/sangre , Caquexia/etiología , Caquexia/metabolismo , Carcinoma Pulmonar de Lewis/complicaciones , Carcinoma Pulmonar de Lewis/tratamiento farmacológico , Carcinoma Pulmonar de Lewis/metabolismo , Carcinoma Pulmonar de Lewis/patología , Interleucina-6/sangre , Masculino , Ratones Endogámicos C57BL , Músculo Esquelético/efectos de los fármacos , Músculo Esquelético/crecimiento & desarrollo , Músculo Esquelético/metabolismo , Atrofia Muscular/etiología , Atrofia Muscular/metabolismo , Triazoles/farmacología , Triazoles/uso terapéutico , Carga Tumoral/efectos de los fármacos , Factor de Necrosis Tumoral alfa/sangre , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
AIM: The selective serotonin (5-HT) and norepinephrine (NE) reuptake inhibitors (SNRIs) are commonly used for the treatment of neuropathic pain and fibromyalgia. Ammoxetine ((±)-3-(benzo[d] [1,3]dioxol-4-yloxy)-N-methyl-3-(thiophen-2-yl)propan-1-amine) has been identified as a novel potent SNRI. In this study, we evaluated the acute analgesic properties of ammoxetine in different animal models of pain, and examined the involvement of monoamines in its analgesic actions. METHODS: The analgesic effects of ammoxetine were assayed using models of acetic acid- and formalin-induced pain in mice, neuropathic pain induced by sciatic nerve injury (SNI), chronic constriction injury (CCI) and reserpine-induced fibromyalgia pain in rats. The contents of 5-HT and NE in brain regions of fibromyalgia rats were measured using HPLC-ECD. In all the experiments, duloxetine was used as a positive control drug. RESULTS: Oral administration of ammoxetine (0.625-10 mg/kg) or duloxetine (2.5-40 mg/kg) dose-dependently decreased the number of acetic acid-induced writhing and formalin-induced first phase and second phase paw licking time in mice. Oral administration of ammoxetine (2.5-10 mg/kg) or duloxetine (10 mg/kg) alleviated mechanical allodynia in SNI and CCI rats and thermal hyperalgesia in CCI rats. The antiallodynic effect of ammoxetine in CCI rats was abolished by pretreatment with para-chlorophenylalanine methyl ester hydrochloride (PCPA, a 5-HT synthesis inhibitor) or α-methyl-para-tyrosine methylester (AMPT, a catecholamine synthesis inhibitor). Oral administration of ammoxetine (30 mg/kg) or duloxetine (50 mg/kg) significantly attenuated tactile allodynia in rats with reserpine-induced fibromyalgia. In the fibromyalgia rats, administration of ammoxetine (10, 30 mg/kg) or duloxetine (30, 50 mg/kg) dose-dependently increased the levels of 5-HT and NE, and decreased the metabolite ratio of 5-HT (5-HIAA/5-HT) in the spinal cord, hypothalamus, thalamus and prefrontal cortex. CONCLUSION: Ammoxetine effectively alleviates inflammatory, continuous, neuropathic and fibromyalgia-related pain in animal models, which can be attributed to enhanced neurotransmission of 5-HT and NE in the descending inhibitory systems.
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Analgésicos no Narcóticos/uso terapéutico , Benzodioxoles/uso terapéutico , Fibromialgia/tratamiento farmacológico , Hiperalgesia/tratamiento farmacológico , Umbral del Dolor/efectos de los fármacos , Propilaminas/uso terapéutico , Neuropatía Ciática/tratamiento farmacológico , Inhibidores de Captación de Serotonina y Norepinefrina/uso terapéutico , Analgésicos no Narcóticos/administración & dosificación , Animales , Benzodioxoles/administración & dosificación , Modelos Animales de Enfermedad , Masculino , Ratones Endogámicos , Estructura Molecular , Dimensión del Dolor , Propilaminas/administración & dosificación , Ratas Sprague-Dawley , Prueba de Desempeño de Rotación con Aceleración Constante , Inhibidores de Captación de Serotonina y Norepinefrina/administración & dosificaciónRESUMEN
OBJECTIVE: To study the effect of carboxyamidotriazole (CAI) on adjuvant arthritis (AA) in rats. METHODS: The rats were randomly divided into normal group,two vehicle groups (polyethylene glycol 400 control and normal sodium control group), CAI-treated groups (10, 20, and 40 mg/kg) and positive control dexamethasone group. Freund's completed adjuvant was used to induce AA in rats. The arthritis index (AI) was scored, and X-ray check of the hind limbs and histopathological examination were performed. The levels of tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, and IL-6 in the inflamed paw tissues were measured. RESULTS: The administration of CAI significantly decreased the AI, restored the body weights, and ameliorated the radiological and histopathological features of joint destruction in AA rats (P<0.05, P<0.01). In addition, CAI reduced the TNF-α, IL-1ß, and IL-6 levels in the inflamed paw tissues (P<0.05, P<0.01). CONCLUSION: CAI has therapeutic effect on AA rats, which may be achieved by decreasing the pro-inflammatory cytokines at the site of inflammation.
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Artritis Experimental , Animales , Adyuvante de Freund , Interleucina-1beta , Interleucina-6 , Ratas , Triazoles , Factor de Necrosis Tumoral alfaRESUMEN
Accumulated evidences suggested that microRNAs (miRs) play an important role in non-small cell lung cancer (NSCLC). However, how miRs perform their functions in lung adenocarcinoma cancer stem cells (CSCs) remains unknown. Notably, most studies pay more attention to the effects of miRNAs on the metastasis traits whereas the growth activities of CSCs are rather undervalued. In our report, using A549CD133+cells, we examined the inhibitory effects and the underlying mechanisms of microRNA-31 (miR-31) on the growth of lung adenocarcinoma CSC-like cells. Initially, we determined the level of miR-31 in A549 and A549CD133+ cells. Over-expression of miR-31 was found in A549CD133+ cells by microarray and real-time quantitative PCR (RTqPCR) assays. Experiments in multiple NSCLC cell lines in vitro and A549CD133+ cells xenograft models in vivo confirmed that down regulation of miR-31 resulted in increase of A549CD133+ cells growth, whereas overexpression of miR-31 led to the inhibition of adenocarcinoma cell proliferation. Also, MET proto-oncogene has been determined to be a direct target of miR-31 by dual luciferase report, RT-qPCR and western blot analysis. Down regulation of MET inhibited viability of A549CD133+ cells. The levels of PI3Kinase, Akt and p-Akt as well as downstream proteins were consequently decreased. These results suggest that miR-31 might inhibit the growth of lung adenocarcinoma cancer stem-like cells via down regulation of the MET-PI3K-Akt signaling pathway.
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Adenocarcinoma/patología , Regulación hacia Abajo , Neoplasias Pulmonares/patología , MicroARNs/metabolismo , Fosfatidilinositol 3-Quinasas/deficiencia , Proteínas Proto-Oncogénicas c-akt/deficiencia , Proteínas Proto-Oncogénicas c-met/deficiencia , Transducción de Señal , Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Animales , Apoptosis , Proliferación Celular , Células Cultivadas , Ensayos de Selección de Medicamentos Antitumorales , Células HEK293 , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , MicroARNs/biosíntesis , MicroARNs/genética , Neoplasias Experimentales/tratamiento farmacológico , Neoplasias Experimentales/metabolismo , Neoplasias Experimentales/patología , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/patología , Fosfatidilinositol 3-Quinasas/biosíntesis , Fosfatidilinositol 3-Quinasas/metabolismo , Proto-Oncogenes Mas , Proteínas Proto-Oncogénicas c-akt/biosíntesis , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas Proto-Oncogénicas c-met/biosíntesis , Proteínas Proto-Oncogénicas c-met/metabolismoRESUMEN
Targeting cancer cell metabolism is a promising strategy against cancer. Here, we confirmed that the anti-cancer drug carboxyamidotriazole (CAI) inhibited mitochondrial respiration in cancer cells for the first time and found a way to enhance its anti-cancer activity by further disturbing the energy metabolism. CAI promoted glucose uptake and lactate production when incubated with cancer cells. The oxidative phosphorylation (OXPHOS) in cancer cells was inhibited by CAI, and the decrease in the activity of the respiratory chain complex I could be one explanation. The anti-cancer effect of CAI was greatly potentiated when being combined with 2-deoxyglucose (2-DG). The cancer cells treated with the combination of CAI and 2-DG were arrested in G2/M phase. The apoptosis and necrosis rates were also increased. In a mouse xenograft model, this combination was well tolerated and retarded the tumor growth. The impairment of cancer cell survival was associated with significant cellular ATP decrease, suggesting that the combination of CAI and 2-DG could be one of the strategies to cause dual inhibition of energy pathways, which might be an effective therapeutic approach for a broad spectrum of tumors.
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Antineoplásicos/farmacología , Glucólisis/efectos de los fármacos , Fosforilación Oxidativa/efectos de los fármacos , Triazoles/farmacología , Animales , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Desoxiglucosa/farmacología , Sinergismo Farmacológico , Glucosa/metabolismo , Humanos , Ratones , Ratones Endogámicos C57BL , Consumo de Oxígeno/efectos de los fármacosRESUMEN
BACKGROUND AND PURPOSE: Pro-inflammatory cytokines are important in rheumatoid arthritis (RA) and their production is mainly regulated by NF-κB and inflammasomes. Carboxyamidotriazole (CAI) exhibits potent anti-inflammatory activities by decreasing cytokines. Here, we have investigated NACHT, LRR and PYD domains-containing protein (NALP) inflammasomes in a rat model of RA and explored the therapeutic effects of CAI in this model and the involvement of NF-κB and inflammasomes in the actions of CAI. EXPERIMENTAL APPROACH: The anti-arthritic effects of CAI were assessed in the adjuvant arthritis (AA) model in rats, using radiological and histological techniques. NALP1 and NALP3 inflammasomes, NF-κB pathway and pro-inflammatory cytokines levels were measured with Western blots, immunohistochemistry and ELISA. KEY RESULTS: CAI decreased the arthritis index, improved radiological and histological changes, and reduced synovial IL-1ß, IL-6, IL-18 and TNF-α levels in rats with AA. Compared with normal rats, the 70 kDa NALP1 isoform was up-regulated, NALP3 was down-regulated, and levels of the 165 kDa NALP1 isoform and the adaptor protein ASC were unchanged in synovial tissue from AA rats. CAI reduced the 70 kDa NALP1 isoform and restored NALP3 levels in AA rats; CAI inhibited caspase-1 activation in AA synovial tissue, but not its enzymic activity in vitro. In addition, CAI reduced expression of p65 NF-κB subunit and IκBα phosphorylation and degradation in AA rats. CONCLUSION AND IMPLICATIONS: NALP1 inflammasomes were activated in synovial tissues from AA rats and appeared to be a novel therapeutic target for RA. CAI could have therapeutic value in RA by inhibiting activation of NF-κB and NALP1 inflammasomes and by decreasing pro-inflammatory cytokines.
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Antiinflamatorios/farmacología , Artritis Experimental/metabolismo , Artritis Reumatoide/metabolismo , Inflamasomas/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Triazoles/farmacología , Animales , Articulación del Tobillo/diagnóstico por imagen , Articulación del Tobillo/efectos de los fármacos , Articulación del Tobillo/metabolismo , Articulación del Tobillo/patología , Antiinflamatorios/uso terapéutico , Artritis Experimental/diagnóstico por imagen , Artritis Experimental/tratamiento farmacológico , Artritis Experimental/patología , Artritis Reumatoide/diagnóstico por imagen , Artritis Reumatoide/tratamiento farmacológico , Artritis Reumatoide/patología , Citocinas/metabolismo , Masculino , Radiografía , Ratas Endogámicas Lew , Membrana Sinovial/metabolismo , Triazoles/uso terapéuticoRESUMEN
Carboxyamidotriazole (CAI) is a non-cytotoxic anti-tumor drug, which also shows considerable anti-inflammatory effects in a variety of animal models of inflammation. The exact target and mechanism of CAI were not clearly understood yet. In the present study, we demonstrate that CAI is a non-selective phosphodiesterase (PDE) inhibitor, which provides comprehensive inhibitions of both adenosine 3',5'-cyclic monophosphate specific PDE (cAMP-PDE) and guanosine 3',5'-cyclic monophosphate specific PDE (cGMP-PDE) isolated from rat brain, mouse pulmonary tissue, primary mouse peritoneal macrophages, RAW264.7 cells, Lewis lung carcinoma (LLC) cells and lymphocytic leukemia cells (L1210) with moderate potencies (IC50≈0.5-30µM). The comprehensive elimination of PDE activities in living LLC cells by CAI results in accumulation of intracellular cAMP and cGMP, which can be visualized by fluorescence resonance energy transfer (FRET)-based cyclic nucleotide sensors. The stimulation by 30µM CAI yielded ~1.5-fold greater cGMP responses compared with 10µM sildenafil citrate, whereas the influence of 30µM CAI on cAMP levels was similar as that of 100µM 3-isobutyl-1-methylxanthine (IBMX). The non-selective inhibitory effect of CAI on cAMP-PDE and cGMP-PDE increases the likelihood for CAI to affect the balance between the levels of intracellular cyclic nucleotides cAMP and cGMP, then a variety of cellular signaling pathways that regulate cell functions and even related disease processes. When examining the widely proven anti-tumor and anti-inflammatory activities of CAI, it is important to affirm its comprehensive inhibitory effect on PDEs, which makes it superior to some selective PDE inhibitors in a way.
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3',5'-AMP Cíclico Fosfodiesterasas/antagonistas & inhibidores , Antiinflamatorios no Esteroideos/farmacología , Antineoplásicos/farmacología , GMP Cíclico/metabolismo , Inhibidores de Fosfodiesterasa/farmacología , Triazoles/farmacología , 3',5'-AMP Cíclico Fosfodiesterasas/metabolismo , Animales , Línea Celular Tumoral , AMP Cíclico/metabolismo , Espacio Intracelular/efectos de los fármacos , Espacio Intracelular/metabolismo , Masculino , Ratones , Ratas , Transducción de Señal/efectos de los fármacosRESUMEN
Carboxyamidotrizole (CAI) has been reported to suppress the production of tumor necrosis factor-α (TNF-α) and interleukin (IL)-1ß and be effective in rats with adjuvant arthritis. The aim of this study was to investigate the role of CAI in inflammatory bowel disease (IBD). We assessed the effect of CAI in dextran sodium sulfate-induced colitis. Inflammation was scored histologically, and potential mediators of IBD were assessed by immunohistochemical and molecular biochemical approaches. CAI-treated colitis animals revealed much fewer signs of colitis with significantly decreased macroscopic and microscopic scores than vehicle-treated animals. CAI inhibited the production of TNF-α, IL-1ß, and IL-6 in serum, supernatant of peritoneal macrophages, and lamina propria. CAI also decreased the expression of intercellular adhesion molecule-1 in colonic tissues. Furthermore, CAI prevented nuclear factor-κB (NF-κB) activation and inhibitor of nuclear factor-κBα phosphorylation and degradation. In addition, CAI showed a beneficial effect on colonic fibrosis, possibly by reducing the production of the fibrogenic cytokine transforming growth factor-ß. The results support that CAI administration is effective in ameliorating experimental colitis and preventing colonic fibrosis. The inhibition of proinflammatory cytokines and adhesion molecules and suppression of NF-κB activation seem to contribute to this effect.
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Colitis/tratamiento farmacológico , Colon/efectos de los fármacos , FN-kappa B/antagonistas & inhibidores , FN-kappa B/metabolismo , Triazoles/farmacología , Animales , Colitis/sangre , Colitis/metabolismo , Colitis/patología , Colon/metabolismo , Colon/patología , Sulfato de Dextran , Fibrosis , Inflamación/sangre , Inflamación/tratamiento farmacológico , Inflamación/metabolismo , Inflamación/patología , Enfermedades Inflamatorias del Intestino/sangre , Enfermedades Inflamatorias del Intestino/tratamiento farmacológico , Enfermedades Inflamatorias del Intestino/metabolismo , Enfermedades Inflamatorias del Intestino/patología , Molécula 1 de Adhesión Intercelular/metabolismo , Interleucina-1beta/sangre , Interleucina-1beta/metabolismo , Interleucina-6/sangre , Interleucina-6/metabolismo , Macrófagos Peritoneales/efectos de los fármacos , Macrófagos Peritoneales/metabolismo , Macrófagos Peritoneales/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Membrana Mucosa/efectos de los fármacos , Membrana Mucosa/metabolismo , Membrana Mucosa/patología , Fosforilación/efectos de los fármacos , Proteolisis/efectos de los fármacos , Factor de Necrosis Tumoral alfa/sangre , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Carboxyamidotriazole (CAI) has not only direct anti-cancer activities, but also anti-inflammation effects in a variety of inflammatory animal models. In the present study, we investigated whether macrophages, which are important both in cancer and inflammation, could be regulated by CAI. The results showed that CAI could inhibit tumour necrosis factor-α (TNF-α) production in macrophages in various environments, including those isolated from peritoneal cavity of adjuvant-induced arthritis (AA) rats, from Lewis lung carcinoma (LLC) transplanted tumours and those induced by LLC cells in vitro. Dexamethasone (DEX), one of the pro-inflammatory cytokines inhibitors, could enhance CAI's inhibition of LLC cells proliferation and invasion in macrophages and LLC cells co-culture systems, as well as the tumour growth in vivo. However, DEX failed to enhance CAI's inhibition of LLC cells proliferation when LLC cells were cultured alone, suggesting that the combination of CAI and DEX exerted great anti-tumour effects probably by acting on macrophages in the tumour environment. Over all, we found CAI could act on macrophages and regulate the production of TNF-α not only in inflammatory diseases but also in tumour microenvironment, which might be another anti-tumour mechanism of CAI.
Asunto(s)
Antiinflamatorios/farmacología , Carcinoma Pulmonar de Lewis/inmunología , Carcinoma Pulmonar de Lewis/metabolismo , Macrófagos/efectos de los fármacos , Neoplasias/tratamiento farmacológico , Triazoles/farmacología , Factor de Necrosis Tumoral alfa/metabolismo , Animales , Protocolos de Quimioterapia Combinada Antineoplásica/sangre , Artritis Experimental/inmunología , Artritis Experimental/metabolismo , Técnicas de Cocultivo , Citocinas , Dexametasona/farmacología , Macrófagos/inmunología , Macrófagos Peritoneales/efectos de los fármacos , Macrófagos Peritoneales/inmunología , Macrófagos Peritoneales/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Trasplante de Neoplasias , Neoplasias/inmunología , Ratas Wistar , Microambiente Tumoral/efectos de los fármacos , Microambiente Tumoral/inmunologíaRESUMEN
OBJECTIVE: To explore the potential anti-inflammatory and analgesic activities of carboxyamidotriazole (CAI). METHODS: A variety of animal models, including the croton oil-induced ear edema, the cotton-induced granuloma, the rat adjuvant-induced arthritis, were used to evaluate anti-inflammatory effect of CAI. Vascular endothelial growth factor (VEGF)--or histamine-stimulated local vascular permeability in mouse modulated by CAI was also determined. In addition, we assessed the effect of CAI on the levels of proinflammatory cytokines tumor necrosis factor-alpha (TNF-alpha) and interleukin-1 beta (IL-beta) at the site of inflammation and in sera. Moreover, antinociceptive effect of CAI on inflammatory pain was assessed using acetic acid-induced writhing model and the formalin test. RESULTS: CAI significantly inhibited acute and chronic phases of inflammation, reduced VEGF or histamine-induced vascular permeability, and showed marked inhibition of proinflammatory cytokines such as TNF-alpha and IL-1 beta. CAI also showed potential therapeutic effect on peripheral inflammatory pain. CONCLUSION: CAI is a promising anti-inflammatory and analgesic agent.