RESUMO
Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest cancers. Late presentation of disease at the time of diagnosis is one of the major reasons for dismal prognostic outcomes for PDAC patients. Currently, there is a lack of clinical biomarkers, which can be used to diagnose PDAC patients at an early resectable stage. This study performed proteomic mass spectrometry to identify novel blood-based biomarkers for early diagnosis of PDAC. Serum specimens from 88 PDAC patients and 88 healthy controls (60 discovery cohort and 28 validation cohort) were analyzed using data independent acquisition high resolution mass spectrometry to identify candidate biomarker proteins. A total of 249 proteins were identified and quantified by the mass spectrometric analysis. Six proteins were markedly (>1.5 fold) and significantly (p < .05; q < 0.1) increased in PDAC patients compared to healthy controls in discovery cohort. Notably, four of these six proteins were significantly upregulated in an independent validation cohort. The top three upregulated proteins (i.e., Polymeric Immunoglobulin Receptor [PIGR], von Willebrand Factor [vWF], and Fibrinogen) were validated using enzyme linked immunosorbent assay, which led to selection of PIGR and vWF as a diagnostic biomarker panel for PDAC. The panel showed high ability to diagnose early stage (stage I and II) PDAC patients (area under the curve [AUC]: 0.8926), which was further improved after the addition of clinically used prognostic biomarker (Ca 19-9) to the panel (AUC: 0.9798). In conclusion, a novel serum protein biomarker panel for early diagnosis of PDAC was identified.
Assuntos
Biomarcadores Tumorais , Carcinoma Ductal Pancreático , Detecção Precoce de Câncer , Neoplasias Pancreáticas , Proteômica , Humanos , Biomarcadores Tumorais/sangue , Neoplasias Pancreáticas/diagnóstico , Neoplasias Pancreáticas/sangue , Feminino , Masculino , Detecção Precoce de Câncer/métodos , Carcinoma Ductal Pancreático/sangue , Carcinoma Ductal Pancreático/diagnóstico , Pessoa de Meia-Idade , Idoso , Proteômica/métodos , Receptores de Imunoglobulina Polimérica/sangue , Fator de von Willebrand/análise , Fator de von Willebrand/metabolismo , Fibrinogênio/análise , Fibrinogênio/metabolismo , Estudos de Casos e Controles , Adulto , Proteínas Sanguíneas/análiseRESUMO
Pancreatic ductal adenocarcinoma (PDAC) patients have late presentation at the time of diagnosis and a poor prognosis. Metal dyshomeostasis is known to play a role in cancer progression. However, the blood and tissue metallome of PDAC patients has not been assessed. This study aimed to determine the levels of essential and toxic metals in the serum and pancreatic tissue from PDAC patients. Serum samples were obtained from PDAC patients before surgical resection. Tissue (tumor and adjacent normal pancreas) were obtained from the surgically resected specimen. Inductively coupled plasma-mass spectrometry (ICP-MS) analysis was performed to quantify the levels of 10 essential and 3 toxic metals in these samples. Statistical analysis was performed to identify dysregulated metals in PDAC and their role as potential diagnostic and prognostic biomarkers. Significantly decreased serum levels of magnesium, potassium, calcium, iron, zinc, selenium, arsenic, and mercury and increased levels of molybdenum were shown to be associated with PDAC. There were significantly decreased levels of zinc, manganese and molybdenum, and increased levels of calcium and selenium in the pancreatic tumor tissue compared with the adjacent normal pancreas. Notably, lower serum levels of calcium, iron, and selenium, and higher levels of manganese, were significantly associated with a poor prognosis (i.e., overall survival) in PDAC patients. In conclusion, this is the first study to comprehensively assess the serum and tissue metallome of PDAC patients. It identified the association of metals with PDAC diagnosis and prognosis.
Assuntos
Biomarcadores Tumorais , Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Humanos , Carcinoma Ductal Pancreático/sangue , Carcinoma Ductal Pancreático/patologia , Carcinoma Ductal Pancreático/metabolismo , Neoplasias Pancreáticas/sangue , Neoplasias Pancreáticas/patologia , Neoplasias Pancreáticas/metabolismo , Feminino , Masculino , Pessoa de Meia-Idade , Idoso , Biomarcadores Tumorais/sangue , Biomarcadores Tumorais/metabolismo , Prognóstico , Metais/sangue , Metais/metabolismo , Metais/análise , Pâncreas/metabolismo , Pâncreas/patologia , Magnésio/sangue , Magnésio/metabolismo , Magnésio/análise , Adulto , Cálcio/sangue , Cálcio/metabolismo , Cálcio/análise , Selênio/sangue , Selênio/análise , Selênio/metabolismo , Ferro/metabolismo , Ferro/sangue , Zinco/sangue , Zinco/metabolismo , Zinco/análise , Molibdênio/sangueRESUMO
A major barrier to successful pancreatic cancer (PC) treatment is the surrounding stroma, which secretes growth factors/cytokines that promote PC progression. Wnt and tenascin C (TnC) are key ligands secreted by stromal pancreatic stellate cells (PSCs) that then act on PC cells in a paracrine manner to activate the oncogenic ß-catenin and YAP/TAZ signaling pathways. Therefore, therapies targeting oncogenic Wnt/TnC cross talk between PC cells and PSCs constitute a promising new therapeutic approach for PC treatment. The metastasis suppressor N-myc downstream-regulated gene-1 (NDRG1) inhibits tumor progression and metastasis in numerous cancers, including PC. We demonstrate herein that targeting NDRG1 using the clinically trialed anticancer agent di-2-pyridylketone-4-cyclohexyl-4-methyl-3-thiosemicarbazone (DpC) inhibited Wnt/TnC-mediated interactions between PC cells and the surrounding PSCs. Mechanistically, NDRG1 and DpC markedly inhibit secretion of Wnt3a and TnC by PSCs, while also attenuating Wnt/ß-catenin and YAP/TAZ activation and downstream signaling in PC cells. This antioncogenic activity was mediated by direct inhibition of ß-catenin and YAP/TAZ nuclear localization and by increasing the Wnt inhibitor, DKK1. Expression of NDRG1 also inhibited transforming growth factor (TGF)-ß secretion by PC cells, a key mechanism by which PC cells activate PSCs. Using an in vivo orthotopic PC mouse model, we show DpC downregulated ß-catenin, TnC, and YAP/TAZ, while potently increasing NDRG1 expression in PC tumors. We conclude that NDRG1 and DpC inhibit Wnt/TnC-mediated interactions between PC cells and PSCs. These results further illuminate the antioncogenic mechanism of NDRG1 and the potential of targeting this metastasis suppressor to overcome the oncogenic effects of the PC-PSC interaction.
Assuntos
Comunicação Celular , Proteínas de Ciclo Celular , Peptídeos e Proteínas de Sinalização Intracelular , Neoplasias Pancreáticas , Células Estreladas do Pâncreas , Tenascina , beta Catenina , Animais , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Camundongos , Metástase Neoplásica , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Células Estreladas do Pâncreas/metabolismo , Células Estreladas do Pâncreas/patologia , Tenascina/genética , Tenascina/metabolismo , beta Catenina/genética , beta Catenina/metabolismo , Neoplasias PancreáticasRESUMO
OBJECTIVES: Postoperative pancreatic fistula (POPF) represents one of the most severe complications following pancreatic surgery. Despite being a leading cause of morbidity and mortality, its pathophysiology is poorly understood. In recent years, there has been growing evidence to support the role of postoperative or post-pancreatectomy acute pancreatitis (PPAP) in the development of POPF. This article reviews the contemporary literature on POPF pathophysiology, risk factors, and prevention strategies. METHODS: A literature search was conducted using electronic databases, including Ovid Medline, EMBASE, and Cochrane Library, to retrieve relevant literature published between 2005 and 2023. A narrative review was planned from the outset. RESULTS: A total of 104 studies fulfilled criteria for inclusion. Forty-three studies reported on technical factors predisposing to POPF, including resection and reconstruction technique and adjuncts for anastomotic reinforcement. Thirty-four studies reported on POPF pathophysiology. There is compelling evidence to suggest that PPAP plays a critical role in the development of POPF. The acinar component of the remnant pancreas should be regarded as an intrinsic risk factor; meanwhile, operative stress, remnant hypoperfusion, and inflammation represent common mechanisms for acinar cell injury. CONCLUSIONS: The evidence base for PPAP and POPF is evolving. Future POPF prevention strategies should look beyond anastomotic reinforcement and target underlying mechanisms of PPAP development.
Assuntos
Fístula Pancreática , Pancreatite , Humanos , Fístula Pancreática/prevenção & controle , Pancreatite/etiologia , Pancreatite/complicações , Doença Aguda , Pâncreas/cirurgia , Pancreatectomia/efeitos adversos , Pancreatectomia/métodos , Fatores de Risco , Complicações Pós-Operatórias/cirurgia , Estudos Retrospectivos , Pancreaticoduodenectomia/efeitos adversosRESUMO
Pancreatic cancer (PaCa) is characterized by dense stroma that hinders treatment efficacy, with pancreatic stellate cells (PSCs) being a major contributor to this stromal barrier and PaCa progression. Activated PSCs release hepatocyte growth factor (HGF) and insulin-like growth factor (IGF-1) that induce PaCa proliferation, metastasis and resistance to chemotherapy. We demonstrate for the first time that the metastasis suppressor, N-myc downstream regulated gene 1 (NDRG1), is a potent inhibitor of the PaCa-PSC cross-talk, leading to inhibition of HGF and IGF-1 signaling. NDRG1 also potently reduced the key driver of PaCa metastasis, namely GLI1, leading to reduced PSC-mediated cell migration. The novel clinically trialed anticancer agent, di-2-pyridylketone 4-cyclohexyl-4-methyl-3-thiosemicarbazone (DpC), which upregulates NDRG1, potently de-sensitized PaCa cells to ligands secreted by activated PSCs. DpC and NDRG1 also inhibited the PaCa-mediated activation of PSCs via inhibition of sonic hedgehog (SHH) signaling. In vivo, DpC markedly reduced PaCa tumor growth and metastasis more avidly than the standard chemotherapy for this disease, gemcitabine. Uniquely, DpC was selectively cytotoxic against PaCa cells, while "re-programming" PSCs to an inactive state, decreasing collagen deposition and desmoplasia. Thus, targeting NDRG1 can effectively break the oncogenic cycle of PaCa-PSC bi-directional cross-talk to overcome PaCa desmoplasia and improve therapeutic outcomes.
Assuntos
Adenocarcinoma/metabolismo , Proteínas de Ciclo Celular/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neoplasias Pancreáticas/metabolismo , Células Estromais/metabolismo , Adenocarcinoma/patologia , Animais , Antineoplásicos/toxicidade , Linhagem Celular , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Feminino , Proteínas Hedgehog/metabolismo , Fator de Crescimento de Hepatócito/metabolismo , Humanos , Fator de Crescimento Insulin-Like I/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neoplasias Pancreáticas/patologia , Piridinas/toxicidade , Células Estromais/efeitos dos fármacos , Tiossemicarbazonas/toxicidade , Proteína GLI1 em Dedos de Zinco/metabolismoRESUMO
Triple negative breast cancer (TNBC) is the most aggressive type of breast cancers which constitutes about 15% of all breast cancer cases and characterized by negative expression of hormonal receptors and human epidermal growth factor receptor 2 (HER2). Thus, endocrine and HER2 targeted therapies are not effective toward TNBCs, and they mainly rely on chemotherapy and surgery for treatment. Despite recent advances in chemotherapy, 40% of TNBC patients develop a metastatic relapse and recurrence. Therefore, understanding the molecular profile of TNBC is warranted to identify targets that can be selected for the development of a new and effective therapeutic approach. Autophagy is an internal defensive mechanism that allows the cells to survive under different stressors. It has been well known that autophagy exerts a crucial role in cancer progression. The critical role of autophagy in TNBC progression is emerging in recent years. This review will discuss autophagic pathway, how autophagy affects TNBC progression and recent therapeutic approaches that can target autophagy as a new treatment modality.
Assuntos
Autofagia , Neoplasias de Mama Triplo Negativas , Animais , Antineoplásicos/uso terapêutico , Autofagia/efeitos dos fármacos , Feminino , Humanos , Neoplasias de Mama Triplo Negativas/tratamento farmacológicoRESUMO
A rigorous exploration of microbial diversity has revealed its presence on Earth, deep oceans, and vast space. The presence of microbial life in diverse environmental conditions, ranging from moderate to extreme temperature, pH, salinity, oxygen, radiations, and altitudes, has provided the necessary impetus to search for them by extending the limits of their habitats. Microbiology started as a distinct science in the mid-nineteenth century and has provided inputs for the betterment of mankind during the last 150 years. As beneficial microbes are assets and pathogens are detrimental, studying both have its own merits. Scientists are nowadays working on illustrating the microbial dynamics in Earth's subsurface, deep sea, and polar regions. In addition to studying the role of microbes in the environment, the microbe-host interactions in humans, animals and plants are also unearthing newer insights that can help us to improve the health of the host by modulating the microbiota. Microbes have the potential to remediate persistent organic pollutants. Antimicrobial resistance which is a serious concern can also be tackled only after monitoring the spread of resistant microbes using disciplines of genomics and metagenomics The cognizance of microbiology has reached the top of the world. Space Missions are now looking for signs of life on the planets (specifically Mars), the Moon and beyond them. Among the most potent pieces of evidence to support the existence of life is to look for microbial, plant, and animal fossils. There is also an urgent need to deliberate and communicate these findings to layman and policymakers that would help them to take an adequate decision for better health and the environment around us. Here, we present a glimpse of recent advancements by scientists from around the world, exploring and exploiting microbial diversity.
RESUMO
Our study aimed to identify a urinary metabolite panel for the detection/diagnosis of pancreatic ductal adenocarcinoma (PDAC). PDAC continues to have poor survival outcomes. One of the major reasons for poor prognosis is the advanced stage of the disease at diagnosis. Hence, identification of a novel and cost-effective biomarker signature for early detection/diagnosis of PDAC could lead to better survival outcomes. Untargeted metabolomics was employed to identify a novel metabolite-based biomarker signature for PDAC diagnosis. Urinary metabolites from 92 PDAC patients (56 discovery cohort and 36 validation cohort) were compared with 56 healthy volunteers using 1 H nuclear magnetic resonance spectroscopy. Multivariate (partial-least squares discriminate analysis) and univariate (Mann-Whitney's U-test) analyses were performed to identify a metabolite panel which can be used to detect PDAC. The selected metabolites were further validated for their diagnostic potential using the area under the receiver operating characteristic (AUROC) curve. Statistical analysis identified a six-metabolite panel (trigonelline, glycolate, hippurate, creatine, myoinositol and hydroxyacetone), which demonstrated high potential to diagnose PDAC, with AUROC of 0.933 and 0.864 in the discovery and validation cohort, respectively. Notably, the identified panel also demonstrated very high potential to diagnose early-stage (I and II) PDAC patients with AUROC of 0.897. These results demonstrate that the selected metabolite signature could be used to detect PDAC and will pave the way for the development of a urinary test for detection/diagnosis of PDAC.
Assuntos
Biomarcadores Tumorais/urina , Carcinoma Ductal Pancreático/diagnóstico , Neoplasias Pancreáticas/diagnóstico , Idoso , Carcinoma Ductal Pancreático/urina , Detecção Precoce de Câncer/métodos , Feminino , Humanos , Masculino , Metabolômica/métodos , Pessoa de Meia-Idade , Neoplasias Pancreáticas/urina , Urinálise/métodosRESUMO
Multidrug resistance (MDR) is a major obstacle in cancer treatment due to the ability of tumor cells to efflux chemotherapeutics via drug transporters (e.g. P-glycoprotein (Pgp; ABCB1)). Although the mechanism of Pgp-mediated drug efflux is known at the plasma membrane, the functional role of intracellular Pgp is unclear. Moreover, there has been intense focus on the tumor micro-environment as a target for cancer treatment. This investigation aimed to dissect the effects of tumor micro-environmental stress on subcellular Pgp expression, localization, and its role in MDR. These studies demonstrated that tumor micro-environment stressors (i.e. nutrient starvation, low glucose levels, reactive oxygen species, and hypoxia) induce Pgp-mediated drug resistance. This occurred by two mechanisms, where stressors induced 1) rapid Pgp internalization and redistribution via intracellular trafficking (within 1 h) and 2) hypoxia-inducible factor-1α expression after longer incubations (4-24 h), which up-regulated Pgp and was accompanied by lysosomal biogenesis. These two mechanisms increased lysosomal Pgp and facilitated lysosomal accumulation of the Pgp substrate, doxorubicin, resulting in resistance. This was consistent with lysosomal Pgp being capable of transporting substrates into lysosomes. Hence, tumor micro-environmental stressors result in: 1) Pgp redistribution to lysosomes; 2) increased Pgp expression; 3) lysosomal biogenesis; and 4) potentiation of Pgp substrate transport into lysosomes. In contrast to doxorubicin, when stress stimuli increased lysosomal accumulation of the cytotoxic Pgp substrate, di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone (Dp44mT), this resulted in the agent overcoming resistance. Overall, this investigation describes a novel approach to overcoming resistance in the stressful tumor micro-environment.
Assuntos
Antineoplásicos/farmacologia , Lisossomos/efeitos dos fármacos , Modelos Biológicos , Neoplasias/tratamento farmacológico , Tiossemicarbazonas/farmacologia , Microambiente Tumoral/efeitos dos fármacos , Subfamília B de Transportador de Cassetes de Ligação de ATP/agonistas , Subfamília B de Transportador de Cassetes de Ligação de ATP/antagonistas & inibidores , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Acridinas/farmacologia , Antineoplásicos/metabolismo , Apoptose/efeitos dos fármacos , Transporte Biológico/efeitos dos fármacos , Hipóxia Celular , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Doxorrubicina/metabolismo , Doxorrubicina/farmacologia , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Peróxido de Hidrogênio/farmacologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/agonistas , Subunidade alfa do Fator 1 Induzível por Hipóxia/antagonistas & inibidores , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Lisossomos/metabolismo , Proteínas de Neoplasias/agonistas , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Neoplasias/metabolismo , Neoplasias/patologia , Biogênese de Organelas , Transporte Proteico/efeitos dos fármacos , Interferência de RNA , Tetra-Hidroisoquinolinas/farmacologiaRESUMO
The iron-regulated metastasis suppressor N-myc downstream-regulated gene 1 (NDRG1) has been shown to inhibit numerous oncogenic signaling pathways in cancer cells. Recent findings have demonstrated that NDRG1 inhibits the ErbB family of receptors, which function as key inducers of carcinogenesis. NDRG1 attenuates ErbB signaling by inhibiting formation of epidermal growth factor receptor (EGFR)/human epidermal growth factor receptor 2 (HER2) and HER2/HER3 heterodimers and by down-regulating EGFR via a mechanism involving its degradation. Understanding the complex interplay between NDRG1, iron, and ErbB signaling is vital for identifying novel, more effective targets for cancer therapy.
Assuntos
Carcinogênese , Proteínas de Ciclo Celular/metabolismo , Receptores ErbB/antagonistas & inibidores , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Modelos Biológicos , Receptor ErbB-2/antagonistas & inibidores , Receptor ErbB-3/antagonistas & inibidores , Transdução de Sinais , Animais , Antígenos CD/metabolismo , Proteínas de Ciclo Celular/química , Vesículas Revestidas por Clatrina , Endocitose , Endossomos/enzimologia , Receptores ErbB/química , Receptores ErbB/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/química , Ferro/metabolismo , Ligantes , Multimerização Proteica , Proteólise , Receptor ErbB-2/agonistas , Receptor ErbB-2/química , Receptor ErbB-2/metabolismo , Receptor ErbB-3/agonistas , Receptor ErbB-3/química , Receptor ErbB-3/metabolismo , Receptores da Transferrina/agonistas , Receptores da Transferrina/metabolismo , Transferrina/metabolismoRESUMO
The metastasis suppressor, N-myc downstream regulated gene-1 (NDRG1), exhibits pleiotropic activity, inhibiting metastasis of various tumor-types, while being correlated with metastasis in others. Notably, NDRG1 phosphorylation and cleavage are associated with its function, although it is unclear if these modifications occur universally, or selectively, in different cancer cell-types and if it contributes to its pleiotropy. Considering the suggested DNA repair role of nuclear NDRG1, the effects of the above post-translational modifications on its nuclear localization was examined. Herein, the full-length (FL) and truncated (T) NDRG1 isoforms were detected using a C-terminus-directed antibody, while only the FL isoform was identified using an N-terminus-directed antibody. For the first time, we demonstrate that the expression of the NDRG1 FL and T forms occurs in all cancer cell-types examined, as does its phosphorylation (p-NDRG1) at Ser330 and Thr346. The FL isoform localized highly in the nucleus compared to the T isoform. Moreover, p-NDRG1 (Ser330) was also markedly localized in the nucleus, while p-NDRG1 (Thr346) was predominantly cytoplasmic in all cell-types. These results indicate the N-terminus region and phosphorylation at Ser330 could be crucial for NDRG1 nuclear localization and function. PTEN silencing indicated that p-NDRG1 (Thr346) could be regulated differentially in different tumor cell-types, indicating PTEN may be involved in the mechanism(s) underlying the pleiotropic activity of NDRG1. Finally, therapeutics of the di-2-pyridylketone thiosemicarbazone class increased nuclear NDRG1 isoforms (FL and T) detected by the C-terminus-directed antibody in HepG2 cells, while having no significant effect in PC3 cells, indicating differential activity depending on the cell-type.
Assuntos
Proteínas de Ciclo Celular/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , PTEN Fosfo-Hidrolase/metabolismo , Neoplasias da Próstata/metabolismo , Proteínas de Ciclo Celular/genética , Células Hep G2 , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Masculino , Metástase Neoplásica , PTEN Fosfo-Hidrolase/genética , Fosforilação/genética , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Transporte Proteico/genéticaRESUMO
The chelatable iron pool (CIP) is a small but chemically significant fraction of total cellular iron. While this dynamic population of iron is limited, it is redox active and capable of generating reactive oxygen species (ROS) that can lead to oxidative stress which is associated with various pathologies. Nitric oxide (â¢NO), is a free radical signalling molecule that regulates numerous physiological and pathological conditions. We have previously shown that macrophages exposed to endogenously generated or exogenously administered nitric oxide (â¢NO) results in its interaction with CIP to form dinitrosyliron complexes with thiol containing ligands (DNICs). In this study we assessed the consequences of DNIC formation in cancer cells as â¢NO is known to be associated with numerous malignancies. Incubation of cancer cells with â¢NO led to a time and dose dependent increase in formation of DNICs. The formation of DNICs results in the sequestration of the CIP which is a major source of iron for redox reactions and reactive oxygen species (ROS) generation. Therefore, we set out to test the antioxidant effect of â¢NO by measuring the ability of DNICs to protect cells against oxidative stress. We observed that cancer cells treated with â¢NO were partially protected against H2O2 mediated cytotoxicity. This correlated to a concomitant decrease in the formation of oxidants when â¢NO was present during H2O2 treatment. Similar protective effects were achieved by treating cells with iron chelators in the presence of H2O2. Interestingly, â¢NO decreased the rate of cellular metabolism of H2O2 suggesting that a proportion of H2O2 is consumed via reactions with cellular iron. When the CIP was artificially increased by supplementation of cells with iron, a significant decrease in the cytoprotective effect of â¢NO was observed. Notably, â¢NO concentrations, at which cytoprotective and antioxidant effects were observed, correlated with concentration-dependent increases in DNIC formation. Collectively, these results demonstrate that â¢NO has antioxidant properties by its ability to sequester cellular iron. This could play a significant role in variety of diseases involving ROS mediated toxicity like cancer and neurodegenerative disorders where â¢NO has been shown to be an important etiologic factor.
Assuntos
Neoplasias da Mama/metabolismo , Neoplasias do Colo/metabolismo , Ferro/metabolismo , Óxido Nítrico/farmacologia , Óxidos de Nitrogênio/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Neoplasias da Mama/patologia , Sobrevivência Celular/efeitos dos fármacos , Neoplasias do Colo/patologia , Feminino , Humanos , Peróxido de Hidrogênio/antagonistas & inibidores , Peróxido de Hidrogênio/metabolismo , Óxido Nítrico/metabolismo , Oxirredução , Células Tumorais CultivadasRESUMO
Nitric oxide (NO) is integral to macrophage cytotoxicity against tumors due to its ability to induce iron release from cancer cells. However, the mechanism for how activated macrophages protect themselves from endogenous NO remains unknown. We previously demonstrated by using tumor cells that glutathione S-transferase P1 (GSTP1) sequesters NO as dinitrosyl-dithiol iron complexes (DNICs) and inhibits NO-mediated iron release from cells via the transporter multidrug resistance protein 1 (MRP1/ABCC1). These prior studies also showed that MRP1 and GSTP1 protect tumor cells against NO cytotoxicity, which parallels their roles in defending cancer cells from cytotoxic drugs. Considering this, and because GSTP1 and MRP1 are up-regulated during macrophage activation, this investigation examined whether this NO storage/transport system protects macrophages against endogenous NO cytotoxicity in two well characterized macrophage cell types (J774 and RAW 264.7). MRP1 expression markedly increased upon macrophage activation, and the role of MRP1 in NO-induced 59Fe release was demonstrated by Mrp1 siRNA and the MRP1 inhibitor, MK571, which inhibited NO-mediated iron efflux. Furthermore, Mrp1 silencing increased DNIC accumulation in macrophages, indicating a role for MRP1 in transporting DNICs out of cells. In addition, macrophage 59Fe release was enhanced by silencing Gstp1, suggesting GSTP1 was responsible for DNIC binding/storage. Viability studies demonstrated that GSTP1 and MRP1 protect activated macrophages from NO cytotoxicity. This was confirmed by silencing nuclear factor-erythroid 2-related factor 2 (Nrf2), which decreased MRP1 and GSTP1 expression, concomitant with reduced 59Fe release and macrophage survival. Together, these results demonstrate a mechanism by which macrophages protect themselves against NO cytotoxicity.
Assuntos
Glutationa S-Transferase pi/antagonistas & inibidores , Isótopos de Ferro/metabolismo , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Proteínas Associadas à Resistência a Múltiplos Medicamentos/antagonistas & inibidores , Óxido Nítrico/metabolismo , Animais , Transporte Biológico , Broncodilatadores/farmacologia , Células Cultivadas , Glutationa/metabolismo , Glutationa S-Transferase pi/fisiologia , Macrófagos/metabolismo , Macrófagos/patologia , Camundongos , Camundongos Knockout , Proteínas Associadas à Resistência a Múltiplos Medicamentos/fisiologia , Óxido Nítrico/toxicidade , Propionatos/farmacologia , Substâncias Protetoras/farmacologia , Quinolinas/farmacologia , RNA Interferente Pequeno/genéticaRESUMO
N-MYC downstream-regulated gene-1 (NDRG1) is a potent growth and metastasis suppressor that acts through its inhibitory effects on a wide variety of cellular signaling pathways, including the TGF-ß pathway, protein kinase B (AKT)/PI3K pathway, RAS, etc. To investigate the hypothesis that its multiple effects could be regulated by a common upstream effector, the role of NDRG1 on the epidermal growth factor receptor (EGFR) and other members of the ErbB family, namely human epidermal growth factor receptor 2 (HER2) and human epidermal growth factor receptor 3 (HER3), was examined. We demonstrate that NDRG1 markedly decreased the expression and activation of EGFR, HER2, and HER3 in response to the epidermal growth factor (EGF) ligand, while also inhibiting formation of the EGFR/HER2 and HER2/HER3 heterodimers. In addition, NDRG1 also decreased activation of the downstream MAPKK in response to EGF. Moreover, novel anti-tumor agents of the di-2-pyridylketone class of thiosemicarbazones, namely di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone and di-2-pyridylketone 4-cyclohexyl-4-methyl-3-thiosemicarbazone, which markedly up-regulate NDRG1, were found to inhibit EGFR, HER2, and HER3 expression and phosphorylation in cancer cells. However, the mechanism involved appeared dependent on NDRG1 for di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone, but was independent of this metastasis suppressor for di-2-pyridylketone 4-cyclohexyl-4-methyl-3-thiosemicarbazone. This observation demonstrates that small structural changes in thiosemicarbazones result in marked alterations in molecular targeting. Collectively, these results reveal a mechanism for the extensive downstream effects on cellular signaling attributed to NDRG1. Furthermore, this study identifies a novel approach for the treatment of tumors resistant to traditional EGFR inhibitors.
Assuntos
Proteínas de Ciclo Celular/metabolismo , Neoplasias do Colo/metabolismo , Receptores ErbB/antagonistas & inibidores , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Sistema de Sinalização das MAP Quinases , Neoplasias Pancreáticas/metabolismo , Piridinas/uso terapêutico , Receptor ErbB-2/antagonistas & inibidores , Receptor ErbB-3/antagonistas & inibidores , Tiossemicarbazonas/uso terapêutico , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Proteínas de Ciclo Celular/agonistas , Proteínas de Ciclo Celular/antagonistas & inibidores , Proteínas de Ciclo Celular/genética , Linhagem Celular Tumoral , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/patologia , Fator de Crescimento Epidérmico/antagonistas & inibidores , Fator de Crescimento Epidérmico/genética , Fator de Crescimento Epidérmico/metabolismo , Receptores ErbB/agonistas , Receptores ErbB/genética , Receptores ErbB/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/agonistas , Peptídeos e Proteínas de Sinalização Intracelular/antagonistas & inibidores , Peptídeos e Proteínas de Sinalização Intracelular/genética , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Camundongos Nus , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/patologia , Piridinas/farmacologia , Interferência de RNA , Distribuição Aleatória , Receptor ErbB-2/agonistas , Receptor ErbB-2/genética , Receptor ErbB-2/metabolismo , Receptor ErbB-3/agonistas , Receptor ErbB-3/genética , Receptor ErbB-3/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Tiossemicarbazonas/farmacologia , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Adenosine monophosphate-activated protein kinase (AMPK) is a cellular energy sensor that monitors ATP levels. There is also evidence that AMPK has onco-suppressive properties. Iron plays a crucial role in cellular energy transducing pathways and tumor cell proliferation. Therefore, metals (e.g., iron) could play an important role in the regulation of AMPK-dependent pathways. Hence, this investigation examined the effect of the iron and copper chelator and potent anti-cancer agent, di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone (Dp44mT), on the AMPK-mediated pathway. These studies demonstrated that Dp44mT, which forms intracellular redox-active complexes with iron and copper, significantly activated AMPK (i.e., p-AMPK/AMPK ratio) in 5 different tumor cell-types. Furthermore, examination of the Dp44mT-metal complexes demonstrated that the effect of Dp44mT on AMPK was due to a dual mechanism: (1) its ability to chelate metal ions; and (2) the generation of reactive oxygen species (ROS). The activation of the AMPK-pathway by Dp44mT was mediated by the upstream kinase, liver kinase B1 (LKB1) that is a known tumor suppressor. Moreover, using AMPKα1-selective silencing, we demonstrated that Dp44mT activated AMPK, resulting in inhibition of acetyl CoA carboxylase 1 (ACC1) and raptor, and activation of Unc-51 like kinase (ULK1). These effects are vital for inhibition of fatty acid synthesis, suppression of protein synthesis and autophagic activation, respectively. Together, this AMPK-mediated repair response aims to rescue the loss of metal ions via chelation and the induction of cytotoxic damage mediated by redox cycling of the Dp44mT-metal ion complex. In conclusion, this study demonstrates for the first time that chelators target the AMPK-dependent pathway.
Assuntos
Proteínas Quinases Ativadas por AMP/genética , Antineoplásicos/farmacologia , Células Epiteliais/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica , Quelantes de Ferro/farmacologia , Tiossemicarbazonas/farmacologia , Quinases Proteína-Quinases Ativadas por AMP , Proteínas Quinases Ativadas por AMP/metabolismo , Acetil-CoA Carboxilase/genética , Acetil-CoA Carboxilase/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Autofagia/efeitos dos fármacos , Proteína Homóloga à Proteína-1 Relacionada à Autofagia/genética , Proteína Homóloga à Proteína-1 Relacionada à Autofagia/metabolismo , Linhagem Celular Tumoral , Metabolismo Energético/efeitos dos fármacos , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Ácidos Graxos/antagonistas & inibidores , Ácidos Graxos/biossíntese , Fibroblastos , Células Endoteliais da Veia Umbilical Humana , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Ferro/metabolismo , Camundongos , Biossíntese de Proteínas/efeitos dos fármacos , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Proteína Regulatória Associada a mTOR , Transdução de SinaisRESUMO
Melanoma has markedly increased worldwide during the past several decades in the Caucasian population and is responsible for 80% of skin cancer deaths. Considering that metastatic melanoma is almost completely resistant to most current therapies and is linked with a poor patient prognosis, it is crucial to further investigate potential molecular targets. Major cell-autonomous drivers in the pathogenesis of this disease include the classical MAPK (i.e., RAS-RAF-MEK-ERK), WNT, and PI3K signaling pathways. These pathways play a major role in defining the progression of melanoma, and some have been the subject of recent pharmacological strategies to treat this belligerent disease. This review describes the latest advances in the understanding of melanoma progression and the major molecular pathways involved. In addition, we discuss the roles of emerging molecular players that are involved in melanoma pathogenesis, including the functional role of the melanoma tumor antigen, p97/MFI2 (melanotransferrin).
Assuntos
Melanoma/genética , Melanoma/patologia , Oncogenes/fisiologia , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/patologia , Animais , Progressão da Doença , Genes ras/fisiologia , Humanos , Sistema de Sinalização das MAP Quinases/fisiologia , Fosfatidilinositol 3-Quinases/metabolismo , Transdução de Sinais/genética , Via de Sinalização Wnt/fisiologia , Quinases raf/fisiologiaRESUMO
The potent and selective anti-tumor agent, di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone (Dp44mT), localizes in lysosomes and forms cytotoxic copper complexes that generate reactive oxygen species (ROS), resulting in lysosomal membrane permeabilization (LMP) and cell death. Herein, the role of lysosomal membrane stability in the anti-tumor activity of Dp44mT was investigated. Studies were performed using molecules that protect lysosomal membranes against Dp44mT-induced LMP, namely heat shock protein 70 (HSP70) and cholesterol. Up-regulation or silencing of HSP70 expression did not affect Dp44mT-induced LMP in MCF7 cells. In contrast, cholesterol accumulation in lysosomes induced by the well characterized cholesterol transport inhibitor, 3-ß-[2-(diethyl-amino)ethoxy]androst-5-en-17-one (U18666A), inhibited Dp44mT-induced LMP and markedly and significantly (p<0.001) reduced the ability of Dp44mT to inhibit cancer cell proliferation (i.e., increased the IC(50)) by 140-fold. On the other hand, cholesterol extraction using methyl-ß-cyclodextrin enhanced Dp44mT-induced LMP and significantly (p<0.01) increased its anti-proliferative activity. The protective effect of U18666A in increasing lysosomal cholesterol and preventing the cytotoxic activity of Dp44mT was not due to induced autophagy. Instead, U18666A was found to decrease lysosomal turnover, resulting in autophagosome accumulation. Moreover, preincubation with U18666A did not prevent the ability of Dp44mT to induce autophagosome synthesis, indicating that autophagic initiation via Dp44mT occurs independently of LMP. These studies demonstrate the significance of lysosomal membrane stability in relation to the ability of Dp44mT to execute tumor cell death and overcome pro-survival autophagy. Hence, lysosomal-dependent cell death induced by Dp44mT serves as an important anti-tumor strategy. These results are important for comprehensively understanding the mechanism of action of Dp44mT.
Assuntos
Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Proliferação de Células/efeitos dos fármacos , Membranas Intracelulares/efeitos dos fármacos , Lisossomos/efeitos dos fármacos , Tiossemicarbazonas/farmacologia , Androstenos/farmacologia , Anticolesterolemiantes/farmacologia , Antineoplásicos/metabolismo , Autofagia/efeitos dos fármacos , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Colesterol/metabolismo , Relação Dose-Resposta a Droga , Feminino , Proteínas de Choque Térmico HSP70/genética , Proteínas de Choque Térmico HSP70/metabolismo , Humanos , Concentração Inibidora 50 , Membranas Intracelulares/metabolismo , Membranas Intracelulares/patologia , Lisossomos/metabolismo , Lisossomos/patologia , Células MCF-7 , Permeabilidade , Interferência de RNA , Tiossemicarbazonas/metabolismo , Transfecção , beta-Ciclodextrinas/farmacologiaRESUMO
Essential metals, such as iron and copper, play a critical role in a plethora of cellular processes including cell growth and proliferation. However, concomitantly, excess of these metal ions in the body can have deleterious effects due to their ability to generate cytotoxic reactive oxygen species (ROS). Thus, the human body has evolved a very well-orchestrated metabolic system that keeps tight control on the levels of these metal ions. Considering their very high proliferation rate, cancer cells require a high abundance of these metals compared to their normal counterparts. Interestingly, new anti-cancer agents that take advantage of the sensitivity of cancer cells to metal sequestration and their susceptibility to ROS have been developed. These ligands can avidly bind metal ions to form redox active metal complexes, which lead to generation of cytotoxic ROS. Furthermore, these agents also act as potent metastasis suppressors due to their ability to up-regulate the metastasis suppressor gene, N-myc downstream regulated gene 1. This review discusses the importance of iron and copper in the metabolism and progression of cancer, how they can be exploited to target tumors and the clinical translation of novel anti-cancer chemotherapeutics.
Assuntos
Antineoplásicos , Quelantes , Cobre/metabolismo , Descoberta de Drogas , Ferro/metabolismo , Metais/metabolismo , Animais , Antineoplásicos/uso terapêutico , Quelantes/uso terapêutico , Descoberta de Drogas/métodos , Descoberta de Drogas/tendências , Humanos , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Neoplasias/patologia , Oxirredução , Espécies Reativas de Oxigênio/metabolismoRESUMO
Autophagy is an evolutionary conserved cellular catabolic degradation process in response to stress which involves lysosomal degradation of unnecessary or damaged organelles and misfolded proteins. This is primarily a pro-survival pathway providing the cell with essential nutrients during stressful conditions. There are number of essential metal ions, which are required for normal physiological functioning of cells. Studies have shown that autophagy can be regulated by cellular metal ion concentrations. On the other hand, autophagy is also shown to regulate intracellular levels of certain metal ions. This review discusses recent advances in the research examining the role of metal ions in the autophagic pathway.
Assuntos
Autofagia , Metais/metabolismo , Animais , Cálcio/metabolismo , Cobre/metabolismo , Humanos , Íons/metabolismo , Ferro/metabolismo , Potássio/metabolismo , Transdução de Sinais , Zinco/metabolismoRESUMO
Metastasis is a complex process that is regulated by multiple signaling pathways, with the focal adhesion kinase (FAK)/paxillin pathway playing a major role in the formation of focal adhesions and cell motility. N-myc downstream regulated gene-1 (NDRG1) is a potent metastasis suppressor in many solid tumor types, including prostate and colon cancer. Considering the antimetastatic effect of NDRG1 and the crucial involvement of the FAK/paxillin pathway in cellular migration and cell-matrix adhesion, we assessed the effects of NDRG1 on this important oncogenic pathway. In the present study, NDRG1 overexpression and silencing models of HT29 colon cancer and DU145 prostate cancer cells were used to examine the activation of FAK/paxillin signaling and the formation of focal adhesions. The expression of NDRG1 resulted in a marked and significant decrease in the activating phosphorylation of FAK and paxillin, whereas silencing of NDRG1 resulted in an opposite effect. The expression of NDRG1 also inhibited the formation of focal adhesions as well as cell migration and cell-collagen adhesion. Incubation of cells with novel thiosemicarbazones, namely di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone and di-2-pyridylketone 4-cyclohexyl-4-methyl-3-thiosemicarbazone, that upregulate NDRG1 also resulted in decreased phosphorylation of FAK and paxillin. The ability of these thiosemicarbazones to inhibit cell migration and metastasis could be mediated, at least in part, through the FAK/paxillin pathway.