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Background/Aims: Extracellular vesicles (EVs) are promising as a biomarker of metabolic dysfunction associated steatotic liver disease (MASLD). The objective is to study EVs and their involvement in MASLD concerning the disease's pathogenesis and progression characteristics. Methods: Male adult Sprague Dawley rats were randomly assigned into two experimental models of MASLD: MASLD-16 and MASLD-28, animals received a choline-deficient high-fat diet (CHFD) and Control-16 and Control-28, animals received a standard diet (SD) for 16 and 28 weeks, respectively. Biological samples from these animal models were used, as well as previously registered variables. EVs from hepatic tissue were characterized using confocal microscopy. EVs were isolated through differential ultracentrifugation from serum and characterized using NanoSight. The data from the EVs were correlated with biochemical, molecular, and histopathological parameters. Results: Liver EVs were identified through the flotillin-1 protein. EVs were isolated from the serum of all groups. There was a decrease of EVs concentration in MASLD-28 in comparison with Control-28 (P < 0.001) and a significant increase in EVs concentration in Control-28 compared with Control-16 (P < 0.001). There was a strong correlation between serum EVs concentration with hepatic gene expression of interleukin (Il)6 (r2 = 0.685, P < 0.05), Il1b (r2 = 0.697, P < 0.05) and tumor necrosis factor-alpha (Tnfa; r2 = 0.636, P < 0.05) in MASLD-16. Moreover, there was a strong correlation between serum EVs size and Il10 in MASLD-28 (r2 = 0.762, P < 0.05). Conclusion: The concentration and size of EVs correlated with inflammatory markers, suggesting their involvement in the systemic circulation, cellular communication, and development and progression of MASLD, demonstrating that EVs have the potential to serve as noninvasive biomarkers for MASLD diagnosis and prognosis.
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Dieta Hiperlipídica , Modelos Animais de Doenças , Vesículas Extracelulares , Ratos Sprague-Dawley , Vesículas Extracelulares/metabolismo , Vesículas Extracelulares/patologia , Animais , Masculino , Ratos , Fígado/metabolismo , Fígado/patologia , Biomarcadores/sangue , Biomarcadores/metabolismo , Hepatopatia Gordurosa não Alcoólica/metabolismo , Hepatopatia Gordurosa não Alcoólica/patologia , Hepatopatia Gordurosa não Alcoólica/etiologia , Fígado Gorduroso/patologia , Fígado Gorduroso/metabolismo , Fígado Gorduroso/etiologia , Mediadores da Inflamação/metabolismo , Mediadores da Inflamação/sangue , Inflamação/patologia , Inflamação/metabolismo , Deficiência de Colina/complicaçõesRESUMO
In 2020, breast cancer (BC) has surpassed lung cancer as the most diagnosed cancer in the world. Tumor microenvironment (TME) plays a critical role in resistance to standard therapies and tumor progression. Two key factors within the TME include adenosine, an immunosuppressive molecule, and glucose, which serves as the primary energy source for tumor cells. In this scenario, inhibiting the purinergic pathway and glucose uptake might be a promising strategy. Therefore, we sought to evaluated different treatment approaches in BC cells (Dapagliflozin, a SGLT2 inhibitor; Paclitaxel, the standard chemotherapy for BC; and ARL67156/APCP, inhibitors of CD39 and CD73, respectively). The expression of some membrane markers relevant to resistance was assessed. BC cell-lines (MCF-7 and MDA-MB-231) were co-treated and cell viability, cell cycle, and annexin/PI assays were performed. Our analysis showed promising results, where the combination of these compounds led to cell death by apoptosis/necrosis and cell cycle arrest. Dapagliflozin showed more impact on early apoptosis, whereas Paclitaxel led to late apoptosis/necrosis as the main mechanism of cell death. Inhibiting purinergic signaling also contributed to reducing cell viability together with the other drugs, suggesting it could have an influence on breast cancer survival mechanisms. Indeed, the overexpression of the NT5E gene in patients with ER+ tumors is strongly associated with reduced overall survival and progression-free interval. However, more studies are needed to fully understand the interactions and mechanism underlying these co-treatment multi-targeting approaches.
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Compostos Benzidrílicos , Neoplasias da Mama , Glucosídeos , Humanos , Feminino , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Paclitaxel/farmacologia , Apoptose , Necrose , Proliferação de Células , Microambiente TumoralRESUMO
Malignant gliomas are highly heterogeneous glia-derived tumors that present an aggressive and invasive nature, with a dismal prognosis. The multi-dimensional interactions between glioma cells and other tumor microenvironment (TME) non-tumoral components constitute a challenge to finding successful treatment strategies. Several molecules, such as extracellular purines, participate in signaling events and support the immunosuppressive TME of glioma patients. The purinergic signaling and the ectoenzymes network involved in the metabolism of these extracellular nucleotides are still unexplored in the glioma TME, especially in lower-grade gliomas (LGG). Also, differences between IDH-mutant (IDH-Mut) versus wild-type (IDH-WT) gliomas are still unknown in this context. For the first time, to our knowledge, this study characterizes the TME of LGG, high-grade gliomas (HGG) IDH-Mut, and HGG IDH-WT patients regarding purinergic ectoenzymes and P1 receptors, focusing on tumor-infiltrating lymphocytes. Here, we show that ectoenzymes from both canonical and non-canonical pathways are increased in the TME when compared to the peripheral blood. We hypothesize this enhancement supports extracellular adenosine generation, hence increasing TME immunosuppression.
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Neoplasias Encefálicas , Glioma , Humanos , Neoplasias Encefálicas/patologia , Linfócitos do Interstício Tumoral/patologia , Isocitrato Desidrogenase/genética , Glioma/patologia , Prognóstico , Mutação , Microambiente TumoralRESUMO
Therapies for wound healing using the secretome and extracellular vesicles (EVs) of mesenchymal stem/stromal cells have been shown to be successful in preclinical studies. This study aimed to characterise the protein content of the secretome from stem cells from human exfoliated deciduous teeth (SHED) and analyse the in vitro effects of SHED-conditioned medium (SHED-CM) and SHED extracellular vesicles (SHED-EVs) on keratinocytes. EVs were isolated and characterised. The keratinocyte viability and migration of cells treated with SHED-EVs and conditioned medium (CM) were evaluated. An HaCaT apoptosis model induced by H2 O2 in vitro was performed with H2 O2 followed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and live/dead assays. Finally, the expression of vascular endothelial growth factor (VEGF) in keratinocytes treated with secretome and EVs was evaluated by immunofluorescence staining and confirmed with RT-qPCR. SHED-EVs revealed a cup-shaped morphology with expression of the classical markers for exosomes CD9 and CD63, and a diameter of 181 ± 87 nm. The internalisation of EVs by HaCaT cells was confirmed by fluorescence microscopy. Proteomic analysis identified that SHED-CM is enriched with proteins related to stress response and development, including cytokines (CXCL8, IL-6, CSF1, CCL2) and growth factors (IGF2, MYDGF, PDGF). The results also indicated that 50% CM and 0.4-0.6 µg/mL EVs were similarly efficient for improving keratinocyte viability, migration, and attenuation of H2 O2 -induced cytotoxicity. Additionally, expression of VEGF on keratinocytes increased when treated with SHED secretome and EVs. Furthermore, VEGF gene expression in keratinocytes increased significantly when treated with SHED secretome and EVs. Both SHED-CM and SHED-EVs may therefore be promising therapeutic tools for accelerating re-epithelialization in wound healing.
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Vesículas Extracelulares , Cicatrização , Humanos , Fator A de Crescimento do Endotélio Vascular/metabolismo , Meios de Cultivo Condicionados/farmacologia , Meios de Cultivo Condicionados/metabolismo , Proteômica , Secretoma , Células-Tronco/metabolismo , Queratinócitos , Vesículas Extracelulares/metabolismo , Dente DecíduoRESUMO
Prostate cancer (PC) is the most diagnosed tumor in males and ranks as the second leading cause of male mortality in the western world. The CD39 and CD73 enzymes play a crucial role in cancer regulation by degrading nucleotides and forming nucleosides. This study aimed to investigate the expression of the CD39 and CD73 enzymes as potential therapeutic targets for PC. The initial part of this study retrospectively analyzed tissue samples from 23 PC patients. Using the TissueFAXSTM cytometry platform, we found significantly higher levels of CD39-labeling its intensity compared to CD73. Additionally, we observed a correlation between the Gleason score and the intensity of CD39 expression. In the prospective arm, blood samples were collected from 25 patients at the time of diagnosis and after six months of treatment to determine the expression of CD39 and CD73 in the serum extracellular vesicles (EVs) and to analyze nucleotide hydrolysis. Notably, the expression of CD39 in the EVs was significantly increased compared to the CD73 and/or combined CD39/CD73 expression levels at initial collection. Furthermore, our results demonstrated positive correlations between ADP hydrolysis and the transurethral resection and Gleason score. Understanding the role of ectonucleotidases is crucial for identifying new biomarkers in PC.
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Bladder cancer (BC) is the most common cancer of the urinary tract. Bozepinib (BZP), a purine-derived molecule, is a potential compound for the treatment of cancer. Purinergic signaling consists of the activity of nucleosides and nucleotides present in the extracellular environment, modulating a variety of biological actions. In cancer, this signaling is mainly controlled by the enzymatic cascade involving the NTPDase/E-NPP family and ecto-5'-nucleotidase/CD73, which hydrolyze extracellular adenosine triphosphate (ATP) to adenosine (ADO). The aim of this work is to evaluate the activity of BZP in the purinergic system in BC cell lines and to compare its in vitro antitumor activity with cisplatin, a chemotherapeutic drug widely used in the treatment of BC. In this study, two different BC cell lines, grade 1 RT4 and the more aggressive grade 3 T24, were used along with a human fibroblast cell line MRC-5, a cell used to predict the selectivity index (SI). BZP shows strong antitumor activity, with notable IC50 values (8.7 ± 0.9 µM for RT4; 6.7 ± 0.7 µM for T24), far from the SI for cisplatin (SI for BZP: 19.7 and 25.7 for RT4 and T24, respectively; SI for cisplatin: 1.7 for T24). BZP arrests T24 cells in the G2/M phase of the cell cycle, inducing early apoptosis. Moreover, BZP increases ATP and ADP hydrolysis and gene/protein expression of the NPP1 enzyme in the T24 cell line. In conclusion, BZP shows superior activity compared to cisplatin against BC cell lines in vitro.
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Bladder cancer is the fourth most common malignancy in men. It can present along the entire continuum of severity, from mild to well-differentiated disease to extremely malignant tumors with low survival rates. Human RAS genes are the most frequently mutated oncogenes in human cancers, and the critical role of aberrant Ras protein function in carcinogenesis is well established. Therefore, considerable efforts have been devoted to the development of anti-Ras inhibitors for cancer treatment. This study presents the biphenyl dihydropyrimidinone LaSOM 335 with high activity against T24 bladder cancer cells (IC50 = 10.73 ± 0.53 µM) and selectivity of cytotoxicity for this cancer cell line compared to two non-cancer cell lines investigated. Furthermore, we also show that this compound reduced vulvar development in the mutant let-60 gene of Caenorhabditis elegans. Let-60 is a homolog of the mammalian Ras gene. In addition, we observed that LaSOM 335 inhibits the enzymatic activity of CD73 and decreases CD73 expression. Possibly, this expression decrease is due to downstream EGFR signaling via the Ras-Raf-ERK pathway, that directly regulates CD73 expression via ERK1/2. Evidence suggests that non-immunomodulating functions of CD73 play an equally important role for cancer cell survival, progression, and migration. Regarding we also notice that LaSOM 335 was safe in the in vivo model of C. elegans. The set of these findings makes this biphenyl dihydropyrimidinone a promising candidate for further investigations in the bladder cancer field.
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Genes ras , Neoplasias da Bexiga Urinária , Masculino , Animais , Humanos , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Linhagem Celular Tumoral , Neoplasias da Bexiga Urinária/tratamento farmacológico , Neoplasias da Bexiga Urinária/metabolismo , 5'-Nucleotidase/genética , 5'-Nucleotidase/metabolismo , Mamíferos/genética , Mamíferos/metabolismoRESUMO
The purinergic signaling has drawn attention from academia and more recently from pharmaceutical industries as a potential therapeutic route for cancer treatment, since ATP may act as chemotactic agent and possess in vitro antineoplastic activity. On the other way, adenosine, produced in extracellular medium by ecto-5'-NT, acts as immunosuppressor and is related to neoangiogenesis, vasculogenesis and evasion to the immune system. Consequently, inhibitors of ecto-5'-NT may prevent tumor progression, reducing adenosine concentrations, preventing escape from the host's immune system and slowing cancer's growth. This review aims to highlight important biochemical and structural features of ecto-5'NT, highlight its expression profile in normal and cancer cell lines detailing compounds which may act as expression regulators and to review the several classes of ecto-5'NT inhibitors developed in the past 12 years, in order to build a general structure-activity relationship model to guide further compound design.
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5'-Nucleotidase , Antineoplásicos , Adenosina/farmacologia , Adenosina/metabolismo , Monofosfato de Adenosina , Antineoplásicos/farmacologia , Linhagem CelularRESUMO
Nanoencapsulation of chemotherapeutics, including doxorubicin, can endow the formulations with unique properties, such as a decrease in adverse effects and toxicity. The chicken embryo model is an alternative and well-accepted strategy for evaluating the toxicity and efficacy of drugs and nanoformulations. Therefore, this study proposes the development of a new lipid nanocarrier for doxorubicin delivery (NanoLip-Dox) and posterior evaluation of toxicological profile and antitumoral efficacy against a breast tumor in chicken embryos. NanoLip-Dox showed a unimodal particle size (< 150 nm), negative zeta potential (-19.5 mV), absence of drug crystals, drug content of 0.099 mg·mL-1, and high entrapment efficiency (95%). NanoLip-Dox did not cause toxicity in the chicken embryos; in contrast, doxorubicin hydrochloride induced moderate irritation in the chorioallantoic membrane (at 862.1 µmol·L-1), a survival rate of 50% (at 1.7 µmol·L-1), and an increase in aspartate aminotransferase (at 862.1, 344.8, and 172.4 µmol·L-1). In addition, NanoLip-Dox (at 1.7 µmol·L-1) showed potent antitumor efficacy with a high tumor remission percentage (40.9 ± 9.7%) compared to the control group (8.6 ± 14.8%). These findings together with the absence of toxicity concerning morphological characteristics, weights of embryos and organs, hematologic parameters, and enzymatic activity (alanine aminotransferase, aspartate aminotransferase, and creatinine) suggest the safety and efficacy of NanoLip-Dox.
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PURPOSE: Although risk-stratified chemotherapy regimens improve B-cell acute lymphoblastic leukemia (B-ALL) clinical outcome, relapse occurs in a significant number of cases. The identification of new therapeutic targets as well as prognostic and diagnostic biomarkers can improve B-ALL patients' clinical outcomes. Purinergic signaling is an important pathway in cancer progression, however the expression of ectonucleotidases and their impact on immune cells in B-ALL lacks exploration. We aimed to analyze the expression of ectonucleotidases in B-ALL patients' lymphocyte subpopulations. METHODS: Peripheral blood samples from 15 patients diagnosed with B-ALL were analyzed. Flow cytometry was used to analyze cellularity, expression level of CD38, CD39, and CD73, and frequency of [Formula: see text], and [Formula: see text] in lymphocyte subpopulations. Plasma was used for cytokines (by CBA kit) and adenine nucleosides/nucleotides detection (by HPLC). RESULTS: Comparing B-ALL patients to health donors, we observed an increase of CD4 + and CD8 + T-cells. In addition, a decrease in CD38 expression in B and Treg subpopulations and an increase in CD39+ CD73+ frequency in Breg and CD8+ T-cells. Analyzing cytokines and adenine nucleosides/nucleotides, we found a decrease in TNF, IL-1ß, and ADO concentrations, together with an increase in AMP in B-ALL patients' plasma. CONCLUSION: As immunomodulators, the expression of ectonucleotidases might be associated with activation states, as well as the abundance of different cellular subsets. We observed a pro-tumor immunity expression profile in B-ALL patients at diagnosis, being associated with cell exhaustion and immune evasion in B-ALL.
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Head and neck squamous cell carcinoma (HNSCC) is an aggressive malignancy. Tumor-derived exosomes (TEX) have immunoregulatory properties. Adenosine triphosphate (ATP) and its immunosuppressive precursor adenosine (ADO) have been found in cancerous tissue. We investigated the effect of TEX on B cells in the presence of ATP. TEX were isolated from human HNSCC cell line (PCI-13) cultures and co-cultured with peripheral blood B cells of healthy donors, with or without TEX in different concentrations and with or without a low (20 µM) or high (2000 µM) ATP dose. We were able to demonstrate that TEX inhibit B-cell proliferation. The addition of TEX to either ATP concentration showed a decreasing trend in CD39 expression on B cells in a dose-dependent manner. High ATP levels (2000 µM) increased apoptosis and necrosis, and analysis of apoptosis-associated proteins revealed dose-dependent effects of ATP, which were modified by TEX. Altogether, TEX exhibited dual immunomodulatory effects on B cells. TEX were immunosuppressive by inhibiting B-cell proliferation; they were immunostimulatory by downregulating CD39 expression. Furthermore, TEX were able to modulate the expression of pro- and anti-apoptotic proteins. In conclusion, our data indicate that TEX play an important, but complex, role in the tumor microenvironment.
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Exossomos , Neoplasias de Cabeça e Pescoço , Intervenção Coronária Percutânea , Humanos , Exossomos/metabolismo , Carcinoma de Células Escamosas de Cabeça e Pescoço/metabolismo , Trifosfato de Adenosina/metabolismo , Neoplasias de Cabeça e Pescoço/metabolismo , Microambiente TumoralRESUMO
Ectonucleotidases modulate inflammatory responses by balancing extracellular ATP and adenosine (ADO) and might be involved in COVID-19 immunopathogenesis. Here, we explored the contribution of extracellular nucleotide metabolism to COVID-19 severity in mild and severe cases of the disease. We verified that the gene expression of ectonucleotidases is reduced in the whole blood of patients with COVID-19 and is negatively correlated to levels of CRP, an inflammatory marker of disease severity. In line with these findings, COVID-19 patients present higher ATP levels in plasma and reduced levels of ADO when compared to healthy controls. Cell type-specific analysis revealed higher frequencies of CD39+ T cells in severely ill patients, while CD4+ and CD8+ expressing CD73 are reduced in this same group. The frequency of B cells CD39+CD73+ is also decreased during acute COVID-19. Interestingly, B cells from COVID-19 patients showed a reduced capacity to hydrolyze ATP into ADP and ADO. Furthermore, impaired expression of ADO receptors and a compromised activation of its signaling pathway is observed in COVID-19 patients. The presence of ADO in vitro, however, suppressed inflammatory responses triggered in patients' cells. In summary, our findings support the idea that alterations in the metabolism of extracellular purines contribute to immune dysregulation during COVID-19, possibly favoring disease severity, and suggest that ADO may be a therapeutic approach for the disease.
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COVID-19 , Adenosina/metabolismo , Difosfato de Adenosina , Trifosfato de Adenosina/metabolismo , Humanos , Purinas , Índice de Gravidade de Doença , Transdução de SinaisRESUMO
Glioblastoma (GBM) is the most aggressive and lethal among the primary brain tumors, with a low survival rate and resistance to radio and chemotherapy. The P2Y12 is an adenosine diphosphate (ADP) purinergic chemoreceptor, found mainly in platelets. In cancer cells, its activation has been described to induce proliferation and metastasis. Bearing in mind the need to find new treatments for GBM, this study aimed to investigate the role of the P2Y12R in the proliferation and migration of GBM cells, as well as to evaluate the expression of this receptor in patients' data obtained from the TCGA data bank. Here, we used the P2Y12R antagonist, ticagrelor, which belongs to the antiplatelet agent's class. The different GBM cells (cell line and patient-derived cells) were treated with ticagrelor, with the agonist, ADP, or both, and the effects on cell proliferation, colony formation, ADP hydrolysis, cell cycle and death, migration, and cell adhesion were analyzed. The results showed that ticagrelor decreased the viability and the proliferation of GBM cells. P2Y12R antagonism also reduced colony formation and migration potentials, with alterations on the expression of metalloproteinases, and induced autophagy in GBM cells. Changes were observed at the cell cycle level, and only the U251 cell line showed a significant reduction in the ADP hydrolysis profile. TCGA data analysis showed a higher expression of P2Y12R in gliomas samples when compared to the other tumors. These data demonstrate the importance of the P2Y12 receptor in gliomas development and reinforce its potential as a pharmacological target for glioma treatment.
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Glioblastoma , Humanos , Ticagrelor/metabolismo , Ticagrelor/farmacologia , Difosfato de Adenosina/metabolismo , Glioblastoma/tratamento farmacológico , Plaquetas , Autofagia , Proliferação de Células , Receptores Purinérgicos P2Y12/metabolismo , Antagonistas do Receptor Purinérgico P2Y/metabolismoRESUMO
The risk stratification of B-acute lymphoblastic leukemia (B-ALL) is based on clinical and biological factors. However, B-ALL has significant biological and clinical heterogeneity and 50% of B-ALL patients do not have defined prognostic markers. In this sense, the identification of new prognostic biomarkers is necessary. Considering different cohorts of childhood B-ALL patients, gene (DPP4/CD38/ENTPD1/NT5E) and protein (CD38/CD39/CD73) expressions of ectonucleotidases were analyzed in silico and ex vivo and the association with prognosis was established. In univariate analyses, expression of NT5E was significantly associated with worse progression-free survival (PFS) in bone marrow (BM) samples. In multivariate analyses, Kaplan-Meier analysis, and log-rank test, higher NT5E expression predicted unfavorable PFS in BM samples. Considering minimal residual disease (MRD), higher levels of cellularity were associated with the high NT5E expression at day 8 of induction therapy. In addition, we observed that white blood cells (WBC) of childhood B-ALL patients had more CD38 compared to the same cell population of healthy donors (HD). In fact, MRD > 0.1% patients had higher CD38 protein expression on WBC in comparison to HD. Noteworthy, we observed higher CD38 expression on WBC than blasts in MRD > 0.1% patients. We suggest that NT5E gene and CD38 protein expression, of the ectonucleotidases family, could provide interesting prognostic biomarkers for childhood B-ALL.
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Leucemia-Linfoma Linfoblástico de Células Precursoras , 5'-Nucleotidase/genética , Biomarcadores , Citometria de Fluxo , Proteínas Ligadas por GPI , Humanos , Neoplasia Residual/tratamento farmacológico , Neoplasia Residual/metabolismo , Leucemia-Linfoma Linfoblástico de Células Precursoras/diagnóstico , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamento farmacológico , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , PrognósticoRESUMO
Glioblastoma is the most common and lethal malignant brain tumor. Despite simvastatin (SVT) showing potential anticancer properties, its antitumoral effect against glioblastoma appears limited when the conventional oral administration route is selected. As a consequence, nose-to-brain delivery has been proposed as an alternative route to deliver SVT into the brain. This study aimed to prepare chitosan-coated simvastatin-loaded lipid-core nanocapsules (LNCSVT-chit) suitable for nose-to-brain delivery and capable of fostering antitumor effects against glioblastoma both in vitro and in vivo. Results showed that the nanocapsules present adequate particle size (mean diameter below 200 nm), narrow particle size distribution (PDI < 0.2), positive zeta potential and high encapsulation efficiency (nearly 100%). In vitro cytotoxicity of LNCSVT-chit was comparable to non-encapsulated SVT in C6 rat glioma cells, whereas LNCSVT-chit were more cytotoxic than non-encapsulated SVT after 72 h of incubation against U-138 MG human glioblastoma cell line. In studies carried out in rats, LNCSVT-chit significantly enhanced the amount of drug in rat brain tissue after intranasal administration (2.4-fold) when compared with free SVT. Moreover, LNCSVT-chit promoted a significant decrease in tumor growth and malignancy in glioma-bearing rats in comparison to control and free SVT groups. Additionally, LNCSVT-chit did not cause any toxicity in treated rats. Considered overall, the results demonstrated that the nose-to-brain administration of LNCSVT-chit represents a novel potential strategy for glioblastoma treatment.
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Quitosana , Glioblastoma , Nanocápsulas , Administração Intranasal , Animais , Encéfalo/metabolismo , Linhagem Celular Tumoral , Quitosana/uso terapêutico , Glioblastoma/tratamento farmacológico , Glioblastoma/metabolismo , Lipídeos , Ratos , SinvastatinaRESUMO
Glioblastoma (GBM) is the most lethal among malignant gliomas. The tumor invasiveness and therapy-resistance are important clinical hallmarks. Growing evidence emphasizes the purinergic signaling contributing to tumor growth. Here we exposed a potential role of extracellular ATPase activity as a key regulator of temozolomide cytotoxicity and the migration process in GBM cells. The inhibition of ATP hydrolysis was able to improve the impact of temozolomide, causing arrest mainly in S and G2 phases of the cell cycle, leading M059J and U251 cells to apoptosis. In addition to eradicating GBM cells, ATP hydrolysis exhibited a potential to modulate the invasive phenotype and the expression of proteins involved in cell migration and epithelial-to-mesenchymal-like transition in a 3D culture model. Finally, we suggest the ATPase activity as a key target to decline temozolomide resistance and the migratory phenotype in GBM cells.
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Neoplasias Encefálicas , Glioblastoma , Adenosina Trifosfatases/genética , Trifosfato de Adenosina/farmacologia , Antineoplásicos Alquilantes/farmacologia , Antineoplásicos Alquilantes/uso terapêutico , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Proliferação de Células , Resistencia a Medicamentos Antineoplásicos , Glioblastoma/patologia , Humanos , Hidrólise , Fenótipo , Temozolomida/farmacologia , Temozolomida/uso terapêuticoRESUMO
Aim: To develop and characterize bozepinib-loaded lipid-core nanocapsules (BZP-LNC+) as a potential treatment for glioblastoma (GBM). Methods: Characterization of nanocapsules was performed by diameter, polydispersity index, Zeta potential, pH and encapsulation efficiency. GBM cell viability, cell cycle and Annexin/PI were evaluated after BZP-LNC+ treatment. Synergism between BZP-LNC+ and temozolomide (TMZ) was performed by CompuSyn software and confirmed in vitro and in vivo. Results: BZP-LNC+ showed adequate particle sizes, positive Zeta potential, narrow size distribution and high encapsulation efficiency. BZP-LNC+ reduces GBM growth by inducing apoptosis. BZP-LNC+ and TMZ showed synergistic effect in vitro and reduced the in vivo glioma growth by approximately 81%. Conclusion: The present study provides proof-of-principle insights for the combination of these drugs for GBM treatment.
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Glioblastoma , Nanocápsulas , Encéfalo , Linhagem Celular Tumoral , Glioblastoma/tratamento farmacológico , Humanos , Nanocápsulas/uso terapêutico , Oxazepinas , PurinasRESUMO
Glioblastoma (GB) is a histological and genetically heterogeneous brain tumor that is highly proliferative and vascularized. The prognosis is poor with currently available treatment. In this study, we evaluated the cytotoxicity and antiangiogenic activity of doxorubicin-loaded-chitosan-coated-arginylglycylaspartic acid-functionalized-poly(ε-caprolactone)-alpha bisabolol-LNC (AB-DOX-LNC-L-C-RGD). The nanoformulation was prepared by self-assembling followed by interfacial reactions, physicochemically characterized and evaluated in vitro against GB cell lines (U87MG and U138MG) and in vivo using the chicken chorioallantoic membrane assay (CAM). Spherical shape nanocapsules had a hydrodynamic mean diameter of 138 nm, zeta potential of +13.4 mV, doxorubicin encapsulation of 65%, and RGD conjugation of 92%. After 24 h of treatment (U87MG and U138MG), the median inhibition concentrations (IC50) were 520 and 490 nmol L-1 doxorubicin-equivalent concentrations, respectively. The treatment induced antiproliferative activity with S-phase cell-cycle arrest and apoptosis in the GB cells. Furthermore, after 48 h of exposure, evaluation of antiangiogenic activity (CAM) showed that the relative vessel growth following treatment with the nanocapsules was 5.4 times lower than that with the control treatment. The results support the therapeutic potential of the nanoformulation against GB and, thereby, pave the way for future preclinical studies.
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Aim: To evaluate the antitumor efficacy of bevacizumab-functionalized nanocapsules in a rat glioblastoma model after the pretreatment with nanocapsules functionalized with a peptide-specific to the epidermal growth factor receptor variant III. Materials & methods: Nanocapsules were prepared, physicochemical characterized and intranasally administered to rats. Parameters such as tumor size, histopathological characteristics and infiltration of CD8+ T lymphocytes were evaluated. Results: The strategy of treatment resulted in a reduction of 87% in the tumor size compared with the control group and a higher infiltration of CD8+ T lymphocytes in tumoral tissue. Conclusion: The block of two different molecular targets using nose-to-brain delivery represents a new and promising approach against glioblastoma.
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Neoplasias Encefálicas , Glioblastoma , Nanocápsulas , Animais , Bevacizumab/uso terapêutico , Encéfalo , Neoplasias Encefálicas/tratamento farmacológico , Linhagem Celular Tumoral , Receptores ErbB , Glioblastoma/tratamento farmacológico , Nanocápsulas/uso terapêutico , Peptídeos/uso terapêutico , RatosRESUMO
Glioblastomas are among the most aggressive tumors, and with low survival rates. They are characterized by the ability to create a highly immunosuppressive tumor microenvironment. Exosomes, small extracellular vesicles (EVs), mediate intercellular communication in the tumor microenvironment by transporting various biomolecules (RNA, DNA, proteins, and lipids), therefore playing a prominent role in tumor proliferation, differentiation, metastasis, and resistance to chemotherapy or radiation. Exosomes are found in all body fluids and can cross the blood-brain barrier due to their nanoscale size. Recent studies have highlighted the multiple influences of tumor-derived exosomes on immune cells. Owing to their structural and functional properties, exosomes can be an important instrument for gaining a better molecular understanding of tumors. Furthermore, they qualify not only as diagnostic and prognostic markers, but also as tools in therapies specifically targeting aggressive tumor cells, like glioblastomas.