Phytochemical study of Magonia pubescens A. St.-Hil. and cytotoxicity of branches aqueous extract on breast cancer and leukemia cells.

Magonia pubescens is a natural species from the Brazilian cerrado biome. Its fruits and seeds are used in the treatment of seborrheic dermatitis, a common inflammatory skin disease. In this work, the known compounds lapachol, stigmasterol, maniladiol and scopoletin were isolated from hexane and dichloromethane extracts of M. pubescens branches. The aqueous extract of this material was fractioned through a liquid-liquid partition and the obtained fractions were analyzed by UHPLC-MS/MS. The results obtained were compared with data from three databases, leading to the putative identification of 51 compounds from different classes, including flavonoids, saponins and triterpenes. The cytotoxicity of aqueous fractions was assayed against breast cancer (MDA-MB-231) and leukemia (THP-1 and K562) cells. The best activity was observed for fraction AE3 against MDA-MB-231 cells (IC50 30.72 µg.mL-1).

Metformin inhibits paclitaxel-induced mechanical allodynia by activating opioidergic pathways and reducing cytokines production in the dorsal root ganglia and thalamus.

It has been shown that AMP-activated protein kinase (AMPK) is involved in the nociceptive processing. This observation has prompted us to investigate the effects of the AMPK activator metformin on the paclitaxel-induced mechanical allodynia, a well-established model of neuropathic pain. Mechanical allodynia was induced by four intraperitoneal (i.p) injections of paclitaxel (2 mg/kg.day) in mice. Metformin was administered per os (p.o.). Naltrexoneandglibenclamide were used to investigate mechanisms mediating metformin activity. Concentrations of cytokines in the dorsal root ganglia (DRG) and thalamus were determined. After a single p.o. administration, the two highest doses of metformin (500 and 1000 mg/kg) attenuated the mechanical allodynia. This response was attenuated by all doses of metformin (250, 500 and 1000 mg/kg) when two administrations, 2 h apart, were carried out. Naltrexone (5 and 10 mg/kg, i.p.), but not glibenclamide (20 and 40 mg/kg, p.o.), attenuated metformin activity. Concentrations of tumor necrosis factor (TNF)-α, interleukin (IL)-1ß and CXCL-1 in the DRG were increased after administration of paclitaxel. Metformin (1000 mg/kg) reduced concentrations of TNF-α, IL-1ß and CXCL-1 in the DRG. Concentration of IL-6, but not TNF-α, in the thalamus was increased after administration of paclitaxel. Metformin (1000 mg/kg) reduced concentration of IL-6 in the thalamus. In summary, metformin exhibits activity in the model of neuropathic pain induced by paclitaxel. This activity may be mediated by activation of opioidergic pathways and reduced production of TNF-α, IL-1ß and CXCL-1 in the DRG and IL-6 in the thalamus.

Alpha-tocopheryl succinate and doxorubicin-loaded liposomes improve drug uptake and tumor accumulation in a murine breast tumor model.

Liposomes composed of a rigid bilayer have high plasma stability; however, they can be challenged in efficacy due to complications in releasing the encapsulated drug as well as being internalized by the tumor cell. On the other hand, fusogenic liposomes may fuse with the plasmatic membrane and release encapsulated material directly into the cytoplasm. In a previous study, fusogenic liposomes composed of alpha-tocopheryl succinate (TS) and doxorubicin (DOX) were developed (pHSL-TS-DOX). These stabilized tumor growth and reduced toxicity compared to a commercial formulation. In the present study, we investigated whether cellular uptake or DOX accumulation in the tumor could justify the better performance of the pHSL-TS-DOX formulation. Release, deformability, and DOX plasmatic concentration studies were also carried out. pHSL-TS-DOX showed an adequate release profile and demonstrated characteristics of a deformable formulation. Data from apoptosis, cell cycle, and nuclear morphology studies have shown that the induction of cell death caused by pHSL-TS-DOX occurred more quickly. Higher DOX cellular uptake and tumor accumulation were observed when pHSL-TS-DOX was administered, demonstrating better drug delivery capacity. Therefore, better DOX uptake as well as tumor accumulation explain the great antitumor activity previously demonstrated for this formulation.

miR-197, miR-26a and miR-27a analysis in chronic lymphocytic leukemia.

Aims: Chronic lymphocytic leukemia (CLL) involves the proliferation and increase of B-lymphocytes in the peripheral blood, bone marrow and lymphoid organs. This study evaluated the microRNAs miR-197, miR-26a and miR-27a as potential biomarkers for CLL. Patients & Methods: Eighty-two patients with CLL and 62 control subjects (CT) were investigated for these targets, using quantitative PCR (qPCR). Results: A significant reduction of all microRNAs was observed in CLL compared to the controls (p < 0.001). Significant negative correlations were observed for the clinical staging groups. After adjusting for multiple logistic regression analysis, miR-197 and miR-26a remained as possible independent risk factors related to the CLL. Conclusions: Our data indicated good performance of this microRNAs as potential biomarkers in CLL.

Chemical Composition of Magonia pubescens Essential Oils and Gamma-Radiation Effects on Its Constituents and Cytotoxic Activity in Leukemia and Breast Cancer Model.

Magonia pubescens A. St.-Hil. is a Brazilian species often used in ethnopharmacology for wound and pain healing and seborrhea treatment. For the first time, essential oils (EOs) obtained from M. pubescens inflorescences were studied. The plant materials (Montes Claros, Brazil, 2018) were submitted to different gamma-radiation doses and their chemical compositions were analyzed by GC/MS and GC-FID. The cytotoxic activity of the EOs was evaluated against K562 and MDA-MB-231 cancer cell lines. A total of 30 components were identified, being 24 compounds detected for the first time in M. pubescens. The main obtained components were hotrienol (35.9 %), cis-linalool oxide (17.0 %) and trans-linalool oxide (10.2 %). The chemical composition of the EO was slightly affected by the applied radiation doses. Irradiated and non-irradiated EOs showed cytotoxic activity against both cell lines and the non-irradiated EO sample was the most active against the K562 cell lines (IC50 =22.10±1.98).

Imines and lactones derived from friedelanes and their cytotoxic activity.

Friedelan-3-one (1) and friedelane-3,16-dione (2) isolated from leaves and branches of Maytenus robusta Reissek were subjected to structural modifications via nucleophilic addition to the carbonyl group and Baeyer-Villiger oxidation in order to synthesize potential cytotoxic compounds. The oximes friedelane-3-hydroxyimino (3) and 3-hydroxyiminofriedelan-16-one (4) together with the lactones friedelane-3,4-lactone (5) and 3,4-lactonefriedelan-16-one (6) were characterized by IR and NMR spectroscopic analyses. Compounds 4 and 6 are reported for the first time. Cytotoxic screening via MTT assay in human leukemia cell lines (THP-1 and K562) demonstrated no significant improvement of compounds 3-6 when compared to the starting materials. Only compounds 3 and 5 demonstrated an improvement against K562 cells. However, the same assay on ovarian and breast cancer cell lines (TOV-21G and MDA-MB-231) showed a reduction in the IC50 for compounds 4-6, indicating that ring A modifications may enhance the biological potential.

Paclitaxel-Loaded Folate-Coated pH-Sensitive Liposomes Enhance Cellular Uptake and Antitumor Activity.

Paclitaxel (PTX) is a microtubule-stabilizing agent widely used to treat breast cancer. Nevertheless, the low solubility of the drug and the side effects of commercial formulations available limit its clinical use. In this way, our group recently described the preparation of PTX-loaded folate-coated long-circulating and pH-sensitive liposomes (SpHL-folate-PTX). Therefore, a proof-of-concept study was designed in order to demonstrate the feasibility of SpHL-folate-PTX against breast tumor cell line MDA-MB-231. Cellular uptake of the liposomes and PTX was evaluated. Apoptosis and cell cycle were analyzed by flow cytometry. In vivo antitumor activity was carried out in MDA-MB-231 tumor-bearing BALB/c nude mice. Cellular uptake assay showed a high cell delivery of PTX by SpHL-folate-PTX, which leads to superior cytotoxicity and activation of apoptosis pathways. The SpHL-folate-PTX treatment induces an expressive increase of cells in the G0/G1 phase compared to free PTX and SpHL-PTX (without folate). In vivo studies showed a significant reduction in the tumor growth and a lower uptake of a radiopharmaceutical in the scintigraphic images for the SpHL-folate-PTX group, suggesting its higher efficacy compared with free PTX and SpHL-PTX. Histomorphometric analyses demonstrated an increase in necrosis and inflammation areas in animals treated with SpHL-folate-PTX. A decrease in the proliferative cells and a higher percentage of apoptotic cells were observed by immunohistochemical analyses after the treatment with SpHL-folate-PTX. Therefore, the data confirmed the potential of SpHL-folate-PTX as an alternative antitumor therapy, especially for breast cancer.

Polymorphisms in exons 6 and 7 of the ABO locus and their association with venous thrombosis in young Brazilian patients.

The A1 and B alleles of the ABO blood system have been associated with high levels of both factor VIII and von Willebrand factor and with a predisposition to venous thromboembolism (VTE). In this study, genotypes of the ABO system were determined by PCR-restriction fragment length polymorphism for 148 young VTE Brazilian patients and 233 unrelated control individuals. The blood group O was more frequent among the controls [odds ratio (OR), 0.21; 95% confidence interval (CI), 0.13-0.34; P = 0.000) and groups A and B (OR, 2.24; 95%, CI, 1.46-3.42; P = 0.000 and OR, 2.52; 95% CI, 1.42-4.48; P = 0.002, respectively) among patients. The patients' group was under Hardy-Weinberg equilibrium, whereas the control group was not (P < 0.0051), suggesting that natural selection might be acting in favor of carriers of the O blood group. When the allelic frequencies were compared through multivariate logistic regression analysis for adjustments of covariates, the alleles A1 (OR, 1.69; 95% CI, 1.17-2.45; P = 0.006), A2 (OR, 2.19; 95% CI, 1.24-3.87; P = 0.010), and B (OR, 2.65; 95% CI, 1.64-4.26; P = 0.000) were independently associated with VTE and may represent important risk factors to the development of VTE among young Brazilian patients. Thus, the inclusion of ABO blood group determination may be helpful to discriminate individuals with high risk for VTE allowing target intervention as well as to manage VTE in young patients.

D-Dimer plasma levels in patients with coronary artery disease

We have previously reported that prothrombin fragment 1+2 levels were not associated to the presence or severity of coronary artery disease (CAD) and do not provide further information on subjects with CAD diagnosed by angiography. Thus, in the present study another marker of hypercoagulability was evaluated in the same subjects. This study aimed at determining D-Dimer plasma levels in a group of subjects undergoing coronary angiography to establish a likely relation between this parameter and the severity of CAD. D-Dimer plasma levels were determined in 17 subjects with no coronary atheromatosis (controls), 12 subjects with mild/moderate atheromatosis and 28 subjects with severe atheromatosis. No significant differences were observed among the three groups. Data analysis enables an inference on a tendency towards an increase in fibrinolytic activity in patients with atheromatosis, reflected by the increase in D-Dimer concentrations in the severe atheromatosis group in subjects with CAD diagnosed by coronary angiography.

Em estudo prévio, os níveis plasmáticos do fragmento 1+2 da protrombina não foram associados com a presença ou com a gravidade da doença arterial coronariana (DAC), não trazendo benefício adicional pelo menos em indivíduos com diagnóstico de DAC estabelecido por angiografia. Desta forma, neste estudo outro marcador de hipercoagulabilidade foi avaliado nos mesmos pacientes. O presente estudo teve como objetivo determinar os níveis plasmáticos do dímero D de um grupo de indivíduos submetidos à angiografia coronariana, buscando estabelecer a possível correlação entre este parâmetro e a gravidade da DAC. Os níveis plasmáticos do dímero D foram determinados em amostras de sangue de 17 indivíduos com ausência de ateromatose nas coronárias (controles), 12 indivíduos apresentando ateromatose leve/moderada e 28 indivíduos apresentando ateromatose grave. Não foram encontradas diferenças estatisticamente significativas entre as médias dos três grupos para o parâmetro avaliado. Uma análise dos dados permite inferir sobre uma tendência ao aumento da atividade fibrinolítica nos pacientes com ateromatose, refletida pela elevação da concentração de dímero D no grupo ateromatose grave em indivíduos com diagnóstico de DAC estabelecido por angiografia coronariana.

Lipoproteína(a) e inibição da fibrinólise na doença arterial coronariana / Lipoprotein(a) and fibrinolysis inhibition in coronary artery disease

O envolvimento da lipoproteína(a) nas doenças ateroscleróticas tem sido alvo de muitos estudos que têm demonstrado que esta lipoproteína é um fator de risco independente para a doença arterial coronariana (DAC). Devido à semelhança estrutural com o plasminogênio, a Lp(a) pode competir com os sítios de ligação deste, diminuindo a geração de plasmina e inibindo a fibrinólise. O trombo formado numa placa aterosclerótica rompida dispara a maioria dos eventos cardiovasculares isquêmicos. Como o trombo é dissolvido através do sistema fibrinolítico, surgiu a hipótese de que uma diminuição da atividade fibrinolítica poderia ser um fator de risco para eventos isquêmicos. Entretanto, alguns estudos demonstraram ausência de associação entre Lp(a) e DAC, enquanto alguns atribuem o real valor preditivo à uma sub-população de Lp(a) com alta afinidade por fibrina. Essa hipótese sugere que alguns fenótipos de Lp(a) não estão associados à aterotrombose. Sendo assim, as pesquisas envolvendo Lp(a) e DAC apresentam resultados controversos, altamente dependentes da população estudada. Na população brasileira, a qual apresenta uma heterogeneidade de etnias, são raros os estudos deste tipo. Diante das notáveis controvérsias, espera-se que a presente revisão possa contribuir para suscitar ânimos no sentido da realização de estudos adicionais envolvendo a dosagem da Lp(a) em pacientes com DAC na nossa população. O conhecimento acumulado ao longo do tempo sobre a associação entre Lp(a) e DAC foi obtido através de estudos conduzidos fora do nosso meio. Assim, estudos fundamentados dentro da nossa realidade poderão trazer respostas mais fidedignas e adequadas para a nossa população.

The role of lipoprotein(a) [Lp(a)] in atherogenesis has been the target of many studies that have demonstrated that this lipoprotein is an independent risk factor for coronary artery disease (CAD). Due to the structural likeness with plasminogen, Lp(a) can compete with binding sites, reducing the plasmin generation and inhibiting fibrinolysis. Thrombus formed due to ruptured atherosclerotic plaque trigger most ischemic cardiovascular events. As the thrombus are dissolved by the fibrinolytic system, the hypothesis that a decrease in fibrinolytic activity might be a risk factor for ischemic events has been suggested. However, some studies have not demonstrated any correlation between Lp(a) and CAD while others attribute the real predictive value to a subpopulation of Lp(a) with high affinity for fibrin. This hypothesis suggests that some Lp(a) phenotypes are not associated with atherothrombosis. Furthermore, research on Lp(a) and CAD is controversial and highly dependent on the studied population. In the Brazilian population, which presents heterogeneity of the ethnic groups, studies of this type are rare. Based on the controversies, it is expected that a contribution of the present review will motivate investigators to develop studies involving the measurement of Lp(a) in patients with CAD in the Brazilian population. The accumulated knowledge on the association between Lp(a) and CAD was obtained through foreign studies. Thus, such studies must be performed in our setting in order to bring a better understanding for a more adequate management of CAD.