Novel rivaroxaban-loaded poly(lactic-co-glycolic acid)/poloxamer nanoparticles: preparation, physicochemical characterization, in vitro evaluation of time-dependent anticoagulant activity and toxicological profile.

Rivaroxaban (RXB), an oral direct factor Xa inhibitor, presents innovative therapeutic profile. However, RXB has shown adverse effects, mainly due to pharmacokinetic limitations, highlighting the importance of developing more effective formulations. Therefore, this work aims at the preparation, physicochemical characterization and in vitro evaluation of time-dependent anticoagulant activity and toxicology profile of RXB-loaded poly(lactic-co-glycolic acid) (PLGA)/poloxamer nanoparticles (RXBNps). RXBNp were produced by nanoprecipitation method and physicochemical characteristics were evaluated. In vitro analysis of time-dependent anticoagulant activity was performed by prothrombin time test and toxicological profile was assessed by hemolysis and MTT reduction assays. The developed RXBNp present spherical morphology with average diameter of 205.5 ± 16.95 nm (PdI 0.096 ± 0.04), negative zeta potential (-26.28 ± 0.77 mV), entrapment efficiency of 91.35 ± 2.40%, yield of 41.81 ± 1.68% and 3.72 ± 0.07% of drug loading. Drug release was characterized by an initial fast release followed by a sustained release with 28.34 ± 2.82% of RXB available in 72 h. RXBNp showed an expressive time-dependent anticoagulant activity in human and rat blood plasma and non-toxic profile. Based on the results presented, it is possible to consider that RXBNp may be able to assist in the development of promising new therapies for treatment of thrombotic disorders.

Oral pentamidine-loaded poly(d,l-lactic-co-glycolic) acid nanoparticles: an alternative approach for leishmaniasis treatment.

Leishmaniasis is a group of diseases caused by a protozoa parasite from one of over 20 Leishmania species. Depending on the tissues infected, these diseases are classified as cutaneous, mucocutaneous and visceral leishmaniasis. For the treatment of leishmaniasis refractory to antimony-based drugs, pentamidine (PTM) is a molecule of great interest. However, PTM displays poor bioavailability through oral routes due to its two strongly basic amidine moieties, which restricts its administration by a parenteral route and limits its clinical use. Among various approaches, nanotechnology-based drug delivery systems (nano-DDS) have potential to overcome the challenges associated with PTM oral administration. Here, we present the development of PTM-loaded PLGA nanoparticles (NPs) with a focus on the characterization of their physicochemical properties and potential application as an oral treatment of leishmaniasis. NPs were prepared by a double emulsion methodology. The physicochemical properties were characterized through the mean particle size, polydispersity index (PdI), zeta potential, entrapment efficiency, yield process, drug loading, morphology, in vitro drug release and in vivo pharmacological activity. The PTM-loaded PLGA NPs presented with a size of 263 ± 5 nm (PdI = 0.17 ± 0.02), an almost neutral charge (-3.2 ± 0.8 mV) and an efficiency for PTM entrapment of 91.5%. The release profile, based on PTM dissolution, could be best described by a zero-order model, followed by a drug diffusion profile that fit to the Higuchi model. In addition, in vivo assay showed the efficacy of orally given PTM-loaded PLGA NPs (0.4 mg kg-1) in infected BALB/c mice, with significant reduction of organ weight and parasite load in spleen (p-value < 0.05). This work successfully reported the oral use of PTM-loaded NPs, with a high potential for the treatment of visceral leishmaniasis, opening a new perspective to utilization of this drug in clinical practice.

A promising oral fucoidan-based antithrombotic nanosystem: Development, activity and safety.

Fucoidan-loaded nanoparticles emerge as great candidates to oral anticoagulant therapy, due to increasing of bioavailability and circulation time of this natural anticoagulant. Crosslink between chitosan chains are performed using glutaraldehyde to confer higher gastric pH resistance to nanoparticle matrices. In this work, chitosan-fucoidan nanoparticles, without (NpCF) and with glutaraldehyde crosslink (NpCF 1% and NpCF 2%), were prepared to evaluate their anticoagulant, antithrombotic and hemorrhagic profile. Nanoparticles were characterized by average diameter, polydispersity index, zeta potential, Fourier transform infrared spectroscopy and fucoidan in vitro release. Anticoagulant and antithrombotic activities were determined by in vitro and in vivo models, respectively. Hemorrhagic profile was in vivo evaluated by tail bleeding assay. Preparations showed nanometric and homogeneous average diameters. Zeta potentials of NpCF and NpCF 1% were stable over gastrointestinal pH range, which was confirmed by low fucoidan release in gastric and enteric media. In pH 7.4, NpCF and NpCF 1% demonstrated fucoidan release of 65.5% and 60.6%, respectively, within the first 24 hours. In comparison to fucoidan, NpCF and NpCF 1% showed increased in vitro anticoagulant activity. A significant difference on oral antithrombotic profile of NpCF 1% was found in comparison to fucoidan. Bleeding profile of NpCF and NpCF 1% showed no differences to control group, indicating the safety of these systems. Surprisingly, oral antithrombotic profile of commercially available fucoidan, from Fucus vesiculosus, has not been previously determined, which reveals new possibilities. In this work, significant advances were observed in anticoagulant and antithrombotic profiles of fucoidan through the preparation of NpCF 1%.

Antileishmanial Thioureas: Synthesis, Biological Activity and in Silico Evaluations of New Promising Derivatives.

Leishmaniasis is a neglected tropical disease caused by protozoan parasites belonging to the genus Leishmania. Currently, the drugs available for treatment of this disease present high toxicity, along with development of parasite resistance. In order to overcome these problems, efforts have been made to search for new and more effective leishmanicidal drugs. The aim of this study was to synthesize and investigate the leishmanicidal effect of N,N'-disubstituted thioureas against Leishmania amazonensis, with evaluation of their in silico pharmacokinetics and toxicity profiles. Our results showed that different thioureas could be obtained in high to moderate yields using simple reaction conditions. Nine thiourea derivatives (3e, 3i, 3k, 3l, 3p, 3q, 3v, 3x and 3z) were active against parasite promastigotes (IC50 21.48-189.10 µM), with low cytotoxicity on mice peritoneal macrophages (CC50>200 µM), except for thiourea 3e (CC50=49.22 µM). After that, the most promising thioureas (3k, 3l, 3p, 3q and 3v) showed IC50 ranging from 70 to 150 µM against L. amazonensis amastigotes in infected macrophages. Except for thiourea 3p, the leishmanicidal activity of the derivatives were independent of nitric oxide (NO) production. Thioureas 3q and 3v affected promastigotes cell cycle without disturbing the mitochondrial membrane potential. Furthermore, our derivatives showed satisfactory theoretical absorption, distribution, metabolism, excretion, toxicity (ADMET) properties. These data indicate that thiourea derivatives are good candidates as leading compounds for the development of new leishmanicidal drugs.

Development and Characterization of Nisin Nanoparticles as Potential Alternative for the Recurrent Vaginal Candidiasis Treatment.

The aim of this work was the development and characterization of nisin-loaded nanoparticles and the evaluation of its potential antifungal activity. Candidiasis is a fungal infection caused by Candida sp. considered as one of the major public health problem currently. The discovery of antifungal agents that present a reduced or null resistance of Candida sp. and the development of more efficient drug release mechanisms are necessary for the improvement of candidiasis treatment. Nisin, a bacteriocin commercially available for more than 50 years, exhibits antibacterial action in food products with potential antifungal activity. Among several alternatives used to modulate antifungal activity of bacteriocins, polymeric nanoparticles have received great attention due to an effective drug release control and reduction of therapeutic dose, besides the minimization of adverse effects by the preferential accumulation in specific tissues. The nisin nanoparticles were prepared by double emulsification and solvent evaporation methods. Nanoparticles were characterized by dynamic light scattering, zeta potential, Fourier transform infrared, X-ray diffraction, differential scanning calorimetry, and scanning electron microscopy. Antifungal activity was accessed by pour plate method and cell counting using Candida albicans strains. The in vitro release profile and in vitro permeation studies were performed using dialysis bag method and pig vaginal mucosa in Franz diffusion cell, respectively. The results revealed nisin nanoparticles (300 nm) with spherical shape and high loading efficiency (93.88 ± 3.26%). In vitro test results suggest a promising application of these nanosystems as a prophylactic agent in recurrent vulvovaginal candidiasis and other gynecological diseases.

Synthesis and antiplatelet activity of antithrombotic thiourea compounds: biological and structure-activity relationship studies.

The incidence of hematological disorders has increased steadily in Western countries despite the advances in drug development. The high expression of the multi-resistance protein 4 in patients with transitory aspirin resistance, points to the importance of finding new molecules, including those that are not affected by these proteins. In this work, we describe the synthesis and biological evaluation of a series of N,N'-disubstituted thioureas derivatives using in vitro and in silico approaches. New designed compounds inhibit the arachidonic acid pathway in human platelets. The most active thioureas (compounds 3d, 3i, 3m and 3p) displayed IC50 values ranging from 29 to 84 µM with direct influence over in vitro PGE2 and TXA2 formation. In silico evaluation of these compounds suggests that direct blockage of the tyrosyl-radical at the COX-1 active site is achieved by strong hydrophobic contacts as well as electrostatic interactions. A low toxicity profile of this series was observed through hemolytic, genotoxic and mutagenic assays. The most active thioureas were able to reduce both PGE2 and TXB2 production in human platelets, suggesting a direct inhibition of COX-1. These results reinforce their promising profile as lead antiplatelet agents for further in vivo experimental investigations.

Structural model of haptoglobin and its complex with the anticoagulant ecotin variants: structure-activity relationship study and analysis of interactions.

Recently the literature described the binding of Haptoglobin (HP) with ecotin, a fold-specific serine-proteases inhibitor with an anticoagulant profile and produced by Escherichia coli. In this work, we used some in silico and in vitro techniques to evaluate HP 3D-fold and its interaction with wild-type ecotin and two variants. Our data showed HP models conserved trypsin fold, in agreement to the in vitro immunological recognition of HP by trypsin antibodies. The analysis of the three ecotin-HP complexes using the mutants RR and TSRR/R besides the wild type revealed several hydrogen bonds between HP and ecotin secondary site. These data are in agreement with the in vitro PAGE assays that showed the HP-RR complex in native gel conditions. Interestingly, the ternary complex interactions varied depending on the inhibitor structure and site-directed mutation. The interaction of HP with TSRR/R involved new residues compared to wild type, which infers a binding energy increase caused by the mutation.

Human thromboxane synthase: comparative modeling and docking evaluation with the competitive inhibitors Dazoxiben and Ozagrel.

Thromboxane synthase (TXAS) is a P450 epoxygenase that synthesizes thromboxane A2 (TXA2), a potent mediator of platelet aggregation, vasoconstriction and bronchoconstriction. This enzyme plays an important role in several human diseases, including myocardial infarction, stroke, septic shock, asthma and cancer. Despite of the increasing interest on developing TXAS inhibitors, the structure and activity of TXAS are still not totally elucidated. In this study, we used a comparative molecular modeling approach to construct a reliable model of TXAS and analyze its interactions with Dazoxiben and Ozagrel, two competitive inhibitors. Our results were compatible with experimental published data, showing feasible cation-π interaction between the iron atom of the heme group of TXAS and the basic nitrogen atom of the imidazolyl group of those inhibitors. In the absence of the experimental structure of thromboxane synthase, this freely available model may be useful for designing new antiplatelet drugs for diseases related with TXA2.

Platelets: still a therapeutical target for haemostatic disorders.

Platelets are cytoplasmatic fragments from bone marrow megakaryocytes present in blood. In this work, we review the basis of platelet mechanisms, their participation in syndromes and in arterial thrombosis, and their potential as a target for designing new antithrombotic agents. The option of new biotechnological sources is also explored.

A further pocket or conformational plasticity by mapping COX-1 catalytic site through modified-mofezolac structure-inhibitory activity relationships and their antiplatelet behavior.

Cyclooxygenase enzymes have distinct roles in cardiovascular, neurological, and neurodegenerative disease. They are differently expressed in different type of cancers. Specific and selective COXs inhibitors are needed to be used alone or in combo-therapies. Fully understand the differences at the catalytic site of the two cyclooxygenase (COX) isoforms is still opened to investigation. Thus, two series of novel compounds were designed and synthesized in fair to good yields using the highly selective COX-1 inhibitor mofezolac as the lead compound to explore a COX-1 zone formed by the polar residues Q192, S353, H90 and Y355, as well as hydrophobic amino acids I523, F518 and L352. According to the structure of the COX-1:mofezolac complex, hydrophobic amino acids appear to have free volume eventually accessible to the more sterically hindering groups than the methoxy linked to the phenyl groups of mofezolac, in particular the methoxyphenyl at C4-mofezolac isoxazole. Mofezolac bears two methoxyphenyl groups linked to C3 and C4 of the isoxazole core ring. Thus, in the novel compounds, one or both methoxy groups were replaced by the higher homologous ethoxy, normal and isopropyl, normal and tertiary butyl, and phenyl and benzyl. Furthermore, a major difference between the two sets of compounds is the presence of either a methyl or acetic moiety at the C5 of the isoxazole. Among the C5-methyl series, 12 (direct precursor of mofezolac) (COX-1 IC50 = 0.076 µM and COX-2 IC50 = 0.35 µM) and 15a (ethoxy replacing the two methoxy groups in 12; COX-1 IC50 = 0.23 µM and COX-2 IC50 > 50 µM) were still active and with a Selectivity Index (SI = COX-2 IC50/COX-1 IC50) = 5 and 217, respectively. The other symmetrically substituted alkoxyphenyl moietis were inactive at 50 µM final concentration. Among the asymmetrically substituted, only the 16a (methoxyphenyl on C3-isoxazole and ethoxyphenyl on C4-isoxazole) and 16b (methoxyphenyl on C3-isoxazole and n-propoxyphenyl on C4-isoxazole) were active with SI = 1087 and 38, respectively. Among the set of compounds with the acetic moiety, structurally more similar to mofezolac (SI = 6329), SI ranged between 1.4 and 943. It is noteworthy that 17b (n-propoxyphenyl on both C3- and C4-isoxazole) were found to be a COX-2 slightly selective inhibitor with SI = 0.072 (COX-1 IC50 > 50 µM and COX-2 IC50 = 3.6 µM). Platelet aggregation induced by arachidonic acid (AA) can be in vitro suppressed by the synthesized compounds, without affecting of the secondary hemostasia, confirming the biological effect provided by the selective inhibition of COX-1. A positive profile of hemocompatibility in relation to erythrocyte and platelet toxicity was observed. Additionally, these compounds exhibited a positive profile of hemocompatibility and reduced cytotoxicity. Quantitative structure activity relationship (QSAR) models and molecular modelling (Ligand and Structure based virtual screening procedures) provide key information on the physicochemical and pharmacokinetic properties of the COX-1 inhibitors as well as new insights into the mechanisms of inhibition that will be used to guide the development of more effective and selective compounds. X-ray analysis was used to confirm the chemical structure of 14 (MSA17).

N, N'-disubstituted Ureas as Novel Antiplatelet Agents: Synthesis, Pharmacological Evaluation and In Silico Studies.

INTRODUCTION: Thrombotic disorders are among the leading causes of morbidity and mortality worldwide. Drugs used in the prevention and treatment of atherothrombosis have pharmacokinetic limitations and adverse effects such as hemorrhagic conditions, highlighting the importance of developing more effective antiplatelet agents. ethod: In this work, we synthesized N,N'-disubstituted ureas 3a-3j and evaluated their antiplatelet profiles through in vitro, ex vivo, and in silico studies. The synthesized derivatives exhibited a selective inhibitory profile against platelet aggregation induced by arachidonic acid (AA) in vitro, without significantly affecting other aspects of primary hemostasis and blood coagulation. The compounds that showed inhibition greater than 85% were submitted to the analysis of their potency by calculating the concentration required to inhibit 50% of platelet aggregation induced by AA (IC50). Urea derivative 3a was the most potent with IC50 of 1.45 µM. Interestingly, this derivative inhibited more than 90% of platelet aggregation induced by AA ex vivo, with a similar effect to acetylsalicylic acid. In the hemolysis assay, most of the urea derivatives presented values below 10% suggesting good hemocompatibility. Additionally, the compounds tested at 100 µM also showed no cytotoxic effects in HepG2 and Vero cells. RESULT: The in silico results suggested that compound 3a may bind to the key residue of COX-1 similar to AA and known COX-1 inhibitors, and the results are also in agreement with our SAR, which suggests that the inhibition of this enzyme is the most likely mechanism of antiplatelet activity. CONCLUSION: Therefore, these results demonstrated that N,N'-disubstituted ureas are promising candidates for the development of novel antiplatelet agents.

Enzymatic synthesis of ascorbyl oleate and evaluation of biological activities.

Compounds that reduce or neutralize free radicals have been evaluated for use as nutraceutical or antioxidant additives in processed foods. This study aimed to enzymatically produce ascorbyl oleate and assess its biological properties. The synthesis was performed under previously maximized conditions (L-ascorbic acid/oleic acid 1:9 molar ratio, 70 °C, 1 h reaction). Immobilized commercial lipase from Candida antarctica (NS 88011) was used as biocatalyst. The reaction product was isolated, and its structure was confirmed by High-Performance Liquid Chromatography and Nuclear Magnetic Resonance. Ascorbyl oleate showed antioxidant and antimicrobial activity, besides no toxicity, did not influencing blood coagulation and also not presenting hemolytic profile. Better storage stability was achieved under refrigerated conditions, and the oxidative stability demonstrated free radicals fighting efficiency, increasing olive oil's shelf life. In vitro gastrointestinal simulation showed that ascorbyl oleate maintained antioxidant potential up to the duodenum stage during the digestive process. Therefore, the synthesized natural compound presented a high potential to be applied in the food and pharmaceutical industries.

Preparation and evaluation of a new nano pharmaceutical excipients and drug delivery system based in polyvinylpyrrolidone and silicate.

PURPOSE: This work describes the preparation of new nanocomposites based on lamellar silicates (AAM-alkyl ammonium montmorillonite) obtained by the intercalation of PVP K30 and glyceril monostearate. METHODS: By XRD, TGA and DSC analysis the AAM was characterized and its compactation characteristics, functionality and toxicity were also tested. The AAM/PVP K-30 and AAM/GME nanocomposite obtained were evaluated to identify the interlamellar spacing values by XRD diffratograms. Tablets were prepared using methyldopa and theophylline as model drugs and the dissolution tests were carried out in simulated gastric fluid and simulated enteric fluid. RESULTS: AAM showed a good compactability and compressibility characteristics for tablets preparation. The intercalation yields (approximately 25%) of the nanocomposites were efficient. The AAM/PVP K-30 nanocomposites were successfully tested as dissolution enhancers and sustained release matrixes. CONCLUSIONS: The results also suggested the promising use of AAM (viscogel B8) and the new nanocomposite prepared by clay/PVP K-30 intercalation as a new matrix for sustained release and the feasibility of using these new nanocomposites as dissolution enhancer.

Development of rivaroxaban microemulsion-based hydrogel for transdermal treatment and prevention of venous thromboembolism.

We have developed a microemulsion (ME)-based hydrogel, containing propylene glycol, Azone®, Labrasol®, isobutanol and water (20:3:18:3:56), for the transdermal delivery of rivaroxaban (RVX). Formulation ME-1:RVX, which was loaded with 0.3 mg/g of RVX, presented as a clear, homogenous fluid with a droplet size of 82.01 ± 6.32 nm and a PdI of 0.207 ± 0.01. To provide gelation properties, 20 % (w/w) of Pluronic® F-127 was added to ME-1:RVX to generate formulation PME-1a. An added benefit was an increased capacity for RVX to 0.4 mg/g (formulation PME-1b). PME-1b displayed spherical droplets with a nanoscale diameter as observed by Transmission Electron Microscopy. The release of RVX from PME-1b was 20.71 ± 0.76 µg/cm2 with a permeation through pig epidermis of 18.32 ± 8.87 µg/cm2 as measured in a Franz Cell for 24 h. PME-1b presented a pseudoplastic behavior, pH value compatible with the skin and good stability over 60 days at room and elevated temperatures. The prothrombin time was assessed for each concentration of RVX obtained in the permeation assay and each demonstrated a relevant anticoagulant activity. PME-1b also presented no cytotoxicity against HaCaT cells. Utilizing GastroPlus® software, an in silico analysis was performed to simulate the delivery of PME-1b through a transdermal system that suggested a minimum dose of RVX for the treatment and prevention of venous thromboembolism could be achieved with an 8 h administration regimen. These results suggest that PME-1b is a promising transdermal formulation for the effective delivery of RVX that could be a viable alternative for the treatment and prevention of venous thromboembolism.

An attempt to chemically state the cross-talk between monomers of COX homodimers by double/hybrid inhibitors mofezolac-spacer-mofezolac and mofezolac-spacer-arachidonic acid.

Cardiovascular diseases (CVDs) account for over 17 million death globally each year, including arterial thrombosis. Platelets are key components in the pathogenesis of this disease and modulating their activity is an effective strategy to treat such thrombotic events. Cyclooxygenase-1 (COX-1) isoenzyme is involved in platelet activation and is the main target of non-steroidal anti-inflammatory drugs (NSAIDs) and new selective inhibitor research. Inhibitors of general formula mofezolac-spacer-mofezolac (mof-spacer-mof) and mofezolac-spacer-arachidonic acid (mof-spacer-AA) were projected to investigate the possible cross-talk between the two monomers (Eallo and Ecat) forming the COX-1 homodimer. Mofezolac was chosen as either one or two moieties of these molecules being the known most potent and selective COX-1 inhibitor and administrated to humans as Disopain™, then arachidonic acid (AA) was used to develop molecules bearing, in the same compound, in addition to the inhibitor moiety (mofezolac) also the natural COX substrate. Depending on the nature of the spacer, COX-1 and COX-2 activity was differently inhibited by mof-spacer-mof set with a preferential COX-1 inhibition. The highest COX-1 selectivity was exhibited by the compound in which the spacer was the benzidine [N,N'-(biphenyl-4,4'-di-yl)bis (2-[3,4-bis(4-methoxyphenyl)isoxazol-5-yl]acetamide) (15): COX-1 IC50 = 0.08 µM, COX-2 IC50 > 50 µM, Selectivity Index (SI) > 625]. In the case of mof-spacer-AA set, the COX inhibitory potency and also the isoform preference changed. (5Z, 8Z, 11Z, 14Z)-N-(4-{2-[3,4-Bis(4-methoxyphenyl)isoxazol-5-yl]acetamido}butyl)icosa-5,8,11,14-tetraenamide (19) and (5Z, 8Z, 11Z, 14Z)-N-(4'-{2-[3,4-bis(4-methoxyphenyl)isoxazol-5-yl]acetamido}-[1,1'-biphenyl]-4-yl)icosa-5,8,11,14-tetraenamide (21), in which the spacer is the 1,2-diaminobutane or benzidine, respectively, selectively inhibited the COX-2, whereas when the spacer is the 1,4-phenylendiamine [(5Z, 8Z, 11Z, 14Z)-N-(4-{2-[3,4-bis(4-methoxyphenyl)isoxazol-5-yl]acetamido}phenyl)icosa-5,8,11,14-tetraenamide) (20) the COX preference is COX-1 (COX-1 IC50 = 0.05 µM, COX-2 IC50 > 50 µM, with a COX-1 selectivity > 1000). Molecular modelling by using FLAP algorithm shows fundamental interactions of the novel compounds at the entry channel of COX and inside its catalytic cavity. The effect of these mof-spacer-mof and mof-spacer-AA in inhibiting in vitro free arachidonic acid-induced platelet aggregation was also determined. A positive profile of hemocompatibility in relation to their influence on the blood coagulation cascade and erythrocyte toxicity was observed. Cytotoxicity and genotoxicity safety were also found for these two novel sets of compounds.

Translational impact of novel widely pharmacological characterized mofezolac-derived COX-1 inhibitors combined with bortezomib on human multiple myeloma cell lines viability.

A set of novel diarylisoxazoles has been projected using mofezolac (1) as a lead compound to investigate structure-inhibitory activity relationships of new compounds and the cyclooxygenases (COXs) catalytic activity. Mofezolac was chosen because is the most potent and selective reversible COX-1 inhibitor [COX-1 IC50 = 0.0079 µM and COX-2 IC50 > 50 µM, with a selectivity index (SI) in favor of COX-1 higher than 6300]. Seventeen new compounds were synthesized in fair to good yields and evaluated for their COXs inhibitory activity and selectivity. SIs ranged between 1 and higher than 1190.3,4-Bis(4-methoxyphenyl)-5-vinylisoxazole (22) has the highest SI with COX-1 IC50 = 0.042 µM and COX-2 IC50 > 50 µM. 1 and 22 were superior to aspirin in inhibiting platelet aggregation (IC50 = 0.45, 0.63 and 1.11 µM, respectively) in human platelet rich plasma (hPRP) assay. They did not induce blood coagulation and hemolysis, and are neither genotoxic nor mutagen. 1 and 22 slightly increase bortezomib cytotoxic effect on multiple myeloma (MM) cell lines (NCI-H929 and RPMI-8226) and affects MM cell cycle and apoptosis when co-administered with the proteasome inhibitor bortezomib, a drug clinically used to treat plasma cell neoplasms including MM. In addition, structure-based binding mode of 1 and 22, through Fingerprints for Ligands and Proteins (FLAG) calculation, allowed to explain the one order of magnitude difference between COX-1 IC50 values of the two compounds. Specifically, the higher inhibitory potency seems due to the formation of a H-bond between COX-1 S530 and the carboxyl, present in 1 and absent in 22.

Synthesis and evaluation of the cytotoxic activity of Furanaphthoquinones tethered to 1H-1,2,3-triazoles in Caco-2, Calu-3, MDA-MB231 cells.

Naphthoquinones and 1,2,3-triazoles are structural pharmacophore that is known to impart several cancer cells. This work shows a synthetic methodology to obtain hybrid molecules involving naphthoquinone and triazol scaffold as multiple ligands. A simple and efficient synthetic route was used to prepare a series of sixteen compounds being eight 2-(1-aryl-1H-1,2,3-triazol-4-yl)-2,3-dihydronaphtho[1,2 b]furan-4,5-diones and eight 2-(1-aryl-1H-1,2,3-triazol-4-yl)-2,3-dihydronaphtho[2,3-b]furan-4,9-diones. These compounds were tested in MDA-MB231, Caco-2 and Calu-3 human cancer cells, and among them 7a was the most selective compound on Caco-2 cells, the most sensitized cell line in this study. In silico study suggest that the blockage of topoisomerase I and IIα may be one of the mechanisms of action responsible for the cytotoxic effect of 7a in Caco-2 cells.

Targeted Nanosystems to Prostate Cancer.

Prostate cancer remains an increasingly common malignancy worldwide. Many advances in drug development have been achieved for the conventional treatments; however, chemotherapeutic agents are distributed nonspecifically in the body where they affect both prostate cancer and healthy cells. Limited dose achievable within the prostate tumor and suboptimal treatment due to excessive toxicities reveal the importance of the development of more specific mechanisms and ways of drug targeting to prostate tumor. In this context, nanotechnology, molecular biology and biochemistry have been applied in the pharmaceutical area for development of new targeted drug delivery nanosystems in order to improve its pharmacokinetic profile, raise the effectiveness of treatment; reduce side effects due to the preferential accumulation in prostate cancer cells, causing low concentrations in healthy tissues; and/or increase the drug chemical stability for improving the prostate cancer therapeutic. Thus, in this review, we will discuss the molecular and biochemical basis of prostate cancer as well as the advantages and disadvantages of conventional clinical treatments, different types and basic characteristics of nanosystems; how these systems can be targeted to prostate cancer, show successful patent examples of prostate cancer targeted nanosystems and present perspectives for the next 10-20 years in this area.

In vitro and in vivo analysis of the antithrombotic and toxicological profile of new antiplatelets N-acylhydrazone derivatives and development of nanosystems: determination of novel NAH derivatives antiplatelet and nanotechnological approach.

BACKGROUND: Cardiovascular diseases are the most frequent cause of morbidity and mortality worldwide. Among the most important cardiovascular diseases are atherothrombosis and venous thromboembolism that present platelet aggregation as a key event. Currently, the commercial antiplatelet agents display several undesirable effects, which prompt the search for new compounds with better therapeutic index, more efficient body distribution and mechanism. METHODS: In this work we characterized in vivo and in vitro the antithrombotic and toxicological profiles of novel antiplatelet N-substituted-phenylamino-5-methyl-1H-1,2,3-triazole-4-carbohydrazides derivatives also comparing them with aspirin. In addition we also analyzed the stability of the more active compound after encapsulation in PLGA or PCL nanoparticles and the release profile of these new nanosystems. RESULTS: The biological results revealed not only the selective effect against arachidonic acid-induced platelet aggregation mainly for compounds 2c, 2e and 2h but also their in vivo active profile on thromboembolism pulmonary animal model with better survival rates (e.g. 82%) than aspirin (33%). The overall toxicological profile was determined by in vitro (MTT reduction tests, neutral red uptake in kidney VERO cells and hemolysis assays) and in vivo (pulmonary embolism) assays that pointed 2c as the most promising derivative with potential as a lead compound. By using the nanoprecipitation technique 2c was loaded into PLGA and PCL nanoparticles showing controlled release profile over 21days according to our drug release tests. CONCLUSION: According to our results compound 2c is the most interesting derivative for further studies as it showed the best activity and toxicological profile also allowing the nanoencapsulation process. Thus 2c may assist in determining a new potential therapy with favorable pharmacokinetics for treatment of thrombotic disorders.

Sulphonamide and sulphonyl-hydrazone cyclic imide derivatives: antinociceptive activity, molecular modeling and in silico ADMET screening.

In this paper, we describe the antinociceptive activity, molecular modeling and in silico ADMET screening of a series of sulphonyl-hydrazone and sulphonamide imidobenzene derivatives. Among these compounds, the sulphonyl-hydrazones 9 and 11 showed the most potent analgesic activity (ID(50) = 5.1 and 6.8 µmol/kg, respectively). Interestingly, all derivatives evaluated in this study have a better analgesic profile than the control drugs, acetyl salicylic acid and acetaminophen. Derivative 9 was the most promising compound; with a level of activity that was 24 times higher than the control drugs. Our SAR study showed a relationship among the distribution of the frontier orbital HOMO coefficients, HOMO-LUMO energy gap of these molecules and their reactivity. The best analgesic compounds (including 6, 9, 10, 11 and 12) fulfilled the Lipinski "rule-of-five", which is theoretically important for good drug absorption and permeation.