Publications

Abstract The discovery of new chemical entities endowed with potent and selective acetylcholinesterase (AChE) and/or butyrylcholinesterase (BChE) inhibitory activity is still a relevant subject for Alzheimer's disease therapy. Therefore, a small library of benzoic based amide nitrones (compounds 24 to 42) was synthesized and screened toward cholinesterase enzymes. SAR studies showed that the tert-butyl moiety is the most favourable nitrone pattern. In general, tert-butyl derivatives effectively inhibited AChE, being compound 33 the most potent (IC50 = 8.3 +/- 0.3 mu M; Ki 5.2 mu M). The data pointed to a non-competitive inhibition mechanism of action, which was also observed for the standard donepezil. None of compounds showed BChE inhibitory activity. Molecular modelling studies provided insights into the enzyme-inhibitor interactions and rationalised the experimental data, confirming that the binding mode of nitrones 33 and 38 towards AChE has the most favourable binding free energy. The tert-butylnitrones 33 and 38 were not cytotoxic on different cell lines (SH-SY5Y and HepG2). Moreover, compound 33 was able to prevent t-BHP-induced oxidative stress in SH-SY5Y differentiated cells. Due to its AChE selectivity and promising cytoprotective properties, as well as its appropriate drug-like profile pointing toward blood-brain barrier permeability, compound 33 is proposed as a valid lead for a further optimization step. (C) 2019 The Authors. Published by Elsevier Masson SAS.

Abstract Pure TiO2 and Fe- and Co-doped TiO2 nanoparticles (NPs) as photocatalysts were synthesized using wet chemical methods (sol-gel + precipitation). Their crystalline structure and optical properties were analyzed using X-ray diffraction (XRD), Raman spectroscopy and Fourier-transform infrared (FTIR) spectroscopy, ultraviolet-visible light (UV-Vis) diffuse reflectance spectroscopy (DRS), and photoluminescence (PL) spectroscopy. The photocatalytic activity of the synthesized nanoparticles was evaluated through degradation of carbamazepine (CBZ) under UV-A and visible-light irradiations. The XRD and Raman analyses revealed that all synthesized nanomaterials showed only the anatase phase. The DRS results showed that the absorption edge was blue-shifted for Fe-doped TiO2 NPs. The decrease in charge recombination was evidenced from the PL investigation for both Co-doped and Fe-doped TiO2 nanomaterials. An enhancement in photocatalytic degradation of carbamazepine in aqueous suspension under both UV-A light and visible-light irradiations was observed for Fe-doped Titania NPs by comparison with pure TiO2. These results suggest that the doping cations could suppress the electron/hole recombination. Therefore, the photocatalytic activity of TiO2-based nanomaterials was enhanced.

Abstract The current pharmacological treatments for Parkinson's disease only offer symptomatic relief to the patients and are based on the administration of levodopa and catechol-O-methyltransferase or monoamine oxidase-B inhibitors (IMAO-B). Since the majority of drug candidates fail in pre- and clinical trials, due largely to bioavailability pitfalls, the use of polymeric nanoparticles (NPs) as drug delivery systems has been reported as an interesting tool to increase the stealth capacity of drugs or help drug candidates to surpass biological barriers, among other benefits. Thus, a novel potent, selective, and reversible IMAO-B (chromone C27, IC50 = 670 +/- 130 pM) was encapsulated in poly(caprolactone) (PCL) NPs by a nanoprecipitation process. The resulting C27-loaded PEGylated PCL NPs (similar to 213 nm) showed high stability and no cytotoxic effects in neuronal (SH-SY5Y), epithelial (Caco-2), and endothelial (hCMEC/D3) cells. An accumulation of PEGylated PCL NPs in the cytoplasm of SH-SY5Y and hCMEC/D3 cells was also observed, and their permeation across Caco-2 and hCMEC/D3 cell monolayers, used as in vitro models of the human intestine and blood-brain barrier, respectively, was demonstrated. PEGylated PCL NPs delivered C27 at concentrations higher than the MAO-B IC50 value, which provides evidence of their relevance to solving the drug discovery pitfalls.

Abstract Imidazopyrazinone is a typical scaffold present in marine bioluminescence, in which thermal energy is converted into excitation energy in an enzyme-catalyzed reaction. In fact, the imidazopyrazinone scaffold is a common link among organisms of eight phyla. The characterization of the light emission mechanism is essential for the development of future applications in bioimaging, bioanalysis and biomedicine. Herein, we have studied the chemiluminescent reaction of three commercially-available imidazopyrazinones (Cypridina luciferin, Coelenterazine and Coelenterazine-e) in several aprotic solvents at different pH. We have found that at acidic pH only DMF and DMSO consistently present high light emission, while chemiluminescence in other solvents is negligible. We have attributed this to the inability of most solvents to allow for the deprotonation of the imidazopyrazinone core, thereby preventing the oxygenation step. We have also observed that increasing the pH of the solution leads to the inhibition of chemiluminescence, which we attributed to the deprotonation of the dioxetanone intermediate, as the neutral species is the one associated with efficient chemiexcitation. We have also observed that the pK(a) of dioxetanone increases with the dielectric constant of the medium. Finally, our work indicated that the chemiexcitation yield increases with increasing polarity of the medium, due to a reduced transition dipole moment associated with S-0 -> S-1 transition.

Abstract The human immunodeficiency virus (HIV) uses the brain as reservoir, which turns it as a promising target to fight this pathology. Nanoparticles (NPs) of poly(lactic-co-glycolic) acid (PLGA) are potential carriers of anti-HIV drugs to the brain, since most of these antiretrovirals, as efavirenz (EFV), cannot surpass the blood-brain barrier (BBB). Forasmuch as the conventional production methods lack precise control over the final properties of particles, microfluidics emerged as a prospective alternative. This study aimed at developing EFV-loaded PLGA NPs through a conventional and microfluidic method, targeted to the BBB, in order to treat HIV neuropathology. Compared to the conventional method, NPs produced through microfluidics presented reduced size (73 nm versus 133 nm), comparable polydispersity (around 0.090), less negative zeta-potential (-14.1 mV versus -28.0 mV), higher EFV association efficiency (80.7% versus 32.7%) and higher drug loading (10.8% versus 3.2%). The microfluidics-produced NPs also demonstrated a sustained in vitro EFV release (50% released within the first 24 h). NPs functionalization with a transferrin receptor-binding peptide, envisaging BBB targeting, proved to be effective concerning nuclear magnetic resonance analysis (delta = -0.008 ppm; delta = -0.017 ppm). NPs demonstrated to be safe to BBB endothelial and neuron cells (metabolic activity above 70%), as well as non hemolytic (1-2% of hemolysis, no morphological alterations on erythrocytes). Finally, functionalized nanosystems were able to interact more efficiently with BBB cells, and permeability of EFV associated with NPs through a BBB in vitro model was around 1.3-fold higher than the free drug.

Abstract Antibiotic resistance has been increasingly reported for a wide variety of bacteria of clinical significance. This widespread problem constitutes one of the greatest challenges of the twenty-first century. Faced with this issue, clinicians and researchers have been persuaded to design novel strategies in order to try to control pathogenic bacteria. Therefore, the discovery and elucidation of the mechanisms underlying bacterial pathogenesis and intercellular communication have opened new perspectives for the development of alternative approaches. Antipathogenic and/or antivirulence therapies based on the interruption of quorum sensing pathways are one of several such promising strategies aimed at disarming rather than at eradicating bacterial pathogens during the course of colonization and infection. This review describes mechanisms of bacterial communication involved in biofilm formation. An overview of the potential of marine bacteria and their bioactive components as QS inhibitors is further provided.

Abstract While photodynamic therapy is known for significant advantages over conventional cancer therapies, its dependence on light has limited it to treating tumors on or just under the skin or on the outer lining of organs/cavities. Herein, we have developed a single-molecule photosensitizer capable of intracellular self-activation and with potential tumor-selectivity due to a chemiluminescent reaction involving only a cancer marker. Thus, the photosensitizer is directly chemiexcited to a triplet excited state capable of generating singlet oxygen, without requiring either a light source or any catalyst/co-factor. Cytotoxicity assays involving the photosensitizer show significant toxicity toward tumor cells, even better than reference drugs, while not inducing toxicity toward normal cells. This work provides a proof-of-concept for a novel type of photosensitizer that eliminates the current restrictions that photodynamic therapy presents regarding tumor size and localization.

Abstract Cancer is a very challenging disease to treat, both in terms of treatment efficiency and side-effects. To overcome these problems, there have been extensive studies regarding the possibility of improving treatment by employing combination therapy, and by exploring therapeutic modalities with reduced side-effects (such as photodynamic therapy (PDT)). Herein, this work has two aims: (i) to develop self-activating photosensitizers for use in light-free photodynamic therapy, which would eliminate light-related restrictions that this therapy currently possesses; (ii) to assess their co-treatment potential when combined with reference chemotherapeutic agents (Tamoxifen and Metformin). We synthesized three new photosensitizers capable of self-activation and singlet oxygen production via a chemiluminescent reaction involving only a cancer marker and without requiring a light source. Cytotoxicity assays demonstrated the cytotoxic activity of all photosensitizers for prostate and breast tumor cell lines. Analysis of co-treatment effects revealed significant improvements for breast cancer, producing better results for all combinations than just for the individual photosensitizers and even Tamoxifen. By its turn, co-treatment for prostate cancer only presented better results for one combination than for just the isolated photosensitizers and Metformin. Nevertheless, it should be noted that the cytotoxicity of the isolated photosensitizers in prostate tumor cells was already very appreciable.

Abstract Persistent bacterial infections and increased resistance to multidrugs are mostly related with biofilm formation which constitute a global concern in nowadays society. Therefore, studies must be conducted in order to discover more efficient antibiofilm agents. Nine compounds with a catecholic moiety (catechol (CAT), veratrol (VER), guaiacol (GUA), 2-ethoxphenol (ETH), 4-methylcatechol (MEC), 4-tert-butylcatechol (TEB), pyrogallol (PYR), 3-methoxycatechol (MET), and o-phenylene-phosphochloridite (OPP)) were investigated for their potential to prevent and eradicate Escherichia coil biofilms. Their action was assessed on biomass and metabolic activity, sessile cells' membrane integrity and culturability, motility and cell surface hydrophobicity (CSH). ETH, MEC, TEB, PYR and OPP exhibited the best antibiofilm activities among the tested catechols, inducing biofilm removal and metabolic inactivation, sessile cells' membrane disruption and death. CAT, TEB and PYR significantly inhibited E. coil swimming motility. Likewise, ETH, MEC, TEB and PYR significantly reduced E. coil CSH. An additive interaction of combined catechols with ciprofloxacin was attained in both control and preventive strategies. From a structure activity relationship (SAR) study, an increase in the hydrocarbon side chain and lipophilicity using ETH and TEB as scaffolds is suggested for increased activity. Hydroxyl groups are also suggested to be particularly related to their antibiofilm activity. All the catechols fulfil the Lipinski's "rule of five" requisites and thus are promising scaffolds in the development of new formulations for therapeutic purposes.

Abstract The comprehension of molecular recognition phenomena demands the understanding of the energetic and kinetic processes involved. General equations valid for the thermodynamic analysis of any observable that is assessed as a function of the concentration of the involved compounds are described, together with their implementation in the AFFINImeter software. Here, a maximum of three different molecular species that can interact with each other to form an enormous variety of supramolecular complexes are considered. The corrections currently employed to take into account the effects of dilution, volume displacement, concentration errors and those due to external factors, especially in the case of ITC measurements, are included. The methods used to fit the model parameters to the experimental data, and to generate the uncertainties are described in detail. A simulation tool and the so called kinITC analysis to get kinetic information from calorimetric experiments are also presented. An example of how to take advantage of the AFFINImeter software for the global multi-temperature analysis of a system exhibiting cooperative 1:2 interactions is presented and the results are compared with data previously published. Some useful recommendations for the analysis of experiments aimed at studying molecular interactions are provided.