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Aging is one of the main risk factors for cardiovascular diseases, and oxidative stress is a key element responsible for the development of age-related pathologies. In addition, the alteration of circadian rhythms also contributes to cardiovascular pathology, but the underlying mechanisms are not well defined. We investigated the aging consequences on the temporal patterns of antioxidant defenses, the molecular clock machinery, and the blood pressure, in the heart of male rats maintained under constant darkness (free running) conditions. Male Holtzman rats from young adult (3-month-old) and older (22-month-old) groups were maintained under constant darkness (12-h dark:12-h dark, DD) condition during fifteen days before the experiment. After the DD period, heart ventricle samples were isolated every 4-h throughout a 24-h period. We observed circadian rhythms of catalase (CAT) and glutathione peroxidase (GPx) mRNA expression, as well as ultradian rhythms of Nrf2 mRNA levels, in the heart of young adult rats. We also found circadian oscillations of CAT and GPx enzymatic activities, reduced glutathione (GSH) and BMAL1 protein in the same group. Interestingly, aging abolished the rhythms of CAT and GPx enzymatic activities, phase-shifted the rhythm's acrophases of GSH and BMAL1 protein levels and turned circadian the ultradian oscillation of Nrf2 expression. Moreover, aging phase-shifted the circadian pattern of systolic blood pressure. In conclusion, aging modifies the temporal organization of antioxidant defenses and blood pressure, probably, as a consequence of a disruption in the circadian rhythm of the clock's transcriptional regulator, BMAL1, in heart.

Bovine mastitis is an infectious disease that affects the mammary gland of dairy cattle with considerable economic losses. Staphylococcus aureus is the main microorganism involved in this highly contagious process, and the treatment is only using antibiotics. Currently, the search for new treatment and/or compounds is still in need due to microbial resistance. In this work, we evaluated the potential of eugenol and thymol derivatives against S. aureus strains from bovine mastitis. On that purpose, nine derivatives were synthesized from eugenol and thymol (1–9), and tested against 15 strains of S. aureus from subclinical bovine mastitis. Initially, the strains were evaluated for the biofilm production profile, and those with strong adherence were selected to the antimicrobial sensitivity determination in the Minimum Inhibitory Concentration (MIC) assays. Herein the compounds toxicity was also evaluated by in silico analysis using Osiris DataWarrior® software. The results showed that 60% of the strains were considered strongly adherent and three strains (S. aureus 4271, 4745 and 4746) were selected for the MIC tests. Among the nine eugenol and thymol derivatives tested, four were active against the evaluated strains (MIC = 32 µg mL−1) within CLSI standard values. In silico analysis showed that all derivatives had cLopP − 4 and TPSA

Heregulin (HRG) signaling has been implicated in the development of an aggressive phenotype in breast cancer (BC) cells, and HER2 overexpression has been associated with a worse prognosis in BC patients. Nevertheless, the molecular mechanisms through which HRG affects the efficiency of anti-HER2 therapies such as trastuzumab (Tz) and trastuzumab-emtansine (T-DM1) are currently unknown. In the present study, we evaluate the molecular action of HRG toward fundamental scaffold proteins and several kinases in the signal transduction pathways triggered via HER2/HER3, which integrate precise and sequential steps to promote changes in cell morphology to impulse BC cell migration. In addition, we evaluate the effectiveness of Tz and T-DM1 on the control of key proteins involved in BC cell motility, since the acquisition of a migratory phenotype is essential to promote invasion and metastasis. We show that HRG induces actin cytoskeleton reorganization and focal adhesion complex formation, and promotes actin nucleation in BT-474 BC cells. This signaling is triggered by HER2/HER3 to c-Src, FAK and paxillin. When paxillin is phosphorylated, it recruits PAK1, which then phosphorylates cortactin. In parallel, paxillin signals to N-WASP, and both signalings regulate Arp2/3 complex, leading to the local reorganization of actin fibers. Our findings reveal an original mechanism by which HRG increases HER2+ BC cell motility, and show that the latter can be abolished by Tz and T-DM1 treatments. These results provide evidence for the molecular mechanisms involved in cell motility and drug resistance. They will be useful to develop new and more specific therapeutic schemes that interfere with the progression and metastasis of HER2+ BC.

To assess adherence to standard clinical practice for the diagnosis and treatment of patients undergoing prostate cancer (PCa) radiotherapy in four European countries using clinical audits as part of the international IROCA project. Multi-institutional, retrospective cohort study of 240 randomly-selected patients treated for PCa (n = 40/centre) in the year 2015 at six European hospitals. Clinical indicators applicable to general and PCa-specific radiotherapy processes were evaluated. All data were obtained directly from medical records. The audits were performed in the year 2017. Adherence to clinical protocols and practices was satisfactory, but with substantial inter-centre variability in numerous variables, as follows: staging MRI (range 27.5–87.5% of cases); presentation to multidisciplinary tumour board (2.5–100%); time elapsed between initial visit to the radiation oncology department and treatment initiation (42–102.5 days); number of treatment interruptions ≥ 1 day (7.5–97.5%). The most common deviation from standard clinical practice was inconsistent data registration, mainly failure to report data related to diagnosis, treatment, and/or adverse events. This clinical audit detected substantial inter-centre variability in adherence to standard clinical practice, most notably inconsistent record keeping. These findings confirm the value of performing clinical audits to detect deviations from standard clinical practices and procedures.

Candidiasis is a serious public health problem that currently affects not only immunodeficient patients with predisposing conditions, but also immunocompetent individuals. Thus, the search for new antifungal agents is required also due to the emergence of resistant strains and to the side effects of the available drugs. The aim of this study is to evaluate the in vitro antifungal profile of nine synthetic 1,2,3-triazole derivatives against four Candida species of medical importance (C. albicans, C. tropicalis, C. parapsilosis, and C. krusei), as well as to identify their in silico structure-activity relationship. Interestingly, the antifungal susceptibility tests showed the compound 5-methyl-1-(phenylamino)-1H-1,2,3-triazol-4-yl-methanol (2b) with the lowest minimal inhibitory concentration value against C. albicans strain (MIC = 8 μg/mL) similar to other promissing compounds described in the literature. According to our in silico evaluation, some stereoelectronic properties (e.g., higher values of log S and lowest unoccupied molecular orbital energy and lower number of atoms, rotatable bonds and Hydrogen bond acceptors) were correlated with the antifungal activity detected. This series reinforced the potential of 1,2,3-triazole as a promising nucleus in the search for new antifungals and may help on designing new drugs for candidiasis.

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.

A molecular modeling study was applied to a data set of 21 analogues (2a–2u) of amodiaquine (2) exhibiting remarkable in vitro activity against the chloroquine- and pyrimethamine-resistant Plasmodium falciparum K1 strain (Guglielmo et al. in Eur J Med Chem 44:5071–5079, 2009). Attempting to correlate structural features with the antiparasitic activity, quantum chemical properties were calculated. In silico ADMET studies were conducted in order to recognize and find the most promising compounds with the potential of becoming new antimalarial agents. The binding mode to the heme group was studied by molecular docking, and the interactions observed in the ligand-heme complex of one of the highly active (2p) and one of the least active (2l) compounds were compared. In order to crosscheck the docking results, compound 2p was submitted to 50 ns of molecular dynamic simulation. According to that result, 2p and amodiaquine share similar binding interaction with the heme group. Additionally, a charged-assisted hydrogen bond between the protonated quinolone and the heme carboxylate group was observed. The introduction of the piperazine moiety increases the antimalarial activity profile of amodiaquine analogues by conformational features that allow these interactions. The results of this study can be used as a guide for the design of new compounds for treatment against malaria.

Despite the large number of current antifungals used for treating fungal infections, the increased resistance of these microorganisms over the years has been one of the main and major challenges for medicine in the new century. Thus, it remains necessary to discover novel antifungal agents. The objective of this study was to identify the stereoelectronic features related to the antifungal activity of a series of triazoles and imidazoles derivates with activity against Candida albicans, also pharmacokinetic and toxicity in silico were evaluated to guide the rational design and identification of new antifungal agents. The results showed that features such as maintenance of imidazole ring, the profile of distribution and density of high occupied molecular orbital and conformational aspects in these derivatives were important for the activity and also demonstrate that all the compounds showed good oral bioavailability and low theoretical toxicity profile when compared to antifungals on the market, presenting a promising profile for future in vivo studies.

The aim of the present work was to use GastroPlus™ software for the prediction of pharmacokinetic profiles and in vitro-in vivo correlation (IVIVC) as tools to optimize the development of new generic medications. GastroPlus™ was used to simulate the gastrointestinal compartment and was based on the advanced compartmental absorption and transit model. Powder dissolution and efavirenz tablet dissolution studies were carried out to generate data from which correlation was established. The simulated plasma profile, based on the physicochemical properties of efavirenz, was almost identical to that observed in vivo for biobatches A and B. A level A IVIVC was established for the dissolution method obtained for the generic candidate using the Wagner-Nelson (r (2) = 0.85) and for Loo-Riegelman models (r(2) = 0.92). The percentage of fraction absorbed indicated that 0.5% sodium lauryl sulfate may be considered a biorelevant dissolution medium for efavirenz tablets. The simulation of gastrointestinal bioavailability and IVIVC obtained from immediate-release tablet formulations suggests that GastroPlus™ is a valuable in silico method for IVIVC and for studies directed at developing formulations of class II drugs.

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.

Herpes simplex virus is an important human pathogen responsible for a range of diseases from mild uncomplicated mucocutaneous infections to life-threatening ones. Currently, the emergence of Herpes simplex virus resistant strains increased the need for more effective and less cytotoxic drugs for Herpes treatment. In this work, we synthesized a series of oxoquinoline derivatives and experimentally evaluated the antiviral activity against acyclovir resistant HSV-1 strain as well as their cytotoxity profile. The most active compound (3b), named here as Fluoroxaq-3b, showed a promising profile with a better cytotoxicity profile than acyclovir. The theoretical analysis of the structure-activity relationship of these compounds revealed some stereoelectronic properties such as lower LUMO energy and lipophilicity, besides a higher polar surface area and number of hydrogen bond acceptor groups as important parameters for the antiviral activity. Fluoroxaq-3b showed a good oral theoretical bioavailability, according to Lipinski rule of five, with a promising profile for further in vivo analysis.

A broad-spectrum antibiotic therapy has led to medical complications and emergence of multiresistant bacteria including Enterococcus faecalis. In this study, we designed, synthesized, and evaluated the antibacterial activity of 13 nor-β-lapachone derivatives against a drug resistant E. faecalis strain. Two triazole substituted compounds (1e = 8 μg/ml and 1c = 16 μg/ml) and the non-substituted derivative (1a = 8 μg/ml) were promising compared to chloramphenicol (12 μg/ml), an antibiotic currently available in the market. We also performed a structure-activity relationship analysis using a molecular modeling approach that pointed the low HOMO energy values; HOMO density concentrated on the nor-β-lapachone ring, lipophilicity, solubility and number HBA as important stereoelectronic features for the antibacterial profile. In addition the triazole compounds presented low theoretical toxicity profile, and drug-score higher than commercial antibiotics also fulfilling the Lipinski "Rule of Five", which pointed them as promising candidates for further studies in infections caused by multiresistant E. faecalis hospital strains.

Ecotin is a bidentate, fold-specific inhibitor of mammalian serine-proteases produced by Escherichia coli. This molecule may be engineered to increase and/or change its affinity and specificity providing significant biotechnological potential. Since ecotin binds tightly to serine proteases of the trypsin fold, it may help to identify the role of these enzymes in different biological processes. In this work, we tested ecotin variants as an affinity purification reagent for identifying enzymes in samples of tumor progression and mammary gland involution. Initially, we used a commercial source of urokinase-type plasminogen activator (u-PA) that remained fully active after elution from an affinity column of the ecotin variant (M84R, M85R). We then successfully identified u-PA from more complex mixtures including lysates from a prostate cancer cell line and involuting mouse mammary glands. Interestingly, a membrane-type serine protease 1 was isolated from the Triton X-100-solubilized PC-3 cell lysates, and surprisingly, haptoglobin, a serine-protease homolog protein, was also identified in mammary gland lysates and in blood. Haptoglobin does not prevent ecotin inhibition of u-PA, but it may act as a carrier within blood when ecotin is used in vivo. Finally, this affinity purification matrix was also able to identify a thrombin-like enzyme from snake venom using an ecotin variant directed against thrombin. Overall, the ecotin variants acted as robust tools for the isolation and characterization of proteins with a trypsin fold. Thus, they may assist in the understanding of the role of these serine proteases and homologous proteins in different biological processes.

Current drugs for treating leishmaniasis are still associated with significant toxicity and failure rates. Thus, new effective and less toxic antileishmanial agents are still in need. Herein, we tested a series of sulfonamide 4-methoxychalcone derivatives against L. amazonensis promastigote and amastigote forms to identify its antileishmanial profile against this species compared to L. braziliensis. In addition, we used molecular modeling tools to determine stereoelectronic features that may lead to the antileishmanial profile. Interestingly, all tested compounds were able to affect L. amazonensis promastigote form in a concentration-dependent manner and with low cytotoxicity, except for derivative 3g. However, our results showed that compound 3f (para-Cl) presents the best profile against both L. amazonensis forms (promastigote and amastigote), differently from that observed for L. braziliensis, when compound 3i was the most active. Structure-activity relationship (SAR) analysis of these derivatives pointed molecular volume, HOMO density, and conformational aspects as important characteristics for parasitic profile. Overall, sulfonamide 4-methoxychalcone derivatives may be pointed out not only as lead compounds for treating leishmaniasis (i.e., 3f) but also as experimental tools presenting parasite-selectivity (i.e., 3i).

Bacterial infections are a significant cause of morbidity and mortality among critically ill patients. The increase of antibiotic resistance in bacteria from human microbiota—such as Staphylococcus epidermidis, an important nosocomial pathogen that affects immunocompromised patients or those with indwelling devices—increased the desire for new antibiotics. In this study we designed, synthesized, and determined the antimicrobial activity of 27 thieno[2,3-b]pyridines (1, 2, 2a–2m, 3, 3a–3m) derivatives against a drug-resistant clinical S. epidermidis strain. In addition, we performed a structure-activity relationship analysis using a molecular modeling approach, and discuss the drug absorption, distribution, metabolism, excretion, and toxicity profile and Lipinski’s “rule of five,” which are tools to assess the relationship between structures and drug-like properties of active compounds. Our results showed that compound 3b (5-(1H-tetrazol-5-yl)-4-(3`-methylphenylamino)thieno[2,3-b]pyridine) was as active as oxacillin and chloramphenicol but with lower theoretical toxicity risks and a better drug likeness and drug score potential than chloramphenicol. All molecular modeling and biological results reinforced the promising profile of 3b for further experimental investigation and development of new antibacterial drugs.

Leishmaniasis is a disease caused by flagellate protozoan Leishmania spp. and represents an emergent illness with high morbidity and mortality in the tropics and subtropics. Since the discovery of the first drugs for Leishmaniasis treatment (i.e., pentavalent antimonials), until the current days, the search for substances with antileishmanial activity, without toxic effects, and able to overcome the emergence of drug resistant strains still remains as the current goal. This article reports the development of new chemotherapies through the rational design of new drugs, the use of products derived from microorganisms and plants, and treatments related to immunity as new alternatives for the chemotherapy of leishmaniasis.

Background: Chagas disease kills 2.5 thousand people per year of 15 million persons infected in Latin America. The disease is caused by the protozoan, Trypanosome cruzi, and vectored by triatomine insects, including Panstrongylus megistus, an important vector in Brazil. Medicines treating Chagas disease have unpleasant side effects and may be ineffective, therefore, alternative control techniques are required. Knowledge of the T. cruzi interactions with the triatomine host needs extending and new targets/strategies for control identified. Serine and cysteine peptidases play vital roles in protozoan life cycles including invasion and entry of T. cruzi into host cells. Peptidase inhibitors are, therefore, promising targets for disease control. Methods: SDS PAGE and chromatograpy detected and isolated a P. megistus serpin which was peptide sequenced by mass spectrometry. A full amino acid sequence was obtained from the cDNA and compared with other insect serpins. Reverse transcription PCR analysis measured serpin transcripts of P. megistus tissues with and without T. cruzi infection. Serpin homology modeling used the Swiss Model and Swiss-PDB viewer programmes. Results: The P. megistus serpin (PMSRP1) has a ca. 40 kDa molecular mass with 404 amino acid residues. A reactive site loop contains a highly conserved hinge region but, based on sequence alignment, the normal cleavage site for serine proteases at P1-P1′ was translocated to the putative position P4′-P5′. A small peptide obtained corresponded to the C-terminal 40 amino acid region. The secondary structure of PMSRP1 indicated nine α-helices and three β-sheets, similar to other serpins. PMSRP1 transcripts occurred in all tested tissues but were highest in the fat body and hemocytes. Levels of mRNA encoding PMSRP1 were significantly modulated in the hemocytes and stomach by T. cruzi infection indicating a role for PMSRP1 in the parasite interactions with P. megistus. Conclusions: For the first time, a constitutively expressed serpin has been characterized from the hemolymph of a triatomine. This opens up new research avenues into the roles of serine peptidases in the T. cruzi/P. megistus association. Initial experiments indicate a role for PMSRP1 in T. cruzi interactions with P. megistus and will lead to further functional studies of this molecule.

Background Vancomycin is an antibiotic of growing importance in the treatment of hospital infections, with particular emphasis on its value in the fight against methicillin-resistant Staphylococcus aureus. However its usage profile must be evaluated to assure maximum benefit and minimum risk. Findings A cross-sectional retrospective study was carried out among inpatients that received vancomycin in a Brazilian quaternary hospital. The occurrence of adverse reactions reported was evaluated in medical records relating to patients taking vancomycin during a one year period. Males comprised 52% (95% CI: 41.7-60.2%) of the sample population, with a mean age of 50.6 (95% CI: 47.2-54.0) years and mean treatment period of 9.7 (95% CI: 8.0-11.5) Days. It was verified that nephrotoxicity occurred in 18.4% (95% CI: 11.3-27.5) of patients, Red man syndrome occurred in 2% (95% CI 0.2-7.2), while the occurrence of thrombocytopenia was 7.1% (95% CI: 2.9-14.2). Conclusions It may be noted that even after 50 years of use, adverse reactions associated with vancomycin continue with high frequency, presenting a public health problem, especially considering its current use in cases of multidrug resistant infections. In this context, we emphasize the importance of intensive pharmacovigilance in hospital as a surveillance tool after drug approval by the sanitary authority.