Drug biophysical profiling using lipid-based colloidal nanosystems and human serum albumin as biomimetic interfaces

The development of new drugs is a highly complex and expensive process, so it is crucial that less promising compounds are rejected early in the discovery phase before progressing to more expensive phases. This scenario impels researchers to refine and speed up the drug discovery process and to seek tools to support decisions related to modifications of the drug chemical structure to improve drugs’ properties and thus increase the probability of success in the process of drug discovery. [1], [2] In the drug discovery process it should be considered that in physiological environment there will be reciprocal interactions between drugs and biological interfaces, such as cell membranes or plasma proteins, and from those interactions different pharmacokinetic profiles can be achieved. [3] Thus, it is important to develop in vitro high throughput methods to evaluate the pharmaceutical profile, consisting in measuring properties such as permeability, lipophilicity, plasma protein binding, and biophysical changes of the membranes, which in turn affect other properties, such as the bioavailability of a drug and its pharmacokinetic profile. [4] Herein, the characterization of a newly synthesized drug (MIT-3) will be based on the measurement of fundamental biophysical properties, which allow inferring about its ADMET profile (absorption, distribution, excretion and toxicity at the membrane level). For this purpose, lipid-based colloidal nanosystems of different compositions were prepared as membrane mimetic models and several biophysical techniques were applied: derivative spectroscopy; quenching of steady-state and time-resolved fluorescence; quenching of intrinsic fluorescence of human serum albumin; synchronous fluorescence; dynamic and electrophoretic light scattering, differential scanning calorimetry and small and wide angle x-ray diffraction. The application of these techniques allowed to predict that MIT-3 has an ubiquitous location at the membrane level, presenting good membrane permeability and a good distribution in the therapeutic target. However, it is also predicted bioaccumulation with distribution in non-therapeutic targets and under conditions of prolonged exposure the drug may cause membrane toxicity as concluded by the impairment of membrane biophysical properties. It is also possible to conclude that the biophysical techniques and the biomimetic models used, constitute a toolbox of strategies for the future evaluation of other drugs.This work was supported by the Portuguese Foundation for Science and Technology (FCT) in the framework of the Strategic Funding UID/FIS/04650/2013. We also acknowledge PEstC/QUI/UI0081/2013, NORTE-01-0145-FEDER-000028 and PTDC/DTP-FTO/2433/2014. F. Cagide and S. Benfeito are thankful for the pos-doctoral and doctoral grants (SFRH/BPD/74491/2010 and SFRH/BD/99189/2013 respectively). Marlene Lúcio acknowledges the exploratory project funded by FCT with the reference IF/00498/2012. Eduarda Fernandes acknowledges COMPETE 2020 “Programa Operacional Competitividade e internacionalização”.info:eu-repo/semantics/publishedVersio

Effect of sitagliptin on cardiovascular outcomes in type 2 diabetes

BACKGROUND: Data are lacking on the long-term effect on cardiovascular events of adding sitagliptin, a dipeptidyl peptidase 4 inhibitor, to usual care in patients with type 2 diabetes and cardiovascular disease. METHODS: In this randomized, double-blind study, we assigned 14,671 patients to add either sitagliptin or placebo to their existing therapy. Open-label use of antihyperglycemic therapy was encouraged as required, aimed at reaching individually appropriate glycemic targets in all patients. To determine whether sitagliptin was noninferior to placebo, we used a relative risk of 1.3 as the marginal upper boundary. The primary cardiovascular outcome was a composite of cardiovascular death, nonfatal myocardial infarction, nonfatal stroke, or hospitalization for unstable angina. RESULTS: During a median follow-up of 3.0 years, there was a small difference in glycated hemoglobin levels (least-squares mean difference for sitagliptin vs. placebo, -0.29 percentage points; 95% confidence interval [CI], -0.32 to -0.27). Overall, the primary outcome occurred in 839 patients in the sitagliptin group (11.4%; 4.06 per 100 person-years) and 851 patients in the placebo group (11.6%; 4.17 per 100 person-years). Sitagliptin was noninferior to placebo for the primary composite cardiovascular outcome (hazard ratio, 0.98; 95% CI, 0.88 to 1.09; P<0.001). Rates of hospitalization for heart failure did not differ between the two groups (hazard ratio, 1.00; 95% CI, 0.83 to 1.20; P = 0.98). There were no significant between-group differences in rates of acute pancreatitis (P = 0.07) or pancreatic cancer (P = 0.32). CONCLUSIONS: Among patients with type 2 diabetes and established cardiovascular disease, adding sitagliptin to usual care did not appear to increase the risk of major adverse cardiovascular events, hospitalization for heart failure, or other adverse events

Bonding to Psychedelics: Synthesis of Molecularly Imprinted Polymers Targeting 4-Bromo-2,5-dimethoxyphenethylamine (2C-B)

The increasing interest in utilizing psychedelics for therapeutic purposes demands the development of tools capable of efficiently monitoring and accurately identifying these substances, thereby supporting medical interventions. 4-Bromo-2,5-dimethoxyphenethylamine (2C-B) has gained significant popularity as one of the most widely used psychedelic compounds in non-medical settings. In this study, we aimed to create a material with selective recognition of 2C-B by synthesizing a series of molecularly imprinted polymers (MIP) using 2C-B as the template and varying ratios of methacrylic acid (MAA) as the functional monomer (1:2, 1:3, and 1:4). Both thermal and microwave-assisted polymerization processes were employed. The molar ratio between the template molecule (2C-B) and functional monomer (MAA) was 1:4, utilizing a microwave-assisted polymerization process. Isotherm studies revealed a Langmuir’s maximum absorption capacity (Bmax) value of 115.6 μmol·mg−1 and Kd values of 26.7 μM for this material. An imprint factor of 4.2 was determined for this material, against the corresponding non-imprinted polymer. The good selectivity against 14 other new psychoactive substances highlighted the material’s potential for applications requiring selective recognition. These findings can contribute to the development of tailored materials for the detection and analysis of 2C-B, supporting advancements in non-medical use monitoring and potential therapeutic models involving psychedelics

Phytochemicals and quaternary phosphonium ionic liquids: Connecting the dots to develop a new class of antimicrobial agents

Introduction: The infections by multidrug-resistant bacteria are a growing threat to human health, and the efficacy of the available antibiotics is gradually decreasing. As such, new antibiotic classes are urgently needed. Objectives: This study aims to evaluate the antimicrobial activity, safety and mechanism of action of phytochemical-based triphenylphosphonium (TPP+) conjugates. Methods: A library of phytochemical-based TPP+ conjugates was repositioned and extended, and its antimicrobial activity was evaluated against a panel of Gram-positive (methicillin-resistant Staphylococcus aureus – MRSA) and Gram-negative bacteria (Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Acinetobacter baumannii) and fungi (Candida albicans, Cryptococcus neoformans var. grubii). The compounds’ cytotoxicity and haemolytic profile were also evaluated. To unravel the mechanism of action of the best compounds, the alterations in the surface charge, bacterial membrane integrity, and cytoplasmic leakage were assessed. Results: Structure-activity-toxicity data revealed the contributions of the different structural components (phenolic ring, carbon-based spacers, carboxamide group, alkyl linker) to the compounds’ bioactivity and safety. Dihydrocinnamic derivatives 5 m and 5n stood out as safe, potent and selective antibacterial agents against S. aureus (MIC  32 µg/mL; HC10 > 32 µg/mL). Mechanistic studies suggest that the antibacterial activity of compounds 5 m and 5n may result from interactions with the bacterial cell wall and membrane. Conclusions: Collectively, these studies demonstrate the potential of phytochemical-based TPP+ conjugates as a new class of antibiotics

Mitochondria-targeted phenolic antioxidants induce ROS-protective pathways in primary human skin fibroblasts

Contains fulltext : 229055.pdf (publisher's version ) (Closed access)Phytochemical antioxidants like gallic and caffeic acid are constituents of the normal human diet that display beneficial health effects, potentially via activating stress response pathways. Using primary human skin fibroblasts (PHSFs) as a model, we here investigated whether such pathways were induced by novel mitochondria-targeted variants of gallic acid (AntiOxBEN(2)) and caffeic acid (AntiOxCIN(4)). Both molecules reduced cell viability with similar kinetics and potency (72 h incubation, IC50 ~23 μM). At a relatively high but non-toxic concentration (12.5 μM), AntiOxBEN(2) and AntiOxCIN(4) increased ROS levels (at 24 h), followed by a decline (at 72 h). Further analysis at the 72 h timepoint demonstrated that AntiOxBEN(2) and AntiOxCIN(4) did not alter mitochondrial membrane potential (Δψ), but increased cellular glutathione (GSH) levels, mitochondrial NAD(P)H autofluorescence, and mitochondrial superoxide dismutase 2 (SOD2) protein levels. In contrast, cytosolic SOD1 protein levels were not affected. AntiOxBEN(2) and AntiOxCIN(4) both stimulated the gene expression of Nuclear factor erythroid 2-related factor 2 (NRF2; a master regulator of the cellular antioxidant response toward oxidative stress). AntiOxBEN2 and ANtiOxCIN4 differentially affected the gene expression of the antioxidants Heme oxygenase 1 (HMOX1) and NAD(P)H dehydrogenase (quinone) 1 (NQO1). Both antioxidants did not protect from cell death induced by GSH depletion and AntiOxBEN(2) (but not AntiOxCIN(4)) antagonized hydrogen peroxide-induced cell death. We conclude that AntiOxBEN(2) and AntiOxCIN(4) increase ROS levels, which stimulates NRF2 expression and, as a consequence, SOD2 and GSH levels. This highlights that AntiOxBEN(2) and AntiOxCIN(4) can act as prooxidants thereby activating endogenous ROS-protective pathways

Prospective Validation of the Prognostic Usefulness of B-Type Natriuretic Peptide in Asymptomatic Patients With Chronic Severe Aortic Regurgitation

ObjectivesThe purpose of this study was to determine the independent and additive prognostic value of B-type natriuretic peptide (BNP) in patients with severe asymptomatic aortic regurgitation and normal left ventricular function.BackgroundEarly surgery could be advisable in selected patients with chronic severe aortic regurgitation, but there are no uniform criteria to identify candidates who could benefit from this strategy. Assessment of BNP has not been studied for this purpose.MethodsWe prospectively evaluated 294 consecutive patients with severe asymptomatic organic aortic regurgitation and left ventricular ejection fraction above 55%. The first 160 consecutive patients served as the derivation cohort and the next 134 patients served as a validation cohort. The combined endpoint was the occurrence of symptoms of congestive heart failure, left ventricular dysfunction, or death at follow-up.ResultsThe endpoint was reached in 45 patients (28%) of the derivation set and in 35 patients (26%) of the validation cohort. Receiver-operator characteristic curve analysis yielded an optimal cutoff point of 130 pg/ml for BNP that was able to discriminate between patients at higher risk in both cohorts. BNP was the strongest independent predictor by multivariate analysis in the derivation set (odds ratio: 6.9 [95% confidence interval: 2.52 to 17.57], p < 0.0001) and the validation set (odds ratio: 6.7 [95% confidence interval: 2.9 to 16.9], p = 0.0001).ConclusionsAmong patients with severe asymptomatic aortic regurgitation and normal left ventricular function, BNP ≥130 pg/ml categorizes a subgroup of patients at higher risk. Because of its incremental prognostic value, we believe BNP assessment should be used in the routine clinical evaluation of these patients