Exploring Free Energies of Specific Protein Conformations Using the Martini Force Field.

Coarse-grained (CG) level molecular dynamics simulations are routinely used to study various biomolecular processes. The Martini force field is currently the most widely adopted parameter set for such simulations. The functional form of this and several other CG force fields enforces secondary protein structure support by employing a variety of harmonic potentials or restraints that favor the protein's native conformation. We propose a straightforward method to calculate the energetic consequences of transitions between predefined conformational states in systems in which multiple factors can affect protein conformational equilibria. This method is designed for use within the Martini force field and involves imposing conformational transitions by linking a Martini-inherent elastic network to the coupling parameter λ. We demonstrate the applicability of our method using the example of five biomolecular systems that undergo experimentally characterized conformational transitions between well-defined structures (Staphylococcal nuclease, C-terminal segment of surfactant protein B, LAH4 peptide, and ß2-adrenergic receptor) as well as between folded and unfolded states (GCN4 leucine zipper protein). The results show that the relative free energy changes associated with protein conformational transitions, which are affected by various factors, such as pH, mutations, solvent, and lipid membrane composition, are correctly reproduced. The proposed method may be a valuable tool for understanding how different conditions and modifications affect conformational equilibria in proteins.

Analysis of pharmacokinetic profile and ecotoxicological character of cefepime and its photodegradation products.

An important problem is the impact of photodegradation on product toxicity in biological tests, which may be complex and context-dependent. Previous studies have described the pharmacology of cefepime, but the toxicological effects of its photodegradation products remain largely unknown. Therefore, photodegradation studies were undertaken in conditions similar to those occurring in biological systems insilico, in vitro, in vivo and ecotoxicological experiments. The structures of four cefepime photodegradation products were determined by UPLC-MS/MS method. The calculated in silico ADMET profile indicates that carcinogenic potential is expected for compounds CP-1, cefepime, CP-2 and CP-3. The Cell Line Cytomotovity Predictor 2.0 tool was used to predict the cytotoxic effects of cefepime and related compounds in non-transformed and cancer cell lines. The results indicate that possible actions include: non-small cell lung cancer, breast adenocarcinoma, prostate cancer and papillary renal cell carcinoma. OPERA models were used to predict absorption, distribution, metabolism and excretion (ADME) endpoints, and potential bioactivity of CP-2, cefepime and CP-4. The results obtained in silico show that after 96h of exposure, cefepime, CP-1, CP-2, and CP-3 are moderately toxic in the zebrafish model, while CP-4 is highly toxic. On the contrary, cefepime is more toxic to T. platyurus (highly toxic) compared to the zebrafish model, similar to products CP-4, CP-3 and CP-2. In vitro cytotoxicity studies were performed by MTT assay and in vivo acute embryo toxicity studies using Danio rerio embryos and larvae. In vitro showed an increase in the cytotoxicity of products with the longest exposure period i.e. for 8 h. Additionally, at a concentration of 200 µg/mL, statistically significant changes in metabolic activity were observed depending on the irradiation time. In vivo studies conducted with Zebrafish showed that both cefepime and its photodegradation products have only low toxicity. Assessment of potential ecotoxicity included Microbiotests on invertebrates (Thamnotoxkit F and Daphtoxkit F), and luminescence inhibition tests (LumiMara). The observed toxicity of the tested solutions towards both Thamnocephalus platyurus and Daphnia magna indicates that the parent substance (unexposed) has lower toxicity, which increases during irradiation. The acute toxicity (Lumi Mara) of nonirradiated cefepime solution is low for all tested strains (<10%), but mixtures of cefepime and its photoproducts showed growth inhibition against all tested strains (except #6, Photobacterium phoreum). Generally, it can be concluded that after UV-Vis irradiation, the mixture of cefepime phototransformation products shows a significant increase in toxicity.

Synthesis of Novel 2-(Cyclopentylamino)thiazol-4(5H)-one Derivatives with Potential Anticancer, Antioxidant, and 11ß-HSD Inhibitory Activities.

In this study, a series of nine new 2-(cyclopentylamino)thiazol-4(5H)-one derivatives were synthesized, and their anticancer, antioxidant, and 11ß-hydroxysteroid dehydrogenase (11ß-HSD) inhibitory activities were tested. Anticancer activity has been assessed using the MTS (MTS: 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) assay against human colon carcinoma (Caco-2), human pancreatic carcinoma (PANC-1), glioma (U-118 MG), human breast carcinoma (MDA-MB-231), and skin melanoma (SK-MEL-30) cancer cell lines. Cell viability reductions, especially in the case of Caco-2, MDA-MB-231, and SK-MEL-30 lines, were observed for most compounds. In addition, the redox status was investigated and oxidative, but nitrosative stress was not noted at a concentration of 500 µM compounds tested. At the same time, a low level of reduced glutathione was observed in all cell lines when treated with compound 3g (5-(4-bromophenyl)-2-(cyclopentylamino)thiazol-4(5H)-one) that most inhibited tumor cell proliferation. However, the most interesting results were obtained in the study of inhibitory activity towards two 11ß-HSD isoforms. Many compounds at a concentration of 10 µM showed significant inhibitory activity against 11ß-HSD1 (11ß-hydroxysteroid dehydrogenase type 1). The compound 3h (2-(cyclopentylamino)-1-thia-3-azaspiro[4.5]dec-2-en-4-one) showed the strongest 11ß-HSD1 inhibitory effect (IC50 = 0.07 µM) and was more selective than carbenoxolone. Therefore, it was selected as a candidate for further research.

A comprehensive pharmacological analysis of fenoterol and its derivatives to unravel the role of ß2-adrenergic receptor in zebrafish.

ß-adrenergic receptors (ßARs) belong to a key molecular targets that regulate the most important processes occurring in the human organism. Although over the last decades a zebrafish model has been developed as a model complementary to rodents in biomedical research, the role of ß2AR in regulation of pathological and toxicological effects remains to elucidate. Therefore, the study aimed to clarify the role of ß2AR with a particular emphasis on the distinct role of subtypes A and B of zebrafish ß2AR. As model compounds selective ß2AR agonists - (R,R)-fenoterol ((R,R)-Fen) and its new derivatives: (R,R)-4'-methoxyfenoterol ((R,R)-MFen) and (R,R)-4'-methoxy-1-naphtylfenoterol ((R,R)-MNFen) - were tested. We described dose-dependent changes observed after fenoterols exposure in terms of general toxicity, cardiotoxicity and neurobehavioural responses. Subsequently, to better characterise the role of ß2-adrenergic stimulation in zebrafish, we have performed a series of molecular docking simulations. Our results indicate that (R,R)-Fen displays the highest affinity for subtype A of zebrafish ß2AR and ß2AAR might be involved in pigment depletion. (R,R)-MFen shows the lowest affinity for zebrafish ß2ARs out of the tested fenoterols and this might be associated with its cardiotoxic and anxiogenic effects. (R,R)-MNFen displays the highest affinity for subtype B of zebrafish ß2AR and modulation of this receptor might be associated with the development of malformations, increases locomotor activity and induces a negative chronotropic effect. Taken together, the presented data offer insights into the functional responses of the zebrafish ß2ARs confirming their intraspecies conservation, and support the translation of the zebrafish model in pharmacological and toxicological research.

Identification of the first-in-class dual inhibitors of human DNA topoisomerase IIα and indoleamine-2,3-dioxygenase 1 (IDO 1) with strong anticancer properties.

Molecular docking of a large set of thiosemicarbazide-based ligands resulted in obtaining compounds that inhibited both human DNA topoisomerase IIα and indoleamine-2,3-dioxygenase-1 (IDO1). To the best of our knowledge, these compounds are the first dual inhibitors targeting these two enzymes. As both of them participate in the anticancer response, the effect of the compounds on a panel of cancer cell lines was examined. Among the cell lines tested, lung cancer (A549) and melanoma (A375) cells were the most sensitive to compounds 1 (IC50=0.23 µg/ml), 2 (IC50=0.83 µg/ml) and 3 (IC50=0.25 µg/ml). The observed activity was even 90-fold higher than that of etoposide, with selectivity index values reaching 125. In-silico simulations showed that contact between 1-3 and human DNA topoisomerase II was maintained through aromatic moieties located at limiting edges of ligand molecules and intensive interactions of the thiosemicarbazide core with the DNA fragments present in the catalytic site of the enzyme.

Cyclodextrin Derivatives as Promising Solubilizers to Enhance the Biological Activity of Rosmarinic Acid.

Rosmarinic acid (RA) is a natural antioxidant with neuroprotective properties; however, its preventive and therapeutic use is limited due to its slight solubility and poor permeability. This study aimed to improve RA physicochemical properties by systems formation with cyclodextrins (CDs): hydroxypropyl-α-CD (HP-α-CD), HP-ß-CD, and HP-γ-CD, which were prepared by the solvent evaporation (s.e.) method. The interactions between components were determined by X-ray powder diffraction (XRPD), differential scanning calorimetry (DSC) and Fourier Transform infrared spectroscopy (FTIR). The sites of interaction between RA and CDs were suggested as a result of in silico studies focused on assessing the interaction between molecules. The impact of amorphous systems formation on water solubility, dissolution rate, gastrointestinal (GIT) permeability, and biological activity was studied. RA solubility was increased from 5.869 mg/mL to 113.027 mg/mL, 179.840 mg/mL, and 194.354 mg/mL by systems formation with HP-α-CD, HP-ß-CD, and HP-γ-CD, respectively. During apparent solubility studies, the systems provided an acceleration of RA dissolution. Poor RA GIT permeability at pH 4.5 and 5.8, determined by parallel artificial membrane permeability assay (PAMPA system), was increased; RA-HP-γ-CD s.e. indicated the greatest improvement (at pH 4.5 from Papp 6.901 × 10-7 cm/s to 1.085 × 10-6 cm/s and at pH 5.8 from 5.019 × 10-7 cm/s to 9.680 × 10-7 cm/s). Antioxidant activity, which was determined by DPPH, ABTS, CUPRAC, and FRAP methods, was ameliorated by systems; the greatest results were obtained for RA-HP-γ-CD s.e. The inhibition of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) was increased from 36.876% for AChE and 13.68% for BChE to a maximum inhibition of the enzyme (plateau), and enabled reaching IC50 values for both enzymes by all systems. CDs are efficient excipients for improving RA physicochemical and biological properties. HP-γ-CD was the greatest one with potential for future food or dietary supplement applications.

The Val34Met, Thr164Ile and Ser220Cys Polymorphisms of the ß2-Adrenergic Receptor and Their Consequences on the Receptor Conformational Features: A Molecular Dynamics Simulation Study.

The gene encoding the ß2-adrenergic receptor (ß2-AR) is polymorphic, which results in possible differences in a primary structure of this protein. It has been shown that certain types of polymorphisms are correlated with some clinical features of asthma, including airways reactivity, whereas the influence of other is not yet understood. Among polymorphisms affecting amino acids at positions 16, 27, 34, 164 and 220, the latter three are present in the crystal structure of ß2-AR, which facilitates studying them by means of molecular dynamics simulations. The current study was focused on investigating to what extent the three polymorphisms of ß2-AR (i.e., Val34Met, Thr164Ile and Ser220Cys) affect the interaction of ß2-AR with its natural molecular environment which includes: lipid bilayer (in the case of all three polymorphs) and Gs protein (which participates in ß2-AR-mediated signaling; in the case of Ser220Cys). We have designed and carried out a series of molecular dynamics simulations at different level of resolution (i.e., either coarse-grained or atomistic simulations), accompanied by thermodynamic integration protocol, in order to identify potential polymorphism-induced alterations in structural, conformational or energetic features of ß2-AR. The results indicate the lack of significant differences in the case of energies involved in the ß2-AR-lipid bilayer interactions. Some differences have been observed when considering the polymorphism-induced alterations in ß2-AR-Gs protein binding, but their magnitude is also negligible in relation to the absolute free energy difference correlated with the ß2-AR-Gs affinity. The Val34Met and Thr164Ile polymorphisms are weakly correlated with alteration of the conformational features of the receptor around polymorphic sites. On the contrary, it has been concluded that the Ser220Cys polymorphism is correlated with several structural alterations located in the intracellular region of ß2-AR, which can induce G-protein binding and, subsequently, the polymorphism-correlated therapeutic responses. More precisely, these alterations involve vicinity of intracellular loops and, in part, are the direct consequence of disturbed interactions of Ser/Cys220 sidechain within 5th transmembrane domain. Structurally, the dynamic structure exhibited by the ß2-ARSer220 polymorph is closer to the Gs-compatible structure of ß2-AR.

Amorphous Inclusion Complexes: Molecular Interactions of Hesperidin and Hesperetin with HP-Β-CD and Their Biological Effects.

This study aimed at obtaining hesperidin (Hed) and hesperetin (Het) systems with HP-ß-CD by means of the solvent evaporation method. The produced systems were identified using infrared spectroscopy (FT-IR), X-ray powder diffraction (XRPD), and differential scanning calorimetry (DSC). Moreover, in silico docking and molecular dynamics studies were performed to assess the most preferable site of interactions between tested compounds and HP-ß-CD. The changes of physicochemical properties (solubility, dissolution rate, and permeability) were determined chromatographically. The impact of modification on biological activity was tested in an antioxidant study as well as with regards to inhibition of enzymes important in pathogenesis of neurodegenerative diseases. The results indicated improvement in solubility over 1000 and 2000 times for Hed and Het, respectively. Permeability studies revealed that Hed has difficulties in crossing biological membranes, in contrast with Het, which can be considered to be well absorbed. The improved physicochemical properties influenced the biological activity in a positive manner by the increase in inhibitory activity on the DPPH radical and cholinoesterases. To conclude the use of HP-ß-CD as a carrier in the formation of an amorphous inclusion complex seems to be a promising approach to improve the biological activity and bioavailability of Hed and Het.

Pharmacological assessment of zebrafish-based cardiotoxicity models.

Cardiotoxicity remains the most common reason for failure during drug development. Recently, the zebrafish (Danio rerio) model has emerged for the evaluation of drug-dependent cardiotoxicity and for the identification of cardioprotective molecules. However, it remains unknown how closely the zebrafish-based results may be translated to humans. To tackle this issue, we established embryonic zebrafish models of doxorubicin-, adrenaline- and terfenadine-induced cardiotoxicity with unified dosing regimen which eventually enabled head-to-head comparison of the drugs. Subsequently, we determined whether human cardioprotective medications - dexrazoxane, metoprolol, carvedilol and valsartan - are able to manage heart dysfunction in zebrafish. Our results indicated that doxorubicin, adrenaline and terfenadine elicited overt signs of cardiotoxicity in fish, and we further showed that the blockade of the renin-angiotensin system and, to a lesser extent, ß-adrenergic system, ameliorated the heart disease in zebrafish. From the drug development standpoint, our work opens the possibility to determine the cardiovascular properties of tested compounds using the rapid and affordable zebrafish model.

The Systems of Naringenin with Solubilizers Expand Its Capability to Prevent Neurodegenerative Diseases.

BACKGROUND: Naringenin (NAR) is a flavonoid with excellent antioxidant and neuroprotective potential that is limited by its low solubility. Thus, solid dispersions with ß-cyclodextrin (ß-CD), hydroxypropyl-ß-cyclodextrin (HP-ß-CD), hydroxypropylmethylcellulose (HPMC), and microenvironmental pH modifiers were prepared. METHODS: The systems formation analysis was performed by X-Ray Powder Diffraction (XRPD) and Fourier-transform infrared spectroscopy (FT-IR). Water solubility and dissolution rates were studied with a pH of 1.2 and 6.8. In vitro permeability through the gastrointestinal tract (GIT) and the blood-brain barrier (BBB) was assessed with the parallel artificial membrane permeability assay (PAMPA) assay. The antioxidant activity was studied with the 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid (ABTS) and cupric ion reducing antioxidant capacity (CUPRAC) assays, while in vitro enzymes studies involved the inhibition of acetylcholinesterase, butyrylcholinesterase, and tyrosinase. For the most promising system, in silico studies were conducted. RESULTS: NAR solubility was increased 458-fold by the solid dispersion NAR:HP-ß-CD:NaHCO3 in a mass ratio of 1:3:1. The dissolution rate was elevated from 8.216% to 88.712% in a pH of 1.2 and from 11.644% to 88.843% in a pH of 6.8 (within 3 h). NAR GIT permeability, described as the apparent permeability coefficient, was increased from 2.789 × 10-6 cm s-1 to 2.909 × 10-5 cm s-1 in an acidic pH and from 1.197 × 10-6 cm s-1 to 2.145 × 10-5 cm s-1 in a basic pH. NAR BBB permeability was established as 4.275 × 10-6 cm s-1. The antioxidant activity and enzyme inhibition were also increased. Computational studies confirmed NAR:HP-ß-CD inclusion complex formation. CONCLUSIONS: A significant improvement in NAR solubility was associated with an increase in its biological activity.

Evaluation of ß-adrenergic ligands for development of pharmacological heart failure and transparency models in zebrafish.

Cardiovascular toxicity represents one of the most common reasons for clinical trial failure. Consequently, early identification of novel cardioprotective strategies could prevent the later-stage drug-induced cardiac side effects. The use of zebrafish (Danio rerio) in preclinical studies has greatly increased. High-throughput and low-cost of assays make zebrafish model ideal for initial drug discovery. A common strategy to induce heart failure is a chronic ß-adrenergic (ßAR) stimulation. Herein, we set out to test a panel of ßAR agonists to develop a pharmacological heart failure model in zebrafish. We assessed ßAR agonists with respect to the elicited mortality, changes in heart rate, and morphological alterations in zebrafish larvae according to Fish Embryo Acute Toxicity Test. Among the tested ßAR agonists, epinephrine elicited the most potent onset of heart stimulation (EC50 = 0.05 mM), which corresponds with its physiological role as catecholamine. However, when used at ten-fold higher dose (0.5 mM), the same compound caused severe heart rate inhibition (-28.70 beats/min), which can be attributed to its cardiotoxicity. Further studies revealed that isoetharine abolished body pigmentation at the sublethal dose of 7.50 mM. Additionally, as a proof of concept that zebrafish can mimic human cardiac physiology, we tested ßAR antagonists (propranolol, carvedilol, metoprolol, and labetalol) and verified that they inhibited fish heart rate in a similar fashion as in humans. In conclusion, we proposed two novel pharmacological models in zebrafish; i.e., epinephrine-dependent heart failure and isoetharine-dependent transparent zebrafish. We provided strong evidence that the zebrafish model constitutes a valuable tool for cardiovascular research.

Molecular Structure of Cefuroxime Axetil Complexes with α-, ß-, γ-, and 2-Hydroxypropyl-ß-Cyclodextrins: Molecular Simulations and Raman Spectroscopic and Imaging Studies.

The formation of cefuroxime axetil+cyclodextrin (CA+CD) complexes increases the aqueous solubility of CA, improves its physico-chemical properties, and facilitates a biomembrane-mediated drug delivery process. In CD-based tablet formulations, it is crucial to investigate the molecular details of complexes in final pharmaceutical preparation. In this study, Raman spectroscopy and mapping were applied for the detection and identification of chemical groups involved in α-, ß-, γ-, and 2-hydroxypropyl-ß-CD (2-HP- ß-CD)+CA complexation process. The experimental studies have been complemented by molecular dynamics-based investigations, providing additional molecular details of CA+CD interactions. It has been demonstrated that CA forms the guest-host type inclusion complexes with all studied CDs; however, the nature of the interactions is slightly different. It seems that both α- and ß-CD interact with furanyl and methoxy moieties of CA, γ-CD forms a more diverse pattern of interactions with CA, which are not observed in other CDs, whereas 2HP-ß-CD binds CA with the contribution of hydrogen bonding. Apart from supporting this interpretation of the experimental data, molecular dynamics simulations allowed for ordering the CA+CD binding affinities. The obtained results proved that the molecular details of the host-guest complexation can be successfully predicted from the combination of Raman spectroscopy and molecular modeling.

Combinations of Piperine with Hydroxypropyl-ß-Cyclodextrin as a Multifunctional System.

Piperine is an alkaloid that has extensive pharmacological activity and impacts other active substances bioavailability due to inhibition of CYP450 enzymes, stimulation of amino acid transporters and P-glycoprotein inhibition. Low solubility and the associated low bioavailability of piperine limit its potential. The combination of piperine with 2-hydroxypropyl-ß-cyclodextrin (HP-ß-CD) causes a significant increase in its solubility and, consequently, an increase in permeability through gastrointestinal tract membranes and the blood-brain barrier. X-ray powder diffraction (XRPD), differential scanning calorimetry (DSC), Fourier-transform infrared spectroscopy (FT-IR), nuclear magnetic resonance (NMR) were used to characterize interactions between piperine and HP-ß-CD. The observed physicochemical changes should be combined with the process of piperine and CD system formation. Importantly, with an increase in solubility and permeability of piperine as a result of interaction with CD, it was proven to maintain its biological activity concerning the antioxidant potential (2,2-diphenyl-1-picryl-hydrazyl-hydrate assay), inhibition of enzymes essential for the inflammatory process and for neurodegenerative changes (hyaluronidase, acetylcholinesterase, butyrylcholinesterase).

Comparison of Carbohydrate Force Fields in Molecular Dynamics Simulations of Protein-Carbohydrate Complexes.

In this paper, we present the results of molecular dynamics simulations aimed at critical comparison of classical, biomolecular force fields (FFs) in the context of their capabilities to describe the structural and thermodynamic features of carbohydrate-protein interactions. We have considered the three main families of FFs (CHARMM, GROMOS, and GLYCAM/AMBER) by applying them to investigate the seven different carbohydrate-protein complexes. The results indicate that although the qualitative pattern of several structural descriptors (intermolecular hydrogen bonding, ligand dynamic location, etc.) is conserved among the compared FFs, there also exists a number of significant divergences (mainly the patterns of contacts between particular amino acid residues and bound carbohydrate). The carbohydrate-protein unbinding free energies also vary from one FF to another, displaying diversified trends in deviations from the experimental data. The magnitude of those deviations is not negligible and indicates the need for refinement in the currently existing combinations of carbohydrate- and protein-dedicated biomolecular force fields. In spite of the lack of explicit functional terms responsible for the corresponding intermolecular forces, all tested FFs are capable of adequately reproducing the CH-π interactions, crucial for carbohydrate-protein binding.

Structural Insights into Ligand-Receptor Interactions Involved in Biased Agonism of G-Protein Coupled Receptors.

G protein-coupled receptors (GPCRs) are versatile signaling proteins that mediate complex cellular responses to hormones and neurotransmitters. Ligand directed signaling is observed when agonists, upon binding to the same receptor, trigger significantly different configuration of intracellular events. The current work reviews the structurally defined ligand - receptor interactions that can be related to specific molecular mechanisms of ligand directed signaling across different receptors belonging to class A of GPCRs. Recent advances in GPCR structural biology allow for mapping receptors' binding sites with residues particularly important in recognition of ligands' structural features that are responsible for biased signaling. Various studies show particular role of specific residues lining the extended ligand binding domains, biased agonists may alternatively affect their interhelical interactions and flexibility what can be translated into intracellular loop rearrangements. Studies on opioid and angiotensin receptors indicate importance of residues located deeper within the binding cavity and direct interactions with receptor residues linking the ortosteric ligand binding site with the intracellular transducer binding domain. Collection of results across different receptors may suggest elements of common molecular mechanisms which are responsible for passing alternative signals from biased agonists.

Tedizolid-Cyclodextrin System as Delayed-Release Drug Delivery with Antibacterial Activity.

Progressive increase in bacterial resistance has caused an urgent need to introduce new antibiotics, one of them being oxazolidinones with their representative tedizolid. Despite the broad spectrum of activity of the parent tedizolid, it is characterized by low water solubility, which limits its use. The combination of the active molecule with a multifunctional excipient, which is cyclodextrins, allows preservation of its pharmacological activity and modification of its physicochemical properties. Therefore, the aim of the study was to change the dissolution rate and permeability through the model membrane of tedizolid by formation of solid dispersions with a cyclodextrin. The research included identification of tedizolid-hydroxypropyl-ß-cyclodextrin (tedizolid/HP-ß-CD) inclusion complex by thermal method (Differential Scanning Colorimetry), spectroscopic methods (powder X-ray diffraction, Fourier-Transform Infrared spectroscopy), and molecular docking. The second part of the research concerned the physicochemical properties (dissolution and permeability) and the biological properties of the system in terms of its microbiological activity. An increase in the dissolution rate was observed in the presence of cyclodextrin, while maintaining a high permeation coefficient and high microbiological activity. The proposed approach is an opportunity to develop drug delivery systems used in the treatment of resistant bacterial infections, in which, in addition to modifying the physicochemical properties caused by cyclodextrin, we observe a favorable change in the pharmacological potential of the bioactives.

Chirality Effects in Biomolecular Systems: Calculation of the Relative Free Energies by Molecular Dynamics Simulations.

Chirality plays an essential role in chemical and biological sciences. At the molecular level, the effects associated with this phenomenon can be studied by using the well-established technique of molecular dynamics simulations. In this work, we present several approaches suited for the molecular dynamics-based free energy calculation in chiral systems. In particular, we have proposed and tested the following strategies relying on the application of general, enhanced sampling methods: (i) biased sampling in the two-dimensional space, along the coordinates defined by the values of the selected torsional angles; (ii) biased sampling in the one- or two-dimensional space, along the path-based coordinate(s); (iii) rational alteration of the system's Hamiltonian in order to enable the interconversion between stereoisomers and reweighting the biased distribution of configurations; (iv) using the free energy landscape generated within approaches (i) or (ii) as time-independent bias in order to further improve sampling efficiency and simultaneously account for multiple chiral centers. All approaches have been tested on a set of model compounds (fenoterol, fructofuranose, and bromochlorofluoromethane), demonstrating the good performance but also some differences in the range of their applicabilities.

Efficient sampling of high-energy states by machine learning force fields.

Regarding their application in the field of molecular sciences, machine learning (ML) methods are capable of combining the high accuracy of ab initio potentials with an efficiency closer to that of classical molecular mechanics. By relying on the reference data (e.g., atomic configurations and corresponding energies), the ML algorithms can reconstruct the potential energy surface for simple molecular systems, which may subsequently serve as a computationally inexpensive force field. The accuracy of such an ML force field is highly dependent on the character of the dataset that was used for its training. In this work, we show that omitting the high-energy states, which results from following the Boltzmann distribution, may lead to a catastrophic loss of accuracy in certain regions of the configurational phase space. To overcome this challenge, we have proposed an alternative solution for generating the ML input data. The most essential step is the biased subsampling of the configurations, aimed at increasing the population of hardly accessible states, usually located on energy barriers. The applicability of the proposed procedure is demonstrated on the example of conformational rearrangements in the two flexible, heterocyclic molecules. This approach provides an essential component required to obtain the ML force fields, accurate within the whole configurational phase space of the system.

Thermodynamic study of new antiepileptic compounds by combining chromatography on the phosphatidylcholine biomimetic stationary phase and differential scanning calorimetry.

Liquid chromatography coupled to spectrophotometric detection of new antiepileptic compounds, 1,2,4-triazole-3-thione derivatives, on immobilized artificial membrane phosphatidylcholine is reported. The curves representing the relationship between ln k versus 1/T generated under isocratic conditions by the use of methanol and acetonitrile-containing eluent systems have been constructed in order to determine the thermodynamic parameters: the enthalpies, entropies and the relative free energies. The hydrocarbon chains of analytes significantly influenced the membrane behavior of the whole molecules. Excellent correlations of the theoretical lipophilicity with the experimental thermodynamic descriptors, have confirmed contribution of the hydrophobic interactions in the retention process. However, presence of sulfur or oxygen as heteroatoms at R1 substituents in the 1,2,4-triazole ring appears to be responsible for more pronounced selectivity of these compounds on the phosphatidylcholine stationary phase. Molecular dynamics simulations revealed the selective preferences of the phosphatidylcholine with respect to the compounds with either ether of sulfide moieties. Experimental and theoretical set-ups resulted in corresponding outcomes.

Development of the 1,2,4-triazole-based anticonvulsant drug candidates acting on the voltage-gated sodium channels. Insights from in-vivo, in-vitro, and in-silico studies.

The treatment of epilepsy remains difficult mostly since almost 30% of patients suffer from pharmacoresistant forms of the disease. Therefore, there is an urgent need to search for new antiepileptic drug candidates. Previously, it has been shown that 4-alkyl-5-substituted-1,2,4-triazole-3-thione derivativatives possessed strong anticonvulsant activity in a maximal electroshock-induced seizure model of epilepsy. In this work, we examined the effect of the chemical structure of the 1,2,4-triazole-3-thione-based molecules on the anticonvulsant activity and the binding to voltage-gated sodium channels (VGSCs) and GABAA receptors. Docking simulations allowed us to determine the mode of interactions between the investigated compounds and binding cavity of the human VGSC. Selected compounds were also investigated in a panel of ADME-Tox assays, including parallel artificial membrane permeability assay (PAMPA), single cell gel electrophoresis (SCGE) and cytotoxicity evaluation in HepG2 cells. The obtained results indicated that unbranched alkyl chains, from butyl to hexyl, attached to 1,2,4-triazole core are essential both for good anticonvulsant activity and strong interactions with VGSCs. The combined in-vivo, in-vitro and in-silico studies emphasize 4-alkyl-5-substituted-1,2,4-triazole-3-thiones as promising agents in the development of new anticonvulsants.