Search

An aminoquinoline (AQ4) was developed and proven to be a new and efficient DNA minor groove fluorescent probe. The specificity for DNA minor groove was attested by comparing it with well-established DNA probes such as Hoechst stain, acridine orange, and ethidium bromide. AQ4 was similar to the Hoechst stain, a classic minor groove probe, and opposite to acridine orange and ethidium bromide, the typical intercalating probes. An advantage of AQ4 to the Hoechst stain was the higher fluorescent signal-to-noise ratio (+DNA/-DNA). The interaction with DNA leads to an exclusive fluorescent band centered at 590 nm. The red-shifted fluorescent band is associated with a new absorption band (490 nm), revealing a ground-state complex formation. The complexation was also evidenced by circular dichroism, anisotropy, and fluorescence lifetime. The complex AQ4-DNA was pH dependent, being favored in an acidic medium. This feature was related to the preferential interaction with a protonated form of AQ4, i.e., AQ4(H+), revealing the role of electrostatic forces, which was corroborated by the strong dependence on the ionic strength of the medium and, particularly, on magnesium ions. The complexes were studied by docking and molecular dynamics and confirmed the stability of AQ4(H+). Additionally, 500 ns simulations were performed by adding salts. The presence of the salts leads to the loss of the binding of AQ4(H+) after 52 ns (NaCl) and 142 ns (MgCl2). Experimental and in silico outcomes showed the advantages of the aminoquinoline over the commercial DNA minor groove stain, the Hoechst dye. Hence, we propose its further application in cell-based assays.

Patients with COVID-19 may experience multiple conditions (e.g., fever, hyperventilation, anorexia, gastroenteritis, acid-base disorder) that may cause electrolyte imbalances. Hypokalemia is a concerning electrolyte disorder that may increase the susceptibility to various kinds of arrhythmia. This study aimed to estimate prevalence, risk factors and outcome of hypokalemia in a cohort of non-critically ill patients. A retrospective analysis was conducted on 290 hospitalized patients with confirmed COVID-19 infection at the tertiary teaching hospital of Modena, Italy.Hypokalemia (Hypokalemia is a frequent disorder in COVID-19 patients and urinary potassium loss may be the main cause of hypokalemia. The disorder was mild in the majority of the patients and was unrelated to poor outcomes. Nevertheless, hypokalemic patients required potassium supplements to dampen the risk of arrhythmias.

Cyclin-Dependent Kinases-2 (CDK2) are members of the serine/threonine protein kinases family. They play an important role in the regulation events of the eukaryotic cell division cycle, especially during the G1 to S phase transition. Experimental evidence indicates that excessive expression of CDK2s should cause abnormal cell cycle regulation. Therefore, since a long time, CDK2s have been considered potential therapeutic targets for cancer therapy. In this work, we collected one-hundred and forty-nine complexes of inhibitors bound in the CDK2-ATP pocket submitting to short MD simulations (10ns) and free energy calculation by means of MM-GBSA. The calculate ∆G values have been compared with experimental data (Ki, Kd, and IC50). Information collected on short MD simulations of protein-ligand complexes has been used to perform molecular modeling approaches that incorporates flexibility into structure-based pharmacophore modeling (Common Hits Approach, CHA,1 and Molecular dYnamics SHAred PharmacophorE, MYSHAPE2 approach) and constraints docking, to enrich the hits list of virtual screening. Short simulations proved to be exhaustive to examine the crucial ligand-protein interactions within the complexes.

Optimization of a new lead promoting the readthrough of the nonsense mutations for CFTR rescue in human CF cells Laura Lentini, Raffaella Melfi, Sara Baldassano, Marco Tutone, Aldo Di Leonardo, Andrea Pace, Ivana Pibiri Department of Biological, Chemical and Pharmaceutical Sciences and Technologies (STEBICEF), University of Palermo Background and rationale Cystic Fibrosis patients with nonsense mutations in the CFTR gene have a more severe form of the disease. Nonsense mutations represent about 10% of the mutations that affect the CFTR gene and they are frequently associated to the classical F508 mutation (1). A potential treatment for this genetic alteration is to promote the translational readthrough of premature termination codons (PTCs) by Translational Read-Through-Inducing Drugs (TRIDs) (2-4). Hypothesis and objectives Our objective is to evaluate the functionality of the CFTR channel after treatment with a new molecule that we individuated in a precedent FFC project, and the activity of new lead molecules in cells stably expressing a nonsense-CFTR-mRNA (ns CFTR) in CF cellular model systems. We want also to study the supramolecular interactions among TRIDs, CFTR mRNA and the ribosomal A-site to identify the biological target and the mechanism of action. Essential methods QSAR, carried out on the basis of our preliminary results, will allow to achieve lead optimization and synthesize then a small library of analogs to be tested and compared to the Lead. We will mutagenize the CFTR cDNA by introducing the most diffuse nonsense mutations. Subsequently, FRT cells engineered with the vector expressing mutagenized nsCFTR, and nonsense-CF-human broncoepithelial cells will be grown in the air-liquid culture system to reproduce in vitro the epithelial organization. CFTR expression after treatments with our molecule will be evaluated by biomolecular techniques. CFTR activity will be revealed by specific CFTR-functionality assays. Finally, in vitro-in vivo (Zebrafish model) analyses of the safety profile for the set of synthesized molecules will complete the study. Preliminary results We screened the activity of several molecules synthetized by us in a precedent FFC project, identifying some molecules that showed high readthrough activity associated to the expression of the CFTR protein in ns CF immortalized cells. Expected final results and their significance We are confident that our findings will provide the validation of molecules with readthrough activity for the recovery of the CFTR function. Moreover, our pre-clinical study will assess the presence of toxic effects caused by the molecules in vivo. References 1. Sermet-Gaudelus I, Boeck KD, Casimir GJ, Vermeulen F, Leal T, Mogenet A, Roussel D, Fritsch J, Hanssens L, Hirawat S, Miller NL, Constantine S, Reha A, Ajayi T, Elfring GL, Miller LL. Ataluren (PTC124) induces cystic fibrosis transmembrane conductance regulator protein expression and activity in children with nonsense mutation cystic fibrosis., Am J RespirCrit Care Med. 2010 Nov 15;182(10):1262-72. 2. Lentini L, Melfi R, Di Leonardo A, Spinello A, Barone G, Pace A, Palumbo Piccionello A, Pibiri I. Towards a rationale for the PTC124 (Ataluren) promoted read-through of premature stop codons: a computational approach and GFP-reporter cell-based assay. Mol. Pharm. 2014 11, 653-664. 3. Pibiri I, Lentini L, Melfi R, Gallucci G, Pace A, Spinello A, Barone G, Di Leonardo A. Enhancement of premature stop codon readthrough in the CFTR gene by Ataluren (PTC124) derivatives European Journal of Medicinal Chemistry 06/2015; 101. 4. Nagel-Wolfrum K, Möller F, Penner I, Baasov T4, Wolfrum U. Targeting Nonsense Mutations in Diseases with Translational Read-Through-Inducing Drugs (TRIDs), BioDrugs. 2016 Apr;30(2):49-74.

Adenosine receptors (AR) belong to the superfamily of G-protein-coupled receptors (GPCRs). They are divided into four subtypes (A1, A2A, A2B, and A3) [1], and can be distinguished on the basis of their distinct molecular structures, distinct tissues distribution, and selectivity for adenosine analogs [2,3]. The hA3R, the most recently identified adenosine receptor, is involved in a variety of intracellular signaling pathways and physiological functions [4]. Expression of A3R was reported to be elevated in cancerous tissues [5], and A3 antagonists have been proposed for therapeutic treatments of cancer. The recent literature availability of crystal structure of hA2A adenosine receptor (PDB code: 3EML) provided us a new template for A3R homology modeling. The validation of the obtained structure model was performed by inspecting the Ramachandran plot (Fig. 1). The modeled protein was optimized using nanosecond scale molecular dynamics simulation. One hundred twenty two active and selective compounds were docked into the obtained model using Induced Fit Docking [6] and used as training set to generate pharmacophore models by means PHASE [7]. Energy-optimized pharmacophore mapping was performed; to each pharmacophore feature site was assigned an energetic value as the sum of the GLIDE XP contributions of the atoms included in the site. This pharmacophore model addresses the prevalent features to be used for the search of new inhibitors. Therefore it was employed as template to screen the ZINC database in the attempt to find new potent and selective human A3R antagonists. [1] B.B. Fredholm, A.P. Ijzerman, K.A. Jacobson, K.N. Klotz, J. Linden, Pharmacol. Rev., 53, 2001, 527. [2] P.G. Baraldi, R. Romagnoli, D. Preti, F. Fruttarolo, M.D. Carrion, M.A. Tabrizi, Curr. Med. Chem., 13, 2006, 3467. [3] M.D.Okusa, Am. J. Physiol. Renal Physiol., 282, 2002, F10. [4] A.Ochaion, S. Bar Yehuda, S. Cohen, F. Barer, R. Patoka, L. Del-Valle, G. Perez-Liz, J. Ophir, R. Galili-Mosberg, T. Reitblat, H. Amital, P. Fishman, Ann. Rheum. Dis., 66, 2007, 446. [5] L.Madi, A. Ochaion, L. Rath-Wolfson, S. Bar-Yehuda, A. Erlanger, G. Ohana, A. Harish, O. Merimski, F. Barer, P. Fishman, Clin. Cancer Res., 10, 2004, 4472. [6] Schrödinger Suite 2009 Induced Fit Docking protocol; Glide version 5.5, Schrödinger, LLC, New York, NY, 2009; Prime version 2.1, Schrödinger, LLC, New York, NY, 2009. [7] Phase, version 3.1, Schrödinger, LLC, New York, NY, 2009.

Cyclin Dependent Kinases-2 (CDK2) are members of serine/threonine protein kinases family. They play an important role in the regulation events of the eukaryotic cell division cycle, especially during the G1 to S phase transition. Experimental evidence indicate that excessive expression of CDK2s should cause abnormal cell cycle regulation. Therefore, since a long time, CDK2s have been considered potential therapeutic targets for cancer therapy. In this work, onehundred and forty-nine complexes of inhibitors bound in the CDK2-ATP pocket were submitted to short MD simulations (10ns) and free energy calculation. Comparison with experimental data (Ki, Kd and pIC50) revealed that short simulations are exhaustive to examine the crucial ligand-protein interactions within the complexes. Information collected on MD simulations of protein-ligand complexes has been used to perform a molecular modelling approach that incorporates flexibility into structure-based pharmacophore modelling (Common Hits Approach, CHA). The high number of pharmacophore models resulting from the MD simulation was thus reduced to a few representative groups of pharmacophore models. The performance of the models has been assessed by using the ROC curves analysis. This definitive set of validated pharmacophore models could be used to screen in-house and/or commercial datasets for detection of new CDK-2 inhibitors. We provide the models to all the researchers involved in this field.

Background: Indicaxanthin, a betaxanthin belonging to the betalain class of compounds, has been recently demonstrated to exert significant antiproliferative effects inducing apoptosis of human melanoma cells through the inhibition of NF-κB as the predominant pathway. Specifically, Indicaxanthin inhibited IκBα degradation in A375 cells. In resting cells, NF-κB is arrested in the cytoplasm by binding to its inhibitor protein IκBα. Upon stimulation, IκBα is phosphorylated by the IKK complex, and degraded by the proteasome, liberating free NF-κB into the nucleus to initiate target gene transcription. Inhibition of the IKK complex leads to the arrest of the NF-κB pathway.Methods: To acquire details at the molecular level of Indicaxanthin’s inhibitory activity against hIKKβ, molecular modeling and simulation techniques including induced-fit docking (IFD), binding pose metadynamics (BPMD), molecular dynamics simulations, and MM-GBSA (molecular mechanics-generalized Born surface area continuum solvation) have been performed.Results: The computational calculations performed on the active and inactive form, and the allosteric binding site of hIKKβ, revealed that Indicaxanthin inhibits prevalently the active form of the hIKKβ. MM-GBSA computations provide further evidence of Indicaxanthin’s stability inside the active binding pocket with a binding free energy of −22.2 ± 4.3 kcal/mol with respect to the inactive binding pocket with a binding free energy of −20.7 ± 4.7 kcal/mol. BPMD and MD simulation revealed that Indicaxanthin is likely not an allosteric inhibitor of hIKKβ.Conclusion: As a whole, these in silico pieces of evidence show that Indicaxanthin can inhibit the active form of the hIKKβ adding novel mechanistic insights on its recently discovered ability to impair NF-κB signaling in melanoma A375 cells. Moreover, our results suggest the phytochemical as a new lead compound for novel, more potent IKKβ inhibitors to be employed in the treatment of cancer and inflammation-related conditions.

Background The benefit of tocilizumab on mortality and time to recovery in people with severe COVID pneumonia may depend on appropriate timing. The objective was to estimate the impact of tocilizumab administration on switching respiratory support states, mortality and time to recovery. Methods In an observational study, a continuous-time Markov multi-state model was used to describe the sequence of respiratory support states including: no respiratory support (NRS), oxygen therapy (OT), non-invasive ventilation (NIV) or invasive mechanical ventilation (IMV), OT in recovery, NRS in recovery. Results Two hundred seventy-one consecutive adult patients were included in the analyses contributing to 695 transitions across states. The prevalence of patients in each respiratory support state was estimated with stack probability plots, comparing people treated with and without tocilizumab since the beginning of the OT state. A positive effect of tocilizumab on the probability of moving from the invasive and non-invasive mechanical NIV/IMV state to the OT in recovery state (HR = 2.6, 95% CI = 1.2–5.2) was observed. Furthermore, a reduced risk of death was observed in patients in NIV/IMV (HR = 0.3, 95% CI = 0.1–0.7) or in OT (HR = 0.1, 95% CI = 0.0–0.8) treated with tocilizumab. Conclusion To conclude, we were able to show the positive impact of tocilizumab used in different disease stages depicted by respiratory support states. The use of the multi-state Markov model allowed to harmonize the heterogeneous mortality and recovery endpoints and summarize results with stack probability plots. This approach could inform randomized clinical trials regarding tocilizumab, support disease management and hospital decision making.

To date, SARS-CoV-2 infectious disease, named COVID-19 by the World Health Organization (WHO) in February 2020, has caused millions of infections and hundreds of thousands of deaths. Despite the scientific community efforts, there are currently no approved therapies for treating this coronavirus infection. The process of new drug development is expensive and time-consuming, so that drug repurposing may be the ideal solution to fight the pandemic. In this paper, we selected the proteins encoded by SARS-CoV-2 and using homology modeling we identified the high-quality model of proteins. A structure-based pharmacophore modeling study was performed to identify the pharmacophore features for each target. The pharmacophore models were then used to perform a virtual screening against the DrugBank library (investigational, approved and experimental drugs). Potential inhibitors were identified for each target using XP docking and induced fit docking. MM-GBSA was also performed to better prioritize potential inhibitors. This study will provide new important comprehension of the crucial binding hot spots usable for further studies on COVID-19. Our results can be used to guide supervised virtual screening of large commercially available libraries.

Cystic fibrosis (CF) patients develop a severe form of the disease when the cystic fibrosis transmembrane conductance regulator (CFTR) gene is affected by nonsense mutations. Nonsense mutations are responsible for the presence of a premature termination codon (PTC) in the mRNA, creating a lack of functional protein. In this context, translational readthrough-inducing drugs (TRIDs) represent a promising approach to correct the basic defect caused by PTCs. By using computational optimization and biological screening, we identified three new small molecules showing high readthrough activity. The activity of these compounds has been verified by evaluating CFTR expression and functionality after treatment with the selected molecules in cells expressing nonsense&ndash, CFTR&ndash, mRNA. Additionally, the channel functionality was measured by the halide sensitive yellow fluorescent protein (YFP) quenching assay. All three of the new TRIDs displayed high readthrough activity and low toxicity and can be considered for further evaluation as a therapeutic approach toward the second major cause of CF.

[Image: see text] Translational readthrough-inducing drugs (TRIDs) rescue the functional full-length protein expression in genetic diseases, such as cystic fibrosis, caused by premature termination codons (PTCs). Small molecules have been developed as TRIDs to trick the ribosomal machinery during recognition of the PTC. Herein we report a computational study to identify new TRID scaffolds. A pharmacophore approach was carried out on compounds that showed readthrough activity. The pharmacophore model applied to screen different libraries containing more than 87000 compounds identified four hit-compounds presenting scaffolds with diversity from the oxadiazole lead. These compounds have been synthesized and tested using the Fluc reporter harboring the UGA PTC. Moreover, the cytotoxic effect and the expression of the CFTR protein were evaluated. These compounds, a benzimidazole derivative (NV2899), a benzoxazole derivative (NV2913), a thiazole derivative (NV2909), and a benzene-1,3-disulfonate derivative (NV2907), were shown to be potential new lead compounds as TRIDs, boosting further efforts to address the optimization of the chemical scaffolds.

We compared the performance of molecular dynamics (MD)-derived pharmacophore modeling approaches, Common Hit Approach (CHA), and the Molecular dYnamics SHAred PharmacophorE (MYSHAPE) approach, with semi-flexible constrained/unconstrained docking. The aim of this work is to enrich the hit-list of a virtual screening on CDK-2 known inhibitors as a case study. Cyclin-dependent kinases (CDKs) deregulation is associated with cancer growth. CDKs are an attractive target for anticancer agents. MD-derived pharmacophore models have been obtained with LigandScout 4.2.1. Docking analysis has been performed through Glide 7.6. The results highlighted the MYSHAPE approach has a better performance when multiple target-ligand complexes are available (ROC5% = 0.99). Moreover, the use of short molecular dynamics simulations improves the performance of the screening (ROC5% = 0.98–0.99) with respect to docking (ROC5% = 0.89–0.94). Outcomes of this work indicate that these approaches are highly suitable for prospective screening by a non-expert, and could be useful for the identification of novel CDK-2 inhibitors.

Molecular dynamics (MD) simulations can be used, prior to virtual screening, to add flexibility to proteins and study them in a dynamic way. Furthermore, the use of multiple crystal structures of the same protein containing different co-crystallized ligands can help elucidate the role of the ligand on a protein's active conformation, and then explore the most common interactions between small molecules and the receptor. In this work, we evaluated the contribution of the combined use of MD on crystal structures containing the same protein but different ligands to examine the crucial ligand-protein interactions within the complexes. The study was carried out on peroxisome proliferator-activated receptor α (PPARα). Findings derived from the dynamic analysis of interactions were then used as features for pharmacophore generation and constraints for generating the docking grid for use in virtual screening. We found that information derived from short multiple MD simulations using different molecules within the binding pocket of the target can improve the early enrichment of active ligands in the virtual screening process for this receptor. In the end we adopted a consensus scoring based on docking score and pharmacophore alignment to rank our dataset. Our results showed an improvement in virtual screening performance in early recognition when screening was performed with the Molecular dYnamics SHAred PharmacophorE (MYSHAPE) approach.

Not always lead compound and/or derivatives are suitable for the specific biological target for which they are designed but, in some cases, discarded compounds proved to be good binders for other biological targets; therefore, drug repurposing constitute a valid alternative to avoid waste of human and financial resources. Our virtual lock-and-key methods, VLKA and Conf-VLKA, furnish a strong support to predict the efficacy of a designed drug a priori its biological evaluation, or the correct biological target for a set of the selected compounds, allowing thus the repurposing of known and unknown, active and inactive compounds.

This review focuses on the use of oxadiazoles as translational readthrough-inducing drugs (TRIDs) to rescue the functional full-length protein expression in mendelian genetic diseases caused by nonsense mutations. These mutations in specific genes generate premature termination codons (PTCs) responsible for the translation of truncated proteins. After a brief introduction on nonsense mutations and their pathological effects, the features of various classes of TRIDs will be described discussing differences or similarities in their mechanisms of action. Strategies to correct the PTCs will be presented, particularly focusing on a new class of Ataluren-like oxadiazole derivatives in comparison to aminoglycosides. Additionally, recent results on the efficiency of new candidate TRIDs in restoring the production of the cystic fibrosis transmembrane regulator (CFTR) protein will be presented. Finally, a prospectus on complementary strategies to enhance the effect of TRIDs will be illustrated together with a conclusive paragraph about perspectives, opportunities, and caveats in developing small molecules as TRIDs.

The skin is the main barrier between the internal body environment and the external one. The characteristics of this barrier and its properties are able to modify and affect drug delivery and chemical toxicity parameters. Therefore, it is not surprising that permeability of many different compounds has been measured through several in vitro and in vivo techniques. Moreover, many different in silico approaches have been used to identify the correlation between the structure of the permeants and their permeability, to reproduce the skin behavior, and to predict the ability of specific chemicals to permeate this barrier. A significant number of issues, like interlaboratory variability, experimental conditions, data set building rationales, and skin site of origin and hydration, still prevent us from obtaining a definitive predictive skin permeability model. This review wants to show the main advances and the principal approaches in computational methods used to predict this property, to enlighten the main issues that have arisen, and to address the challenges to develop in future research.

Ataluren, also known as PTC124, is a 5-(fluorophenyl)-1,2,4-oxadiazolyl-benzoic acid suggested to suppress nonsense mutations by readthrough of premature stop codons in the mRNA. Potential interaction of PTC124 with mRNA has been recently studied by molecular dynamics simulations highlighting the importance of H-bonding and stacking π−π interactions. A series of non-acidic analogues of PTC124 were selected from a large database via a ligand-based virtual screening approach. Eight of them were synthesized and tested for their readthrough activity using the Fluc reporter harboring the UGA premature stop codon. The most active compound was further tested for suppression of the UGA nonsense mutation in the bronchial epithelial IB3.1 cell line carrying the W1282X mutation in the CFTR gene.

Background:Telomerase, a reverse transcriptase, maintains telomere and chromosomes integrity of dividing cells, while it is inactivated in most somatic cells. In tumor cells, telomerase is highly activated, and works in order to maintain the length of telomeres causing immortality, hence it could be considered as a potential marker to tumorigenesis.A series of 1,3,4-oxadiazole derivatives showed significant broad-spectrum anticancer activity against different cell lines, and demonstrated telomerase inhibition.Methods:This series of 24 N-benzylidene-2-((5-(pyridine-4-yl)-1,3,4-oxadiazol-2yl)thio)acetohydrazide derivatives as telomerase inhibitors has been considered to carry out QSAR studies. The endpoint to build QSAR models is determined by the IC50 values for telomerase inhibition, i.e., the concentration (μM) of inhibitor that produces 50% inhibition. These values were converted to pIC50 (- log IC50) values. We used the most common and transparent method, where models are described by clearly expressed mathematical equations: Multiple Linear Regression (MLR) by Ordinary Least Squares (OLS).Results:Validated models with high correlation coefficients were developed. The Multiple Linear Regression (MLR) models, by Ordinary Least Squares (OLS), showed good robustness and predictive capability, according to the Multi-Criteria Decision Making (MCDM = 0.8352), a technique that simultaneously enhances the performances of a certain number of criteria. The descriptors selected for the models, such as electrotopological state (E-state) descriptors, and extended topochemical atom (ETA) descriptors, showed the relevant chemical information contributing to the activity of these compounds.Conclusion:The results obtained in this study make sure about the identification of potential hits as prospective telomerase inhibitors.

Indicaxanthin is a bioactive and bioavailable betalain pigment from the Opuntia ficus-indica fruits. In this in vivo study, kinetic measurements showed that indicaxanthin is revealed in the rat brain within 1 h from oral administration of 2 μmol/ kg, an amount compatible with a dietary consumption of cactus pear fruits in humans. A peak (20 ± 2.4 ng of indicaxanthin per whole brain) was measured after 2.5 h; thereafter the molecule disappeared with first order kinetics within 4 h. The potential of indicaxanthin to affect neural activities was in vivo investigated by a microiontophoretic approach. Indicaxanthin, administered in a range between 0.085 ng and 0.34 ng per neuron, dose-dependently modulated the rate of discharge of spontaneously active neurons of the hippocampus, with reduction of the discharge and related changes of latency and duration of the effect. Indicaxanthin (0.34 ng/neuron) showed inhibitory effects on glutamate-induced excitation, indicating activity at the level of glutamatergic synapses. A molecular target of indicaxanthin is suggested by in silico molecular modeling of indicaxanthin with N- methyl-D-aspartate receptor (NMDAR), the most represented of the glutamate receptor family in hippocampus. Therefore, at nutritionally compatible amounts indicaxanthin (i) crosses the rat BBB and accumulates in brain; (ii) can affect the bioelectric activity of hippocampal neurons locally treated with amounts comparable with those measured in the brain; and (iii) modulates glutamate-induced neuronal excitation. The potential of dietary indicaxanthin as a natural neuromodulatory agent deserves further mechanistic and neurophysiologic investigation.

RATIONALE: The study of self-assembly processes of surfactant molecules in the gas phase is of great interest for several theoretical and technological reasons related to their possible exploitation as drug carriers, protein shields and cleaning agents in the gas phase. METHODS: The stability and fragmentation patterns of singly and multiply charged (either positively or negatively) aggregates of the surfactant (1R,2S)-dodecyl(2-hydroxy-1-methyl-2-phenylethyl)dimethyl ammonium bromide (DMEB) in the gas phase have been studied by ion mobility mass spectrometry and tandem mass spectrometry. Molecular dynamics (MD) simulations of positively and negatively singly and multiply charged DMEB aggregates have been performed to obtain structural and energetics information. Finally, in order to ascertain some clues on the DMEB growth mechanism, quantum mechanics calculations were carried out. RESULTS: It has been evidenced that positively and negatively singly charged aggregates at low collision energy decompose preferentially by loss of only one DMEB molecule. Increasing the collision energy, the loss of neutrals becomes increasingly abundant. Multiply charged DMEB aggregates are unstable and decompose forming singly charged monomers or dimers. MD simulations show reverse micelle-like structures with polar heads somewhat segregated into the aggregate interior. Finally, a good correlation between experimental and calculated collisional cross sections (CCS) was found. CONCLUSIONS: The fragmentation pathways of DMEB charged species evidenced for singly charged aggregates exhibit features similar to that of other detergent aggregates, but multiply charged aggregates showed a system-specific behavior. QMcalculations on the optimized structures (21+, 31+, 11- and 21-) indicate that the most determinant interactions are due to an OH---Br hydrogen bonding that is also involved in the link between monomericDMEB units. TheMDmodels gave CCS values in good agreement with experimental ones, evidenced by a less strict reverse micelle-like structure and a reasonably spread bromine anion distribution.

Surfactants are an interesting class of compounds characterized by the segregation of polar and apolar domains in the same molecule. This peculiarity makes possible a whole series of microscopic and macroscopic effects. Among their features, their ability to segregate particles (fluids or entire domains) and to reduce the surface/interfacial tension is the utmost important. The interest in the chemistry of surfactants never weakened; instead, waves of increasing interest have occurred every time a new field of application of these molecules has been discovered. All these special characteristics depend largely on the ability of surfactants to self-assemble and constitute supramolecular structures where their chemical properties are amplified. The possibility to obtain structural and energy information and, above all, the possibility of forecast the self-organizing mechanisms of surfactants have had a significant boost via computational chemistry. The molecular dynamics models, initially coarse-grained and subsequently (with the increasing computer power) using more accurate models, allowed, over the years, to better understand different aspects of the processes of dispersion, self-assembly, segregation of surfactant. Moreover, several other aspects have been investigated as the effect of the counterions of many ionic surfactants in defining the final supramolecular structures, the mobility of side chains, and the capacity of some surfactant to envelope entire proteins. This review constitutes a perspective/prospective view of these results. On the other hand, some comparison of in silico results with experimental information recently acquired through innovative analytical techniques such as ion mobility mass spectrometry which have been introduced.

In a previous work, we developed the in house Virtual Lock-and-Key Approach (VLKA) in order to evaluate target assignment starting from molecular descriptors calculated on known inhibitors used as an information source. This protocol was able to predict the correct biological target for the whole dataset with a good degree of reliability (80%), and proved experimentally, which was useful for the target fishing of unknown compounds. In this paper, we tried to remodel the previous in house developed VLKA in a more sophisticated one in order to evaluate the influence of 3D conformation of ligands on the accuracy of the prediction. We applied the same previous algorithm of scoring and ranking but, this time, combining it with a structure-based approach as docking. For this reason, we retrieved from the RCSB Protein Data Bank (PDB), the available 3D structures of the biological targets included into the previous work, and we used them to calculate poses of the 7352 dataset compounds in the VLKA biological targets. First, docking protocol has been used to retrieve docking scores, then, from the docked poses of each molecule, 3D-descriptors were calculated (Conf-VLKA), While the use of the simple docking scores proved to be inadequate to improve compounds classification, the Conf-VLKA showed some interesting variations compared to the original VLKA, especially for targets whose ligands present a high number of rotamers. This work represent a first preliminary study to be completed using other techniques such as induced fit docking or molecular dynamics structure clustering to take into account the protein side chains adaptation to ligands structures.

The cyclin dependent kinases (CDKs) are a small family of serine/threonine protein kinases that can act as a potential therapeutic target in several proliferative diseases, including cancer. This short review is a survey on the more recent research progresses in the field achieved by using in silico methods. All the "armamentarium" available to the medicinal chemists (docking protocols and molecular dynamics, fragment-based, de novo design, virtual screening, and QSAR) has been employed to the discovery of new, potent, and selective inhibitors of cyclin dependent kinases. The results cited herein can be useful to understand the nature of the inhibitor-target interactions, and furnish an insight on the structural/molecular requirements necessary to achieve the required selectivity against cyclin dependent kinases over other types of kinases.

In a recent paper, we presented a new virtual screening workflow that addresses the arising issues of molecular docking and pharmacophore modeling when using a single set of coordinates and a single active ligand [1]. MD simulations were carried out and ligand-protein interactions were analyzed and collected together with their appearance frequency. A pharmacophore model was then created using only the common feature patterns that all the ligands exhibited during MD simulations. This ‘Molecular dYnamics SHAred PharmacophorE’ was then used for virtual screening on active and inactive molecules library. MYSHAPE was also used as constraints for the creation of the docking grid. The application of the MYSHAPE model showed an interesting increase of the screening capability both in terms of sensitivity of the model and specificity when compared to the PDB models. This work [1] was a first essay for a workflow that should be applied to different proteins. In the present study we tried to apply the MYSHAPE approach to other three different ligand-protein systems (ERα; RXRα, and MAPKp38) with the aim to optimize the method to each different biological target taking in consideration the early recognition. The obtained results for these new targets confirmed that it is mandatory, to optimize the virtual screening campaign, the selection of dynamic features and constraints for docking. In particular, the addition of the constraints derived from MD simulation leads to an improvement in the model selectivity for RXRα and ERα in standard precision docking mode. For MAPKp38, validation metrics such as ROC, BEDROC, and AUAC are higher in extra precision mode. For the pharmacophore modeling, the addition of the features derived from the common interactions in MD simulations guarantee an improvement in the AUC for RXRα (37%), and ERα (77%), but light improvement for MAPKp38. MD simulation derived common interactions revealed fundamental for docking selectivity, while they are applied to pharmacophore modeling only when the number of final features in the common and dynamic pharmacophore is higher than the starting static pharmacophore. The strength behind the protocol is the ease of use related to the improvement of results. It also could represent a valid alternative to use very time-consuming techniques such as XP docking with constraints. Reference: 1. Perricone, U., Wieder, M., Seidel, T., Langer, T., Padova, A., Almerico, A. M., & Tutone, M. (2017). A Molecular Dynamics–Shared Pharmacophore Approach to Boost Early-Enrichment Virtual Screening: A Case Study on Peroxisome Proliferator-Activated Receptor α. ChemMedChem, 2017. DOI: 10.1002/cmdc.201600526

In efforts to find agents with improved biological activity against cancer cells, recent years have seen an increased interest in the study of small molecules able to bind the deoxyribonucleic acid (DNA) when it assumes secondary structures known as G-quadruplexes (G4s) preferring them over the B form. Currently, several compounds reported in literature have already shown to be good candidates as G4s DNA stabilizers. Even though some specific features for the G4s affinity are known, such as a π-delocalized system able to stack at the top/end of a G-tetrad and positively charged substituents able to interact with the grooves, it is not clear yet what kind of structural features affect more the G4 arrangement. This is mainly due to the structure heterogeneity of both the G4 stabilizer compounds and the DNA G4s isoforms. In this review, we aim to classify some known G4 binders by analyzing them from a new perspective surprisingly never approached up to date: the symmetry features. Molecular symmetry could be responsible for the specific binding mode to the G4-DNA but could also be crucial in determining different isoform affinity. We propose to classify the G4s stabilizers in five main point group symmetry classes. This classification could be useful to design new ligands able to stabilize a specific G-quadruplex isoform, in order to increase the selectivity of new potential anticancer G-quadruplex targeting drugs, a goal yet highly sought by researchers.

The dopamine-amino acid conjugate DA-Phen was firstly designed to obtain a useful prodrug for the therapy of Parkinson's disease, but experimental evidence shows that it effectively interacts with D1 dopamine receptors (D1DRs), leading to an enhancement in cognitive flexibility and to the development of adaptive strategies in aversive mazes, together with a decrease in despair-like behavior. In this paper, homology modelling, molecular dynamics, and site mapping of D1 receptor were carried out with the aim of further performing docking studies on other dopamine conjugates compared with D1 agonists, in the attempt to identify new compounds with potential dopaminergic activity. Two new conjugates (DA-Trp 2C, and DA-Leu 3C) have been identified as the most promising candidates, and consequently synthesized. Preliminary evaluation in terms of distribution coefficient (DpH7.4), stability in rat brain homogenate, and in human plasma confirmed that DA-Trp (2C), and DA-Leu (3C) could be considered as very valuable candidates for further in vivo studies as new dopaminergic drugs.

p53 is a powerful anti-tumoral molecule frequently inactivated by mutations or deletions in cancer. However, half of all human tumors expresses wild-type p53, and its activation, by antagonizing its negative regulator Mdm2, might offer a new strategy for therapeutic protocol. In this work, we present a molecular dynamics study on Mdm2 structure bound to two different known inhibitors with the aim to investigate the structural transitions between apo-Mdm2 and Mdm2-inhibitor complexes. We tried to gain information about conformational changes binding a benzodiazepine derivative inhibitor with respect the known nutlin and the apo form. The conformational changes alter the size of the cleft and were mainly in the linker regions, suggesting that the overall dynamic nature of Mdm2 is related to dynamic movements in these regions.

Parasitic diseases caused by protozoarian agents are still relevant today more than ever. Recently, we synthesized several polycondensed diazine derivatives by means 1,3-dipolar cycloaddition reactions. A broad selection of these compounds were submitted to in vitro biological screening against Plasmodium falciparum , Leishmania infantum , Trypanosoma brucei , and Trypanosoma cruzi , resulting active at micromolar level. Induced Fit Docking/MM-GBSA studies were performed giving interesting indications about the probable mechanism of action of the most active compounds

The IKK kinases family represents a thrilling area of research because of its importance in regulating the activity of NF-kB transcription factors. The discovery of the central role played by IKK-β in the activation of transcription in response to apoptotic or inflammatory stimuli allowed to considerate its modulation as a promising tool for the treatment of chronic inflammation and cancer. To date, several IKK-β inhibitors have been discovered and tested. In this work, an analysis of the interactions between different classes of inhibitors and their biological target was performed, through the application of Molecular Docking and Pharmacophore/3D-QSAR approaches to a set of 141 inhibitors included in the Binding Database. In order to overcome the difficulty due to the lack of crystallographic data for IKK-β, a homology model of this protein has been built and validated. The results allowed to study in depth the structural bases for the interaction of each family of inhibitors and provided clues for further modifications, with the aim of improving the activity and selectivity of designed drugs targeting this enzyme.

One of the major problems in the fight against cancer is drug-resistance, which, at a molecular level, can be acquired through mutations able to deactivate apoptosis. In particular, proteins in the Bcl-2 family are central regulators of programmed cell death, and members that inhibit apoptosis, such as Bcl-xl and Bcl-2, are overexpressed in many tumours. The development of new inhibitors of these proteins as potential anticancer therapeutics represents a new frontier. In this work, we carried out an in-silico screening of compounds from a free database of more than 2 million structures (ZINC database), which allowed us to identify 17 sulfonamide derivatives as new potential inhibitors; these are currently undergoing biological evaluation.

In this paper we describe a comparative analysis between multivariate and docking methods in the study of the drug resistance to the reverse transcriptase and the protease inhibitors. In our early papers we developed a simple but efficient method to evaluate the features of compounds that are less likely to trigger resistance or are effective against mutant HIV strains, using the multivariate statistical procedures PCA and DA. In the attempt to create a more solid background for the prediction of susceptibility or resistance, we carried out a comparative analysis between our previous multivariate approach and molecular docking study. The intent of this paper is not only to find further support to the results obtained by the combined use of PCA and DA, but also to evidence the structural features, in terms of molecular descriptors, similarity, and energetic contributions, derived from docking, which can account for the arising of drug-resistance against mutant strains.

Background: Recently, we have shown anti-proliferative and pro-apoptotic effects of indicaxanthin associated with epigenetic modulation of the onco-suppressor p16INK4a in the human colon cancer cell line CACO2. In the present study, the epigenetic activity of indicaxanthin and the mechanisms involved were further investigated in other colorectal cancer cell lines. Methods: LOVO1, CACO2, HT29, HCT116, and DLD1 cells were used to evaluate the potential influence of consistent dietary concentrations of indicaxanthin on DNA methylation, and the epigenetic mechanisms involved were researched. Results: Indicaxanthin exhibited anti-proliferative activity in all cell lines but HT29, induced demethylation in the promoters of some methylation-silenced onco-suppressor genes involved in colorectal carcinogenesis (p16INK4a, GATA4, and ESR1), and left unchanged others which were basally hypermethylated (SFRP1 and HPP1). In apparent contrast, cell exposure to indicaxanthin increased DNMT gene expression, although indicaxanthin appeared to be an inhibitor of DNMT activity. Indicaxanthin also increased the expression of genes involved in DNA demethylation. Finally, an in silico molecular modelling approach suggested stable binding of indicaxanthin at the DNMT1 catalytic site. Conclusions: Our findings contribute to new knowledge in the field of phytochemicals and specifically suggest dietary indicaxanthin as a potential epigenetic agent to protect colon cells against tumoral alterations.

Health benefits of dietary phytochemicals have been suggested in recent years. Among 1000s of different compounds, Betalains, which occur in vegetables of the Cariophyllalae order (cactus pear fruits and red beet), have been considered because of reducing power and potential to affect redox-modulated cellular processes. The antioxidant power of Betalains is strictly due to the dissociation rate of the acid moieties present in all the molecules of this family of phytochemicals. Experimentally, only the pK avalues of betanin were determined. Recently, it was evidenced it was evidenced as the acid dissociation, at different environmental pHs, affects on its electron-donating capacity, and further on its free radical scavenging power. The identical correlation was studied on another Betalains family compound, Betalamic Acid. Experimental evidences showed that the free radical scavenging capacity of this compound drastically decreases at pH>5, but pK a values were experimentally not measured. With the aim to justify the Betalamic Acid behavior as free radical scavenger, in this paper we tried to predict in silico the pK a values by means different approaches. Starting from the known experimental pK as of acid compounds, both phytochemicals and small organic, two empirical approaches and quantum-mechanical calculation were compared to give reliable prediction of the pK as of Betalamic Acid. Results by means these computational approaches are consistent with the experimental evidences. As shown herein, in silico, the totally dissociated species, at the experimental pH>5 in solution, is predominant, exploiting the higher electron-donating capability (HOMO energy). Therefore, the computational estimated pK a values of Betalamic Acid resulted very reliable.