Search Results (303)

Background: The Nerve Growth Factor (NGF) is essential for neuronal survival and function and represents a key therapeutic target for pain and inflammation-related disorders, as well as for neurodegenerative diseases. Small-molecule antagonists of human NGF (hNGF) offer advantages over monoclonal antibodies, including oral availability and reduced immunogenicity. However, their development is often hindered by solubility challenges, necessitating the use of solvents like dimethyl sulfoxide (DMSO). This study investigates whether DMSO directly interacts with hNGF and affects its receptor-binding properties. Methods: Integrative/hybrid computational and experimental biophysical approaches were used to assess DMSO-NGF interaction by combining machine-learning tools and Nuclear Magnetic Resonance (NMR), Fourier Transform Infrared (FT-IR) spectroscopy, Differential Scanning Fluorimetry (DSF) and Grating-Coupled Interferometry (GCI). These techniques evaluated binding affinity, conformational stability, and receptor-binding dynamics. Results: Our findings demonstrate that DMSO binds hNGF with low affinity in a specific yet non-disruptive manner. Importantly, DMSO does not induce significant conformational changes in hNGF nor affect its interactions with its receptors. Conclusions: These results highlight the importance of considering solvent–protein interactions in drug discovery, as these low-affinity yet specific interactions can affect experimental outcomes and potentially alter the small molecules binding to the target proteins. By characterizing DMSO-NGF interactions, this study provides valuable insights for the development of NGF-targeting small molecules, supporting their potential as effective alternatives to monoclonal antibodies for treating pain, inflammation, and neurodegenerative diseases. Full article

Background: Anthralin is known for its efficacy in treating psoriasis and acne, possessing poor solubility. Addressing these limitations, the present study endeavors to develop a microemulgel formulation of anthralin aimed at enhancing solubility. Method: The solubility study was performed in various solvents. An o/w (oil-in-water) emulsion was formed using the water titration method, which was optimized by statistical experimental design half-run CCD. The final optimized batch was evaluated for physicochemical and in vitro properties Result: The final optimized batch showed a particle size (PS) of 417 nm, −25.2 mV zeta potential (ZP) and pH 5.8, which remained stable upon centrifugation, heating–cooling and freeze–thawing cycle. Furthermore, microemulsion with Carbopol 943 5% w/v was selected as the gel base for the formation of microemulgel characterized by PS, ZP, pH, and viscosity of 230 nm, −50.6 mV, 6.9 and 14,200 cps, respectively, that ensured it a high enough stability. In silico molecular docking between ligand and protein provides the binding energies validating the interaction. Hence, the in silico study was performed for psoriasis and P. acne proteins. An in vitro antibacterial activity study on Propionibacterium revealed a significant efficiency of the formulation and MTT assay using L929 cell line in the presence of the drug-loaded microemulgel indicated an inhibition of growth proving that formulation has anti-psoriatic activity. Conclusions: Combination therapy with Clindamycin might improve efficacy while reducing antibiotic resistance risks. Full article

Background/Objectives: Sustained drug exposure is a key factor in the treatment of patients infected with human immunodeficiency virus (HIV) or hepatitis B virus (HBV) in order to achieve the intended virological response. Although influenced also by other parameters, adherence to the treatment scheme is the most important for adequate drug exposure. This can be assessed by therapeutic drug monitoring (TDM). Tenofovir (TFV) is a nucleotide analogue used in the treatment of both HIV and HBV. Although various analytical methods for the quantification of tenofovir prodrugs have been published, there is limited literature on methods for simultaneous TFV and its active metabolite, tenofovir diphosphate (TFVDP) direct determination. Methods: In this study, we describe a novel micro-liquid-chromatography-mass spectrometry (micro-LC-MS/MS) method for TDM of TFV and TFVDP in biological matrices (whole blood, plasma). The challenging separation of the high-polarity analytes was resolved on an amino stationary phase, eluted in HILIC (hydrophilic interaction liquid chromatography) mode. The sample preparation included a clean-up step with hexane for the removal of lipophilic compounds and then protein precipitation with organic solvent. Results: The achieved low limits of quantification in blood were 0.25 ng/mL for TFV, and 0.5 ng/mL for TFVDP. Linearity, accuracy (91.63–109.18%), precision (2.48–14.08), and stability were validated for whole blood matrix, meeting the guidelines performance criteria. Samples collected from treated patients were analyzed, with results being in accordance with the reported pharmacokinetics. Conclusions: The new method is adequate for analyzing samples in a clinical set-up. The measurement of both TFV and TFVDP improves clinical decision by an in-depth evaluation of long-term adherence, and together with viral load and resistance data helps guiding the treatment towards the intended virological suppression. Full article

The pharmaceutical industry faces mounting pressure to reduce its environmental impact while maintaining innovation in drug development. Artificial intelligence (AI) has emerged as a transformative tool across healthcare and drug discovery, yet its potential to drive sustainability by improving molecular design remains underexplored. This review critically examines the applications of AI in molecular design that can support in advancing greener pharmaceutical practices across the entire drug life cycle—from design and synthesis to waste management and solvent optimisation. We explore how AI-driven models are being used to personalise dosing, reduce pharmaceutical waste, and design biodegradable drugs with enhanced environmental compatibility. Significant advances have also been made in the predictive modelling of pharmacokinetics, drug–polymer interactions, and polymer biodegradability. AI’s role in the synthesis of active pharmaceutical compounds, including catalysts, enzymes, solvents, and synthesis pathways, is also examined. We highlight recent breakthroughs in protein engineering, biocatalyst stability, and heterogeneous catalyst screening using generative and language models. This review also explores opportunities and limitations in the field. Despite progress, several limitations constrain impact. Many AI models are trained on small or inconsistent datasets or rely on computationally intensive inputs that limit scalability. Moreover, a lack of standardised performance metrics and life cycle assessments prevents the robust evaluation of AI’s true environmental benefits. In particular, the environmental impact of AI-driven molecules and synthesis pathways remains poorly quantified due to limited data on emissions, waste, and energy usage at the compound level. Finally, a summary of challenges and future directions in the field is provided. Full article

Although enormous efforts have been made to prepare tasty and soluble steviol glycosides (SGs), the structure–property relationship of SGs still remains unclear, neither in experiment fact nor in the mechanism, such as the influence of linkage type and position of substituted glucosyl on physiochemical properties and sensory features of SGs. The favorable SGs, rebaudioside D (RD) and rebaudioside A (RA), possess good edulcorant quality, poor solubility, and other significantly different physical properties. This research chose two pairs of isomeric SGs, RA and its isomer rebaudioside E (RE) and RD and its isomer RA1G (a synthetic SG, α-1,6-mono-glucosylated RA), to conduct a comparative study, aiming to reveal the structure–property relevance on their solubility, sweetness, stability, and crystal structure. The RA1G presents an aqueous solubility 13 times that of RA and 137 times that of RD and exhibits better edulcorant quality than that of RA, similar to RD. The results indicate that the glucosyl linkage type and position have a stronger impact on the properties of the SGs than the number of glucosyl moieties. The underlying mechanism of their structure–property relevance was elucidated by analyzing the interaction energies between the SGs with solvent and human receptor proteins, respectively. Full article

Raman spectroscopy is one of the best techniques for obtaining information concerning the physical–chemical interactions between a lipid and a solvent. Phospholipids in water are the main elements of cell membranes and, by means of their chemical and physical structures, their cells can interact with other biological molecules (i.e., proteins and vitamins) and express their own biological functions. Phospholipids, due to their amphiphilic structure, form biomimetic membranes which are useful for studying cellular interactions and drug delivery. Synthetic systems such as DPhPC-based liposomes replicate the key properties of biological membranes. Among the different models, phospholipid mimetic membrane models of lamellar vesicles have been greatly supported. In this work, a biomimetic system, a deuterium solution (50 mM) of the synthetic phospholipid 1,2-diphytanoyl-sn-glycero-3-phosphocholine (DPhDC), is studied using μ-Raman spectroscopy in a wide temperature range from −181.15 °C up to 22.15 °C, including the following temperatures: −181.15 °C, −146.15 °C, −111.15 °C, −76.15 °C, −61.15 °C, −46.15 °C, −31.15 °C, −16.15 °C, −1.15 °C, 14.15 °C, and 22.15 °C. Based on the Raman evidence, phase transitions as a function of temperature are shown and grouped into five classes, where the corresponding Raman modes describe the stretching of the (C−N) bond in the choline head group (gauche) and the asymmetric stretching of the (O−P−O) bond. The acquisition temperature of each Raman spectrum characterizes the rocking mode of the methylene of the acyl chain. These findings enhance our understanding of the role of artificial biomimetic lipids in complex phospholipid membranes and provide valuable insights for optimizing their use in biosensing applications. Although the phase stability of DPhPC is known, the collected Raman data suggest subtle molecular rearrangements, possibly due to hydration and second-order transitions, which are relevant for membrane modeling and biosensing applications. Full article

The carnivorous plant Sarracenia purpurea has been traditionally used in various ethnobotanical applications, including treatments for type 2 diabetes and tuberculosis-like symptoms. This study investigates the cytotoxic effects of S. purpurea root extract (Sp-R) on human non-small-cell lung cancer (NSCLC) cell lines, including H1975, H838, and A549, focusing on its impact on cell survival, apoptosis, proliferation, and migration. Additionally, its ability to inhibit the single-stranded DNA-binding activity of human RPA32 (huRPA32), a key protein in DNA replication, was evaluated. Extracts from different plant parts (leaf, stem, and root) were prepared using various solvents (water, methanol, ethanol, and acetone) and screened for apoptosis-inducing potential using the chromatin condensation assay. Among these, the acetone-extracted root fraction (Sp-R-A) exhibited the most potent pro-apoptotic effects. The MTT assay demonstrated a dose-dependent cytotoxic effect on NSCLC cells, with IC₅₀ values of 33.74 μg/mL for H1975, 60.79 μg/mL for H838, and 66.52 μg/mL for A549. Migration and clonogenic assays further revealed that Sp-R-A significantly inhibited cancer cell migration and colony formation in a dose-dependent manner. Moreover, Sp-R-A enhanced apoptosis when combined with the EGFR inhibitor afatinib, suggesting a potential synergistic effect. The electrophoretic mobility shift assay confirmed that Sp-R-A significantly inhibited the DNA-binding activity of huRPA32, with an IC₅₀ of 13.6 μg/mL. AlphaFold structural prediction and molecular docking studies indicated that major bioactive compounds in S. purpurea, including α-amyrin, ursolic acid, and betulinaldehyde, strongly interact with the DNA-binding domain of huRPA32, potentially contributing to its inhibitory effect. Overall, these findings suggest that huRPA32 is a potential molecular target of Sp-R-A and the anticancer potential of S. purpurea root extract against NSCLC is highlighted, supporting further investigation into its therapeutic applications. Full article

The chemical interaction of salicylic acid, formaldehyde, and sulfuric acid produced a disalicylic ligand (3,3′-dicarboxy-2,2′-dihydroxydiphenylmethane, DCM), which was then allowed to coordinate with copper (II) ions. The solid compounds’ chemical structures were determined using elemental analysis, UV-Vis, FT-IR, MS, ¹H-NMR, PXRD, SEM, TEM, magnetic studies, as well as molecular modeling based on DFT (density functional theory) calculations. It was proposed that the ligand coordinates in a tetradentate fashion with the copper ion to give a square-planar binuclear complex. A significant difference in the diffraction patterns between Cu(II)–DCM (amorphous) and DCM (crystalline) was displayed using an X-ray diffraction analysis. Spherical granules were identified throughout through morphology analysis using SEM and TEM. UV-Vis spectra were used to quantify the optical characteristics such as the energy gap, optical conductivity, refractive index, and penetration depth. The band gap values that lie within the semiconductor region suggested that the compounds could be used for electronic applications. The optimized structure of the synthesized Cu(II)–DCM complex was investigated using DFT and TD-DFT (time-dependent density functional theory) at the B3LYP/6-31G(d, p) level, with the LANL2DZ basis set for Cu in an ethanol solvent and the gas environment modeled by CPCM. The experimental data suggest a square-planar geometry of the Cu(II) binuclear complex. The theoretical calculations support the proposed structure of the compound. The cytotoxicity of the DCM against HCT–116 (human colon cancer) cells was tested, and the outcome exhibited good inhibitions of growth. A molecular docking (MD) examination was carried out to illustrate the binding mode/affinity of the prepared compounds (DCM and Cu(II)–DCM) in the active site of the receptor protein [CDK2 enzyme, PDB ID: 6GUE]. The compounds formed hydrogen bonds with the amino acid residues of the protein, increasing the binding affinity from −7.2 to −9.3 kcal/mol through the coordination process. The information from this current study, particularly the copper complex, is beneficial for exploring new compounds that have anticancer potential. Full article

Proteins and their surrounding hydration water engage in a dynamic interplay that is critical for maintaining structural stability and functional integrity. However, the intricate coupling between protein dynamics and the structural order of hydration water remains poorly understood. Here, we employ all-atom molecular dynamics simulations to investigate this relationship across four representative proteins. Our results reveal that protein residues with greater flexibility or solvent exposure are surrounded by more disordered hydration water, akin to bulk water, whereas rigid and buried non-polar residues are associated with structurally ordered hydration shells. Due to their strong hydrogen bonding and electrostatic interactions, charged residues exhibit the most disordered hydration water, while non-polar residues are associated with the structurally most ordered hydration water. We further uncovered a positive correlation between the relaxation dynamics of protein residues and their hydration water: slower (faster) protein relaxation is coupled with slower (faster) relaxation of the structural order of hydration water. Notably, this coupling weakens with increasing residue flexibility or solvent exposure, with non-polar residues displaying the strongest coupling, and charged residues the weakest. To further uncover their coupling mechanism, we elucidate residue-specific coupled fluctuations between protein residues and hydration water by generating scatter plots. These findings provide a comprehensive understanding of the mechanisms underlying protein–water interactions, offering valuable insights into the role of hydration water in protein stability, dynamics, and function. Full article

The tetrameric protein transthyretin (TTR) transports the hormone thyroxine in plasma and cerebrospinal fluid. Certain point mutations of TTR, including the Val122Ile mutation investigated here, destabilize the tetramer leading to its dissociation, misfolding, aggregation, and the eventual buildup of amyloid fibrils in the myocardium. Cioffi et al. reported the design and synthesis of a novel TTR kinetic stabilizing ligand, referred to here as TKS14, that inhibited TTR dissociation and amyloid fibril formation. In this study, molecular dynamics simulations were used to investigate the binding of TKS14 and eight TSK14 derivatives to the Val122Ile TTR mutant. For each complex, the ligand’s solvent accessible surface area (SASA), ligand–receptor hydrogen-bonding interactions, and the free energy of ligand-binding to TTR were investigated. The goal of this study was to identify the TSK14 functional groups that contributed to TTR stabilization. TKS14 was found to form a stable, two-point interaction with TTR by hydrogen bonding to Ser-117 residues in the inner receptor binding pocket and interacting through hydrogen bonds and electrostatically with Lys-15 residues near the receptor’s surface. The free energy of TKS14-TTR binding was −18.0 kcal mol⁻¹ and the ligand’s average SASA value decreased by over 80% upon binding to the receptor. The thermodynamic favorability of TTR binding decreased when TKS14 derivatives contained either methyl ester, amide, tetrazole, or N-methyl functional groups that disrupted the above two-point interaction. One derivative in which a tetrazole ring was added to TKS14 was found to form hydrogen bonds with Thr-106, Thr-119, Ser-117, and Lys-15 residues. This derivative had a free energy of TTR binding of −21.4 kcal mol⁻¹. Overall, the molecular dynamics simulations showed that the functional groups within the TKS14 structural template can be tuned to optimize the thermodynamic favorability of ligand binding. Full article

Because of the involvement of π-electron cyclic constituents in their side chains, the so-called aromatic residues give rise to a number of strong, narrow, and well-resolved lines spread over the middle wavenumber (1800–600 cm⁻¹) region of the Raman spectra of peptides and proteins. The number of characteristic aromatic markers increases with the structural complexity (Phe → Tyr → Trp), herein referred to as (F_i = 1, …, 6) in Phe, (Y_i = 1, …, 7) in Tyr, and (W_i = 1, …, 8) in Trp. Herein, we undertake an overview of these markers through the analysis of a representative data base gathered from the most structurally simple tripeptides, Gly-Xxx-Gly (where Xxx = Phe, Tyr, Trp). In this framework, off-resonance Raman spectra obtained from the aqueous samples of these tripeptides were jointly used with the structural and vibrational data collected from the density functional theory (DFT) calculations using the M062X hybrid functional and 6-311++G(d,p) atomic basis set. The conformation dependence of aromatic Raman markers was explored upon a representative set of 75 conformers, having five different backbone secondary structures (i.e., β-strand, polyproline-II, helix, classic, and inverse γ-turn), and plausible side chain rotamers. The hydration effects were considered upon using both implicit (polarizable solvent continuum) and explicit (minimal number of 5–7 water molecules) models. Raman spectra were calculated through a multiconformational approach based on the thermal (Boltzmann) average of the spectra arising from all calculated conformers. A subsequent discussion highlights the conformational landscape of conformers and the wavenumber dispersion of aromatic Raman markers. In particular, a new interpretation was proposed for the characteristic Raman doublets arising from Tyr (~850–830 cm⁻¹) and Trp (~1360–1340 cm⁻¹), definitely excluding the previously suggested Fermi-resonance-based assignment of these markers through the consideration of the interactions between the aromatic side chain and its adjacent peptide bonds. Full article

Background: Dengue virus (DENV) is the fatal pathogenic arthropod-borne virus (arboviruses) that belongs to the Flaviviridae family, which transmits to humans through mosquito bites from infected Aedes aegypti and Aedes albopictus mosquitoes or maternal-fetal transmission. Despite antigenic differences, the four serotypes of DENV (DENV-1 to DENV-4) share 65–78% of their genome. Non-structural (NS) proteins amongst serotypes show analogous functions. Among NS proteins, NS3 and NS5 are frequently used as targets for antiviral drugs due to their multifunctional roles. Methods: To identify potential inhibitors of DENV, we created a phytochemical library of 898 compounds derived from 17 medicinal plants recognized for their medicinal and antiviral properties. The phytochemicals library has been docked against the target proteins. Phytochemicals with a docking score greater than −8.0 kcal/mol were selected for further evaluation using a machine learning approach. Further, molecular dynamics (MD) simulations were conducted to evaluate the root mean square deviation, root mean square fluctuation, solvent-accessible surface area, radius of gyration, and hydrogen bond count of the compounds. Results: From the docking results, Silibinin, Rubiadin, and Ellagic acid showed binding affinities of −8.5, −8.3, and −8.2 kcal/mol, respectively, for NS3, and NSC 640467, Bisandrographolide A, and Andrographidin A showed binding affinities of −9.3, −10.1, and −9.3 kcal/mol, respectively, for NS5 target proteins. These compounds exhibited strong interactions with target proteins. MD simulation results confirmed the stable formation of protein–ligand complexes. Further, absorption, distribution, metabolism, excretion, and toxicity (ADMET) and bioactivity predictions confirmed their pharmacological safety. Conclusions: Despite global public health concerns, DENV still lacks specific drug treatments. Our identified new drug candidates might help for developing effective antiviral inhibitors against the DENV. However, further confirmation is needed through in vivo and in vitro research. Full article

Polycyclic aromatic hydrocarbons (PAHs) and global warming impact aquatic ecosystems, eventually interacting. Monolayer (2D) cultures of cell lines, such as the rainbow trout liver RTL-W1, are employed for unveiling toxicological effects in fish. Nonetheless, three-dimensional (3D) models constitute an alternate paradigm, better emulating in vivo responses. Here, ultra-low attachment (ULA) plates were used to generate ten-day-old RTL-W1 spheroids for exposure to a control, a solvent control (0.1% DMSO) and the model PAH benzo[k]fluoranthene (BkF) at 10 and 100 nM and at 18 and 23 °C (thermal stress). After a 4-day exposure, spheroids were analyzed for viability (alamarBlue and lactate dehydrogenase), biometry (area, diameter and sphericity), histocytology (optical and electron microscopy), and mRNA levels of the detoxification-related genes cytochrome P450 (CYP)1A, CYP3A27, aryl hydrocarbon receptor (AhR), glutathione S-transferase (GST), uridine diphosphate–glucuronosyltransferase (UGT), catalase (CAT), multidrug resistance-associated protein 2 (MRP2) and bile salt export protein (BSEP). Immunocytochemistry (ICC) was used to assess CYP1A protein expression. Neither temperature nor BkF exposure altered the spheroids’ viability or biometry. BkF modified the cell’s ultrastructure. The expression of CYP1A was augmented with both BkF concentrations, while AhR’s increased at the higher concentration. The CYP1A protein showed a dose-dependent increase. Temperature and BkF concurrently modelled UGT’s expression, which increased in the 100 nM condition at 23 °C. Conversely, CYP3A27, MRP2, and BSEP expressions lowered at 23 °C. CAT and GST mRNA levels were uninfluenced by either stressor. Overall, BkF and temperature impacted independently or interactively in RTL-W1 spheroids. These seem to be useful novel tools for studying the liver-related effects of temperature and PAHs. Full article

Deep eutectic solvents (DESs) have shown promise as a medium for extracting polar volatile fatty acids (VFAs) and in situ esterification of the extracted molecules using lipases. This solvent enhanced biocatalysis process can potentially streamline VFA separation from fermentation broth by integrating conversion and extraction steps. Two commercial lipases from Aspergillus oryzae (AoL) and Candida rugosa (CrL) were evaluated in reaction systems containing hydrophilic or hydrophobic DESs using a newly optimized lipase assay. The optimal pH for both lipases was around 5.0, with a slight reduction in activity at pH 8.0 and a significant inhibition at pH 2.0. The impact of DES concentration on lipase activity varied depending on the specific DES–lipase pairs. Most hydrophilic DESs show good compatibility with the tested lipases. Specifically for choline chloride/ethylene glycol (1:2) and choline chloride/levulinic acid (1:2), taking into account the influence of pH, CrL activity increased with DES concentration. However, the hydrophobic DES thymol/2,6-dimethoxyphenol (1:2) demonstrated enhanced inhibitory effects on both lipases. Docking simulation helped explain the ligand–protein interactions but showed limited capability in predicting the compatibility of specific DES–lipase pairs due to its constraints in simulating flexible protein structures and the complex interactions between DES components and water. Full article

Botulinum toxin A (BoNT-A) is widely used in both therapeutic and aesthetic settings; however, the formation of neutralizing antibodies (NAbs) remains a critical concern, leading to treatment failure. Immunogenic responses are known to vary between individuals due to HLA polymorphisms. Although some claim that neurotoxin-associated proteins (NAPs) shield BoNT-A from immune detection or are themselves immunogenic, there is limited molecular evidence supporting either view. This study applies computational immunogenetics to explore BoNT-A immunogenicity, focusing on HLA binding and the influence of accessory proteins. Epitope mapping, molecular docking, and HLA binding predictions were used to evaluate interactions between BoNT-A epitopes and selected class II HLA alleles (HLA-DQA1*01:02, HLA-DQA1*03:03, HLA-DQB1*06:04, HLA-DQB1*03:01, and HLA-DRB1*15:01). To assess the potential immunomodulatory role of NAPs, molecular dynamics (MD) simulations, solvent-accessible surface area (SASA) analysis, and electrostatic potential mapping were also conducted. Key epitopes—L11, N25, and C10—showed strong binding affinities to HLA-DQA1*01:02, HLA-DQB1*06:04, and HLA-DQA1*03:03, indicating a potential immunodominant role. NAPs did not obstruct these epitopes but slightly increased their exposure and appeared to stabilize the toxin structure. Electrostatic mapping and binding free energy calculations suggested no significant immunogenic shift in the presence of NAPs. BoNT-A immunogenicity appears to be influenced by HLA allele variability, reinforcing the value of patient-specific genetic profiling. The presumed immunogenic role of NAPs remains unsubstantiated at the molecular level, underscoring the need for evidence-based evaluation over commercial rhetoric. While these findings provide valuable molecular insight, it is important to acknowledge that they are derived entirely from in silico analyses. As such, experimental validation remains essential to confirm the immunological relevance of these predicted interactions. Nonetheless, this computational framework offers a rational basis for guiding future clinical research and the development of HLA-informed BoNT-A therapies. Full article