Microwave-assisted Green Synthesis and Integrated Bioinformatics Study Reveal Curcumin Analogs Dibenzylidene-cyclohexanones as Novel Potential Anti-Tuberculosis Agents

Studying noncanonical intermolecular interactions between a ligand and a protein constitutes an emerging research field. Identifying synthetically accessible molecular fragments that can engage in intermolecular interactions is a key objective in this area. Here, it is shown that so‐called “π‐hole interactions” are present between the nitro moiety in nitro aromatic ligands and lone pairs within protein structures (water and protein carbonyls and sulfurs). Ample structural evidence was found in a PDB analysis and computations reveal interaction energies of about −5 kcal mol−1 for ligand–protein π‐hole interactions. Several examples are highlighted for which a π‐hole interaction is implicated in the superior binding affinity or inhibition of a nitro aromatic ligand versus a similar non‐nitro analogue. The discovery that π‐hole interactions with nitro aromatics are significant within protein structures parallels the finding that halogen bonds are biologically relevant. This has implications for the interpretation of ligand–protein complexation phenomena, for example, involving the more than 50 approved drugs that contain a nitro aromatic moiety.

Multidrug-resistant tuberculosis (MDR-TB) is one of the world's most devastating contagious diseases and is caused by the MDR-Mycobacterium tuberculosis (MDR-Mtb) bacteria. It is therefore essential to identify novel anti-TB drug candidates and target proteins to treat MDR-TB. Here, in vitro and in silico studies were used to investigate the anti-TB potential of two newly sourced actinomycins, actinomycin-X2 (act-X2) and actinomycin-D (act-D), from the Streptomyces smyrnaeus strain UKAQ_23 (isolated from the Jubail industrial city of Saudi Arabia). The anti-TB activity of the isolated actinomycins was assessed in vitro using the Mtb H37Ra, Mycobacterium bovis (BCG), and Mtb H37Rv bacterial strains, using the Microplate Alamar Blue Assay (MABA) method. In silico molecular docking studies were conducted using sixteen anti-TB drug target proteins using the AutoDock Vina 1.1.2 tool. The molecular dynamics (MD) simulations for both actinomycins were then performed with the most suitable target proteins, using the GROningen MAchine For Chemical Simulations (GROMACS) simulation software (GROMACS 2020.4), with the Chemistry at HARvard Macromolecular Mechanics 36m (CHARMM36m) forcefield for proteins and the CHARMM General Force Field (CGenFF) for ligands. In vitro results for the Mtb H37Ra, BCG, and Mtb H37Rv strains showed that act-X2 had minimum inhibitory concentration (MIC) values of 1.56 ±0.0, 1.56 ±0.0, and 2.64 ±0.07 µg/mL and act-D had MIC values of 1.56 ±0.0, 1.56 ±0.0, and 1.80 ±0.24 µg/mL respectively. The in silico molecular docking results showed that protein kinase PknB was the preferred target for both actinomycins, while KasA and pantothenate synthetase were the least preferred targets for act-X2and act-D respectively. The molecular dynamics (MD) results demonstrated that act-X2 and act-D remained stable inside the binding region of PknB throughout the simulation period. The MM/GBSA (Molecular Mechanics/Generalized Born Surface Area) binding energy calculations showed that act-X2 was more potent than act-D. In conclusion, our results suggest that both actinomycins X2 and D are highly potent anti-TB drug candidates. We show that act-X2is better able to antagonistically interact with the protein kinase PknB target than act-D, and thus has more potential as a new anti-TB drug candidate.

Type 2 diabetes mellitus (T2DM) is a global health challenge that requires new therapeutic approaches. Natural compounds from botanical sources offer promise as alternative treatments due to their multifaceted bioactivity and favourable safety profiles. This study employs an in-silico methodology to evaluate their therapeutic potential by targeting genes associated with T2DM. Bioactive natural compounds were screened and sourced from Natural Product Activity and Species Source Database (NPASS) and ZINC12 databases against 14 T2DM-associated genes (GPD2, IRS1, PPARG, IAPP, GCK, ABCC8, MAPK8, MTNR1B, AKT2, PTPN1, INSR, AMPK, GAA, and SLC2A4). Molecular docking assessed binding affinities, while pharmacokinetic and ADMET profiles were predicted. Comparative analyses with approved drugs from DrugBank, network pharmacology approaches, and molecular dynamics simulations further evaluated their therapeutic potential. 72 natural compounds exhibited superior or comparable binding affinities to standard drugs of which, 17 ligands —Moracin D, Moracin P, Plantagineoside A, Pyrene (carcinogenic), Curcumin, Rohitukine, Berberine Chloride, Berberrubine, Apigenin, Emodin, Chelerythrine, Alvocidib, A-443,654, Xambioona, Altertoxin I, Ursolic Acid, and Oleanolic Acid —were selected as top candidates for further analysis. ADME/T analyses highlighted Pyrene, Guggulsterone, Melatonin, Gefitinib, Apigenin, Rotenone, Curcumin, Bavachinin A, Bavachinin, and Quinidine as particularly promising in terms of superior ADME and oral bioavailability. Four ligands—2-Tert-Butyl-6-[(3-Tert-Butyl-2-Hydroxy-5-Methylphenyl)Methyl]-4-Methylphenol, 4-(2-Phenylpropan-2-Yl)Phenol, Phenothiazine, and 2-Naphthalen-1-Ylacetic Acid— showed notable minimal toxicity. Chelerythrine, Emodin, Rohitukine, A-443,654, and Alvocidib demonstrated multi-target networking interactions. Molecular dynamics simulations of the 17 highest-ranked complexes (500 ns) demonstrated stable RMSD values (4.39–5.33 Å), strong hydrogen bonding, and favorable energetic profiles. Particularly, Moracin P, Moracin D, Plantagineoside A, Chelerythrine, Alvocidib, and Ursolic Acid showed the most stable MD trajectories and highly favorable free energy as promising lead candidates. This study highlights the effectiveness of in-silico techniques to identify natural products as prospective alternative or adjunct therapies for T2DM. Further experimental validation is necessary to confirm compounds’ efficacy and safety, paving the way for future clinical investigations.Supplementary InformationThe online version contains supplementary material available at 10.1038/s41598-025-27173-9.

Halogen bonding motifs for anion recognition

Tuberculosis remains a global health threat, with increasing infection rates and mortality despite existing anti-TB drugs. The present work focuses on the research findings regarding the development and evaluation of thiadiazole-linked thiazole derivatives as potential anti-tuberculosis agents. We present the synthesis data and confirm the compound structures using spectroscopic techniques. The current study reports twelve thiazole-thiadiazole compounds (5 a-5 l) for their anti-tuberculosis and related bioactivities. This paper emphasizes compounds 5 g, 5 i, and 5 l, which exhibited promising MIC values, leading to further in silico and interaction analysis. Pharmacophore mapping data included in the present analysis identified tubercular ThyX as potential drug targets. The compounds were evaluated for anti-tubercular activity using standard methods, revealing significant MIC values, particularly compound 5 l, with the best MIC value of 7.1285 μg/ml. Compounds 5 g and 5 i also demonstrated moderate to good MIC values against M. tuberculosis (H37Ra). Structural inspection of the docked poses revealed interactions such as hydrogen bonds, halogen bonds, and interactions containing Pi electron cloud, shedding light on conserved interactions with residues like Arg 95, Cys 43, His 69, and Arg 87 from the tubercular ThyX enzyme.

Background/Objectives: Epilepsy is characterized by unpredictable seizures and drug resistance, along with rising antimicrobial resistance (AMR), highlighting the urgent need for innovative dual-action therapies. This study aimed to design, develop, and evaluate novel pyrazolone derivatives for a dual antimicrobial and antiepileptic potential. Methods: Novel pyrazolone derivatives were designed, synthesized (using 2,4-dinitrophenylhydrazine/semicarbazide condensation with ethyl acetoacetate), and evaluated through molecular docking against antimicrobial (4URM, 3FYV, 3FRA) and neuronal targets (4COF, 5TP9, 5L1F). The in vitro antimicrobial activity was assessed against Gram-positive (S. aureus) and in vitro Gram-negative (E. coli, P. aeruginosa) strains via agar cup plate assays, while in vivo antiepileptic efficacy was tested in a PTZ-induced seizure model in Swiss albino mice. Results: Compound IIa showed potent dual activity, inhibiting E. coli (9 mm zone at 80 μg/mL) and S. aureus (9.5 mm at 80 μg/mL), alongside a significantly delayed seizure onset in the PTZ-induced mouse model (100% survival rate, 45 sec delayed seizure onset, p < 0.001). Compounds Ia and Id showed selective activity against E. coli (6 mm at 80 μg/mL) and P. aeruginosa (7 mm at 80 μg/mL), respectively. Docking studies revealed that compound IIa has a superior binding affinity (-7.57 kcal/mol for 3FYV) compared to standards, driven by hydrogen bonds (SER X: 49) and hydrophobic interactions (LEU X: 20). Conclusions: This study presents a novel approach by proposing a rationally designed pyrazolone scaffold exhibiting both antimicrobial and antiepileptic activity, which integrates in silico modeling with experimental validation. Compound IIa emerged with preliminary dual biological activities, exhibiting strong antibacterial activity, a superior binding affinity toward both bacterial and neuronal targets, and notable seizure prevention in vivo. These findings show the potential of multifunctional pyrazolone derivatives as a new treatment strategy for addressing drug-resistant infections linked to epilepsy and support further optimization toward clinical development.

A new rhodamine based OFF–ON fluorescent chemosensors for selective detection of Hg2+ and Al3+ in aqueous media

Network Pharmacology in Traditional Chinese Medicine

Background/Objectives: Fructose-driven metabolic disorders, such as obesity, non-alcoholic fatty liver disease (NAFLD), dyslipidemia, and type 2 diabetes, are significant global health challenges. Ketohexokinase C (KHK-C), a key enzyme in fructose metabolism, is a promising therapeutic target. α-Mangostin, a naturally occurring prenylated xanthone, has been identified as an effective KHK-C inhibitor, prompting exploration of its analogs for enhanced efficacy. This study aimed to identify α-Mangostin analogs with improved inhibitory properties against KHK-C to address these disorders. Methods: A library of 1383 analogs was compiled from chemical databases and the literature. Molecular docking, binding free energy calculations, pharmacokinetic assessments, molecular dynamics simulations, and quantum mechani-cal analyses were used to screen and evaluate the compounds. α-Mangostin's binding affinity (37.34 kcal/mol) served as the benchmark. Results: Sixteen analogs demonstrated binding affinities superior to α-Mangostin (from -45.51 to -61.3 kcal/mol), LY-3522348 (-45.36 kcal/mol), and reported marine-derived inhibitors (from -22.74 to -51.83 kcal/mol). Hits 7, 8, 9, 13, and 15 not only surpassed these benchmarks in binding affinity, but also exhibited superior pharmacokinetic properties compared to α-Mangostin, LY-3522348, and marine-derived inhibitors, indicating strong in vivo potential. Among these, hit 8 emerged as the best performer, achieving a binding free energy of -61.30 kcal/mol, 100% predicted oral absorption, enhanced metabolic stability, and stable molecular dynamics. Conclusions: Hit 8 emerged as the most promising candidate due to its superior binding affinity, favorable pharmacokinetics, and stable interactions with KHK-C. These findings highlight its potential for treating fructose-driven metabolic disorders, warranting further experimental validation.

Targeting EGFR by Newer 1-(3,5-Bis((E)-4‑hydroxy-3-methoxystyryl)-1H-pyrazol-1-yl)-2-((substituted phenyl)amino)ethan-1-one Analogues for the Treatment of Cancer: Synthesis, In-vitro and In-silico Studies

Aging is a complex natural process and an inherent part of the life cycle. The continuous aging and accumulation of cells contribute to the degradation of the extracellular matrix, which further accelerates the aging process. Various topical agents have been utilized to combat aging, primarily through anti-aging treatments. Examples include the use of topical agents such as retinoids (retinol), vitamin C, hyaluronic acid, alpha hydroxy acids (AHA), and beta hydroxy acids (BHA). This study aims to explore the potential anti-aging properties of compounds found in the quinine-producing plants Cinchona ledgeriana (Ledger) and Cinchona succirubra (Succi) through a molecular approach using the molecular docking method to assess their activity against aging-related proteins. The tested ligands in this study include quinine, cinchonine, quinidine, and cinchonidine, with retinol as the antagonist ligand. In silico analysis suggests that quinine, cinchonine, quinidine, and cinchonidine from Cinchona ledgeriana (Ledger) and Cinchona succirubra (Succi) exhibit strong potential as anti-aging agents by targeting the proteins hyaluronidase receptor (1FCV), elastase (1Y93), collagenase (2D1N), tyrosinase (2Y9X), and HSP90 (5XRB). Molecular docking simulations using MOE 2015.10 revealed quinine's superior binding affinity, with free energy values ranging from -4.9 to -6.8 kcal/mol across target proteins. Critical interactions with His263 (tyrosinase) and Asp206 (elastase)were identified through LigPlot analysis. The docking analysis confirmed the reliability of the methodology, demonstrating that quinine, quinidine, cinchonine, and cinchonidine strongly bind to key skin-ageing enzymes. This study demonstrates the potential of Cinchona bark-derived phytochemicals as anti-aging agents, with quinine exhibiting superior inhibitory activity against tyrosinase. Quinine demonstrated the strongest inhibition against tyrosinase (ΔG = -6.84 kcal/mol), outperforming reference ligands and other Cinchona alkaloids. These findings provide a scientific basis for further exploration of these compounds in mitigating skin aging and enhancing dermatological health. Keywords: Antiaging, in silico, molecular docking, quinine

D-Pantothenic acid (vitamin B5) has wide applications in the feed, food, chemical, and pharmaceutical industries. Its biological production routes which employ pantothenate synthetase (PS) as the key enzyme are attractive since they avoid the tedious and time-consuming optical resolution process. However, little data is available on the activity and kinetics of this enzyme, hampering the rational selection of an efficient enzyme for the biological production of D-pantothenic acid. In this study, six phylogenetically distant PS-encoding genes, from Escherichia coli, Corynebacterium glutamicum, Bacillus subtilis, Bacillus thuringiensis, Bacillus cereus, and Enterobacter cloacae, were expressed in E. coli. The PS from C. glutamicum exhibited a specific activity of 205.1U/mg and a turnover number of 127.6s-1, which to our best knowledge are the highest values ever reported. The addition of substrates (D-pantoic acid and β-alanine) to the E. coli strain harboring this enzyme during the early log phase of fermentation resulted in the production of 97.1g/L of D-pantothenic acid within 32h, corresponding to a conversion yield of 99.1% and a productivity of 3.0g/L/h. To the best of our knowledge, this is the highest productivity reported to date.

Urinary tract infections persist as recurring maladies in human health, triggered by diverse bacterial species. The rise of antibiotic resistance necessitates novel therapeutic agents. This investigation delves into the experimental and theoretical exploration of three compounds—Methyl ganoderate B (A1), 12-acetoxy-15-hydroxy-3,7,11,23-tetraoxolanost-8-en-26-oic acid (A2), and 15-hydroxy-3,7,11,23-tetraoxolanost-8,20-dien-26-oic acid (A3)—via Density Functional Theory (DFT). Leveraging geometrical optimization, spectroscopic (FT-IR, LC–MS) analysis, electronic property studies in polar (water) and non-polar (cyclohexane) solvents, we uncover their solvent-dependent stability and reactivity. Quantum descriptors reveal A1’s elevated reactivity (−7.113 eV energy gap), while A2 showcases enhanced stability (−4.981 eV energy gap). Molecular docking investigations employing significant Escherichia coli adhesion proteins (PDB: 5LNE and 5LNE) spotlight the compounds’ superior binding affinities over the standard drug (sulfamethoxazole). ADMET studies unveil the compounds’ enhanced druglikeness against E. coli-caused urinary tract infections. Notably, predicted toxicity evaluation assigns A1, A2, and A3 LD50 values of 5000 mg/kg, 6802 mg/kg, and 500 mg/kg, respectively, aligning with toxicity classes 5, 6, and 4. Demonstrating non-hepatotoxic, non-cytotoxic, non-carcinogenic, and non-mutagenic attributes, this study underlines the substantial potential of the investigated compounds as robust agents against urinary tract infections.

With 19.3 million new cases and almost 10 million deaths in 2020, cancer has become a leading cause of death today. Curcumin and its analogues were found to have promising anticancer activity. Inspired by curcumin’s promising anticancer activity, we prepared three semi-synthetic analogues by chemically modifying the diketone function of curcumin to its pyrazole counterpart. The curcumin analogues (3a–c) were synthesized by two different methods, followed by their DFT analyses to study the HOMO/LUMO configuration to access the stability of compounds (∆E = 3.55 to 3.35 eV). The curcumin analogues (3a–c) were tested for antiproliferative activity against a total of five dozen cancer cell lines in a single (10 µM) and five dose (0.001 to 100 µM) assays. 3,5-Bis(4-hydroxy-3-methoxystyryl)-1H-pyrazole-1-yl-(phenoxy)ethanone (3b) and 3,5-bis(4-hydroxy-3-methoxystyryl)-1H-pyrazole-1-yl-(2,4-dichlorophenoxy)ethanone (3c) demonstrated the most promising antiproliferative activity against the cancer cell lines with growth inhibitions of 92.41% and 87.28%, respectively, in a high single dose of 10 µM and exhibited good antiproliferative activity (%GIs > 68%) against 54 out of 56 cancer cell lines and 54 out of 60 cell lines, respectively. The compound 3b and 3c demonstrated the most potent antiproliferative activity in a 5-dose assay with GI50 values ranging between 0.281 and 5.59 µM and 0.39 and 0.196 and 3.07 µM, respectively. The compound 3b demonstrated moderate selectivity against a leukemia panel with a selectivity ratio of 4.59. The HOMO-LUMO energy-gap (∆E) of the compounds in the order of 3a > 3b > 3c, was found to be in harmony with the anticancer activity in the order of 3c ≥ 3b > 3a. Following that, all of the curcumin analogues were molecular docked against EGFR, one of the most appealing targets for antiproliferative activity. In a molecular docking simulation, the ligand 3b exhibited three different types of interactions: H-bond, π-π-stacking and π-cationic. The ligand 3b displayed three H-bonds with the residues Met793 (with methoxy group), Lys875 (with phenolic group) and Asp855 (with methoxy group). The π-π-stacking interaction was observed between the phenyl (of phenoxy) and the residue Phe997, while π-cationic interaction was displayed between the phenyl (of curcumin) and the residue Arg841. Similarly, the ligand 3c displayed five H-bonds with the residue Met793 (with methoxy and phenolic groups), Lys845 (methoxy group), Cys797 (phenoxy oxygen), and Asp855 (phenolic group), as well as a halogen bond with residue Cys797 (chloro group). Furthermore, all the compound 3a–c demonstrated significant binding affinity (−6.003 to −7.957 kcal/mol) against the active site of EGFR. The curcumin analogues described in the current work might offer beneficial therapeutic intervention for the treatment and prevention of cancer. Future anticancer drug discovery programs can be expedited by further modifying these analogues to create new compounds with powerful anticancer potentials.

In the tumor microenvironment (TME), the accumulation of adenosine inhibits anti-tumor immune responses, thereby reducing the effectiveness of immune-based cancer therapies. CD39 is involved in inducing excessive adenosine production in tumor cells and immune cells, causing an immune-suppressive environment. As a result, cancer cells are able to evade the immune system, and immune cells are impaired in their ability to kill cancer cells. Therefore, the discovery of outstanding antibodies targeting CD39 has great potential for cancer immunotherapy. To this end, we have discovered antibodies that effectively inhibit the hydrolysis of ATP to adenosine.We confirmed the binding affinity of the anti-CD39 mAb to the CD39 antigen. BLI measurements revealed that anti-CD39 mAb had binding kinetics of KD = 9.40 x 10-11 M, which is better than those of comparative antibodies, competitor 1 (2.39x10-10 M) and competitor 2 (2.27x10-9 M). The cell binding affinity of anti-CD39 mAb was evaluated using SK-MEL-28 cells, which endogenously express high levels of surface CD39. The results showed that the anti-CD39 mAb exhibited superior cell binding affinity compared to the two comparative antibodies. Furthermore, anti-CD39 mAb demonstrated greater efficacy in inhibiting ATP hydrolysis by membrane CD39 and soluble CD39 when compared to the comparative antibodies. In vitro functional assays using human PBMCs demonstrated that inhibiting CD39 enhances T cell proliferation. The anti-CD39 mAb showed superior efficacy in promoting proliferation in CD4+ cells compared to the competitor antibody. Similarly, in CD8+ cells, anti-CD39 mAb exhibited more efficacy compared to competitor 1 antibody. In conclusion, our data indicates that anti-CD39 mAb exhibits excellent characteristics, including superior antigen binding affinity, inhibitory efficacy of ATP hydrolysis and promotion of T cell proliferation. In the future, we will validate the efficacy and stability of this antibody in preclinical studies. The use of this antibody is expected to contribute to immune-based cancer therapy by inhibiting the generation of adenosine in the tumor microenvironment. Citation Format: Seonmi Yu, Wanki Park, Jaeho Song, Sooyoung Kim, Bumchan Park. Discovery and in vitro efficacy validation of anti-CD39 monoclonal antibody for improving the tumor microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB348.