Binding Affinity Research Articles

GATA3-Driven ceRNA Network in Lung Adenocarcinoma Bone Metastasis Progression and Therapeutic Implications

Background/Objectives: Bone metastasis is a common and severe complication of lung adenocarcinoma (LUAD), impacting prognosis and treatment outcomes. Understanding the molecular mechanisms behind LUAD bone metastasis (LUADBM) is essential for developing new therapeutic strategies. The interactions between long non-coding RNAs (lncRNAs), microRNAs (miRNAs), and mRNAs in the competing endogenous RNA (ceRNA) network are crucial in cancer progression and metastasis, but the regulatory mechanisms in LUADBM remain unclear. Methods: Microarray analysis was performed on clinical samples, followed by weighted gene co-expression network analysis (WGCNA) and construction of a ceRNA network. Molecular mechanisms were validated using colony formation assays, transwell migration assays, wound healing assays to assess cell migration, and osteoclastogenesis assays to evaluate osteoclast differentiation. Potential therapeutic drugs and their binding affinities were predicted using the CMap database and Kdeep. The interaction between the small-molecule drug and its target protein was confirmed by surface plasmon resonance (SPR) and drug affinity responsive target stability (DARTS) assays. Mechanistic insights and therapeutic efficacy were further validated using patient-derived organoid (PDO) cultures, drug sensitivity assays, and in vivo drug treatments. Results: Our results identified the XLOC_006941/hsa-miR-543/NPRL3 axis as a key regulatory pathway in LUADBM. We also demonstrated that GATA3-driven Th2 cell infiltration creates an immunosuppressive microenvironment that promotes metastasis. Additionally, we confirmed that the inhibitor E7449 effectively targets NPRL3, and its combination with the IL4R-blocking antibody dupilumab resulted in improved therapeutic outcomes in LUADBM. Conclusions: These findings offer new insights into the molecular mechanisms of LUADBM and highlight potential therapeutic targets, including the XLOC_006941/miR-543/NPRL3 axis and GATA3-driven Th2 cell infiltration. The dual-target therapy combining E7449 with dupilumab shows promise for improving patient outcomes in LUADBM, warranting further clinical evaluation.

Effects of Montelukast on Neuroinflammation in Parkinson's Disease: An Open Label Safety and Tolerability Trial with CSF Markers and [11C]PBR28 PET.

Dysregulated leukotriene signaling is proposed to be involved in pathogenesis of Parkinson's disease (PD). The objective was to examine the safety and tolerability of montelukast, a cysteinyl-leukotriene receptor1 and GPR17 antagonist, in patients with PD. Secondary outcomes were target engagement, effects on PD signs/symptoms, and central neuroinflammation. Fifteen PD patients were recruited to a 12-week open-label trial of 20 mg bi-daily montelukast treatment. Patients underwent ratings with the Movement Disorder Society Unified Parkinson Disease Rating Scale (MDS-UPDRS), the Montreal Cognitive Assessment (MoCA), Beck's Depression Inventory (BDI), Parkinson's Disease Questionnaire-39 (PDQ-39), [11C]PBR28-PET, and lumbar punctures before and during montelukast treatment. All patients completed the study. Three patients reported loose stool. No serious adverse events related to treatment were reported. MDS-UPDRS-Total scores improved by 6.9 points. Very low levels of montelukast were detected in all cerebrospinal fluid (CSF) samples and resulted in a reduction in inflammation/metabolism markers. [11C]PBR28 binding was lowered in high, but not mixed, affinity binders after montelukast. Montelukast crosses the blood-brain barrier at very low levels and is well tolerated and safe in PD patients. Preliminary effects on neuroinflammation and clinical scores motivate a future randomized controlled trial (RCT) in PD. © 2025 The Author(s). Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

Insilico, network pharmacology and neural network studies on Sida acuta Burm f.- based phytochemicals targeting NADH-ubiquinone oxidoreductase

The biochemical activities of phytochemicals obtained from S. acuta Burm f. have been described in several forms by various scientists globally. This work has revealed the antimalarial activities of S. acuta Burm f. using various methods such as density functional theory method, induced fit docking technique, quantitative structure activities relationship (QSAR) via neural network approach and pharmacokinetics studies using ADMETLab software. Compound F10 showed a stronger ability to interact in terms of HOMO energy and energy gap while Compound F12 exhibited a high tendency to accept electrons from nearby compounds. Furthermore, compound F13 demonstrated greater potential to inhibit NADH-Ubiquinone Oxidoreductase. We observed that the derivatives in F10, F12 and F13 enhanced the reactivity and inhibiting activities of the parent compound. The prediction of binding affinity for compounds F1 to F16 using neural network was found to be accurate and dependable. Also, ADMET study was executed and reported appropriately. Our research could pave the way for the creation of a library of effective phytochemicals from S. acuta Burm f -based drugs with potential anti-malaria activity.

Development and evaluation of [11C]DPA-813 and [18F]DPA-814: novel TSPO PET tracers insensitive to human single nucleotide polymorphism rs6971.

The translocator protein 18kDa (TSPO) is a widely used marker for imaging neuroinflammation via Positron Emission Tomography (PET). However, the vast majority of reported TSPO PET tracers display low binding affinity to a common isoform of human TSPO (rs6971; A147T), making them unsuitable for universal use in the general population. In this study, we have developed and preclinically validated two novel tracers designed to image TSPO in patients of all genotypes. Novel analogues of known TSPO ligands were synthesised, evaluated for TSPO binding affinity in vitro (membranes prepared from transfected HEK-293T cells expressing wild-type (WT) or A147T TSPO) and radiolabelled with carbon-11 or fluorine-18. They were evaluated in situ (autoradiography on genotyped human brain tissue) and in vivo (rat, both WT and clinically relevant experimental autoimmune encephalomyelitis (EAE) neuroinflammation model) as potential polymorphism-insensitive TSPO PET tracers. Two new TSPO ligands, DPA-813 and DPA-814, displayed equivalent single-digit nanomolar binding affinities in vitro towards both human TSPO isoforms. [11C]DPA-813 and [18F]DPA-814 were synthesised in moderate radiochemical yields, high radiochemical purity, and high molar activity. Autoradiography on human MS tissues showed high specific binding for both tracers, irrespective of the TSPO isoform. The tracers demonstrated high plasma stability after 45min and no brain metabolism with > 99% intact tracer. Biodistribution in WT animals indicated good brain uptake for both tracers (0.28 and 0.41%ID/g for [18F]DPA-814 and [11C]DPA-813, respectively). PET imaging in the clinically relevant EAE neuroinflammation model in rats showed significantly higher uptake of [11C]DPA-813 and [18F]DPA-814 in the spinal cord of the EAE rats compared to the controls. We have developed two novel PET tracers that display indiscriminately high binding affinity to both common isoforms of human TSPO, show favourable metabolic stability and brain penetration in rats, and significantly higher uptake in the spinal cord of a neuroinflammatory rat model of multiple sclerosis. Going forward, first-in-human clinical validation will mark a critical juncture in the development of these tracers, which could offer substantial improvements over existing imaging tools for detecting neuroinflammation, irrespective of genetic variations.

Nano-Polymers as Cas9 Inhibitors

Despite wide applications of CRISPR/Cas9 technology, effective approaches for CRISPR delivery with functional control are limited. In an attempt to develop a nanoscale CRSIPR/Cas9 delivery platform, we discovered that several biocompatible polymers, including polymalic acid (PMLA), polyglutamic acid (PGA), and polyaspartic acid (PLD), when conjugated with a trileucine (LLL) moiety, can effectively inhibit Cas9 nuclease function. The Cas9 inhibition by those polymers is dose-dependent, with varying efficiency to achieve 100% inhibition. Further biophysical studies revealed that PMLA-LLL directly binds the Cas9 protein, resulting in a substantial decrease in Cas9/sgRNA binding affinity. Transmission electron microscopy and molecular docking were performed to provide a possible binding mechanism for PMLA-LLL to interact with Cas9. This work identified a new class of Cas9 inhibitor in nano-polymer form. These biodegradable polymers may serve as novel Cas9 delivery vehicles with a potential to enhance the precision of Cas9-mediated gene editing.

Exploring precision therapeutics: computational design of antisense oligonucleotides targeting AXL gene transcripts in multiple sclerosis treatment management

Multiple sclerosis (MS) is a chronic autoimmune illness characterized by demyelination, neurodegeneration, and inflammation in the central nervous system. The AXL gene, which codes for a receptor tyrosine kinase, has emerged as a promising therapeutic target due to its involvement in neuroinflammation and oligodendrocyte dysfunction. In the current study, we employed in silico techniques to design Antisense Oligonucleotides (ASOs) that selectively target AXL gene transcripts to modulate AXL expression and mitigate MS pathology. Three ASOs, A1, A2, and A3, were designed to specifically target the 5′ untranslated region (5′UTR) and coding region of the AXL gene transcripts. The ASOs were optimized with a focus on stability, binding affinity, and specificity towards AXL mRNA while minimizing off-target effects. To investigate ASO-mRNA interactions and gauge their ability to alter AXL expression, Molecular Docking was performed. Our analyses showed that A1, A2, and A3 had substantial interactions with AXL mRNA, with binding affinities of −9.5 kcal/mol, −10.8 kcal/mol, and −10.6 kcal/mol, respectively. The targeting of AXL gene transcripts through ASOs shows promise in reducing MS symptoms. Precision ASO-based therapies could effectively manage MS by targeting the essential pathways involved in the disease. ASOs provide a highly targeted approach for treating MS and offer a precise therapeutic strategy for this debilitating condition. The study lays the groundwork for future in vitro and in vivo studies to confirm the therapeutic potential of these ASOs for the treatment of MS.

Mycochemistry, antioxidant, anticancer activity, and molecular docking of compounds of F12 of ethyl acetate extract of Astraeus asiaticus with BcL2 and Caspase 3

Globally the current research is going for cancer disease management by different strategies as frequency and severity of this disease are increasing day by day. The fruit body of edible and tasty Astraeus asiaticus mushroom was extracted in ethyl acetate solvent (EA) and quantitative analysis revealed that it contained a significant amount of total phenols, flavonoids, and ascorbic acids. The FT-IR study revealed different functional groups in the extracts with different characteristic peak values. The GC/MS profile of AAEA (Astraeus asiaticus ethyl acetate) extract exhibited 61 compounds. The column chromatography of AAEA extract was performed and the F12 fraction demonstrated the greatest radical scavenging activity with an EC50 of 25.65 ± 4.82 µg. mLˉ¹. Mycochemistry (GC and mass spectrum) analysis showed that F12 was a mixture of six important compounds like Hexadecanoic acid, 3,4,5,6 Tetramethyloctane, 9,12-Octadecadienoic acid, 9,12-Octadecadienoic acid, methyl ester, 1-cyclododecyne, cis-9,10-Epoxyoctadecan-1-ol. The chemical properties of all six compounds were screened by SwissADME and pkCSM and AdmetSAR predictors. Out of them Hexadecanoic acid, 9,12-Octadecadienoic acid and 3,4,5,6 Tetramethyloctane, exhibited suitable properties for drug -preparation and they showed anticancer activity and antioxidant activity as per NIST data base and library search. We have tried to focus on anticancer compounds derived from the partial purification (F12) of mushroom extract (AAEA) from this edible mushroom against cancer (cervical, lung, and breast) cell lines. After 24 h of treatment, the percentages of cell growth inhibition of HeLa, MCF-7, and A549 cell lines by highest concentration (1500 µg. mL− 1) of F12 were 92.03 ± 6.21 a, 90.38 ± 4.53a, and 87.51 ± 5.36a % respectively and the IC50 values were 701 ± 11.54, 728.71 ± 10.53, and 806.88 ± 11.52 µg. mL− 1 respectively but the growth of normal cell HEK 293T was inhibited slightly (3.0%). The mechanism of anticancer effect of F12 (AAEAE) on cancer cell lines included induction of apoptosis, LDH leakage, and up regulation of gene expression levels of Caspase 3, Caspase 9, P53, and down regulation of BcL2 of all three cell lines. Molecular docking of the three important compounds (Hexadecanoic acid, 3,4,5,6 Tetramethyloctane and 9,12-Octadecadienoic acid), with apoptotic protein Caspase 3 and antiapoptotic protein BcL2 was done to find out the binding affinity, stability and drug- likeness properties of these chemicals. In conclusion, F12 fraction of AAEA extract of this mushroom containing six bioactive compounds was a promising antioxidant and anticancer agent and the use of this fraction in cancer treatment will be a novel study for future drug development.

Synthesis, Characterization, Cytotoxicity, Biological Evaluation, DFT Calculations, and Molecular Docking of a Novel Schiff Base and Its Pt(IV) Complex

ABSTRACTA novel Schiff base ligand derived from 1,2,4‐triazole‐3‐thione was synthesized and complexed with platinum (IV) ion to enhance its bioactivity. Characterization techniques, including FTIR, NMR, UV‐Vis, and mass spectrometry, confirmed the formation of Schiff base molecule and revealed that its Pt(IV) complex has an octahedral geometry. The antibacterial and antifungal activity of the synthesized compounds was evaluated against Staphylococcus epidermidis, Pseudomonas aeruginosa, Candida albicans, and Aspergillus flavus, with the metal complex outperforming the ligand. The antioxidant properties were evaluated using a DPPH assay, with the Pt(IV) complex demonstrating superior radical‐scavenging ability. Cytotoxicity studies via MTT assays on MCF‐7 breast cancer cells showed dose‐dependent reductions in cell viability, with the Pt(IV) complex inducing enhanced antiproliferative effects compared with the ligand. DFT calculations were performed to investigate the electronic properties of the compounds, and molecular docking studies were conducted against the alpha estrogen receptor (PDB ID: 3ERT) to predict their binding affinities. The results demonstrated that the Pt(IV) complex exhibited higher biological activity than the free ligand, suggesting its potential as an effective therapeutic agent.

Clinical and Research Applications of FAPI Tracers: A Review.

In the last decade, fibroblast activation protein-α inhibitors (FAPIs), which target the cancer-associated fibroblasts of the tumour microenvironment, have become a topic of great interest. In oncology, FAPI PET/CT imaging has repeatedly demonstrated a higher lesion detection rate than other conventional imaging modalities such as CT or 18F-FDG PET/CT for several tumours. In some cases, the initial staging and therapeutic management may even change. Some FAPI radioligands may also be labelled with therapeutic radionuclides for theranostic applications. It is thus possible to treat certain metastatic cancers with FAPI radioligand therapy (FAPI-RLT), which is generally well tolerated with little toxicity. Recently, new FAPIs have been developed with the particularity of having a higher binding affinity the target, which further improves the lesion detection rate on PET/CT and clinical outcomes following FAPI-RLT. This review provides recent updates in the clinical use of FAPI PET/CT and FAPI-RLT, and discusses potential emergent applications, including in inflammation imaging.

Effect of cubebin against streptozotocin-induced diabetic nephropathy rats via inhibition TNF-α/NF-κB/TGF-β: in vivo and in silico study

Cubebin, a dibenzyl butyrolactone lignan belonging to several distinct families, including Aristolochiaceae, Myristicaceae, Piperaceae, and Rutaceae, and possesses several pharmacological activities, including analgesic, anti-inflammatory, antioxidant, and vasodilatory. The current study aimed to evaluate the effect of cubebin on streptozotocin (STZ)-evoked diabetic nephropathy (DN). DN is a well-identified complication of diabetes mellitus (DM) characterized by renal hypertrophy that progressively declines kidney function. Wistar rats were randomly divided into groups- normal, STZ control (65 mg/kg/body weight), and STZ + cubebin (10 and 20 mg/kg). Biochemical parameters such as glucose levels, kidney parameters, lipid profile, oxidative stress, endogenous antioxidant markers, inflammatory cytokines and histopathology were performed. Molecular docking [(PDB ID: TNF-α (7JRA), NF-κB (1SVC), TGF-β1 (3TZM)] and dynamic simulation (MDS) were also performed with the selected target. STZ-induced DN was changes in these parameters. In contrast, DN + cubebin at 10 and 20 mg/kg doses improved the biochemical parameters and histological changes. Furthermore, molecular docking and simulation studies showed a binding affinity with negative binding energy with TNF-α (7jra, − 11.342 kcal/mol), TGF-β1 (3tzm, − 9.162 kcal/mol) and NF-κB (1svc, − 6.665 kcal/mol). The results of MDS provided insight into the mechanisms that associate proteins TNF-α, NF-κB, and TGF-β1 in conformational dynamics upon binding to cubebin. In conclusion, these findings exhibit a potential effect of cubebin in STZ-evoked DN rats.

Investigating the aging-modulatory mechanism of Rasayana Churna, an Ayurvedic herbal formulation, using a computational approach.

This study investigates the impact and mechanisms of Rasayana Churna, an Ayurvedic poly-herbal formulation, in treating aging-related disorders through text mining, network pharmacology, molecular docking simulation, Super-MMPBSA, and density functional theory. The text mining of Rasayana Churna highlighted the diverse therapeutic potential of Phyllanthus emblica, Tinospora cordifolia, and Tribulus terrestris in managing aging-related disorders through their antidiabetic, antioxidant, and anti-inflammatory properties. Using network pharmacology, 17 bioactive compounds and 137 corresponding potential targets of Rasayana Churna were identified and used to construct protein-protein interaction and hub gene networks. Key targets such as AKT1, BCL2, ESR1, and GSK3B were linked to aging-related pathways, with GO and KEGG enrichment analyses highlighting processes like apoptosis, oxidative stress response, and pathways like PI3K-Akt signaling. Molecular docking analysis identified 14 compounds with strong binding affinity toward the key aging target AKT1. Three bioactive compounds-Kaempferol, N-Caffeoyltyramine, and Multifidol glucoside-exhibited superior stability and binding interactions in MD simulations, confirmed by RMSD, RMSF, Rg, hydrogen bonding, SASA, PCA, and free energy landscape analysis. Super-MMPBSA (last 30ns) calculation was performed to analyze dynamic behavior and protein-ligand stability, revealing significantly lower ΔG binding free energy values for the three hit compounds (-177.871, -164.855, -199.649kJ/mol, respectively) compared to the AKT1-reference complex (-109.463kJ/mol). DFT analysis revealed favorable electronic properties and kinetic stability for these compounds. Integrating traditional Ayurvedic knowledge with computational techniques suggests Rasayana Churna could prevent and manage aging-related conditions. However, further in vitro, in vivo, and clinical studies are needed to validate its aging-modulatory potential.

Essential Oils From Chromolaena odorata (L.) R. M. King and H. Robinson Stem Barks and Leaves: Chemical Analysis, Biological Activity and In Silico Approach.

The current study first describes the chemical profiles of essential oils from Vietnamese Chromolaena odorata fresh stem barks and leaves. The gas chromatography-flame ionization detection/mass spectrometry (GC-FID/MS) analysis revealed that α-pinene (6.97%-38.91%), β-pinene (5.68%-11.64%), geijerene (10.60%-26.68%), germacrene D (9.33%-16.87%), and (E)-caryophyllene (5.03%-9.12%) were the major compounds. Essential oils showed strong anti-inflammation against NO (nitric oxidative) production in lipopolysaccharide (LPS)-stimulated RAW264.7 cells with IC50 values of 42.44-49.01µg/mL. Two essential oils also exhibited cytotoxicity against four cancer cell lines, K562, HeLa, HepG2, and MCF-7, antioxidant activity to capture DPPH radicals, and antidiabetics in α-glucosidase inhibition. Both samples showed varying levels in the antimicrobial experiment, especially the leaf essential oil, which showed strong antibacterial activity against the bacterium Bacillus subtilis with a MIC value of 64µg/mL. From the molecular docking investigation, geijerene had a higher affinity for cyclooxygenase-2 (COX-2) than the other main compounds, whereas germacrene D had the strongest binding affinity for two protein targets, COX-2, and B. subtilis FtsZ. It was discovered that the main factor stabilizing the bindings of these compounds to the target proteins was hydrophobic interactions.

Identifying natural inhibitors against FUS protein in dementia through machine learning, molecular docking, and dynamics simulation

Dementia, a complex and debilitating spectrum of neurodegenerative diseases, presents a profound challenge in the quest for effective treatments. The FUS protein is well at the center of this problem, as it is frequently dysregulated in the various disorders. We chose a route of computational work that involves targeting natural inhibitors of the FUS protein, offering a novel treatment strategy. We first reviewed the FUS protein's framework; early forecasting models using the AlphaFold2 and SwissModel algorithms indicated a loop-rich protein—a structure component correlating with flexibility. However, these models showed limitations, as reflected by inadequate ERRAT and Verify3D scores. Seeking enhanced accuracy, we turned to the I-TASSER suite, which delivered a refined structural model affirmed by robust validation metrics. With a reliable model in hand, our study utilized machine learning techniques, particularly the Random Forest algorithm, to navigate through a vast dataset of phytochemicals. This led to the identification of nimbinin, dehydroxymethylflazine, and several other compounds as potential FUS inhibitors. Notably, dehydroxymethylflazine and cleroindicin C identified during molecular docking analyses—facilitated by AutoDock Vina—for their high binding affinities and stability in interaction with the FUS protein, as corroborated by extensive molecular dynamics simulations. Originating from medicinal plants, these compounds are not only structurally compatible with the target protein but also adhere to pharmacokinetic profiles suitable for drug development, including optimal molecular weight and LogP values conducive to blood-brain barrier penetration. This computational exploration paves the way for subsequent experimental validation and highlights the potential of these natural compounds as innovative agents in the treatment of dementia.

Investigation of Pyrrole Analogues Inhibition on Main Protease by Spectroscopic Analysis, Molecular Docking, and Enzyme Activity.

The main protease (Mpro) is a cysteine enzyme and represents a vital target for antiviral drug screening. In this work, 25 pyrrole derivatives were synthesized and screened by enzyme activity experiments. Results indicate that six pyrrole derivatives can bind to Mpro and have an inhibitory effect on Mpro. The binding mode involves a static quenching process. Among them, 1d exhibits the highest binding affinity. The interactions between pyrrole derivatives and Mpro are investigated by spectra and molecular docking. The interaction between 1d and Mpro is spontaneous and enthalpy-driven. Hydrogen bonding is identified as the primary binding force for 1d. Furthermore, nitro groups are important for the binding between pyrrole analogs and Mpro. This study elucidates the mechanism of interactions and provides direction and valuable insights for developing novel Mpro inhibitors.

Column Chromatography-Free Synthesis of Spirooxindole and Spiroindanone-Based Naphthalimides as Potent c-MYC G4 Stabilizers and HSA Binders for Elevating Anticancer Potential.

G-quadruplex (G4) DNA plays a pivotal regulatory role in fundamental biological processes, integral for governing cellular functions such as replication, transcription, and repair in living cells. Within cancer cells, G4 DNA exerts an impact on the expression of crucial genes such as c-MYC, effectively repressing its activity when structured within its promoter region. Therefore, employing molecular scaffolds to target these structures offers an attractive strategy for altering their functions. In our pursuit of potent and selective G-quadruplex binders, herein we report a series of spironaphthalimide-pyrrolidine analogues that demonstrate the ability to stabilize c-MYC G4 formation and subsequently inhibit c-MYC expression. These analogues are evaluated for their anticancer activity against 60 human cancer cell lines at 10 μM. The most potent analogues 8j and 21c underwent additional testing at five dose concentrations (10-4-10-8 M) where low MG-MID GI50 values are observed for both the analogues 8j (9.98 μM) and 21c (2.49 μM). To correlate with the antiproliferative activity, the mechanism is explored in vitro by performing Pu27 DNA binding studies through multispectroscopic techniques, and the results are compared with Pu22, human telomere, and calf thymus DNA. Additionally, insights from molecular docking suggested stacking over the G-tetrad of G4 structures of both analogues, with quantum mechanical studies further reinforcing the rationale for the stability of this quadruplex secondary structure. The analogues are also evaluated for their binding affinity to human serum albumin, revealing their robust capability to effectively bind and potentially facilitate targeted delivery to specific sites. Amidst the abundance of G4s across the human genome, the above findings underscore the significance of spiro analogues, with potent multitargeting anticancer attributes, marking a transformative leap forward in G4-ligand innovation, promising frontiers in the quest for effective anticancer modalities.

Chemosensory Proteins Protect Nilaparvata lugens from Imidacloprid by Sequestering the Insecticide and Facilitating Metabolic Detoxification.

The involvement of chemosensory proteins (CSPs) in binding to insecticides has been implicated. However, our understanding of CSP-mediated insecticide resistance remains limited. Herein, 15 CSP genes were identified and characterized from Nilaparvata lugens. Expression analysis identified six CSPs with overexpression in the imidacloprid-resistant strain, whose involvement in imidacloprid resistance was validated by RNA interference. Among them, four CSPs were successfully expressed using a prokaryotic expression system, and their binding affinities to imidacloprid were confirmed through fluorescence competitive binding assays. Knockdown of them impaired the capacity of N. lugens to metabolize imidacloprid and inhibited the activity of metabolic detoxification pathways, while their overexpression in Escherichia coli enhanced bacterial metabolic efficiency toward imidacloprid. Furthermore, the transcriptional regulation of CSP2 and CSP15 was found to be mediated by AhR/ARNT and CncC/MafK. These findings suggest that the overexpression of CSPs in N. lugens promotes imidacloprid resistance by sequestering the insecticide and enhancing metabolic detoxification.

In silico, in vitro, and in vivo characterization of thiamin-binding proteins from plant seeds.

Thiamin, an essential micronutrient, is a cofactor for enzymes involved in the central carbon metabolism and amino acid pathways. Despite efforts to enhance thiamin content in rice by incorporating thiamin biosynthetic genes, increasing thiamin content in the endosperm remains challenging, possibly due to a lack of thiamin stability and/or a local sink. The introduction of storage proteins has been successful in several biofortification strategies, and similar efforts targeting thiamin have been performed, leading to a 3-4-fold increase in white rice. However, only one thiamin-binding protein (TBP) sequence has been described in plants, more specifically from sesame seeds. Therefore, we aimed to identify and characterize TBPs, as well as to evaluate the effect of their expression on thiamin concentration, using a comprehensive approach integrating in silico, in vitro, and in vivo methods. We identified the sequences of putative TBPs from Oryza sativa (Os, rice), Fagopyrum esculentum (Fe, buckwheat), and Zea mays (Zm, maize) and pinpointed the thiamin-binding pockets through molecular docking. FeTBP and OsTBP contained one pocket with binding affinities similar to the Escherichia coli TBP, a well-characterized TBP, supporting their function as TBPs. In vivo expression studies of TBPs in tobacco leaves and rice callus resulted in increased thiamin levels, with FeTBP and OsTBP showing the most pronounced effects. Additionally, thermal shift assays confirmed the thiamin-binding capabilities of FeTBP and OsTBP, as observed by the significant increases in melting temperatures upon thiamin binding, indicating protein stabilization. These findings offer new insights into the diversity and function of plant TBPs and highlight the potential of FeTBP and OsTBP to modulate thiamin levels in crop plants.

Network pharmacology and experimental verification to investigate the mechanism of isoliquiritigenin for the treatment of Alzheimer’s disease

Isoliquiritigenin (ISL), a flavone isolated from licorice, has been demonstrated to exhibit anti-inflammatory and antioxidant properties in the treatment of Alzheimer’s disease (AD). However, the molecular details of the contribution of ISL to AD remain largely elusive. The present study aimed to investigate the molecular mechanisms of ISL against AD. In this study, AD targets and ISL targets were collected via different databases. The overlapped targets between AD and ISL were generated with Venny. Then we performed Gene Ontology (GO) and Kyoto Encyclopaedia of Genes and Genomes (KEGG) pathway enrichment analyses on these common targets. The protein-protein interaction (PPI) network was constructed and clusters were obtained using the Molecular Complex Detection (MCODE) and the Cytohubba plugins. Further, molecular docking study was performed for these core targets. Subsequently, the receiver operating characteristic (ROC) curve analysis and the assessment of hub gene expression levels between AD and healthy individuals were used to estimate a possible link between target genes in AD. Finally, experiments were conducted to verify the therapeutic mechanism of ISL in lipopolysaccharide (LPS)-induced BV2 microglial cells. GO and KEGG pathway analysis found that ISL was significantly enriched in regulation of mitogen-activated protein kinase (MAPK) signaling pathway. The PPI network manifested 7 key targets including albumin (ALB), epidermal growth factor receptor (EGFR), solute carrier family 2 member 1 (SLC2A1), insulin-like growth factor 1 (IGF1), mitogen-activated protein kinase 1 (MAPK1), peroxisome proliferator activated receptor alpha (PPARA) and peroxisome proliferator activated receptor gamma (PPAR-γ, PPARG). Molecular docking showed that ISL had high binding affinity with these key targets. The experimental results revealed that ISL decreased extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation and increased the expression of PPAR-γ, and suppressed the production of proinflammatory mediators. Our work revealed that ISL might be an effective treatment strategy in the treatment of AD by its anti-inflammatory effect towards microglia through the ERK/PPAR-γ pathway.

Insilico targeting of virus entry facilitator NRP1 to block SARS-CoV2 entry.

The entry and infectivity of a virus are determined by its interaction with the host. SARS-CoV-2, the virus responsible for COVID-19, utilizes the spike (S) protein to attach to and enter host cells. Recent studies have identified neuropilin-1 (NRP1) as a crucial facilitator for the entry of SARS-CoV-2. The binding of the spike protein to the b1 domain of NRP1 has been shown to enhance viral infection twofold. Consequently, targeting NRP1 to disrupt this interaction represents a promising strategy to mitigate viral infection. In this study, a small molecule library of approximately 10,000 compounds was screened to identify those that could inhibit the interaction between NRP1 and the spike protein by targeting the b1 domain of NRP1. The crystallographic structure of the b1 domain of human NRP1 (PDB entry: 7JJC) was used for this purpose. Following virtual screening, docking studies, and evaluation of binding affinity and ADMET properties, 10 compounds were shortlisted. The top two candidates, AZD3839 and LY2090314, were selected for molecular dynamics simulation studies over 100 ns to assess binding stability. MM/GBSA calculations indicated that both AZD3839 and LY2090314 exhibited strong and stable binding to the b1 domain of NRP1. Computational modeling of the interaction between the b1 domain of NRP1 and the receptor-binding domain of the spike protein suggested that AZD3839 and LY2090314 could effectively hinder the NRP1-spike interaction. Therefore, these compounds may serve as potential drug candidates to reduce SARS-CoV-2 infectivity.

Harnessing crRNA Transformer for Facile and Specific Nucleic Acid Detection.

CRISPR/Cas systems have emerged as promising tools for nucleic acid detection. However, their practical applications have been limited by poor specificity and the need for additional preprocessing steps. Inspired by the concept of transformers, we found that changing the forms of crRNA with spatial arrangement may endow CRISPR/Cas with an enhanced performance for nucleic acid detection. Specifically, we rationally designed two crRNA transformers─swap crRNA and split crRNA─and found that they direct the CRISPR/Cas system for cis- and trans- cleavage with enhanced specificity and decreased Cas binding affinity and possess both DNA and RNA detection abilities. Based on these findings, our strategy enabled the identification of clinical prostatic cancer in a one-step reaction, with a remarkable sensitivity of 90.0% and specificity of 96.0%. Our study deepens the understanding of CRISPR/Cas systems and introduces a promising strategy for simple nucleic acid detection with enhanced specificity, sensitivity, and functionality in clinical molecular diagnosis.