Search Results (1,272)

Type 2 diabetes (T2D) is a metabolic syndrome frequently associated with obesity and endoplasmic reticulum stress-mediated inflammation, which can trigger the unfolded protein response (UPR), impair insulin signaling, and promote apoptosis. To identify potential natural therapeutic candidates, this study investigated the mechanisms of action of 14 compounds from Uncaria tomentosa (UT), a medicinal plant from the Amazon rainforest, using in silico modeling. The study focused on the UPR, TRAF2/JNK pro-inflammatory signaling pathway, and insulin signaling pathways, which play key roles in T2D. Some of the UT compounds were docked against several human proteins involved in these pathways, and molecular dynamics simulations confirmed stable interactions between the target proteins (PERK, TRAF2, JNK, TNF-α, IRS-1, PI3K, AKT, GSK3β, and PPARγ) and four of the UT compounds, 5-Carboxystrictosidine, Cinchonain, Epicatechin and Mitraphylline. Additionally, absorption, distribution, metabolism, excretion, and toxicity (ADMET) properties analyses were conducted to predict the four compounds, revealing suitable pharmacokinetic properties. These findings suggest that specific UT compounds may be used in experimental tests to whether investigate their therapeutic potential in managing T2D by modulating signaling pathways related to the conditions UPR, inflammation, and insulin resistance. Full article

Intestinal inflammation can disrupt epithelial integrity, immune homeostasis, and nutrient utilization in laying hens. This entirely in silico study investigated the molecular associations between reported Fraxini Cortex constituents and intestinal inflammation by integrating network pharmacology, chicken-specific bioinformatics, molecular docking, molecular dynamics (MD) simulation, and MM/GBSA analysis. After chemical standardization and SwissADME screening, 38 candidate constituents were retained, yielding 528 nonredundant predicted human targets. Intersection with 1452 intestinal inflammation-related genes identified 170 shared human targets, which were subsequently mapped to 133 nonredundant Gallus gallus homologues. A chicken-specific protein–protein interaction network contained 117 non-isolated nodes and 683 interactions, and combined cytoHubba and MCODE analyses identified 13 core candidate targets, including BCL2, SRC, HIF1A, MMP9, ESR1, EGFR, ALB, TLR4, STAT1, PTGS2, SIRT1, JUN, and PPARG. GO and KEGG enrichment associated these targets with receptor and kinase signaling, innate immune recognition, epithelial adhesion and survival, extracellular-matrix remodeling, lipid-mediator metabolism, apoptosis, autophagy, and related processes. Molecular docking of ten candidate compounds against six prioritized targets generated 60 ligand–target combinations, with AutoDock Vina scores ranging from −7.930 to −3.412 kcal/mol. Caffeic acid–MMP9 and scopoletin–ESR1 were further evaluated using three independent 100 ns MD simulations, which showed broadly reproducible conformational behavior across replicates. MM/GBSA calculations yielded mean endpoint binding-energy estimates of −19.92 ± 0.30 kcal/mol for caffeic acid–MMP9 and −21.53 ± 0.32 kcal/mol for scopoletin–ESR1. These findings computationally prioritize caffeic acid, scopoletin, and a group of chicken homologous targets and pathways potentially associated with intestinal inflammation in laying hens and provide testable hypotheses for subsequent chicken-specific experimental validation. Full article

Heritability in Parkinson’s disease (PD) may be driven by rare variants (RVs), but genetic data for Chinese cohorts remain limited. We investigated the relationship between RVs and the risk and phenotype of PD by performing targeted exome sequencing of 29 PD candidate genes in 311 late-onset, sporadic Chinese PD patients and 699 local controls. A total of 174 RVs (that were likely deleterious using in silico prediction tools) were identified in 27 of the 29 genes sequenced. Mean RV burden was higher (1.178 vs. 0.478, p = 1.26 × 10⁻²⁴) and HLA-DRB5 p.Val104ArgfsTer26 was enriched in PD patients (0.207 vs. 0.000, p < 0.0003) compared with controls. Gene-based RV carrier status in PD patients was then examined for correlation with phenotypic characteristics. The RV carrier status of LRRK2 and HLA-DRB5 was associated with tremor, SREBF1 with gait disturbance, and SYNJ1 and SCARB2 with motor fluctuations. Earlier onset age and neuropsychiatric symptoms were associated with GBA1 variants. Olfactory deficit, autonomic disturbance, anxiety and depression were associated with variants in SLC44A1, HLA-DRB5, and SCARB2 respectively. Our results demonstrated a prominent RV burden in Chinese PD and illustrated the importance of genetic influence on the risk and phenotype of sporadic PD. Further studies are warranted to correlate these genes with putative pathogenic pathways. Full article

Although the Pacific oyster (Crassostrea gigas) is a premium marine protein source, its neuroprotective peptidome remains largely uncharacterized. This study established an integrated in silico and in vitro pipeline to discover acetylcholinesterase (AChE)-targeting peptides with cellular AChE-regulating and neuroprotective peptides from simulated gastrointestinal digests of oyster. Peptidomic profiling identified 18,292 sequences, which were filtered down to seven candidates predicted to have favorable blood–brain barrier (BBB) permeability and to be non-toxic and non-allergenic (VPYPR, VPVHF, HHTF, PVHF, GPKPW, HWF, and KYW) via multi-step virtual screening. In cellular assays, simulated H₂O₂ injury (500 μM) reduced PC12 cell viability to 47.53 ± 4.53%. Compared with the model group, pretreatment with the three most potent candidates—HHTF, VPYPR, and VPVHF (200 μM)—significantly rescued injured cells, restoring cell viability to 88.31 ± 7.83%, 85.12 ± 3.35%, and 82.00 ± 3.47%, respectively (p < 0.05). These peptides effectively fortified cellular antioxidant defenses by increasing glutathione (GSH) levels to 24.24, 30.11, and 26.83 nmol/mg protein (from 20.22 nmol/mg protein in the model group) and superoxide dismutase (SOD) activity to 151.41, 153.97, and 151.96 U/mg protein (from 119.33 U/mg protein), while suppressing malondialdehyde (MDA) accumulation to 0.088, 0.064, and 0.086 nmol/mg protein (from 0.193 nmol/mg protein). Crucially, the peptides alleviated cholinergic dysfunction by normalizing the H₂O₂-induced elevation of intracellular AChE activity (11.39 nmol/min/mg protein) down to 7.02, 6.22, and 7.14 nmol/min/mg protein, respectively. Specifically, VPYPR (200 μM) restored AChE activity to a level (6.22 nmol/min/mg protein) that was not significantly different from that in the normal control group (p > 0.05). Molecular dynamics (MD) simulations (100 ns) and molecular mechanics Poisson–Boltzmann surface area (MM-PBSA) calculations identified VPYPR as the leading candidate with a remarkably low binding free energy of −49.74 ± 3.58 kcal/mol. This study demonstrates that oyster gastrointestinal digests are valuable reservoirs of multi-target neuroprotective ingredients and provides an efficient strategy for marine bioactive peptide discovery. Full article

Exposure to ambient particulate matter (PM) has been linked to renal dysfunction, particularly in diabetic populations, but the underlying mechanisms remain unclear. We performed bidirectional Mendelian randomization to assess causal relationships between PM exposure and estimated glomerular filtration rate (eGFR), integrated transcriptomic datasets to identify PM-related genes overlapping with diabetic kidney disease (DKD) differentially expressed genes, and applied machine learning approaches to select key feature genes and construct diagnostic models. Single-cell and spatial transcriptomic analyses were used to characterize cell-type and region-specific expression patterns, while in silico knockout analysis explored potential functional associations. PM2.5–10 exposure was causally associated with decreased eGFR, particularly among individuals with diabetes, with no evidence of reverse causality. Transcriptomic integration identified 168 shared PM-DKD genes enriched in inflammatory, immune, and metabolic pathways, including AGE-RAGE, IL-17, TNF, and PI3K-Akt signaling. Seven feature genes (AVPI1, DUSP1, FOSB, JUNB, PDK2, TPPP3, and VIM) showed good diagnostic performance across training and external validation cohorts, and machine learning models and nomogram analyses demonstrated consistent predictive performance. Single-cell and spatial transcriptomic analyses revealed distinct cell-type and region-specific expression patterns, with VIM enriched in interstitial and fibrotic regions, TPPP3 mainly detected in podocytes, and other genes distributed across tubular or immune cell populations. In silico knockout analysis suggested potential associations of these genes with mitochondrial metabolism, oxidative stress, tubular function, and inflammatory processes. Database-based therapeutic exploration identified VIM as a potential candidate target, with sanguinarine showing favorable predicted binding affinity. Collectively, these findings suggest that PM2.5–10 exposure may contribute to DKD susceptibility through inflammatory, metabolic, and oxidative stress-related mechanisms, and provide candidate molecular markers for further investigation. Full article

Background/Objectives: Ephedrae Herba (EH) is widely used in traditional East Asian medicine, but safety concerns regarding its adverse events remain. This study aimed to investigate EH-associated adverse events using clinical pharmacovigilance data and to perform exploratory in silico analyses to propose potential molecular mechanisms underlying these adverse events. Methods: A disproportionality analysis was performed using individual case safety reports from the Korea Adverse Event Reporting System database. EH-containing products were compared with other herbal medicine products using reporting odds ratios (RORs), proportional reporting ratios, and information components. Network pharmacology identified adverse event-related genes and pathways, and protein–protein interaction networks were constructed. Molecular docking predicted direct adverse event-associated targets of ephedrine and compared mechanisms with control compounds (aconitine and spinosin). Results: Four adverse-event signals were detected in the primary analysis: sleep disorder, dry mouth, insomnia, and palpitations. Sensitivity analysis identified four signals, with three (dry mouth, insomnia, and palpitations) consistent across both analyses; constipation emerged only in the sensitivity analysis. Conclusions: Adverse event-associated network analysis predicted key pathways: Neuroactive ligand–receptor interaction, Pathways of neurodegeneration, and Dopaminergic synapse. Molecular docking predicted that ephedrine may act on downstream signaling mechanisms shared by neurotransmitter systems, including the dopaminergic system, distinct from control compounds. Full article

Oxidative stress is a key mechanism underlying nicotine-induced male reproductive dysfunction, yet evidence-based phytopharmacological interventions remain limited. Uncaria gambir Roxb., a catechin-rich medicinal plant from West Sumatra, Indonesia, exhibits strong antioxidant potential; however, a comprehensive multi-regional evaluation combined with in vivo androgenic assessment is lacking. This study aimed to characterize the antioxidant activity, phytochemical composition (HPLC-DAD), and antimicrobial properties of gambir extracts from three production regions (Halaban, Mungka, and Pesisir Selatan), to evaluate catechin pharmacokinetics in silico (PASS Online; SwissADME), and to assess dose-dependent effects in male Wistar rats exposed to nicotine (n = 36). The Halaban extract showed the highest antioxidant activity (IC₅₀ = 6.99 ± 0.28 µg mL⁻¹) and catechin content (7.91 ± 0.07 mg g⁻¹). All extracts demonstrated broad-spectrum antimicrobial activity. PASS analysis predicted strong membrane integrity agonism (Pa = 0.950) and HMOX1 induction (Pa = 0.778), while SwissADME indicated favorable pharmacokinetic properties, including full Lipinski compliance and high gastrointestinal absorption. In the in vivo study, gambir Halaban extract at 300 mg kg⁻¹ day⁻¹ significantly increased follicle-stimulating hormone, luteinizing hormone, and testosterone levels compared to controls (p < 0.05), exceeding baseline values and indicating HPG axis stimulation. A non-linear (hormetic) response was observed, with reduced efficacy at higher doses. These findings highlight bioactive-rich gambir as a promising multi-target natural antioxidant for mitigating oxidative-stress-related reproductive dysfunction, warranting further mechanistic and translational studies. Full article

Background: Esophageal squamous cell carcinoma (ESCC) remains highly aggressive and continues to limit clinical treatment for substantial cancer-associated morbidity and mortality worldwide. Despite advances in therapeutic interventions the lack of effective molecularly targeted treatments continues to restrict clinical management beyond conventional chemotherapy and radiotherapy. Among these therapeutic targets the ribosomal S6 kinase 4 (RSK4) has gained considerable attention because of its critical involvement in ESCC progression, survival and proliferation, suggesting its potential as a potential target for anticancer drug development. Methods: A series of thiazole-clubbed pyrimidine linked hydrazone hybrids (1–14) were synthesized via 4-aminothiazole-5-carbohydrazide functionalized intermediates and fully characterized and evaluated for their inhibitory activity against RSK4. Results: Biological assessment demonstrated that the synthesized analogues exhibited remarkable potency, with IC₅₀ values between 15.32 ± 1.35 and 54.61 ± 2.17 nM compared with the reference inhibitor BI-D1870 (IC₅₀ = 33.16 ± 1.34 nM). Among the evaluated compounds, 2, 8, 9, 13 and 14 emerged as the most potent candidates and showed pronounced activity towards RSK4. For further insights into the molecular basis of their activity the lead candidates were subjected to computational investigations, such as molecular docking, molecular dynamics simulations, in silico ADMET and ProTox-3.0 characterization. The computational analyses provided structural and pharmacological insights into experimentally observed RSK4 inhibitory activity, like predicted interactions, stability dynamically and preliminary ADMET/toxicity characteristics. This study supports the need for further optimization and experimental validation of the identified RSK4 active lead candidates. Conclusions: These findings highlight the thiazole-clubbed pyrimidine-linked hydrazone scaffold as a potential chemotype for promising scaffolds targeting RSK4 lead discovery, and the identified candidates warrant further investigation into ESCC cellular models to establish anticancer efficacy and pathway-level activity. Full article

COVID-19 remains a relevant area of biomedical investigation because its pathogenesis involves complex virus–host interactions. This study aimed to explore, through purely in silico and hypothesis-generating insights, the predicted molecular associations between benzyl isothiocyanate (BITC) from Salvadora persica and COVID-19-associated host-response pathways. BITC-associated targets were collected from compound-target databases, while COVID-19-associated targets were obtained from disease-gene databases and transcriptomic datasets. Overlapping targets were analyzed using protein–protein interaction network construction, hub-gene prioritization, Gene Ontology and KEGG enrichment analyses, and molecular docking. A total of 271 unique BITC-associated targets and 1890 COVID-19-associated targets were identified, with 36 candidate targets overlapping. PPI analysis generated a connected network of 24 nodes and 39 edges. Hub-gene analysis prioritized ACE, JUN, MAOA, CDK1, MAOB, HCK, CCNA2, ACHE, GADD45A, and ADRA2A. Enrichment analysis indicated associations with inflammatory response, vascular regulation, calcium homeostasis, monoamine oxidase activity, NF-κB signaling, serotonergic synapse, and tryptophan metabolism. Validated active-site docking of six targets yielded comparative Vina scores ranging from −5.380 to −6.437 kcal/mol. These preliminary findings provide theoretical target–pathway associations supporting further investigation of BITC as a potential immunomodulatory candidate within COVID-19-related host-response pathways. Full article

Proteases are widely studied hydrolases as “green technologies” for degrading structures within complex matrices, although combining different classes poses challenges for catalytic stability. To the best of our knowledge, this is the first study to specifically evaluate the biochemical control of Cryptosporidium spp. oocysts using plant- and microbial-derived proteases. This study evaluated the compatibility and sanitizing potential of the microbial serine protease subtilisin Carlsberg (HPF-1SCA) and the plant cysteine protease papain (1PPP), both individually and in a 15% (w/v) combination, hypothesizing a synergistic effect due to their distinct catalytic specificities, as a potential biochemical approach to reduce contamination by these zoonotic protozoa of global importance, which are highly resilient to conventional disinfectants. The experimental design comprised five distinct treatment groups: a negative control (distilled water), a positive chemical control (0.04% v/v NaClO), treatment with isolated papain (15% w/v), treatment with isolated microbial HPF formulation (15% w/v), and a combined treatment using both enzymes simultaneously (15% w/v each). Concurrently, an in silico molecular docking investigation was conducted to elucidate the predicted binding scores, structural compatibility, and preferential binding mechanisms of these enzymes toward the Cryptosporidium Oocyst Wall Protein (COWP). In vitro compatibility assays revealed an immediate antagonistic effect due to mutual proteolysis, reducing overall proteolytic activity by 45% after 72 h. Despite this antagonism, the enzyme mixture (G5) and the isolated microbial protease (G4) achieved a 93% reduction in oocysts, equivalent to the conventional 0.04% NaClO treatment (G2), while papain (G3) achieved 65%. All these results represented statistically significant efficacy (p < 0.01) compared to the negative control (G1). In silico molecular docking studies provided a structural predictive basis for the experimental data, suggesting that subtilisin Carlsberg (HPF-1SCA) exhibits more favorable predicted binding scores and structurally compatible interfaces, compared to papain, particularly toward the major oocyst structural proteins COWP8 (estimated ΔG = −15.2 kcal·mol⁻¹) and COWP6 (estimated ΔG = −13.3 kcal·mol⁻¹). This provides robust evidence for initial target recognition and molecular anchoring, although these static models do not definitively confirm catalytically productive cleavage conformations. In summary, this in vitro proof-of-concept demonstrates that microbial proteases show promise for the biochemical destabilization of oocysts without generating toxic chlorinated byproducts. While these baseline findings align with the One Health concept, future field-scale validations assessing enzyme stability under variable environmental conditions and in vivo infectivity assays are required before practical application in sanitation protocols. Full article

Background/Objectives: Colorectal cancer (CRC) remains a leading cause of cancer-associated mortality worldwide. The current therapeutic interventions are heavily constrained by the development of resistance and severe systemic toxicity. To address these challenges, this study integrated a multi-disciplinary framework involving high-throughput in silico screening followed by in vitro experimental validation to identify novel genetic targets of CRC and evaluate the efficacy of FDA-approved drugs. The primary objective was to identify safe and selective therapeutic agents capable of modulating their effect. Methods: The methodology employed a systematic screening of recent large-scale Genome-Wide Association Studies (GWASs) to pinpoint novel targets, followed by in silico pathogenicity prediction, homology modelling and high-throughput virtual screening of over 1615 FDA-approved drugs. The prioritized candidates were validated in vitro using MTT cytotoxicity assays and differential gene expression analysis across CRC cell lines (HCT116 and HT29) and a non-tumorigenic control, Human embryonic kidney cell line HEK293. Results: In silico analysis identified CLUH, CLSTN3 and SLC11A2 as novel potential targets. Based on in silico predicted deleterious mutations and subsequent molecular docking-based virtual screening, Telmisartan, Dutasteride and Venetoclax were prioritized. This prioritization was supported by their high binding affinity and dose-dependent cytotoxicity in MTT assays; thus, suggesting their repurposing potential for CRC treatment. Telmisartan exhibited a superior therapeutic profile not only in terms of the statistically significant cytotoxicity (p < 0.01), but also its selective effect on HCT116 and HT29 when compared to high safety profile in HEK293. This was further validated when Telmisartan selectively downregulated CLUH and SLC11A2 in CRC cell lines, HCT116 and HT29 while maintaining expression levels in the non-cancerous HEK293 cell line remained significantly unaffected. Furthermore, a 100 ns molecular dynamics simulation confirmed the stable binding conformation and structural reliability of the SLC11A2 (Trp179Ser)–Telmisartan complex. Conclucions: Our findings conclude that Telmisartan is a promising candidate for drug repurposing for CRC treatment and capable of modulating selected novel biomarkers CLUH and SLC11A2. However, further multi-omics-based confirmatory studies and pre-clinical validation studies are needed in the future to confirm the long-term efficacy of this repositioning strategy. Full article

The orthoflavivirus capsid (C) protein is a multifunctional protein that plays essential roles throughout the viral life cycle. Besides viral RNA encapsidation for nucleocapsid assembly, it associates with lipid droplets, interacts with host proteins, and translocates to the nucleus, although its nuclear functions are still poorly understood. How these diverse activities are coordinated remains an open question. Post-translational modifications (PTMs), which are key regulators of protein function, have emerged as critical modulators of the infection cycle in many RNA viruses. However, little is known about the occurrence and functional significance of PTMs in orthoflavivirus C proteins. Here, we review the current evidence on PTMs in orthoflavivirus C proteins and integrate insights from studies of other RNA viruses to propose mechanisms by which PTMs may regulate C protein function. To complement this review, we performed a comparative in silico analysis of predicted PTM sites in the C proteins of dengue, Zika, West Nile, and Japanese encephalitis viruses. By integrating PTM predictions with experimentally validated modification sites, residue conservation, and structural mapping, we identified conserved regulatory hotspots that represent promising targets for future experimental validation. Together, these findings highlight PTMs as an underexplored regulatory mechanism in orthoflavivirus capsid biology and provide a framework for future mechanistic investigations. Full article

Background: Therapeutic vaccination against human papillomavirus type 16 (HPV16) is frequently limited by the weak HLA-A*02:01 presentation of native E6/E7 oncoprotein epitopes. This study aims to utilize an integrated computational-experimental pipeline to design and evaluate anchor-optimized altered peptide ligands (APLs) for improving peptide presentation and HPV16-specific T-cell responses. Methods: Anchor-residue substitutions were introduced into three wild-type E6/E7 epitopes. Candidates were prioritized in silico based on predicted presentation, affinity, immunogenicity, and toxicity, and subsequently evaluated via in vitro functional assays using HLA-A*02:01-positive donor cells and molecular dynamics (MD) simulations. Results: Anchor optimization successfully converted weak binders into strong binders; for instance, E6apl improved predicted affinity from 329.33 to 6.21 nM. Crucially, candidate E7apl1 exhibited normal CD8⁺ T-cell expansion but reduced IFN-γ secretion, revealing a distinct binding–immunogenicity dissociation. MD simulations suggested that altered peptide conformational dynamics may contribute to differences in functional activity. Subsequently, an optimized six-peptide formulation (three APLs and three wild-type epitopes) was assembled. The resulting multi-epitope HPV-specific cytotoxic T lymphocytes (meHPV-CTLs) mediated target-specific cytotoxicity against cervical cancer cells, achieving 74.1% ± 5.1% specific lysis at an effector-to-target ratio of 30:1, which was largely abrogated by HLA class I blockade. Conclusions: These proof-of-concept findings demonstrate that stable peptide–MHC binding is a necessary but insufficient condition for optimal T-cell activation. The experimentally characterized multi-epitope formulation provides a proof-of-concept strategy for further preclinical evaluation of HPV16-targeted peptide-based immunotherapies. Moreover, because these anchor-optimized APLs are defined at the sequence level, these sequence-defined APLs may potentially be explored in alternative vaccine delivery platforms, including mRNA-based approaches, in future studies. Full article

Background: Cordyceps militaris (C. militaris) is a medicinal mushroom recognized for its diverse pharmacological activities, largely attributed to its principal bioactive nucleosides, cordycepin and adenosine. Although accumulating evidence supports their neuroprotective potential, the molecular mechanisms underlying their effects against Alzheimer’s disease (AD) remain incompletely understood. This study investigated the neuroprotective effects of cordycepin and adenosine against amyloid-β (Aβ₄₂)-induced neurotoxicity and explored their potential molecular mechanisms using integrated experimental and computational approaches. Methods: SH-SY5Y neuroblastoma cells were pretreated with cordycepin (COR), adenosine (ADE), or donepezil (DNPZ) prior to Aβ₄₂ exposure, and cell viability was assessed using the MTT assay. Drug-likeness and absorption, distribution, metabolism, excretion, and toxicity (ADMET) properties were evaluated in silico, followed by network pharmacology to identify potential therapeutic targets and enriched biological pathways. Molecular docking and molecular dynamics simulations were performed to elucidate the interactions of the compounds with selected Alzheimer’s disease-related proteins. Results: COR and ADE significantly attenuated Aβ₄₂-induced cytotoxicity and improved SH-SY5Y cell viability. Network pharmacology identified 84 shared molecular targets, including 9 AD-associated genes. Protein–protein interaction analysis revealed hub genes involved in signal transduction, epigenetic regulation, and purine metabolism, while Gene Ontology and KEGG enrichment analyses highlighted pathways associated with neuroactive ligand–receptor interaction, calcium signaling, and inflammatory regulation. ADMET analysis predicted favorable pharmacokinetic properties for both compounds, although cordycepin was predicted to be AMES-positive. Molecular docking and molecular dynamics simulations demonstrated stable interactions of COR and ADE with liver X receptors (LXRα and LXRβ), whereas donepezil exhibited stronger binding affinity toward β-secretase (BACE1). Conclusions: COR and ADE exert neuroprotective effects through coordinated modulation of multiple AD-related signaling pathways rather than a single molecular target. These findings provide mechanistic insights into the neuroprotective activities of C. militaris-derived nucleosides and support further investigation of their potential as multi-target therapeutic candidates for AD and other neurodegenerative disorders. Full article

Pseudomonas aeruginosa (PA) is a clinically relevant opportunistic pathogen whose persistence and antimicrobial tolerance are largely driven by biofilm formation and quorum-sensing (QS)-regulated virulence. Targeting QS has therefore emerged as an antivirulence strategy that attenuates pathogenicity without exerting strong selective pressure on bacterial growth. This systematic review (2020–2024) analyzes recent advances in the identification of QS inhibitors against PA, emphasizing studies that integrate in silico approaches. Forty-six studies met the inclusion criteria. All employed molecular docking, and 36.9% (n = 17) incorporated molecular dynamics simulations. While valuable for initial detection, these computational predictions have inherent limitations in accurately estimating binding energy and conformational dynamics, requiring empirical validation to confirm actual biological activity. Approximately one-fifth of the studies were exclusively computational, whereas the remainder combined in silico screening with in vitro and, in some cases, in vivo assays. The most frequently investigated QS regulators were LasR, PqsR, and RhlR, alongside additional virulence-associated proteins. The evaluated compounds encompassed phytochemicals, synthetic molecules, nanomaterials and natural product-derived compounds, several of which demonstrated experimental evidence of biofilm attenuation and reduction in QS-regulated virulence factors. Overall, the findings highlight the value of integrating computational and experimental strategies to rationally prioritize antivirulence candidates. However, the intrinsic complexity and redundancy of the QS network suggest that future research should increasingly focus on multitarget approaches and on the exploration of chemically diverse and previously underexplored compound libraries to improve efficacy against PA biofilms. Full article