Pharmaceuticals doi: 10.3390/ph19091370

Authors: Daniel Miron Brie Cristian Mornoș Alexandru Tîrziu Roxana Popescu Alina Diduța Brie

Background: Chronic coronary syndrome (CCS) is a major source of morbidity and healthcare utilization, and a substantial proportion of patients continue to experience angina despite contemporary revascularization and guideline-directed medical therapy. Trimetazidine, a metabolic anti-ischemic agent that does not affect heart rate or blood pressure, has been used for decades as a non-hemodynamic add-on. Its position in international guidelines, however, has shifted. We aimed to synthesize current evidence on the clinical efficacy, safety, and guideline status of trimetazidine in CCS and coronary artery disease (CAD) and to delineate clinical circumstances in which a symptom-directed trial may be considered. Methods: We performed a structured narrative review of randomized controlled trials, network and conventional meta-analyses, and large observational studies indexed in PubMed/MEDLINE, Embase, and the Cochrane Library between January 1990 and March 2026. Reference lists of eligible articles, international clinical practice guidelines, and regulatory communications from European and national agencies were also examined. Results: Across pooled analyses, trimetazidine modestly but reproducibly improved exercise tolerance and reduced angina frequency and short-acting nitrate use, with efficacy comparable to other non–heart-rate-lowering antianginal agents. Improvements in left ventricular function were reported in small, single-center studies in ischemic cardiomyopathy and selected high-risk subgroups; however, these findings have not been validated in contemporary large-scale trials, and their clinical significance against modern guideline-directed therapy is unknown. The ATPCI trial confirmed safety but showed no reduction in major adverse cardiovascular events after percutaneous coronary intervention. Rare extrapyramidal effects prompted the 2012 European Medicines Agency referral and several prescribing restrictions. Conclusions: Trimetazidine remains a reasonable symptom-directed adjunct for residual angina in carefully selected CCS patients, particularly those in whom further hemodynamic up-titration is limited or poorly tolerated. The 2024 ESC class IIb recommendation may partly reflect a broader emphasis on prognostic benefit, although the neutral post-PCI outcome evidence and regulatory safety restrictions also support a more cautious position.

Pharmaceuticals doi: 10.3390/ph19091369

Authors: Wael Jalloul Cristina Mariana Uritu Despina Jalloul Vlad Ghizdovat Andreia Vranceanu Ciobanu Bogdan Ionel Tamba Cipriana Stefanescu Irena Cristina Grierosu

Parkinsonian syndromes comprise overlapping neurodegenerative and non-degenerative disorders, making aetiological diagnosis difficult, while existing procedural guidance does not fully integrate tracer-specific biology with clinical decision-making, multimodal strategies, and emerging quantitative and pathology-directed biomarkers. This review synthesises evidence on the molecular targets, radiopharmaceutical characteristics, biological interpretation, and clinical applications of dopaminergic single-photon emission computed tomography (SPECT) and positron emission tomography (PET), focusing on the dopamine transporter (DAT), aromatic L-amino acid decarboxylase (AADC), vesicular monoamine transporter type 2 (VMAT2), dopamine D2/D3 receptors, differential diagnosis, semiquantification, kinetic modelling, and artificial intelligence-assisted interpretation. The evidence confirms that DAT SPECT with [123I]ioflupane ([123I]FP-CIT) remains the most established method for demonstrating or arguing against presynaptic nigrostriatal dysfunction, but an abnormal result cannot establish aetiology, and a normal result can redirect evaluation towards non-degenerative mimics; target-specific PET provides complementary biological information, although availability, standardisation, and prospective validation remain limiting. By additionally integrating genetic and prodromal applications, question-driven multimodal imaging, emerging acquisition approaches, α-synuclein imaging and seed amplification assays, and contemporary biological staging frameworks, this review extends procedural guidance and positions dopaminergic imaging as a targeted functional biomarker selected according to the unresolved clinical question rather than as a stand-alone disease label.

Pharmaceuticals doi: 10.3390/ph19091368

Authors: Xin Zeng Wen-Feng Du Wen-Hao Yin Jun-Yao Zhu Yu-Bin Yang Wei-Jun Guo Wen-Liang Li Hao Gong Zi-Zhong Yang Yi Li

Background: Orientothele washanensis is a venomous spider with considerable ecological and scientific importance. Its venom, characterized by complex composition and ease of collection, serves as a valuable resource for the discovery of natural peptide drugs. Conventional wet-lab screening methods are limited by rigorous experimental conditions, high resource consumption, and long research cycles, which hinder the efficient identification of functional peptides from the venom gland transcriptome of this spider. Methods: To address these technical bottlenecks, this study developed a novel deep learning model named PepPI-DRN for peptide-protein interaction prediction. The model integrated a residual equivariant graph neural network, a residual 1-dimensional convolutional neural network, and a dual-modal attention mechanism by leveraging both sequence and structural features of peptides and proteins. Results: Results on an independent test set indicated that PepPI-DRN achieved the competitive or superior performance compared with state-of-the-art methods on multiple key evaluation metrics. Candidate peptides with high interaction probabilities against targets were obtained from the venom gland transcriptome of Orientothele washanensis. Furthermore, lead peptides with high binding strength and good structural stability were identified from candidate peptides by molecular docking, and molecular dynamics simulation. Conclusions: These results showed that the pipeline with PepPI-DRN, molecular docking and molecular dynamics simulation enabled efficient and reliable identification of lead peptides from the venom gland transcriptome of Orientothele washanensis, providing a robust and effective strategy for the discovery and development of natural peptide drugs from the spider venom.

Pharmaceuticals doi: 10.3390/ph19091367

Authors: Xi-Yuan Feng Meng Gao Fei-Long Liu Ming-Ze Li Xiao-Li Cui Meng-Yang Wang Zhi-Hong Zhang He Li Chun-Mei Wang Jing-Hui Sun

Objectives: This study focuses on exploring the mechanism by which Schisandrin B (Sch B) regulates metabolic dysfunction-associated steatotic liver disease (MASLD) mice induced by a high-fat methionine–choline-deficient (MCD) diet through the activation of peroxisome proliferator-activated receptor γ (PPARγ). Methods: Male C57BL/6 mice were fed a high-fat MCD diet for 8 weeks to establish a mouse MASLD model, and the effects of Sch B on MASLD and the mechanisms were investigated. PPARγ overexpression (OE) was induced by adeno-associated virus (AAV) administration via intrahepatic portal vein injection in mice, and a negative control (NC-OE) was also established. Body weight; wet liver weight; hepatic index; serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and interleukin-1β (IL-1β); and hepatic triglyceride (TG) levels were measured in the mice. The histopathology and lipid deposition were observed by hematoxylin and eosin (H&E) staining and Oil Red O staining, while the fibrosis was assessed using Masson staining. Western blot was employed to detect the expression levels of PPARγ, sterol regulatory element-binding protein 1c (SREBP-1c), carnitine palmitoyltransferase 1A (CPT1A), transforming growth factor β1 (TGF-β1), α-smooth muscle actin (α-SMA), collagen type I (collagen I), Smad family members 2/3 (Smad2/3), c-Jun N-terminal kinase (JNK), p38 mitogen-activated protein kinase (p38), and extracellular signal-regulated kinase 1/2 (ERK1/2), along with the phosphorylation activation status of these kinases. Results: It was confirmed that Sch B caused effects similar to those induced by PPARγ overexpression, reducing the hepatic index, AST, and ALT levels while alleviating lipid accumulation and fibrosis; and upregulating PPARγ and CPT1A while inhibiting SREBP-1c; and the phosphorylation of the TGF-β/Smad and MAPK pathways were involved in the mechanisms. Conclusions: Sch B can alleviate high-fat MCD-induced MASLD by activating PPARγ in mice.

Pharmaceuticals doi: 10.3390/ph19091366

Authors: Samridhi Kurl Neeraj Mittal Rajwinder Kaur Balakumar Chandrasekaran Gurpreet Kaur

Background/Objectives: Psychobiotics are live microorganisms that, when administered in appropriate amounts, can enhance mental health. Through the production of neuroactive metabolites, regulation of immune responses, and interactions along the gut–brain axis, they modulate host neurophysiology. However, significant strain-specific variability exists in their metabolic capabilities, colonization potential, and functional interactions, necessitating a systematic approach for psychobiotic strain selection. Methods: In the present study, an integrated in silico systems biology approach combining comparative genomics, genome-scale metabolic modeling, network pharmacology, molecular docking, molecular dynamics, and ADMET analysis was employed to computationally evaluate the therapeutic potential and molecular interactions of a quercetin-psychobiotic co-delivery system for managing depression by modulating the gut–brain axis. Results: Comparative genomic analysis of candidate psychobiotic strains from the genera Bifidobacterium and Lactobacillus predicted the presence of key functional genes involved in neuroactive metabolite production, immune modulation, oxidative stress tolerance, and adhesion. The presence of acetate-producing pathways alongside the absence of classical butyrate biosynthesis genes indicated an indirect contribution to short-chain fatty acid production via microbial cross-feeding interactions. Genome-scale metabolic Modeling under the conditions of quercetin supplementation predicted a significant inter-strain variation in terms of predicted biomass flux, nutrient utilization, and metabolic adaptability. Results of molecular docking showed that the binding energies were in the range of −5.854 to −9.489 kcal/mol. Conclusion: Collectively, these in silico findings provided a computational basis for the selection of psychobiotic strains and predicted that a combination of psychobiotics and quercetin could present a promising strategy for the modulation of the gut–brain axis.

Pharmaceuticals doi: 10.3390/ph19091365

Authors: Yiğit Erkmen Zekiye Ceren Arıtuluk Aydın Engin Koçak Hasya Nazlı Gök Emirhan Nemutlu Merve Yüzbaşıoğlu Baran

Background/Objectives: Berberis species are recognized for their ethnomedicinal importance and phytochemical richness associated with antidiabetic activity. Although B. crataegina is widely distributed in the Turkish flora and has a history of traditional use, its phytochemical properties and biological activities remain less extensively investigated than those of other Berberis species. This study aimed to comparatively evaluate the phytochemical profiles, in vitro antidiabetic potential, and cytoprotective effects of extracts prepared from different parts of B. crataegina in a high-glucose-induced SH-SY5Y cell-based model of diabetic neuropathy. Methods: Crude hydroethanolic extracts (70% ethanol) were prepared from the leaves, flowers, shoots, roots, and fresh fruits of B. crataegina collected from Kızılcahamam, Ankara, Türkiye. The total phenolic and flavonoid contents, antioxidant capacities and α-glucosidase inhibitory activities of leaf, flower, shoot, root, and fruit extracts were determined. The cytoprotective effects of the crude extracts were evaluated in a high-glucose-induced SH-SY5Y cell model. The metabolomic profiles of the extracts were comparatively analyzed using liquid chromatography–quadrupole time-of-flight mass spectrometry (LC-QTOF-MS). The most active fruit extract was fractionated by reversed-phase vacuum liquid chromatography (RP-VLC), and the phytochemical contents and α-glucosidase inhibitory activities of the fractions were evaluated. The major phenolic compounds in the most active fruit extract were quantitatively determined using high-performance liquid chromatography–diode array detection (HPLC-DAD). The relationships between metabolites and biological activities were investigated through correlation analysis. Results: The fruit extract was distinguished by its α-glucosidase inhibitory activity, total phenolic content, antioxidant capacity, and cytoprotective effect in the high-glucose-induced SH-SY5Y cell model. It showed no marked cytotoxicity at concentrations ranging from 25 to 400 µg/mL and, at 25 µg/mL, attenuated the high-glucose-induced reduction in cell viability, restoring viability to a level close to that of the normal control. Untargeted metabolomic analysis identified 191 metabolites detected in at least two crude extracts, while the metabolites detected exclusively in the fruit extract were predominantly flavonoids and flavonoid glycosides. Fractionation showed that α-glucosidase inhibitory activity was mainly concentrated in the medium-polarity fractions; however, none of the fractions reached the activity level of the crude extract. Chlorogenic acid, rutin, caffeic acid, protocatechuic acid, quercetin, and quercetin-3-O-glucoside were detected and quantified in the fruit extract by HPLC-DAD. Correlation analysis indicated that the observed biological activities may be associated with phenolic acids, flavonoids, and other polyphenolic metabolites. Conclusions: The fruit extract of B. crataegina showed promising α-glucosidase inhibitory activity and cytoprotective effects in the high-glucose-induced SH-SY5Y cell model. The findings suggest that these effects may arise from the combined contribution of multiple constituents within a polyphenolic matrix rather than from a single compound; however, synergistic interactions were not experimentally demonstrated. These results support further evaluation of the fruit extract through mechanistic, in vivo, and standardization studies.

Pharmaceuticals doi: 10.3390/ph19091364

Authors: Sonu Kamal Y. Thajudeen Saad Ali Alshehri Mohammed Muqtader Ahmed Mayur Porwal Sagar Verma

Background: The current study uses a computational technique to investigate synthesized thiazine compounds that exhibit potential interactions with the GABAA receptor. A series of novel thiazine derivatives was synthesized via the cyclization reaction of chalcone with thiourea. Methods: For the synthesized compounds, spectroscopic characterization, namely FT-IR, 1H NMR, 13C NMR, and mass spectrometry, was carried out. In addition, the AutoDock Vina 4.2 software was applied to carry out an in silico docking study. Results: The synthesized compounds C3 and C6 exhibited comparable predicted docking scores (−9.0 and −9.1 kcal/mol, respectively), with favorable predicted interactions at the GABAA receptor binding site. Furthermore, in silico ADMET analysis provided insights into the drug-likeness and pharmacokinetic profiles of the synthesized compounds; however, potential safety liabilities, including Ames positivity, hERG II liability, and hepatotoxicity, were predicted for some derivatives. Conclusions: It was determined that the synthesized compounds’ various physiologically significant and physiologically relevant parameters fell within the range of Lipinski’s rule of five. The synthesized compounds (C1–C10) were tested for anxiolytic activity by the elevated plus maze test, and compound C6 was found to be the most potent. Additionally, the in vitro GABA-AT enzyme activity assay showed that compound C6 had the highest inhibitory potential, with an IC50 of 13.29 ± 1.622 µM.

Pharmaceuticals doi: 10.3390/ph19091363

Authors: Chunlai Feng Qiuqi Feng Shengnan She Ruojing Yang Mengru Li Lu Gong Mengjie Rui

Background/Objectives: Breast cancer exhibits substantial molecular heterogeneity, resulting in diverse therapeutic vulnerabilities and limiting the efficacy of single-agent therapies. Although multi-target strategies may offer improved therapeutic benefit, the systematic identification of synergistic higher-order target combinations remains challenging. This study aimed to identify and validate effective higher-order target combinations for heterogeneous breast cancer. Methods: DeepMDS, a previously developed deep learning-based multi-compound synergy prediction model, was used to prioritize candidate target combinations for breast cancer. Exhaustive two-target and three-target combinations were ranked in the gene-expression contexts of ER-positive luminal-like MCF-7 and triple-negative MDA-MB-231 breast cancer cells. The top-ranked target combinations were evaluated using single, pairwise, and triple small interfering RNA (siRNA) perturbations. Representative inhibitors were subsequently assessed in fixed-ratio combinations in MCF-7, MDA-MB-231, and 4T1 cells and in a 4T1 syngeneic mouse experiment. Results: BIRC5-NAMPT-TOP1 ranked first in both cell lines. Triple siRNA co-transfection targeting BIRC5, NAMPT, and TOP1 produced the strongest antiproliferative effects, with inhibition rates of 62.94% in MCF-7 cells and 55.62% in MDA-MB-231 cells. The corresponding inhibitors, LQZ-7I, FK866, and topotecan, exhibited synergistic antiproliferative activity at multiple molar ratios, with combination index values below 1. The optimized 2.5:10:1 molar ratio showed strong synergy in MCF-7, MDA-MB-231, and 4T1 cells, with combination index values of 0.26, 0.19, and 0.15, respectively. In tumor-bearing mice model, the triple-inhibitor regimen achieved a tumor inhibition rate of 62.05%. Topotecan-containing groups showed hematological alterations, whereas no statistically detectable elevations were observed in the measured terminal serum hepatic or renal biomarkers. Conclusions: The BIRC5-NAMPT-TOP1 combination showed reproducible phenotypic activity in the tested genetic and pharmacological models. These findings support the use of DeepMDS as a hypothesis-generation tool for higher-order target prioritization.

Pharmaceuticals doi: 10.3390/ph19091362

Authors: Qin Liu Huiyi Peng Min Su Zhoujin Tan Maijiao Peng

Background: Shenling Baizhu Powder (SLBZP) is a traditional Chinese medicine formula used to treat diarrhea. This study investigated its therapeutic effects and potential mechanisms in mice with fatigue- and high-fat diet-induced diarrhea, focusing on gut microbiota, bile acid metabolism, and farnesoid X receptor (FXR)-related signaling. Methods: Male Kunming mice were used to establish a model of diarrhea, followed by treatment with SLBZP. Fecal moisture content, serum levels of diamine oxidase and liver enzymes, and intestinal and hepatic histopathological changes were assessed. Small-intestinal microbiota composition, bile acid profiles in colonic contents, and FXR-related signaling in the ileum and liver were further analyzed. Results: SLBZP significantly reduced fecal moisture content. It also modulated the small-intestinal microbiota, with a significant increase in Faecalibaculum. In colonic contents, SLBZP reduced total bile acid levels, increased the secondary-to-primary bile acid ratio, and altered multiple bile acid species. These changes were accompanied by increased ileal FXR expression and changes in downstream regulators involved in hepatic bile acid synthesis. Correlation analysis further showed associations of selected bacterial taxa and FXR protein expression with colonic bile acid profiles. Conclusions: SLBZP alleviates diarrhea induced by fatigue combined with a high-fat diet in mice. Its effects may involve coordinated modulation of the gut microbiota, bile acid homeostasis, and FXR-related signaling, providing experimental support for the pharmacological basis of this traditional formula in diet-related diarrhea.

Pharmaceuticals doi: 10.3390/ph19091361

Authors: Jayoung Oh Ji Ho Choi

Background: Obstructive sleep apnea (OSA) remains under-treated because continuous positive airway pressure, though efficacious, is frequently limited by poor tolerance and adherence. Long considered a purely anatomical disease refractory to drugs, OSA is now understood as the shared endpoint of four measurable endotypes—a collapsible upper airway, poor pharyngeal dilator-muscle responsiveness, high loop gain, and a low arousal threshold—each a distinct pharmacological target. This review synthesizes the mechanistic rationale and clinical evidence for targeting them. Methods: In this narrative review, we searched PubMed/MEDLINE, Embase, and the Cochrane Central Register of Controlled Trials (January 2000 to February 2026) for randomized trials, meta-analyses, and mechanistic endotyping studies, appraising the evidence narratively. Results: Tirzepatide became the first drug approved for OSA, acting through weight loss to reduce obesity-related mechanical loading, and the noradrenergic–antimuscarinic combination AD109 met its primary endpoint in a pivotal phase 3 trial by targeting dilator-muscle activity independently of weight. Both, however, used AHI-based endpoints in clinically defined rather than endotype-selected populations; AD109’s placebo-corrected effect was modest (−4.0 events/h), and 21.2% versus 3.1% of participants discontinued for adverse events. They are therefore best described as mechanism-informed rather than endotype-validated. Carbonic anhydrase inhibition with acetazolamide or sultiame and selected sedating agents address ventilatory instability and a low arousal threshold, while solriamfetol and pitolisant relieve residual sleepiness. Conclusions: Mechanism-informed pharmacotherapy for OSA is now clinically credible, but scalable bedside endotyping, attention to tolerability, and trait-stratified trials are needed before treatment can be called endotype-guided.

Pharmaceuticals doi: 10.3390/ph19091360

Authors: Dongsheng Wei Han Li Mei Zhao Mai Liu Yongyue Liu Menglan Zhao Huiming Cao Xiaoqing Zhang

Background: Pathological cardiac remodeling is a major driver of heart failure progression and is closely associated with fibroblast activation, myocardial inflammation, oxidative injury, and impaired autophagic homeostasis. Epidermal growth factor receptor (EGFR) signaling participates in adverse remodeling through activation of MAPK pathways, but the relationship between pharmacologically modulated EGFR turnover and cardiac remodeling remains insufficiently defined. Methods: This study investigated the protective effects of cycloastragenol (CAG), a triterpenoid sapogenin derived from Astragalus membranaceus, in Angiotensin II (Ang II)-induced cardiac remodeling and examined changes in EGFR turnover and associated MAPK signaling and autophagy-related processes. Circulating soluble EGFR was associated with heart failure severity after adjustment for age and sex. Separately, an independent exploratory transcriptomic analysis of left ventricular samples from GSE161472 showed that myocardial EGFR expression distinguished HFrEF from non-failing samples with an AUC of 0.870. Male C57BL/6 mice subjected to continuous Ang II infusion at 500 ng/kg/min for 28 days were used to establish the in vivo cardiac remodeling model and evaluate the effects of CAG on cardiac function, myocardial injury, fibrosis, inflammation, oxidative stress, apoptosis, and autophagy-related changes, with enalapril used as a positive control. Primary cardiac fibroblasts were further used to assess proliferation, migration, collagen production, EGFR membrane accumulation, lysosomal trafficking, MAPK signaling, and autophagic flux. Circulating soluble EGFR increased with heart failure severity and was associated with BNP elevation, ventricular dilation, and reduced ejection fraction. Results: CAG improved cardiac performance, reduced myocardial injury markers, alleviated fibrosis and hypertrophy, and suppressed inflammatory and oxidative responses in Ang II-treated mice. In cardiac fibroblasts, CAG reduced sustained EGFR membrane accumulation, increased EGFR recovery in pan-ubiquitin immunoprecipitates and lysosomal localization, and accelerated EGFR protein turnover, consistent with a predominantly lysosome-associated pathway. The K716R mutation attenuated CAG-associated changes in EGFR turnover, MAPK phosphorylation, and autophagic flux. Conclusions: These findings suggest that CAG attenuates Ang II-induced cardiac remodeling and is associated with enhanced EGFR lysosomal turnover, accompanied by changes in MAPK signaling and autophagy-related processes.

Pharmaceuticals doi: 10.3390/ph19091359

Authors: Enikő Rauch Csilla Ari Dominic P. D’Agostino Zsolt Kovács

Background/Objectives: Inflammatory processes, including elevated pro-inflammatory cytokines such as interleukin 1β (IL-1β), have been implicated in the pathophysiology of anxiety disorders. The ketone body β-hydroxybutyrate (βHB) was shown to mitigate inflammatory processes induced by lipopolysaccharide (LPS) administration. It was also suggested that exogenous ketone supplements, such as ketone esters (KEs) and ketone salts (KSs), can decrease anxiogenic effects through increased blood ketone levels in the body (ketosis) in preclinical animal models. Methods: Thus, we investigated whether standard rodent food supplemented with KEKS (KEKS food: mix of a KE R/S-1,3-butanediol-acetoacetate diester and a KS Na+- and Ca2+- R/S-βHB salt in 1:1 ratio) for 7 days can alleviate the intraperitoneal (i.p.) 1 mg/kg LPS-generated anxiogenic effect in female Wistar Albino Glaxo/Rijswijk (WAG/Rij) rats. For the evaluation of anxiogenic effects, the light–dark box (LDB) test was used. Results: KEKS food significantly increased the blood R-β-hydroxybutyrate (R-βHB) level and decreased the blood glucose level compared to the baseline. Moreover, the LPS significantly (i) decreased the time spent in the light compartment, the number of chamber transitions, the number of re-entries to the light compartment and the number of rearing in the light compartment, and also (ii) increased the time of latency to the first re-entry to the light compartment compared to control. Nevertheless, KEKS food treatment abolished LPS-generated anxiety-like behavior. Conclusions: The current study demonstrated that a 7-day KEKS-supplemented diet can increase blood R-βHB levels and abolish acute LPS administration-evoked increases in anxiety levels in female WAG/Rij rats. Moreover, our results support the important role of neuroinflammation in pathological processes resulting in anxiety. Thus, these results suggest that exogenous ketone supplement-generated ketosis may be a promising therapeutic tool in the treatment of inflammatory process-mediated diseases, such as anxiety disorders.

Pharmaceuticals doi: 10.3390/ph19091358

Authors: Xiao-Hui Chi Ming-Feng Liao Ya-Qun Zhou

Although the underlying mechanisms of renal IRI have been extensively studied, the corresponding effective treatments are still lacking. HMGB1, an important nuclear factor that is secreted outside cells when experiencing stress conditions, acts as a DAMP and exerts crucial effects on renal IRI. Many studies have suggested that the effect of HMGB1 on kidney damage is mediated mainly through the interaction of HMGB1 with pattern recognition receptors such as TLR4 and RAGE, which then results in the aggravation of local inflammatory response, increased infiltration of leukocytes, and finally renal tubular damage. Preclinical studies using animal models have demonstrated that inhibition of HMGB1 and downstream signal pathways can attenuate renal injury. This review critically evaluates HMGB1 in renal IRI across redox state, subcellular localization, temporal and cell-specific release, receptor usage, integrated stress pathways, autophagy, and regulated cell death. It also compares direct neutralization, inhibition of release or translocation, epigenetic/RNA-based regulation, and receptor-directed strategies. The evidence is predominantly derived from short-term rodent studies, with one large-animal antibody study and no therapeutic human trials. Accordingly, HMGB1 is best regarded as a biologically compelling but clinically unvalidated target whose therapeutic value will depend on redox- and phase-selective inhibition, kidney-directed delivery, and rigorous pharmacokinetic and safety evaluation.

Pharmaceuticals doi: 10.3390/ph19091357

Authors: Artyom Savelyev Dmitriy Khrustalev Irina Losseva Azamat Yedrissov Anastassiya Khrustaleva Vladimir Kazantsev Sofiya Shapovalenko Marlen Kiikbayev Polina Rusyaeva Kristina Perepelitsyna

Background: Topical semi-solid dosage forms are crucial for treating inflammatory skin disorders, wounds, burns, and superficial infections. Despite their widespread clinical use, comprehensive pharmaceutical market analyses of registered topical medicines remain limited in Central Asia. The aim of this study was to characterize the pharmaceutical market of topical semi-solid dosage forms registered in the Republic of Kazakhstan, including their dosage-form distribution, pharmacotherapeutic composition, manufacturer structure, and structural diversity of active pharmaceutical ingredients (APIs), with particular attention to heterocyclic compounds. Methods: A cross-sectional descriptive analysis was conducted using the State Register of Medicinal Products of the Republic of Kazakhstan. Registration records of topical semi-solid dosage forms were identified according to predefined inclusion criteria and classified by dosage form, pharmacotherapeutic category, manufacturer, country of origin, and API chemical structure. Descriptive statistical analysis, market concentration indices, and structural classification of heterocyclic compounds were applied. Therapeutic recommendations for dermatitis management from selected CIS countries were reviewed to provide a contextual interpretation of the observed market structure. Results: A total of 1046 registered topical semi-solid dosage forms met the inclusion criteria. Ointments (39.5%), gels (28.9%), and creams (25.4%) accounted for 93.8% of all registered products, indicating a highly concentrated dosage-form structure. Anti-inflammatory agents represented the largest pharmacotherapeutic group, whereas wound-healing and combination products accounted for smaller proportions of the market. Structural classification demonstrated broad representation of established heterocyclic chemical classes, while no registered medicinal products containing dihydropyrimidine-2(1H)-thione (DHPM-thione) derivatives were identified in the registry. Comparison with regional clinical recommendations indicated differences between recommended therapeutic approaches and the distribution of registered pharmacotherapeutic categories. Conclusions: This study provides the first comprehensive descriptive assessment of the registered market of topical semi-solid dosage forms in Kazakhstan. The results establish a reference dataset describing the current pharmaceutical landscape, including dosage forms, therapeutic categories, manufacturers, and API chemical classes. The absence of registered DHPM-thione-containing medicinal products indicates that this chemical class is currently not represented in the national medicines registry. Together with previously published preclinical evidence, this observation supports consideration of DHPM-thione derivatives as a subject for future pharmaceutical research, although the present study does not permit conclusions regarding their clinical effectiveness, market demand, or future commercialization.

Pharmaceuticals doi: 10.3390/ph19091356

Authors: Aleksandr Subbotin Holger A. Volk Sebastian Meller Gemma Mazzuoli-Weber Kristin Elfers

Background/Objectives: Antiseizure drugs (ASDs) are the primary therapeutic approach for epilepsy in small animals. Although ASDs are primarily used to modulate central neuronal excitability, they are commonly administered systemically, most often by the oral route, and may therefore influence neuronal populations outside the central nervous system. Nevertheless, their functional effects on peripheral neuronal populations, including enteric and dorsal root ganglion (DRG) neurons, remain incompletely characterized at a comparative pharmacological level. This study aimed to perform a comparative functional neuropharmacological profiling of commonly used ASDs in primary cultured myenteric and DRG neurons. Methods: Changes in neuronal activity were assessed in primary cultured guinea pig myenteric and DRG neurons using voltage-sensitive dye imaging with Di-8-ANEPPS following direct ASD application under standardized in vitro conditions. Results: ASDs exerted distinct drug- and neuron-type-specific effects on peripheral neuronal excitability. Topiramate induced the most pronounced reduction in neuronal excitability in myenteric neurons, whereas phenobarbital and levetiracetam produced only minor changes compared with buffer control. Potassium bromide induced mainly excitatory effects in both enteric and DRG neurons. Overall, most ASDs predominantly increased neuronal excitability in DRG neurons. Conclusions: These findings demonstrate distinct functional response profiles of ASDs in enteric and sensory neuronal populations. This comparative in vitro approach may provide a basis for future studies investigating peripheral neuronal drug effects and may help relate experimental pharmacological profiling to clinically relevant challenges associated with ASD treatment across different disorders.

Pharmaceuticals doi: 10.3390/ph19091355

Authors: Tamás Gáll Dávid Pethő Szilárd Póliska József Balla György Balla

Background/Objectives: Hypoxia is a major contributor to the development of retinal diseases by promoting metabolic adaptation and pathological angiogenesis through hypoxia-inducible factor (HIF) signaling. HIF-prolyl hydroxylase inhibitors (HIF-PHIs), which stabilize HIF under normoxic conditions, are widely used to treat anemia associated with chronic kidney disease. However, it remains unclear whether pharmacological HIF stabilization fully reproduces the cellular responses induced by physiological hypoxia. Methods: In this study, ARPE-19 cells were exposed to either physiological hypoxia or HIF-PHI treatment and analyzed using RNA sequencing, pathway enrichment, and pharmacological inhibition of PI3K/Akt, mTOR, and HIF-related signaling pathways. Results: Both treatments elicited a common transcriptional response marked by the induction of glycolytic and hypoxia-responsive genes together with increased vascular endothelial growth factor a VEGFA expression. Despite these similarities, only a small number of genes differed between the two conditions, suggesting that physiological hypoxia activates additional regulatory mechanisms beyond HIF stabilization alone. Rapamycin inhibited VEGFA production only under physiological hypoxia, further supporting mechanistic differences between the two models. Functionally, conditioned media from hypoxic ARPE-19 cells promoted endothelial tube formation, which was significantly reduced by pathway inhibition. Conclusions: Collectively, these findings show that HIF-PHIs reproduce many, but not all, aspects of the physiological hypoxic response in RPE cells.

Pharmaceuticals doi: 10.3390/ph19091354

Authors: Oscar Salvador Barrera-Vázquez Gil Alfonso Magos-Guerrero Juan Luis Escobar-Ramírez Rubén San Miguel-Chávez Maira Huerta-Reyes

Chiranthodendron pentadactylon Larreat (“flor de manita”) is traditionally used for gastrointestinal, cardiovascular, and neurological disorders. Objective: This study comprehensively integrated phytochemical characterization, in silico toxicological profiling, an exposure-informed Integrative Toxicity Prediction Model (ITPM), uncertainty analysis, and acute testing of a fresh flower aqueous extract (FFAE). Methods: Thirty-six phytochemicals were screened for hepatotoxicity, nephrotoxicity, Ames mutagenicity, carcinogenicity, hERG inhibition, reproductive-effects alerts, and acute oral toxicity. Five organ- or effect-specific endpoints were integrated into an exploratory Toxicological Alert Score (TAS), whereas reproductive alerts and predicted LD50 were reported separately. Seventeen compounds quantified in the FFAE were compared with a 20-compound literature-informed profile through 50,000 Monte Carlo iterations. Male CD-1 mice received a single FFAE dose of 300–5000 mg/kg and were observed for 14 days. Results: Positive predictions occurred for hERG inhibition in 33 compounds (91.7%), hepatotoxicity in 29 (80.6%), nephrotoxicity and carcinogenicity in 28 each (77.8%), and mutagenicity in 21 (58.3%). Six compounds had predicted LD50 values ≤ 1000 mg/kg. The proportional ITPM toxicity-evidence index was 6.91% for the experimental profile and 19.34% for the literature-informed profile. Monte Carlo means were 6.98% (95% uncertainty interval, 5.65–8.44%) and 19.32% (16.76–22.37%), respectively. No mortality, overt clinical signs, or treatment-related body weight differences occurred up to 5000 mg/kg. Conclusions: Compound-level predictions identified priorities for confirmatory testing, while the extract-specific model yielded a lower comparative toxicity-evidence proportion, and the animal study showed low overall acute toxicity under the evaluated conditions. These findings do not establish general or long-term safety, calibrated risk probabilities, or biological neutralization of toxicological liabilities.

Pharmaceuticals doi: 10.3390/ph19091353

Authors: Saied A. Aboushanab Denis A. Oberiukhtin Irina G. Danilova Pavel M. Vassiliev Ksenia V. Sokolova Kewei Chen Elena G. Kovaleva Alicia C. Mondragon Ekaterina I. Demicheva Jose M. Miranda

Background/Objectives: Type 2 diabetes mellitus (T2DM) is a multifactorial metabolic disorder characterized by chronic hyperglycemia, oxidative stress, protein glycation, low-grade inflammation, and dysregulation of signaling pathways, including those involving nuclear factor kappa B (NF-κB). Isoflavones from Fabaceae species exhibit diverse biological activities and may represent multifunctional candidates for addressing the interconnected mechanisms underlying T2DM. This study performed a consensus in silico pharmacodynamic screening of twelve representative Fabaceae isoflavones to prioritize compounds with predicted antiglycating, deglycating, antidiabetic, antioxidant, and NF-κB-related anti-inflammatory activities. Methods: Twelve representative Fabaceae isoflavones were evaluated using a multimethod computational workflow. Antiglycating and deglycating activities were predicted using IT Microcosm, antidiabetic and hypoglycemic activities were assessed using Prediction of Activity Spectra for Substances (PASS), antioxidant potential was evaluated using similarity-based prediction and quantum-chemical calculations, and NF-κB-related anti-inflammatory activity was investigated using similarity-based prediction combined with molecular docking. Molecular docking was applied specifically to the NF-κB-related anti-inflammatory analysis and did not contribute to the hypoglycemic/antidiabetic prediction. The individual predictions were integrated using a consensus scoring strategy to prioritize compounds across multiple pharmacodynamic domains. Results: The investigated isoflavones demonstrated moderate but broad predicted multifunctional activities. Malonylgenistin, biochanin A, and formononetin showed the most favorable predicted antiglycating profiles. Genistin, acetyldaidzin, acetylglycitin, and puerarin ranked highest in terms of predicted antidiabetic activity, whereas genistein exhibited the strongest predicted antioxidant profile. Daidzin and genistin received the highest integrated NF-κB-related computational rankings, although these predictions apply to intact glycosides rather than their expected in vivo hydrolysis products. Conclusions: This study presents a consensus computational framework for the comparative prioritization of Fabaceae isoflavones based on their predicted multifunctional pharmacodynamic profiles. The results provide a rational basis for selecting candidate compounds for future biochemical, cell-based, pharmacokinetic, and in vivo investigations, although it is emphasized that the present findings are based exclusively on computational predictions and require experimental validation.

Pharmaceuticals doi: 10.3390/ph19091352

Authors: Pasquale Degennaro Imane Ghafir El Idrissi Rosa Purgatorio Annalisa Fanizzi Mariagrazia Rullo Leonardo Pisani Eleonora Macchia Luisa Torsi Angela Stefanachi Francesco Leonetti

Proteolysis-Targeting Chimeras (PROTACs) are heterobifunctional molecules that induce the selective degradation of a protein of interest by recruiting an E3 ubiquitin ligase, thereby triggering ubiquitination and proteasomal clearance. As part of the broader targeted protein degradation (TPD) paradigm, PROTACs offer a powerful strategy to eliminate pathogenic proteins that are difficult to modulate with traditional occupancy-based inhibitors. However, their clinical translation is often limited by poor aqueous solubility, suboptimal cellular permeability, and off-target effects. Notably, some PROTACs retain potent biological activity despite limited membrane permeability, owing to their catalytic mechanism of action, which allows even a small number of molecules reaching the target site to drive substantial protein degradation and produce important pharmacological effects. Growing evidence supports the application of PROTAC-based approaches in neurodegenerative diseases, where the selective removal of toxic or misfolded proteins is particularly attractive. This review summarizes recent advances in chemical protein degradation strategies for neurodegenerative disorders and highlights potential future perspectives of multifunctional PROTACs for therapeutic development.

Pharmaceuticals doi: 10.3390/ph19091351

Authors: Yi-Xuan Li Xiao-Li Zhou Jing-Wen Xu Chen Zhao You-Yang Li Hai-You Zhao Liang-Peng Li Xin-Xin Tan Yu-Xiang Zhang Bei-Bei Yang Li Li Li-Hui Yin

Background/Objectives: Rotigotine is a conformationally flexible chiral dopamine agonist, but its solution-phase chiroptical behavior and conformational heterogeneity have not been systematically assessed. This study characterized its electronic circular dichroism (ECD), optical rotatory dispersion (ORD), and vibrational circular dichroism (VCD) responses and evaluated their consistency with its absolute configuration. Methods: ECD and ORD data were acquired for rotigotine ((S)-1) and its simplified analog (S)-2 in acetonitrile and methanol, and VCD spectra were recorded in appropriate deuterated solvents. Conformational searches, Boltzmann weighting, and density functional theory/time-dependent density functional theory calculations were performed. A principal-component-analysis/torsion (PCA-Tor) workflow selected a compact but structurally representative conformer ensemble for (S)-1. Results: In acetonitrile, (S)-1 exhibited a positive Cotton effect (CE) at 208.5 nm and a negative CE at 194.0 nm. The Boltzmann-weighted ECD spectrum reproduced the intense negative band below 200 nm and the positive band near 210 nm. The calculated VCD spectrum agreed with the signs and positions of the main experimental bands in the 1500–1100 cm−1 region. Experimental ORD curves of both compounds were negative in acetonitrile and methanol. The calculations reproduced the sign and overall trend, although their magnitudes were protocol- and population-sensitive. Ring puckering strongly affected the calculated ECD and ORD data, whereas side-chain flexibility expanded the accessible conformational space. Conclusions: The combined results support the known S configuration of rotigotine and demonstrate the value of representative conformational sampling for flexible chiral pharmaceuticals. This study also defines the practical utility and limitations of simplified molecular models in chiroptical analysis.

Pharmaceuticals doi: 10.3390/ph19091350

Authors: Jingyi Shen Xiaoya Liu Xuanyan Yan Tao Zhang Xianfang Zhang Huiquan Gu Weimin Chen Zhengwen Wang Qiang Liu

Hepatocellular carcinoma (HCC) remains difficult to control because recurrence, impaired hepatic reserve, and treatment resistance limit durable benefit. Natural products and traditional Chinese medicine (TCM) provide resources that range from drug-lead discovery to adjunctive multicomponent therapy. This review integrates pharmacological and clinical evidence for purified compounds, semisynthetic derivatives, extracts, formulas, and delivery systems. It focuses on metabolic reprogramming and redox homeostasis, stress responses and regulated cell death, tumor cell plasticity and vascular remodeling, and the immune microenvironment and host response. Recent studies have strengthened selected mechanistic claims through chemical probes, functional perturbation, and resistance models. Clinical research has concentrated on recurrence control after surgery or minimally invasive treatment and on combinations with transarterial chemoembolization, targeted agents, and immunotherapy. Randomized trials and prospective cohorts suggest potential benefit in specific settings, although product standardization, external validation, and long-term follow-up remain limited. Major translational barriers include uncertain active constituents, inadequate batch comparability, missing tumor-exposure data, and sparse herb–drug interaction studies. Future development should match target validation, pharmacokinetics, safety assessment, and clinical endpoints to each product class and clarify whether a candidate is best positioned as a drug lead, adjunctive therapy, or supportive intervention.

Pharmaceuticals doi: 10.3390/ph19091349

Authors: Zhihao Zeng Yanchang Liu Xiaoli Bi Wanchun Chen Yunhui Ouyang Jingnian Zhang Weitao Chen Guanlin Xiao

Background/Objective: Traditionally derived from the dried leaves of Microcos paniculata L., Microctis folium (M. folium) has been employed to manage various inflammatory and metabolic conditions. Despite its reported hypolipidemic potential, the active constituents contributing to this effect and their molecular mechanisms of action remain to be determined. Therefore, this study aimed to systematically identify the active hypolipidemic constituents of M. folium and elucidate their mechanisms of action through an integrated strategy combining chemical profiling, pharmacokinetic screening, network pharmacology, and experimental validation. Methods: UPLC fingerprint and UPLC-QQQ-MS/MS were employed to characterize and quantify the chemical constituents of M. folium. Pharmacokinetic analysis was conducted to identify systemically absorbed compounds. Network pharmacology was applied to predict potential targets and signaling pathways. The lipid-lowering effects were subsequently validated in vitro using an oleic acid/palmitic acid (OA/PA)-induced lipid accumulation model in HepG2 cells and in vivo in a Triton WR-1339-induced hyperlipidemia mouse model. Results: UPLC fingerprint analysis identified 15 common peaks among 21 batches of M. folium, with similarity values ranging from 0.885 to 0.990, indicating good chemical consistency. Eighteen representative compounds were quantified, among which flavone C-glycosides and phenolic acids were predominant. Pharmacokinetic results demonstrated that multiple M. folium’s constituents were absorbed into systemic circulation. Network pharmacology analysis identified 73 potential targets related to hyperlipidemia and highlighted the PI3K-Akt signaling pathway as a key regulatory pathway. In vitro experiments showed that M. folium’s compounds significantly reduced intracellular lipid accumulation and oxidative stress in OA/PA-induced HepG2 cells. Furthermore, in vivo studies demonstrated that vitexin, ferulic acid, isoferulic acid, and N-trans-feruloyltyramine significantly reduced serum and hepatic lipid levels, including TC, TG, and LDL-c, alleviated hepatic steatosis, and improved liver injury and oxidative stress markers in Triton WR-1339-induced hyperlipidemic mice. Conclusions: M. folium exerts significant lipid-lowering effects through multi-component and multi-target mechanisms involving regulation of lipid metabolism and oxidative stress, partly mediated by activation of the PI3K-Akt signaling pathway. These findings provide a scientific basis for the development of M. folium’s natural agents for the treatment of hyperlipidemia.

Pharmaceuticals doi: 10.3390/ph19091348

Authors: Qiuqi Zhuang Zhitao Yang Yan Liu

Objective: This study aimed to systematically evaluate the efficacy and safety of cadonilimab in the treatment of digestive system neoplasms. Methods: PubMed, Embase, Cochrane Library, Web of Science, Wiley Online Library, CNKI, Wanfang Data, CBM, and VIP databases were systematically searched to collect randomized controlled trials, non-randomized studies, and real-world studies investigating cadonilimab for the treatment of digestive system neoplasms. The initial search spanned from the inception of the databases to 4 February 2026, with an update search conducted in June 2026. Meta-analysis was performed using Stata 16.0 to systematically assess efficacy and safety outcomes. Results: A total of 10 studies, predominantly single-arm or retrospective, were included in this meta-analysis. Regarding the primary outcomes, because of the clinical heterogeneity across tumor types, pooled estimates should be interpreted as a summary across diverse diseases rather than as a single-tumor estimate. The overall pooled disease control rate (DCR) was 84% (95% CI = 76–91%), with tumor-specific DCRs of 87% in ESCC, 79% in HCC, 88% in G/GEJ adenocarcinoma, and 86% in PDAC. The pooled incidence of any-grade treatment-related adverse events (TRAEs) was 98% (95% CI = 95–100%), and the pooled incidence of any-grade immune-related adverse events (irAEs) was 32% (95% CI = 18–47%). For secondary outcomes: The partial response (PR) rate was 33% (95% CI = 21–46%), and the progressive disease (PD) rate was 11% (95% CI = 5–18%). The pooled incidence of grade ≥ 3 TRAEs was 52% (95% CI = 38–66%), and the pooled incidence of grade ≥ 3 irAEs was 11% (95% CI = 8–16%). However, incomplete reporting of low-frequency AEs across studies likely underestimates the pooled irAE rate. Conclusions: This study suggests that cadonilimab has the potential for antitumor activity in digestive system neoplasms. The observed safety profile was consistent with the known toxicities of immunotherapy, with notably high rates of TRAEs warranting vigilant monitoring. These findings should be considered hypothesis-generating and are insufficient to establish clinical benefit.

Pharmaceuticals doi: 10.3390/ph19091347

Authors: Zhihan Zhu Yu Liu Linglin Feng

Objective: Conventional progesterone (P4) formulations suffer from low bioavailability, severe local irritation, and poor patient adherence due to P4’s poor aqueous solubility. This work aimed to develop and screen a biodegradable PLGA/NMP (Poly(lactic-co-glycolic acid)/N-methyl-2-pyrrolidone) injectable in situ gel for sustained P4 delivery to overcome these clinical limitations. Significance: Commercial oral, vaginal, and oil-based intramuscular P4 preparations cannot maintain stable long-term drug exposure, and they cause injection-site pain/inflammation. The screened in situ depot system reduces administration frequency and local tissue irritation, supporting convenient luteal phase support and pregnancy maintenance. Methods: Nine formulations with variable P4 loading (10–50% w/w) and PLGA concentration (15–55% w/w) were fabricated. Formulations were screened via three core endpoints: injectability (injection force and discharge rate), in vitro sustained release in 10% Hydroxypropyl-β-cyclodextrin (HP-β-CD)-Phosphate-buffered saline (PBS) sink medium, and 7-day subcutaneous histocompatibility in rats. high-performance liquid chromatography (HPLC) was validated for progesterone quantification; Hematoxylin and eosin (H&E) staining assessed local inflammatory responses. Results: Formulations with progesterone ≤ 30% w/w and PLGA ≤ 35% w/w exhibited acceptable injectability (injection force < 50 N; discharge rate > 79%). Higher PLGA concentrations suppressed initial burst release (16.74% at 8 h for 35% PLGA vs. 29.8% for 20% PLGA). All formulations formed stable ellipsoidal subcutaneous depots and completed progesterone release within 4 days. Histopathology revealed only mild local inflammation (histological score = 1) without severe necrosis, superior to highly irritating oil injections in formulation control groups. Conclusions: The screened PLGA-based progesterone in situ gel resolves critical drawbacks of traditional progesterone dosage forms. This low-irritation, sustained-release injectable platform provides a scalable industrial formulation candidate for long-acting hormone therapy.

Pharmaceuticals doi: 10.3390/ph19091346

Authors: Mehmet Fatih Dasiran Hassen Daghmoura Ahmet Akbaş Yavuz Selim Angın Bakiye Akbaş Hatice Aygun Yiğit Uyanıkgil Oytun Erbas

Aim: Radiation-induced intestinal injury is one of the major dose-limiting complications of abdominal radiotherapy and is closely associated with oxidative stress, inflammation, and disruption of epithelial barrier integrity. The present study aimed to investigate the protective effects of verapamil on radiation-induced small-intestinal injury in rats by evaluating histopathological alterations together with tissue levels of TNF-α, 5-LOX, Nrf2, and ZO-1 as markers related to inflammatory, redox-associated, and tight-junction-associated responses. Materials and Methods: Thirty female Wistar albino rats were divided into three groups: Normal Control (n = 10), Radiation + Saline (RAD, n = 10), and Radiation + Verapamil (n = 10). Whole-abdomen irradiation was performed using a single 12 Gy dose delivered by a 6 MV linear accelerator. Verapamil (10 mg/kg/day, s.c.) treatment was started 5 days before irradiation and continued for 5 days afterward. Histopathological injury in jejunal tissues was evaluated using hematoxylin–eosin staining. Tissue levels of TNF-α, 5-LOX, Nrf2, and ZO-1 were measured using ELISA. Results: Whole-abdomen irradiation caused marked histopathological injury characterized by villus disruption, glandular degeneration, and mucosal architectural damage. Irradiation significantly increased small-intestinal TNF-α and 5-LOX levels while decreasing Nrf2 and ZO-1 levels compared with the Normal Control group (p < 0.05). Verapamil treatment significantly attenuated radiation-induced histopathological injury and reduced TNF-α and 5-LOX levels. TNF-α returned to values comparable with the Normal Control group, whereas 5-LOX remained significantly elevated. Verapamil also partially restored Nrf2 and ZO-1 levels compared with the RAD group (p < 0.05), although both remained significantly lower than control levels. Conclusions: Verapamil attenuated radiation-induced small-intestinal injury and was associated with favorable modulation of TNF-α, 5-LOX, Nrf2, and ZO-1 levels. The protective response was partial, with persistent differences in 5-LOX, Nrf2, and ZO-1 compared with controls. These biomarker changes should not be interpreted as direct evidence of functional pathway activation or preservation of intestinal barrier function. Further studies incorporating direct mechanistic and functional assessments are warranted before clinical translation.

Pharmaceuticals doi: 10.3390/ph19091345

Authors: Cody O’Rear Alejandro Chapa-Rodriguez Arminder Singh Safi Afzal Roshan Acharya Madalyn Wickham Usman Younus

Background: Direct oral anticoagulants (DOACs) are widely used among hospitalized patients, yet real-world inpatient bleeding data remains relatively sparse. Most evidence derives from randomized trials enrolling ambulatory, clinically stable patients, or those being treated for a single specific indication such as acute venous thromboembolism (VTE), which may not reflect the risk profile of acutely ill inpatients with multimorbidity, procedural exposures, and fluctuating renal function. This study describes the incidence and types of bleeding events in a real-world inpatient cohort receiving apixaban or rivaroxaban at a single tertiary care center. Methods: We conducted a single-center, retrospective cohort study of 867 hospital encounters involving apixaban (n = 722, 83.3%) or rivaroxaban (n = 145, 16.7%) between 2019 and 2021. The primary outcome was a non-adjudicated, non-standardized composite of any documented bleeding (gastrointestinal bleeding, hematuria, intracranial hemorrhage, vaginal bleeding, or epistaxis) during the index hospitalization; International Society on Thrombosis and Haemostasis (ISTH) major bleeding or clinically relevant non-major (CRNM) bleeding definitions were not applied. Secondary outcomes included individual bleeding subtypes, hemoglobin drop ≥2 g/dL, transfusion requirement, ICU/CCU admission, vasopressor use, and bleeding-related mortality. All comparisons are unadjusted and descriptive. Between-group comparisons used Fisher’s exact test and the Mann–Whitney U test. Results: Bleeding was documented in 13 encounters (1.5% overall; apixaban 1.39%, rivaroxaban 2.07%); given reliance on clinical documentation rather than systematic adjudication, true incidence may be higher. The composite outcome was driven predominantly by gastrointestinal bleeding (n = 10, 76.9% of events). No hemoglobin drop ≥2 g/dL, transfusion, ICU/CCU admission, vasopressor use, or bleeding-related mortality occurred in either group. No statistically significant between-agent difference was observed (p = 0.47); all comparisons are unadjusted and descriptive. Median length of stay (LOS) differed between drug groups (8.6 vs. 6.2 days; p = 0.006), an exploratory finding confounded by patient class imbalance. Conclusions: In this single-center, retrospective cohort, bleeding was documented in 1.5% of encounters; ascertainment limitations mean true incidence may be higher. Non-adjudicated, non-standardized outcome definitions and the small number of events limit comparability with the literature and preclude definitive inference. The absence of a statistically significant between-agent difference should not be interpreted as evidence of equivalence. These findings are hypothesis-generating and should not be used for comparative inference. Prospective, multicenter studies using ISTH-standardized outcomes, indication-level data, and formal adjudication are needed.

Pharmaceuticals doi: 10.3390/ph19091344

Authors: Danielle Alice Vieira da Silva Micaely Cristina dos Santos Tenório Fabiana Andréa Moura Nassib Bezerra Bueno Nathálya da Silva Severino Alexandra Rodrigues Bezerra Bianca Gomes de Souza Marilene Brandão Tenório Fragoso Tauane Alves Dutra Orlando Roberto Pimentel de Araújo Amylly Sanuelly da Paz Martins Marília Oliveira Fonseca Goulart Alane Cabral Menezes de Oliveira

Background/Objectives: This study aimed to evaluate the effects of oral administration of N-acetylcysteine (NAC) in pregnant women with preeclampsia (PE), using serum markers of redox imbalance and cytokine levels and perinatal outcomes. Methods: A double-blind, randomized, placebo-controlled pilot study was carried out from 2021 to 2023. The study was retrospectively registered with REBEC (Brazilian Clinical Trials Registry). After screening, women were randomized into 2 groups according to the type of supplementation: 600 mg of NAC (NAC group) or capsules containing 2% colloidal silicon dioxide and 98% microcrystalline cellulose (placebo group) until delivery. Blood samples were collected to quantify biomarkers of redox imbalance and cytokine levels. Questionnaires containing socioeconomic, clinical, obstetric, and anthropometric data were applied. Results: A total of 51 pregnant women with PE were included (26 in the placebo group and 25 in the NAC group). The use of NAC was associated with a higher gestational age at birth (GA) of the conceptus (placebo group: 37.39 weeks ± 1.96 SD vs. NAC group: 38.45 weeks ± 1.51 SD; p = 0.044). A moderate effect size was observed for this variable (Cohen’s d = 0.52). However, treatment allocation (NAC vs. control) was not independently associated with gestational age at birth after adjustment for covariates (F(1,28) = 0.144; p = 0.707; η2p = 0.005). The adjusted mean difference between groups was 0.92 weeks (95% CI: −0.16 to 2.00). No other significant effects on redox and inflammatory markers were identified using NAC. Conclusions: NAC supplementation did not impact the mode of delivery between the NAC and placebo groups. As a secondary exploratory finding, a possible trend toward prolonged gestational age was observed in the NAC group; however, this result should be interpreted with caution, given the exploratory nature of the analysis, the neutrality of the primary outcome, and the limited sample size.

Pharmaceuticals doi: 10.3390/ph19091343

Authors: Sıtkı Ün Ramazan Sabırlı İbrahim Türkçüer Gergana Lengerova Martina Bozhkova Steliyan Petrov Aylin Köseler

Background: Acute renal colic is a common urological emergency characterized by substantial interindividual variability in analgesic response. Pharmacogenetic variation in OPRM1, COMT, and ABCB1 may contribute to differences in opioid efficacy and pain control. This study primarily evaluated the associations of OPRM1 A118G (rs1799971), COMT Val158Met (rs4680), and ABCB1 C3435T (rs1045642) polymorphisms with opioid analgesic response in patients with acute renal colic. As a secondary exploratory analysis, genotype and allele frequencies were compared between patients and healthy controls. Methods: This prospective case–control study included 150 patients with acute renal colic and 100 healthy controls. Genotyping was performed using TaqMan SNP Genotyping Assays based on real-time polymerase chain reaction. Genotype frequencies were compared between groups using dominant and recessive genetic models, and Hardy–Weinberg equilibrium was assessed. In addition, genotype–phenotype associations were evaluated using pain severity, early analgesic response, initial opioid dose, rescue analgesic requirement, and multivariable logistic regression analyses. Results: In the secondary exploratory case–control analysis, no statistically significant differences in genotype or allele frequencies of OPRM1 rs1799971, COMT rs4680, or ABCB1 rs1045642 were observed between patients with acute renal colic and healthy controls. Within the patient cohort, however, genotype–phenotype analyses identified differences in early analgesic outcomes. Baseline-adjusted 30 min VAS differed according to OPRM1, COMT, and ABCB1 genotype, with the most pronounced difference observed for ABCB1 rs1045642. Patients with the ABCB1 TT genotype had higher adjusted 30 min VAS scores and showed a pattern of greater opioid requirement and more frequent rescue analgesia. In exploratory multivariable analysis, the ABCB1 TT genotype was associated with higher odds of inadequate early analgesic response (adjusted OR = 2.74, 95% CI 1.18–6.37; p = 0.019). Given the limited number of outcome events, this adjusted association should be considered preliminary and hypothesis-generating. Conclusions: No significant differences in the distributions of the polymorphisms investigated were observed between patients with acute renal colic and healthy controls. Within the patient group, ABCB1 genetic variation was associated with early opioid analgesic response, although this finding should be considered preliminary and requires confirmation in larger prospective pharmacogenetic studies before clinical implementation. Any potential future pharmacogenetic application should be considered as an adjunct to established first-line renal–colic management and specifically in patients for whom opioid therapy is clinically indicated.

Pharmaceuticals doi: 10.3390/ph19091342

Authors: Laura Schönfelder Shaoning An Peter Reusch Pedro M. Faustmann Timo Jendrik Faustmann Fatme S. Ismail

Background/Objectives: Glia-mediated inflammation contributes to a wide range of central nervous system (CNS) disorders, including epilepsy. Pregabalin (PGB) and cannabidiol (CBD) are CNS-acting drugs prescribed for various neuropsychiatric conditions, especially seizures. This study investigated the effects of PGB and CBD on glial properties in an astrocyte–microglia co-culture model of inflammation. Methods: Physiological (M5, containing 5–10% microglia) and pathological inflammatory astrocyte–microglia co-cultures (M30, containing 30–40% microglia) were collected from the postnatal brain hemispheres of Wistar rats (P0-P2) according to an established protocol and treated with different concentrations of PGB (3, 10, 30 and 60 µg/mL) for 24 h or CBD (50, 500 and 1000 ng/mL) for 1 h or 24 h. Metabolic activity was assessed by the MTT assay. Microglial phenotypes and astroglial connexin (Cx)43 expression were detected by immunocytochemistry. Results: In M5 co-cultures, short-term incubation (1 h) with high concentrations (1000 ng/mL) of CBD significantly reduced glial viability (p < 0.05), while no significant changes were observed in M30 co-cultures. After 24 h of incubation, M5 cultures exhibited a significant increase in metabolic activity at 50 ng/mL (p < 0.0001) and 500 ng/mL (p < 0.05), and a reduction at 1000 ng/mL (p < 0.05), suggesting impaired viability at high concentrations under physiological conditions. The distribution of microglial phenotypes in physiological M5 co-cultures incubated with CBD remained unchanged. In M30 co-cultures, CBD incubation for 1 and 24 h significantly reduced microglial activation and promoted a shift from reactive, phagocytic to homeostatic, ramified microglial phenotype (p < 0.05, p < 0.01, p < 0.0001). In contrast, PGB did not affect glial cell viability, microglial phenotypes or Cx43 expression in physiological and inflammatory co-cultures, indicating that the mechanisms of action of PGB probably do not include modulation of glial cells in vitro. Conclusions: The inhibition of microglial reactivity by CBD suggests potential positive effects on the neuroinflammatory component involved in the pathogenesis of CNS disorders such as epilepsy.

Pharmaceuticals doi: 10.3390/ph19091341

Authors: Xingli Zhang Huili Zhu Chengshu Yu Shuang Huang Dongdong Fang

Background/Objectives: Ovarian cancer is a prevalent gynecological malignancy with high mortality rates worldwide. Harmine hydrochloride (HMH), a tricyclic β-carboline alkaloid derived from the seeds of the medicinal plant Peganum harmala L., has demonstrated anti-tumor activity. This study investigated the effect of HMH on ovarian cancer and elucidated its underlying molecular mechanism. Methods: The anti-ovarian cancer effects of HMH were evaluated using the CCK-8 assay, cell cycle and apoptosis analyses, as well as a cell migration assay. The RNA-seq was utilized to explore the mechanisms of HMH. Western blot, qRT-PCR and IF were employed for further validation of the expression levels of relevant molecules. The Kaplan–Meier (KM) curve was conducted to evaluate the possible link between NFAT1 levels and the prognosis of ovarian cancer. The biological functions of NFAT1 were explored through siRNA knockdown experiments. Auto-dock molecular docking and DARTS were used to analyze the direct interaction between HMH and NFAT1. The in vivo efficacy of HMH was examined through bioluminescence imaging of intraperitoneal xenografts established with luciferase-expressing HEY cells. Results: HMH induced apoptosis, arrested the cell cycle, and suppressed migration of HEY and CAOV3 cells. NFAT1 was highly expressed in the nucleus of HEY and CAOV3 cells and promoted ovarian cancer progression. In addition, low NFAT1 expression levels had significantly improved overall survival and progression-free survival. Moreover, HMH reduced NFAT1 expression and reversed Ion-induced NFAT1 levels in ovarian cancer cell nuclei. The molecular docking and DARTS results suggested that HMH binds to NFAT1. Treatment with MG132 effectively blocked HMH-induced NFAT1 downregulation. Furthermore, HMH effectively blocked intraperitoneal xenograft development, and extended the life span of animals bearing HEY/Luc cells, when used alone or in combination with CDDP. Conclusions: The research indicated that HMH exerts anti-ovarian cancer effects, at least in part, by targeting NFAT1. Additionally, our research revealed that NFAT1 promotes ovarian cancer progression, and plays a significant role in HMH’s anti-cancer activity. These results may aid in the development of novel candidates for ovarian cancer and identify a new target for ovarian cancer treatment.

Pharmaceuticals doi: 10.3390/ph19091338

Authors: Zheming Xiong Yijie Su Luping Shi Shuyuan Ge Ge Wang Fangyu Li Xu Liu Jianguang Li Zejiang Ma

Background: Metabolic dysfunction-associated fatty liver disease (MAFLD) is a prevalent chronic liver disorder with limited pharmacological therapies, highlighting the need for novel multi-target agents from traditional medicinal plants. Artemisia rupestris L. has hepatoprotective effects, but its active constituents and mechanisms against MAFLD remain unexplored. Methods: The chemical profile of an ultrasonically-assisted 70% ethanol extract of Artemisia rupestris L. (ARE) was determined using UPLC-Q-TOF-MS. Network pharmacology, molecular docking, 100-ns molecular dynamics (MD) simulations, and in vitro experiments in oleic acid-induced HepG2 cells were integrated to identify active compounds, core targets, and pathways, and to validate lipid-lowering and hepatoprotective effects. Results: Seventy-eight compounds were identified, mainly flavonoids, phenolic acids, and terpenoids. Network analysis highlighted AKR1B10 and MAPK14 as core targets and isorupestonic acid and rupestonic acid as key components. MD simulations supported the stable binding of isorupestonic acid to AKR1B10 (RMSD ~1.6–3.2 Å), driven by van der Waals and electrostatic interactions. In vitro, ARE dose-dependently reduced intracellular triglycerides, total cholesterol, and LDL-C, decreased lipid droplets, and lowered AST/ALT levels, with a high-dose efficacy comparable to atorvastatin. Conclusions: Through in vitro experiments and network pharmacology analyses, this study preliminarily elucidated the beneficial effects of ARE on lipid deposition in hepatocytes and its potential mechanism, thereby providing an experimental basis and potential avenues for subsequent in vivo studies on pharmacodynamics, pharmacokinetics, and toxicology.

Pharmaceuticals doi: 10.3390/ph19091340

Authors: Musun Park Hyeun-Kyoo Shin Yujin Choi

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.

Pharmaceuticals doi: 10.3390/ph19091339

Authors: Zike Xu Hongxia Luo Yimin Zhao Xuhui Wang Sha Chen

Background/Objectives: Cerebral ischemia–reperfusion injury (CIRI) poses therapeutic challenges because of oxidative stress, blood–brain barrier disruption, and neuronal apoptosis, limiting current treatments. Paeoniflorin (PF) from Paeonia lactiflora has neuroprotective potential, but its multi-target mechanisms remain unclear. This study investigated the role and mechanisms of PF in CIRI, focusing on neuron–endothelial crosstalk. Methods: Oxygen–glucose deprivation/reoxygenation (OGD/R) models were established using SH-SY5Y (human neuroblastoma) and HCMEC/D3 cells (human cerebral microvascular endothelial). Network pharmacology was used to predict potential PF targets and pathways. RNA sequencing, molecular docking, and molecular dynamics simulation were performed to screen and evaluate PF binding characteristics with key targets, and MTT, flow cytometry, Western blotting, and co-cultures were employed to detect paracrine interactions. Results: Network pharmacology and transcriptomics identified VEGF/PI3K-AKT/mTOR pathway enrichment. Molecular docking confirmed stable PF binding to VEGF-A (−8.4 kcal/mol), AKT (−5.5 kcal/mol), and mTOR (−9.6 kcal/mol). PF (10–80 μM) showed no cytotoxicity and reduced OGD/R injury in a concentration-dependent manner (maximal at 40 μM). PF activated VEGF/PI3K-AKT/mTOR signaling, reducing apoptosis by 57% (SH-SY5Y) and 33% (HCMEC/D3); PI3K inhibitor LY294002 abolished these effects. PF-treated HCMEC/D3-conditioned media enhanced OGD/R neuronal viability, verifying paracrine crosstalk. Conclusions: PF alleviated CIRI by directly protecting neurons and indirectly modulating neuron–endothelial crosstalk through VEGF/PI3K-AKT/mTOR activation, supporting its multi-target therapeutic potential.

Pharmaceuticals doi: 10.3390/ph19091337

Authors: Vladislav S. Sukhachev Sergey M. Ivanov Dmitry A. Filimonov Anastasia V. Rudik Vladimir V. Poroikov

Background/Objectives: Adverse drug reactions (ADRs) caused by drug–drug interactions (DDIs) represent an important problem in pharmacotherapy, especially in patients receiving multiple medications. Most computational approaches to DDI-associated ADR prediction formulate the task as a binary classification, but they do not explicitly consider the severity of adverse reactions. Our study aims to develop structure-based models that predict the severity of ADRs associated with specific drug pairs. Methods: Datasets were generated using DrugMAP as the source of drug pair–ADR associations with annotated severity categories, and TwoSides was used as an additional source to generate conditionally negative examples. The drug pairs were represented using PoSMNA descriptors, which encode pair-specific structural features derived from the molecular structures of both compounds. Predictive models were built using PASS DDI software. Model performance was evaluated using a modified cross-validation procedure that excluded compound-level overlap between the training and test sets, thereby reducing information leakage caused by the repeated occurrence of the same drugs in different pairs. Results: Models were developed for 14 clinically relevant ADR types, including cardiovascular, hepatotoxic, nephrotoxic, hemorrhagic, metabolic, and neurological effects. The unweighted class-level macro-average AUC values ranged from 0.830 for the Major category to 0.911 for the Minor category, while balanced accuracy ranged from 0.776 to 0.857. Predictive performance varied significantly between ADR types and severity categories. Higher accuracy was observed for some ADR types that were better captured by the structure-based descriptors used in this study, whereas complex multifactorial reactions, such as hepatotoxicity, were less accurately predicted. Case-based assessment using clinically documented drug combinations showed that the predicted severity profiles were generally consistent with the expected clinical risk patterns. Conclusions: The proposed approach demonstrates that the PoSMNA descriptors of drug pairs can be used for preliminary prediction of DDI-associated ADR severity. The developed models can help to filter out potentially dangerous drug combinations at an early stage and are implemented in the AdverDDIPred web-application.

Pharmaceuticals doi: 10.3390/ph19091336

Authors: Shini Zhang Xiao-Li Luo Hangxing Yang Ganghua Huang Jing Leng Tao Yang

Background: Alzheimer’s disease (AD) is a prevalent neurodegenerative disorder for which effective treatments remain limited. Liubao tea, a distinctive post-fermented dark tea, develops unique aged and areca nut aromas through prolonged storage. However, comprehensive studies on its anti-Alzheimer’s disease effects are lacking. This study aimed to identify the bioactive constituents of Liubao tea with varying storage durations and aroma profiles, and to further evaluate its neuroprotective potential against Alzheimer’s disease. Methods: Chromatography and spectrophotometry were employed to quantitatively determine the primary bioactive components in various Liubao tea samples. A D-galactose-induced Alzheimer’s disease zebrafish (Danio rerio) model was established to evaluate the neuroprotective effects of the aqueous extract of Liubao tea. Results: The contents of theabrownins, theaflavins, tea pigments, and catechins showed potential associations with the anti-AD capacity of Liubao tea. Variations in these bioactive components appeared to correlate with the neuroprotective activity. Conclusions: This study elucidates the anti-AD potential of aged Liubao tea and associates its characteristic constituents with neuroprotective effects. The findings provide essential theoretical support for developing Liubao tea as a promising natural product candidate worthy of further anti-AD research.

Pharmaceuticals doi: 10.3390/ph19091334

Authors: Yoon-Mi Lee Sumin Han Gyun Seok Park Judy Gopal Jae-Wook Oh

Background/Objectives: Colorectal cancer (CRC) remains a major cause of cancer-related mortality worldwide, largely owing to metastasis, recurrence, and resistance to conventional therapies. Natural-product-derived compounds represent an important source of anticancer drug candidates. This study investigated the anticancer activity of purified narciclasine, which was tentatively annotated as a major detected constituent of Lycoris sanguinea extract, and examined its association with HELLS-associated DNA damage responses in CRC cells. Methods: Human CRC cell lines HT29 and HCT116 were used to evaluate the cytotoxic and mechanistic effects of L. sanguinea extract and narciclasine. Cell viability, clonogenic growth, apoptosis, mitochondrial membrane potential, cell cycle distribution, migration, and invasion were assessed. Western blotting was performed to analyze apoptosis-, cell cycle-, epithelial–mesenchymal transition (EMT)-, and DNA damage-related proteins. UPLC-QTOF/MS analysis was used to identify major phytochemical constituents. HELLS knockdown and γ-H2AX immunofluorescence staining were conducted to examine the involvement of HELLS in narciclasine-associated DNA damage and apoptosis. Results: L. sanguinea extract reduced CRC cell viability, suppressed clonogenic growth, induced mitochondrial dysfunction and caspase-dependent apoptosis, and promoted G2/M cell cycle arrest with decreased cyclin B1 and increased p53/p21 expression. The extract also inhibited migration and invasion, accompanied by reduced β-catenin, N-cadherin, vimentin, slug, and snail expression and increased E-cadherin expression. UPLC-QTOF/MS tentatively annotated narciclasine as a major detected constituent. Purified narciclasine recapitulated several anticancer effects of the extract, accompanied by HELLS downregulation, increased γ-H2AX accumulation, reduced cyclin B1 expression, and enhanced PARP and caspase-3 cleavage. HELLS knockdown further sensitized CRC cells to narciclasine-associated γ-H2AX accumulation, G2/M arrest-associated signaling, and apoptosis. Conclusions: Narciclasine treatment was associated with HELLS downregulation, DNA damage-associated signaling, G2/M cell cycle arrest-associated changes, EMT suppression, and apoptosis in CRC cells. These findings suggest that narciclasine is a promising natural-product-derived lead compound for further preclinical evaluation in HELLS-associated CRC vulnerabilities.

Pharmaceuticals doi: 10.3390/ph19091335

Authors: Giorgio Cameli Alessia Piergentili Eleonora Spinozzi Alessia Tombesi Riccardo Petrelli Loredana Cappellacci Maria Beatrice Morelli

Background: Glioblastoma (GBM) is the most aggressive subtype of malignant glioma. Current therapeutic options remain limited, highlighting the need for the development of novel compounds capable of improving the efficacy of standard treatments. This study aimed to evaluate the biological activity of a series of pyrazolone-based hydrazone compounds (TPPs) in vitro GBM cell lines. Methods: The eight TPPs were synthesized by a nucleophilic addition reaction of different substituted hydrazines with 1-(5-hydroxy-3-methyl-1-phenyl-1H-pyrazol-4-yl)-2-phenylethan-1-one and tested on two human GBM cell lines, T98 and U87. The cytotoxic effects were evaluated via MTT assay. The most active compound was further investigated at IC50 and IC25 concentrations to evaluate mechanisms of cellular damage, including reactive oxygen species (ROS) production and mitochondrial membrane potential (ΔΨm) changes. Additional assays included colony formation, cell cycle analysis, and evaluation of DNA damage and apoptosis markers. Results: TPP25 exhibited the highest activity with IC50 values of 11.01 μM (95% CI: 10.42 to 11.64) and 13.12 μM (95% CI: 10.23 to 16.87) on T98 and U87 lines, respectively. Treatment induced early ROS production and mitochondrial depolarization, along with a significant reduction in colony formation. Cell cycle analysis revealed accumulation in the sub-G0 phase, consistent with increased cell death, supported by propidium iodide uptake. Furthermore, the results suggest the involvement of an apoptotic-like mechanism as supported by Annexin V positivity, γ-H2AX upregulation and transient caspase-3 activation. Conclusions: TPP25 demonstrates significant in vitro cytotoxicity, likely driven by a pro-apoptotic mechanism. This profile positions it as a potential lead compound for further preclinical evaluation, supporting its future transition into in vivo GBM models.

Pharmaceuticals doi: 10.3390/ph19091333

Authors: Adina Hamiti Yingying Geng Pengfei Huang Jinghui Xie Tingting Cui

Vitiligo is a chronic acquired depigmenting disorder characterized by progressive melanocyte loss. Its pathogenesis involves a multifactorial interplay among oxidative stress, impaired melanocyte regeneration, immune-mediated cytotoxicity, and neuroendocrine dysregulation. In recent years, traditional Chinese medicine (TCM)-derived active ingredients have attracted increasing research interest as potential interventions for vitiligo, although most isolated compounds remain at the preclinical stage. This review summarizes pharmacological and natural compounds applied in vitiligo treatment and highlights their regulatory mechanisms on core pathogenic modules. A comprehensive analysis of natural and semisynthetic active ingredients is provided, covering their preclinical and clinical evidence, mechanisms of action, and therapeutic relevance. Multiple ingredients, including baicalein, quercetin, curcumin, paeoniflorin, kaempferol, glycyrrhizin, and epigallocatechin-3-gallate (EGCG), have demonstrated antioxidant, anti-inflammatory, immunomodulatory, and neuroprotective effects that are involved in the pathogenesis of vitiligo. Some TCM-derived interventions, such as Ginkgo biloba L. extract and compound glycyrrhizin, have been evaluated clinically, whereas most isolated active compounds, including baicalein and quercetin, remain at the preclinical stage. Overall, TCM-derived active ingredients represent a promising therapeutic strategy for vitiligo. However, continued translational and clinical research is still required to optimize formulations, dosing regimens, and safety profiles, thereby facilitating their integration into routine clinical practice.

Pharmaceuticals doi: 10.3390/ph19091332

Authors: Jungha Min Sueun Shin Sohyeon Park Jeongha Yun Kaylnn Park Sandy Jeong Rhie

Background/Objectives: Real-world antidepressant use for depressive disorder varies across clinical settings. This study aimed to characterize prescribing pathways and evaluate the comparative safety of frequently prescribed antidepressants in a large multicenter cohort with newly diagnosed depressive disorder. Methods: We conducted a retrospective multicenter observational study using electronic health record data standardized to the OMOP Common Data Model (1999–2026) in adults initiating antidepressants. Prescribing patterns among 13,381 patients were visualized using sunburst plots and Sankey diagrams. Comparative safety, escitalopram vs. sertraline and trazodone, was evaluated across 18 hospitals (n = 44,542) for nine clinical safety outcomes using propensity score-matched Cox proportional hazards models and empirically calibrated random-effects meta-analysis. Results: Selective serotonin reuptake inhibitors, particularly escitalopram (21.4%, n = 2868), were the preferred antidepressant. Common adjunctive medications included trazodone (14.6%, n = 794) and quetiapine (14.0%, n = 761), though utilization varied across hospitals. No clear differences were detected for most safety outcomes between cohorts. However, empirically calibrated QTc prolongation risk was significantly increased with sertraline relative to escitalopram (HR 1.75, 95% CI 1.10–2.78), a signal requiring further confirmation. Conclusions: This study demonstrates substantial institutional variation exists in real-world antidepressant treatment pathways and combination strategies. While comparative safety profiles were broadly comparable across evaluated outcomes, the isolated QTc prolongation signal for sertraline warrants targeted external confirmation. Furthermore, trazodone and escitalopram cohorts should not be regarded as clinically interchangeable first-line options, supporting individualized, evidence-based antidepressant selection in routine clinical practice.

Pharmaceuticals doi: 10.3390/ph19091331

Authors: Goksu Kasarci-Kavsara Sinem Bireller Baris Ertugrul Bedia Cakmakoglu

Background/Objectives: Drug resistance is a major challenge in cancer therapy, and mitochondria contribute to this process by controlling both metabolic adaptability and cell survival signaling. Mitophagy, the selective lysosomal removal of dysfunctional mitochondria, has been implicated in therapy resistance, yet its role in sunitinib-resistant renal cancer remains poorly defined. Methods: In this study, acquired sunitinib resistance was established in ACHN renal cancer cells through eight months of stepwise dose escalation. Initial selection conditions were determined using CCK-8 viability and crystal violet colony assays in parental ACHN cells, whereas sustained proliferation under continuous sunitinib exposure was used as the operational criterion for the resistant phenotype. Resistant and parental sensitive cells were treated with 25 µM and 50 µM succinic acid, alone or in combination with sunitinib. Gene expression of BNIP3, NIX, FUNDC1, LC3, PINK1, Parkin, PGAM5, SRC, LONP1, and ATP5F1A was measured by RT-qPCR, and BNIP3 and NIX protein levels were assessed by ELISA. Results: Resistant cells showed significant upregulation of receptor-mediated mitophagy components BNIP3, NIX and FUNDC1 (p < 0.05), with no significant change in LC3, alongside suppression of PINK1, Parkin, and mitochondrial homeostasis-associated genes LONP1, PGAM5, and ATP5F1A (p < 0.05). Succinic acid predominantly reduced BNIP3 and NIX protein levels in both cell lines and suppressed BNIP3, NIX, and LC3 mRNA expression in resistant cells. In contrast, the sunitinib + 50 µM succinic acid combination selectively increased PARKIN, PGAM5, LONP1, and ATP5F1A expression in resistant cells (2.49- to 5.98-fold; p < 0.005), a pattern not observed in parental cells. Conclusions: These findings indicate that sunitinib resistance in ACHN cells is associated with upregulated transcription of receptor-mediated mitophagy components and downregulated transcription of PINK1/Parkin pathway genes, and that exogenous succinic acid selectively upregulates PARKIN and other mitochondrial homeostasis-related gene expression in resistant, but not parental, cells.

Pharmaceuticals doi: 10.3390/ph19091330

Authors: Ibrahim M. Aziz Rawan M. Alshalan Reem M. Aljowaie

Background/Objectives: The increasing global prevalence of chronic diseases has intensified the search for potential natural therapeutic agents. Fagonia arabica is a medicinal plant widely used in traditional medicine; however, comparative information on the phytochemical composition and biological activities of its leaves and seeds remains limited. To the best of our knowledge, this study represents the first comparative investigation of the phytochemical profiles and bioactivities of methanolic leaf (FALE) and seed (FASE) extracts of F. arabica collected from Riyadh, Saudi Arabia. Methods: Methanolic extracts were analyzed by gas chromatography–mass spectrometry (GC–MS), and their total phenolic content (TPC), total flavonoid content (TFC), antioxidant, antimicrobial, anticancer, and antidiabetic activities were evaluated using standard in vitro assays. Results: GC–MS analysis identified 60 and 68 compounds in FALE and FASE, respectively, with (9Z,12Z,15Z)-octadeca-9,12,15-trienoic acid, hexadecanoic acid, dimethyl-2,3,4,7,8,9,11,12,14,15,16,17-dodecahydro-1H-cyclopenta[a]phenanthren-3-ol, and (1R,3aR,5aR,5bR,7aR,9S,11aR,11bR,13aR,13bR)-3a,5a,5b,8,8,11a-Hexamethyl-1-(prop-1-en-2-yl)-1,2,3,4,5,6,7,7a,9,10,11,11b,12,13,13a,13b-hexadecahydrocyclopenta[a]chrysen-9-ol as major constituents. FASE exhibited higher TPC and TFC values (104.13 mg GAE/g and 62.16 mg QE/g, respectively) than FALE. Both extracts demonstrated antioxidant activity, with FASE showing greater potency against DPPH and ABTS radicals (IC50 = 63.3 and 42.4 μg/mL, respectively). FASE also displayed broad-spectrum antimicrobial activity, particularly against Staphylococcus aureus (MIC = 7.81 μg/mL). Furthermore, FASE exhibited stronger concentration-dependent cytotoxicity against MCF-7 and HepG2 cells, with IC50 values of 106.8 and 89.5 μg/mL, respectively, compared with 112.5 and 103.1 μg/mL for FALE. Apoptosis induction in MCF-7 and HepG2 cells was associated with upregulation of caspase-3, caspase-8, caspase-9, and Bax, alongside downregulation of Bcl-2 and Bcl-xL. In addition, FASE exhibited potent α-amylase and α-glucosidase inhibitory activities (IC50 = 61.62 and 67.38 μg/mL, respectively), comparable to acarbose. Conclusions: These findings highlight F. arabica seed extract as a promising source of multifunctional bioactive compounds with potential pharmaceutical application.

Pharmaceuticals doi: 10.3390/ph19091329

Authors: Ekaterina I. Alexeeva Natalia M. Kondrateva Tatyana M. Dvoryakovskaya Dmitry A. Kudlay Ivan A. Kriulin Maria S. Botova Irina T. Tsulukiya Elizaveta A. Krekhova Meiri S. Shingarova Maria Y. Kokina Anna N. Fetisova Tatiana Y. Kriulina Kseniya B. Isaeva Aleksandra M. Chomakhidze Christina V. Chibisova Mikhail M. Kostik

Introduction: Despite the growing spectrum of modern drugs for the treatment of juvenile idiopathic arthritis (JIA), patients with a refractory, progressive course of the disease remain a clinical challenge. B-cell-depleting therapy has demonstrated efficacy in adult rheumatoid arthritis and several pediatric rheumatic diseases, encouraging us to evaluate rituximab (RTX) for children with refractory JIA. Objective: This study aimed to assess the long-term efficacy and safety of rituximab therapy in children with refractory juvenile idiopathic arthritis. Methods: This retrospective, single-center, observational study included 106 patients with JIA who had failed previous therapy. The efficacy and safety of the therapy were evaluated at baseline and at 3, 6, 12, 24, 36, and 48 months post-initial rituximab infusion using the American College of Rheumatology Pediatric Criteria (ACRpedi 30/50/70/90), C. Wallace criteria for inactive disease/remission, and the JADAS71 activity index until patient completion or withdrawal from the study. The primary endpoint was achieving an ACRpedi50 response by 3 months, and the secondary endpoint was achieving inactive disease status according to C. Wallace criteria by 6 months of follow-up. Results: An ACRpedi50 response was achieved in 78/100 (78%) patients by month 3 of rituximab therapy, and inactive disease status was reached in 48/93 (52%) patients by month 6. More than half of the patients demonstrate inactive disease or remission at each observation point. During rituximab therapy, oral glucocorticoids were discontinued in 18/57 (32%) patients, and 28/57 (49%) patients were able to halve their corticosteroid dose (p = 0.03). Rituximab demonstrated an acceptable safety profile, with a total of 351 adverse events (non-serious AEs: 304/351, 87%) and a serious AE rate of 0.8 per 100 patient-years. Conclusions: Rituximab can reduce disease activity and lower the dose of oral glucocorticoids in patients with refractory JIA, serving as a viable therapeutic option with an acceptable safety profile.

Pharmaceuticals doi: 10.3390/ph19091328

Authors: Sara Vasović Lucija Vasović Nikola Martić Somyot Chirasatitsin Velibor Vasović Saša Vukmirović Nebojša Pavlović

Peptide therapeutics occupy a unique position between small organic compounds and large protein biomolecules, combining high specificity, strong pharmacological efficacy, and favourable safety profiles. Consequently, they have emerged as important therapeutic agents for a wide range of diseases, including metabolic and oncological disorders. However, oral administration of peptide drugs remains a major challenge due to extensive enzymatic degradation, low intestinal permeability, mucus entrapment, and presystemic metabolism within the gastrointestinal tract. This review provides a comprehensive overview of contemporary strategies for improving oral peptide delivery, with special emphasis on emerging pharmaceutical formulation technologies, bioinspired delivery systems and ingestible device-assisted approaches. A qualitative literature search was conducted using major scientific databases and included relevant publications available up to May 2026. The analysis identified the main barriers responsible for low oral bioavailability of peptide drugs, as well as promising approaches to overcoming these obstacles, including peptide modification, enzyme inhibition, permeation enhancement, mucolytic strategies, and advanced carrier systems. Special attention is given to multifunctional carrier systems, ingestible medical devices and bile acid-inspired technologies as emerging directions in oral peptide delivery. The convergence of pharmaceutical sciences, bioinspired formulation strategies and biomedical engineering is expected to accelerate the clinical translation of oral peptide formulations and enable their therapeutic potential to be fully exploited.

Pharmaceuticals doi: 10.3390/ph19091327

Authors: Mervenur Yavuz Firli R. P. Dewi İlknur Keskin Turan Demircan

Background: Drug repurposing represents an accelerated and cost-effective approach to discovering novel oncologic therapeutics. Here, we investigated the anticancer potential and underlying mechanisms of marbofloxacin (MBF), a veterinary fluoroquinolone (FQ), against breast cancer (BC) cells. Methods: The cellular impacts of MBF on cell viability, anchorage-dependent growth, tumorigenicity, migration, apoptosis, proliferation, senescence, and mitochondrial function were thoroughly characterized. To further elucidate its mechanistic activity, real-time qRT-PCR, untargeted LC-MS/MS-based metabolomics, network pharmacology, and molecular docking analysis were integrated. Results: MBF suppressed BC cell growth by inhibiting cellular proliferation and migration, disrupting mitochondrial membrane potential, and inducing ROS-mediated apoptosis and irreversible cellular senescence. These phenotypic impacts were accompanied by upregulation of tumor suppressors such as CDKN1A and PUMA and downregulation of oncogenes including MKI67, BIRC5, and BCL-2. Metabolomic analysis revealed broad suppression of biosynthesis-related metabolic pathways, characterized by the depletion of critical polyamines and nucleotide pathways. Network pharmacology and molecular docking analyses identified EGFR and HSP90AA1 as putative hub proteins potentially associated with the observed anticancer phenotype. Conclusions: These results provide initial evidence that MBF induces metabolic and molecular rewiring in BC, highlighting its promise as a repositionable therapeutic candidate.

Pharmaceuticals doi: 10.3390/ph19091326

Authors: Nawaf Alshammari Reyaz Hassan Mitesh Patel Mohd Adnan

Neurodegenerative diseases represent a major cause of disability and death, but early diagnosis, prognosis, and therapeutic monitoring are challenging due to biological heterogeneity and the absence of disease-specific biomarkers. Neurofilament light chain (NfL) is a highly sensitive fluid biomarker of neuroaxonal injury with well-established clinical utility in selected neurological disorders, especially in disease monitoring and prognostic evaluation. However, since NfL is not disease-specific, the interpretation has to be integrated with complementary molecular, imaging, and clinical biomarkers. Recent advances in genomics, epigenomics, transcriptomics, proteomics, metabolomics, microbiome profiling, and neuroimaging provide complementary information about the molecular and biological processes underlying neurodegeneration. Artificial intelligence (AI) and machine-learning approaches also allow the integration of these heterogeneous datasets for multimodal biomarker modeling. This review examines the biological and clinical relevance of NfL across major neurodegenerative diseases and critically discusses its combination with multi-omics, neuroimaging, and AI-based approaches. Special emphasis is placed on disease monitoring, prognosis, patient stratification, and therapeutic-response modeling, distinguishing established clinical applications from emerging research directions. The present review also addresses ongoing methodological challenges, including assay standardization, data harmonization, model interpretability, multicenter validation, and clinical translation. Finally, future potential is discussed for NfL-based multimodal biomarker frameworks in precision neurology.

Pharmaceuticals doi: 10.3390/ph19091325

Authors: Deniz Öğütmen Koç İbrahim Sarbay Melike Mercan Başpınar Lütfi Mangal Burcu Eda Arda Hande Sipahi

Background/Objectives: Orlistat is a gastrointestinal lipase inhibitor used for weight management. Although its established safety profile is largely characterised by fat malabsorption-related gastrointestinal events, post-marketing reports suggest a broader spectrum of adverse events. Given the increasing intersection between obesity pharmacotherapy and the management of non-alcoholic fatty liver disease/metabolic dysfunction-associated steatotic liver disease (NAFLD/MASLD), this study evaluated gastrointestinal, hepatobiliary, pancreatic, and anorectal adverse-event reporting signals associated with orlistat in the U.S. Food and Drug Administration (FDA) Adverse Event Reporting System (FAERS) and assessed concordance with regulatory prescribing information. Methods: Quarterly FAERS files from 2004 Q1 through 2026 Q1 were analysed; some retained reports had Initial FDA Received Date (FDA_DT) values dating back to 1999. Descriptive analyses included all orlistat-associated reports, whereas disproportionality analyses were restricted to primary-suspect reports. Sixty-five prespecified Medical Dictionary for Regulatory Activities (MedDRA) preferred terms (PTs) were screened. Reporting odds ratios (RORs), 95% confidence intervals (CIs), and proportional reporting ratios (PRRs) were calculated. A signal was defined as ROR > 2.0, lower bound of the 95% CI > 1.0, and n ≥ 3. Two-sided p values calculated using Fisher’s exact test on the corresponding 2 × 2 contingency tables were adjusted using the Benjamini–Hochberg false discovery rate procedure to address multiple testing. Signal-positive PTs were cross-referenced with prescribing information from three jurisdictions. Results: Overall, 30,454 deduplicated orlistat-associated reports were identified, including 16,684 primary-suspect reports. Thirty-eight of the 65 prespecified MedDRA PTs met the signal criteria, and all remained statistically significant after Benjamini–Hochberg false discovery rate correction (q < 0.05). The strongest signals were rectal discharge (ROR 1175.53; 95% CI 1093.44–1263.78) and steatorrhoea (ROR 1147.99; 95% CI 1058.12–1245.50); change in bowel habit also showed a high ROR (47.32; 95% CI 38.27–58.51). Unlabelled signal-positive PTs included constipation, faeces hard, cholecystitis, and irritable bowel syndrome; change in bowel habit showed jurisdiction-specific labelling discordance. Conclusions: Orlistat-associated FAERS signals included both expected fat-malabsorption-related events and additional gastrointestinal, hepatobiliary, pancreatic, and anorectal PTs, several of which were not explicitly represented in the evaluated regulatory labels. These findings reflect disproportionate reporting rather than incidence, absolute risk, or causality and require confirmation in independent clinical or epidemiological data sources. They do not, by themselves, support regulatory labelling changes.

Pharmaceuticals doi: 10.3390/ph19081324

Authors: Simona Ardelean Ioana Ciopănoiu Ioana Cuc-Hepcal Anda O. J. Samoila Corina Morodan Mihaela Borlea Silviu L. Constantinescu Oana Koppandi Sorina Ciurlea Carmen Tomoroga Adriana Ledeți Livia C. Borcan George A. Drăghici Paul Albu Cristina A. Dehelean

Most active pharmaceutical ingredients (APIs) reach their target by passive systemic distribution, so the dose required for efficacy at the lesion is set by what healthy tissue can tolerate; conventional dosage forms consequently produce pharmacokinetic profiles that oscillate between toxic peaks and sub-therapeutic troughs. Nanoparticulate carriers (liposomes, lipid nanoparticles, polymeric and inorganic systems, and biomimetic carriers) and hydrogels (natural, synthetic, supramolecular, and microgel-assembled) have emerged as the dominant strategies to address this, increasingly combined as hybrid nanoparticle–hydrogel constructs in which the gel provides locoregional retention and the nanoparticles provide cargo protection, intracellular delivery and stimuli responsiveness. Stimuli-responsive chemistries (pH, redox, enzyme, ROS, hypoxia, temperature, light, magnetic, ultrasound, glucose, and multi-stimuli logic) translate the molecular signatures of a disease into spatiotemporally controlled cargo release. This narrative review consolidates the state of the art (prioritizing 2022–2026) and departs from the conventional carrier-type survey in one respect: the literature is read along an explicit chain—disease cue, sensing chemistry, carrier architecture, release mechanism and kinetics, administration route, and clinical readiness—which exposes a variable that classification by carrier type conceals. Across all three material classes, what governs release behavior is not primarily the carrier chemistry but the identity of the released species (dissolved drug, drug from an embedded nanoparticle, an intact nanoparticle, and a matrix fragment) and the transport step that limits it. This is why power-law exponent analysis developed for dissolved drug fits particulate release poorly, why statistical goodness-of-fit cannot by itself establish a release mechanism, and why carrier class predicts clinical readiness less well than administration route and regulatory product type. Translational hurdles—CMC complexity, regulatory fragmentation, anti-PEG immunogenicity, and the structural mismatch between preclinical promise and clinical efficacy—are critically appraised in light of previously reported <1% delivery efficiency analysis. This review identifies converging strategies that could move stimuli-responsive controlled release from an aspirational outcome to a routine clinical reality.

Pharmaceuticals doi: 10.3390/ph19081323

Authors: Mujeeb Ul Naeem Syeda Farwa Naqvi Yousaf Khan Samina Aslam Syed Aminullah Azmatullah Khan Thoraya A. Farghaly Wajid Rehman

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.

Pharmaceuticals doi: 10.3390/ph19081322

Authors: Alaa M. Elsayad Khaled A. Elsayad

Background/Objectives: Human carbonic anhydrase (CA) isoforms are clinically relevant zinc metalloenzymes, but their highly conserved catalytic Zn2+ site makes isoform-selective inhibitor design challenging. This study developed an explainable multi-isoform QSAR workflow for prioritizing selective inhibitors of human CA I, CA II, CA IX, and CA XII, integrating PubChem experimental concordance and applicability-domain (AD) analysis to distinguish model-supported candidates from exploratory extrapolative hypotheses. Methods: Curated ChEMBL Ki datasets were standardized to pKi and modeled in KNIME using 4185 two-dimensional descriptors, Random Forest-based feature selection, and H2O.ai AutoML stacked ensembles. A matched 3200-compound four-isoform matrix supported direct selectivity profiling. Potent ChEMBL inhibitors (Ki < 10 nM) seeded 90% PubChem similarity expansion; analogues were scored across all four isoform-specific models, while PubChem records were filtered to retain only human isoform-specific Ki evidence. AD was calibrated using Morgan fingerprint similarity, descriptor-space distance, and leverage analysis. Candidates were prioritized by predicted potency, selectivity, SAR plausibility, SwissADME profile, structural alerts, and AD membership. Results: Models achieved held-out R2 values of 0.727, 0.719, 0.652, and 0.607 for CA I, CA II, CA IX, and CA XII, respectively. PubChem concordance identified 1098 prioritized compounds with assay records, including 847 with direct Ki evidence and 428 with complete four-isoform coverage; same-target agreement showed Pearson r > 0.86 and linear-fit R2 > 0.74. Test-set error rose from very-high-AD to low/out-of-domain classes. Conclusions: The workflow integrates QSAR prediction, PubChem concordance, explainable SAR, ADME triage, and AD-based reliability assessment. CA II and CA XII candidates showed the strongest support, CA IX showed intermediate support, and CA I candidates require cautious interpretation as low-domain exploratory hypotheses.

Pharmaceuticals doi: 10.3390/ph19081321

Authors: Sandro La Vignera Rosita Condorelli

Background/Objectives: Male hypogonadism affects 35–50% of men with type 2 diabetes mellitus (T2DM) and obesity. The relationship between testosterone deficiency and response to incretin-based therapies (GLP-1 and GIP receptor agonists) remains incompletely understood. This systematic literature review, conducted following PRISMA 2020 guidelines, examines whether male hypogonadism represents a risk factor for poor response to incretin therapy or, conversely, whether these agents offer therapeutic benefits for this population. Methods: A comprehensive systematic literature search was conducted on 31 December 2024, across PubMed/MEDLINE, Scopus, and Web of Science using three search domains: (1) hypogonadism and incretin therapy response; (2) testosterone deficiency and GLP-1/GIP receptor agonists; (3) incretin response and testosterone. Eligibility criteria included human studies (randomized controlled trials [RCTs], observational studies, cohort studies, cross-sectional studies, systematic reviews, and meta-analyses) in adult men reporting testosterone levels and/or incretin response. Animal studies, pediatric populations, case reports with n < 5, editorials, and letters without data were excluded. Study selection followed PRISMA 2020 guidelines with independent dual screening. Risk of bias was assessed using the Cochrane Risk-of-Bias 2.0 tool (ROB2) for RCTs, the Newcastle–Ottawa Scale (NOS) for observational studies, and AMSTAR-2 for systematic reviews and meta-analyses. Due to substantial heterogeneity in study designs, populations, interventions, and outcome measures, a meta-analysis was not feasible; therefore, a narrative synthesis was performed. Results: From 326 records identified, 29 studies were included after deduplication and screening (primary studies: 14; systematic reviews/meta-analyses: five; narrative reviews/expert opinion: 10). Risk-of-bias assessment revealed moderate-to-high overall risk: RCTs had small sample sizes (n = 12–42) and short follow-up (12–24 weeks), raising concerns about statistical power; observational studies were of fair-to-good quality (NOS 4–7 stars) but subject to confounding; and systematic reviews were of low-to-moderate confidence (AMSTAR-2). Primary evidence from RCTs and observational studies demonstrates that GLP-1 receptor agonists (GLP-1RAs)—particularly semaglutide and liraglutide—and the dual GIP/GLP-1 receptor agonist tirzepatide significantly increase total testosterone levels in men with obesity-related functional hypogonadism (mean increase 2.5–5.2 nmol/L). This effect is largely mediated by weight loss and improvement of insulin resistance rather than direct androgenic action. Evidence regarding whether baseline hypogonadism impairs glycemic or weight-loss response to incretin therapy is limited and indirect; no adequately powered comparative trials stratified by baseline testosterone status were identified. Conclusions: Incretin-based therapies, particularly GLP-1 receptor agonists and the dual GIP/GLP-1 receptor agonist tirzepatide, appear to improve testosterone levels in men with obesity-related functional hypogonadism, an effect that is largely mediated by weight loss and improvement of insulin resistance rather than a direct androgenic action. However, direct comparative evidence between hypogonadal and eugonadal men regarding glycemic or weight-loss response to incretin therapy remains insufficient to draw firm conclusions. The hypothesis that male hypogonadism does not impair incretin therapy response is biologically plausible but is currently supported only by indirect evidence. Prospective, adequately powered trials stratified by baseline testosterone status are needed to resolve this question.

Pharmaceuticals doi: 10.3390/ph19081320

Authors: Anh Tuan Do Quoc Long Pham Huong Thi Thu Phung Minh Quan Pham

Background/Objectives: Cyclin-dependent kinase 2 (CDK2) is a key regulator of cell-cycle progression and a potential anticancer target. This study aimed to identify alkaloid-derived CDK2 ligands using an integrated computational workflow and to obtain preliminary evidence of their effects on cancer-cell viability. Methods: Molecular docking with mVina and fast pulling of ligand (FPL) simulations were benchmarked using 20 experimentally characterized CDK2 inhibitors. A library of 2692 PubChem-derived alkaloids was screened, followed by ADMET evaluation, 100 ns molecular dynamics simulations, and FPL-based relative-affinity re-ranking. The three prioritized compounds were evaluated in HepG2 and HGC-27 cells using an MTT assay after 48 h of exposure. Results: Docking and FPL showed correlations with experimental affinity data of RDock = 0.549 ± 0.180 and RW = −0.676 ± 0.119, respectively. CID 636885, CID 46184320, and CID 101691758 were prioritized for detailed evaluation. All three compounds reduced cell viability, with lower IC50 values observed in HepG2 cells than in HGC-27 cells. CID 101691758 exhibited the highest growth-inhibitory activity among the tested compounds, with IC50 values of 15.37 ± 0.46 µg mL−1 in HepG2 cells and 52.64 ± 1.33 µg mL−1 in HGC-27 cells. Conclusions: The workflow identified three preliminary alkaloid hits, with CID 101691758 showing the most favorable combined computational and cell-viability profile. However, the MTT assay does not establish direct CDK2 inhibition or kinase selectivity. Biochemical CDK2 inhibition, target-engagement, and kinase-panel studies are therefore required.

Pharmaceuticals doi: 10.3390/ph19081319

Authors: Ziying Xu Wei Xia Haiqiang Wu Haiping Zhang

Background/Objectives: Blood–brain barrier (BBB) permeability is a major practical obstacle in central nervous system (CNS) drug discovery, because only a small fraction of drug-like molecules achieve sufficient brain exposure. Methods: We present DeepBBB, a graph-based, data-composition-aware screening workflow for predicting BBB permeability and for constructing BBB-focused screening libraries from commercial chemical space. Rather than introducing a new graph-learning architecture, the workflow combines standard graph convolutional and graph-transformer models with deliberate control of training-set composition, commercial-library filtering, chemical-space profiling, and prospective experimental evaluation. Three model variants were trained on the Blood–Brain Barrier Database (B3DB): a baseline classifier/regressor pair (DeepBBB_V1_BC/RG), a variant trained with a more strongly negative-enriched configuration (DeepBBB_V2_BC), and a graph-transformer counterpart (DeepBBB_trans_BC/RG). Because the sample-level split assignments and per-compound predictions from the original runs were not recoverable, the archived summary metrics are reported descriptively in the main text and are not used to support calibration, scaffold-level validity, or generalization. Applying the workflow to the ChemDiv collection (~1.5 million compounds) and the Enamine REAL lead-like space (~1.7 billion compounds) produced three progressively more stringently filtered BBB-focused libraries (21,991; 4,808,885; and 151,790 compounds). Results: Analysis of available processed data indicated that the predicted BBB-permeable set occupies a compact, BBB-compatible property region. Physicochemical, fragment, and scaffold summaries were interpreted descriptively at the constructed-library level. In a first prospective campaign, one of 12 tested candidate compounds was PAMPA-BBB-positive (all-tested molecular-level positive fraction 8.3%; exact 95% CI 0.2–38.5%). In a second campaign, five of 35 tested candidate compounds were PAMPA-BBB-positive (14.3%; exact 95% CI 4.8–30.3%); 13 compounds were not quantifiable and were not treated as ordinary CNS-negative measurements, and the two campaigns differed in compound source, selection strategy, and assay setting, so the numerical difference is reported descriptively rather than causally. Conclusions: Together, these results support the feasibility of BBB-focused computational filtering and a PAMPA-BBB evaluation workflow for CNS-oriented discovery.

Pharmaceuticals doi: 10.3390/ph19081318

Authors: Normand García-Hernández Israel Ramírez-Sánchez Rosa María Ordoñez-Razo

The study of plant-derived secondary metabolites has advanced the field of pharmacochemistry [...]

Pharmaceuticals doi: 10.3390/ph19081316

Authors: Georgios D. Panos

Background: Faricimab 6 mg and aflibercept 8 mg permit extended treatment intervals in neovascular age-related macular degeneration (nAMD), but their pivotal programmes used different loading and maintenance schedules. This study compared treatment burden under the same loading and treat-and-extend protocol and included an exploratory longitudinal analysis of published aggregate visual and anatomical outcomes. Methods: Treatment-specific Dirichlet distributions represented persistent Q8W, Q12W and Q16W capability. Both treatments received injections at weeks 0, 4, 8 and 16, followed by identical four-week extensions from Q8W to Q16W. The primary analysis comprised 30,000 evidence draws, 100,000 paired virtual eyes, a 1.5-million-pair scenario grid and a nested five-million-pair probabilistic analysis. Exploratory multilevel meta-regressions estimated BCVA and change in retinal thickness through week 52; IRF, SRF and complete retinal dryness were analysed separately. Results: Mean injections with faricimab and aflibercept 8 mg were 7.255 and 7.212 at week 52 and 11.522 and 11.144 at week 104. The nested 104-week difference was −0.393 injection (95% uncertainty interval −0.682 to −0.107), but broad source-weight uncertainty included no difference and credible source analyses changed the direction of the contrast. Exploratory week-52 BCVA gains were 5.65 and 6.29 letters, respectively; the difference was 0.64 letter (95% uncertainty interval −1.41 to 2.70) and should be interpreted in the context of three aflibercept 8 mg study families. The expanded retinal-thickness sensitivity model estimated changes of −171.8 and −147.2 micrometres, while the between-treatment direction varied in the three-dose-only analysis. Complete retinal dryness was projected in 57.0% and 69.0% of eyes, respectively, based on three study families per treatment. Conclusions: Use of the same protocol predicted essentially equal one-year burden and a small two-year fixed-weight difference that varied across source choices. The exploratory longitudinal projections, informed by three independent aflibercept 8 mg study families at one year, provide supportive context for the visual and anatomical findings but should be interpreted as hypothesis-generating rather than confirmatory.

Pharmaceuticals doi: 10.3390/ph19081317

Authors: Ji Hye Hwang Chul Jung

Background/Objectives: Skin aging involves interconnected inflammatory, oxidative, hormonal, extracellular matrix (ECM), and cellular senescence-related mechanisms, supporting the need for multitarget approaches. This study aimed to evaluate the effects of a multicomponent natural product-based injectable formulation developed in Korean medicine practice, Dong-An Pharmacopuncture (DAP), on skin aging-related pathways. Methods: A network pharmacology approach was used to identify the active compounds, predicted molecular targets, and signaling pathways associated with DAP. Sixty-two active compounds from 11 constituent materials were screened, and 70 final targets were identified using STITCH-based prediction, intersection with GeneCards-derived skin aging-related targets, and quality filtering. Results: Herb-compound-target network analysis yielded 225 compound–target interactions across 292 edges. Protein-protein interaction analysis identified a highly connected network with 1,334 edges, and hub analysis converged on 10 core targets: TNF, IL6, ESR1, TP53, AKT1, PPARG, EGFR, PTGS2, CASP3, and PPARA. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses identified four major mechanistic axes: inflammatory and oxidative stress regulation, hormonal skin homeostasis, tissue repair and ECM remodeling, and cellular senescence-related regulation. Because DAP is administered by injection, a Good Laboratory Practice-compliant single-dose subcutaneous toxicity study was additionally conducted in Sprague–Dawley rats, which showed no mortality, abnormal clinical signs, or histopathological findings attributable to DAP at 1.0 mL/head. Conclusions: The findings in this study provide a systems-level framework for the predicted multitarget mechanisms of DAP in skin aging-related pathways and support the need for further experimental validation of its predicted mechanisms and repeated-dose safety.

Pharmaceuticals doi: 10.3390/ph19081315

Authors: Zineb Ariba Houda Zentar Mohammed Zerrouk Safaa Hidaoui Mohammed Er-rajy Meryem Hrimla Mustapha Ait El Had Rachid Chahboun Juan Sainz Rachid Ouarsal Mohammed Lachkar Fernando J. Reyes-Zurita

Background/Objectives: Metal-based coordination compounds are attracting attention in medicinal inorganic chemistry because changes in the metal centre can influence their physicochemical properties and biological responses. This study aimed to evaluate two isostructural ammonium-M(II) hydrogenophosphite hydrates, (NH4)2[Mg(H2O)6]3(HPO3)4 (complex 1) and (NH4)2[Co(H2O)6]3(HPO3)4 (complex 2), as bioactive systems with potential anticancer and anti-inflammatory properties. Methods: The complexes were synthesized and characterized by X-ray powder diffraction (XRPD), Fourier-transform infrared spectroscopy (FTIR), thermogravimetric analysis (TGA), density functional theory (DFT) calculations, and Hirshfeld surface analysis. Their cytotoxic activity was evaluated against B16-F10, HT29, HepG2, and HL-60 tumour cell lines. For complex 2, apoptosis, cell-cycle distribution, and mitochondrial membrane potential were analysed by flow cytometry. Nitric oxide production was measured in LPS-stimulated RAW 264.7 macrophages. Results: Complex 2 showed the highest cytotoxic activity, particularly against HL-60 leukaemia cells (IC50 = 36.98 μg/mL), whereas complex 1 displayed weaker activity. Complex 2 increased apoptotic cell populations, altered cell-cycle distribution, and induced mitochondrial membrane depolarization in HL-60 cells. Both complexes reduced nitric oxide production, with complex 2 showing the strongest effect (IC50 NO = 22.46 μg/mL), exceeding that of diclofenac under the experimental conditions, while complex 1 showed comparable activity to the reference drug. DFT descriptors indicated higher electronic reactivity for complex 2. Conclusions: Replacement of Mg(II) by Co(II) enhances the biological activity of this hydrogenophosphite framework. Complex 2 combines cytotoxic activity against HL-60 cells with apoptosis-associated mitochondrial dysfunction and NO-inhibitory activity in activated macrophages, supporting hydrogenophosphite-based metal complexes as bioactive coordination compounds with potential as multifunctional agents.

Pharmaceuticals doi: 10.3390/ph19081314

Authors: Elena Coyago-Cruz Gabriela Méndez Johana Zúñiga-Miranda Alejandro Porras Carlos Barba-Ostria Ivanna Sarabia Linda Guamán Celina Coello Jorge Heredia-Moya Blanca Naranjo María Segovia-Salcedo

This exploratory study compared the flowers, leaves, and roots of Hypochaeris sessiliflora to evaluate physicochemical properties, bioactive compounds, in vitro chemical antioxidant activity, antimicrobial activity, and locomotion-disrupting effects on Caenorhabditis elegans. Bioactive compounds were determined by liquid chromatography; in vitro chemical antioxidant activity by microplate spectrophotometry; antimicrobial activity against a range of ATCC and multidrug-resistant microorganisms; and locomotion-disrupting effects on Caenorhabditis elegans. The results revealed significant differences between plant organs. The leaves contained the highest concentrations of calcium (1163.1 mg/100 g DW), potassium (3256.6 mg/100 g DW), vitamin C (39.2 mg/100 g DW), organic acids (1213.5 mg/100 g DW), and total quantified chlorophylls (89.0 mg/100 g DW). The flowers had the highest levels of total quantified carotenoids (152.1 mg/100 g DM) and total quantified phenolic compounds (12,341.1 mg/100 g DM). They exhibited the highest in vitro chemical antioxidant activity, as measured by the ABTS assay (6.1 mmol TE/100 g DW). The extracts exhibited antimicrobial activity that depended on the plant organ and the microorganism tested, with the root extract being the most active against Staphylococcus epidermidis (MIC = 8.2 mg/mL). No activity was detected against Candida species or against multidrug-resistant bacteria under the experimental conditions used. Furthermore, the extracts under the experimental conditions used did not significantly alter C. elegans’ locomotion. Overall, this study provides baseline evidence of the organ-specific chemical composition and biological properties of H. sessiliflora, supporting further investigation of the compounds responsible for the observed in vitro chemical antioxidant and selective and antimicrobial activities.

Pharmaceuticals doi: 10.3390/ph19081313

Authors: Caterina Nela Dumitru Teodora Marcu Alina Oana Dumitru Simona Steliana Tudor Ionela Daniela Ferțu Alina-Mihaela Elisei Larisa Goroftei

Background: Berberine, an isoquinoline alkaloid present in Berberis spp., Coptis chinensis and Hydrastis canadensis, is among the most widely consumed metabolic-health supplements, popularized as “nature’s Ozempic”. Concurrent, often undisclosed use with prescription drugs is common in older adults, yet berberine is far from inert. Objective: To synthesize the evidence on berberine as a perpetrator of supplement–drug interactions, propose a four-axis mechanistic taxonomy, with product quality treated separately as a modifier of exposure rather than as a mechanism, and derive a clinically actionable risk-stratification framework. Methods: Structured narrative review, prepared per the SANRA quality criteria; PubMed/MEDLINE, Scopus, Web of Science and Embase were searched up to May 2026. Results: Despite very low systemic exposure (oral bioavailability 0.68% in rats; low ng/mL plasma concentrations in humans), high luminal, enterocytic and hepatic concentrations generate interaction liability, documented in humans for a few pairs and mechanistic for most, along four mechanistic axes: inhibition, and transcriptional induction, of CYP3A4, with CYP2D6/CYP2C9 inhibition that is quasi-irreversible through a metabolite-intermediate complex; transporter modulation (P-glycoprotein, OCT1/OCT2, and MATE1); pharmacodynamic additivity (hypoglycemia, hypotension, and QT prolongation); and microbiome- and gut-barrier-mediated effects, the last of these being a candidate axis rather than a demonstrated one. Product-quality variability is treated separately, as a modifier of exposure. The clinical anchor is increased cyclosporine exposure in renal-transplant recipients (AUC +34.5%; trough 29.3% above control). These elements are integrated into a three-tier risk-stratification framework that combines perpetrator potency, victim-drug vulnerability, and patient vulnerability, with each tier being linked to a defined pharmacy action. Conclusions: In patients on multiple medications, and particularly when berberine is co-administered with drugs of narrow therapeutic index, it should be managed as an active pharmacological perpetrator rather than as an inert supplement. Unstandardized product quality and an unsettled European regulatory framework, under which national limits differ by more than an order of magnitude, further widen the uncertainty around the dose actually delivered. Berberine use should therefore be elicited routinely at medication reconciliation and stratified by mechanism, by victim-drug vulnerability, and by patient risk, with particular attention to metabolic self-medication in the GLP-1 era.

Pharmaceuticals doi: 10.3390/ph19081312

Authors: Yukiko Kurihara Hiroki Kurihara

G protein-coupled receptors (GPCRs) are dynamic allosteric proteins whose signaling properties are governed by reciprocal communication between extracellular ligand-binding sites and intracellular transducer interfaces. Although classical pharmacological models established the concept that ligand binding and G protein coupling are thermodynamically linked, recent structural, biophysical, and computational studies have revealed a far more complex picture in which GPCRs exist as ensembles of interconverting conformational states. Accumulating evidence indicates that G proteins function not only as downstream signaling effectors but also as endogenous allosteric modulators. By reshaping receptor conformational landscapes, G protein coupling can influence the structure and dynamics of orthosteric ligand-binding pockets, thereby regulating ligand affinity, binding kinetics, and receptor selectivity. These findings support a bidirectional model of GPCR signaling in which information is transmitted not only from ligand-binding sites to intracellular signaling partners but also in the reverse direction through receptor-wide allosteric networks. Disease-associated mutations of endothelin A receptor (ETAR) provide in vivo evidence that structural perturbations located far from orthosteric ligand-binding sites can alter ligand recognition through long-range allosteric communication. In addition, emerging studies of positive allosteric modulators demonstrate the therapeutic potential of selectively stabilizing ligand–receptor–G protein complexes. These observations suggest that ligand recognition, receptor activation, and transducer coupling should be viewed as integrated properties of a dynamic receptor–ligand–transducer ensemble. This perspective provides a conceptual framework that links classical GPCR pharmacology, structural biology, disease mechanisms, and next-generation drug discovery.

Pharmaceuticals doi: 10.3390/ph19081311

Authors: Kunal Verma Jaskiran Kaur Mohit Kumar Ankit Awasthi Dinesh Kumar Neeraj Choudhary Emad M. Abdallah

Background/Objectives: The aim of the present study was to investigate the mechanisms in Parkinson’s disease (PD), a progressive neurodegenerative disorder characterized by loss of dopaminergic neurons, aggregation of α-synuclein, mitochondrial dysfunction, oxidative stress, neuroinflammation, gut dysbiosis, and blood–brain barrier (BBB) impairment. Although there are several approved therapies that have been developed, their aqueous solubility, oral bioavailability, first-pass metabolism, and inability to penetrate the BBB make them less effective over time. This review is intended to critically analyze the potential of self-nanoemulsifying drug delivery systems (SNEDDSs) as a pathogenesis-related approach to enhance the delivery and therapeutic activity of repurposed drugs and conventional drugs for PD. Methods: A comprehensive literature search was conducted to address the pathogenic mechanisms of PD, the deficiencies of current pharmacotherapy, recent developments in SNEDDS formulation strategies and their application in improving oral bioavailability, lymphatic transport, BBB penetration and targeted brain delivery. A special focus was dedicated to drug repurposing, functionalized SNEDDSs, PEGylation, and gut–brain axis modulation. Results: SNEDDSs significantly enhance the water solubility, stability, intestinal absorption and systemic exposure of poorly water-soluble therapeutic agents and, to a certain extent, lymphatic uptake to avoid first-pass metabolism. These systems include improved brain delivery, decreased pharmacokinetic variability, and prolonged drug levels within the therapeutic range. Moreover, SNEDDSs can be used to deliver multiple molecules that are found to be neuroprotective, antioxidant, anti-inflammatory and probiotic, all at once, which can act on multiple pathogenic mechanisms associated with PD. Functionalized and PEGylated SNEDDSs add further to formulation stability, extend systemic circulation and increase efficiency of brain targeting. Conclusions: SNEDDSs are a promising translational nanomedicine platform for enhancing the effectiveness of conventional and repurposed therapeutics in PD, which address key pharmacokinetic and biological challenges. The next generation of oral therapies with targeted surface engineering, precision drug repurposing and clinical validation will be expected to bring about a faster advancement of drugs that can alter the course of disease rather than giving only symptomatic relief.

Pharmaceuticals doi: 10.3390/ph19081310

Authors: Marina Bilotta Adriana Gargano Roberta Rocca Valentina Maggisano Stefania Bulotta Stefano Alcaro

Background: Tankyrase 1 (TNKS1) is a poly(ADP-ribose) polymerase involved in Wnt/β-catenin signaling, telomere maintenance, and genomic stability, making it an attractive therapeutic target in oncology. This study aimed to develop and apply an integrated computational workflow to identify novel TNKS1 inhibitor candidates. Methods: A curated dataset of experimentally validated TNKS1 inhibitors and property-matched DUD-E decoys was used to develop a consensus supervised machine learning (ML) model prioritization framework for ligand-based virtual screening, integrating Morgan fingerprints with three complementary classifiers. The model screened more than 700,000 compounds, and prioritized hits were evaluated by structure-based virtual screening (SBVS), Prime MM-GBSA binding free-energy refinement, and 500 ns molecular dynamics simulations (MDs). The top candidates were subsequently tested in an in vitro TNKS1 enzymatic inhibition assay. Results: The consensus ML framework prioritized 670 compounds, yielding five candidates for experimental testing. Compound 3 displayed the most favorable computational profile and was experimentally confirmed as a TNKS1 inhibitor candidate, exhibiting approximately 80% TNKS1 inhibition at 0.1 μM, whereas the remaining candidates showed only limited activity. Conclusions: The proposed workflow efficiently reduced a large chemical space to a focused set of TNKS1 inhibitor candidates while substantially reducing the experimental screening burden. Compound 3 represents a promising starting point for future structure–activity relationship studies and lead optimization in the context of TNKS1 inhibition. Moreover, this work highlights the value of integrating consensus ML, SBVS, and experimental validation to accelerate early-stage hit discovery for TNKS1 and other therapeutic targets.

Pharmaceuticals doi: 10.3390/ph19081309

Authors: Muhammad Ishfaq Shahi Jahan Shah Imran Khalid Mashail M. M. Hamid Muhammad Zahir Kota Abdul Ahad Ghaffar Khan Mohammed Ibrahim Samuel Ebele Udeabor Abosofyan Salih Atta Elfadeel Mohamed Salih Chidozie Ifechi Onwuka

Background: Mitogen-activated protein kinase 3 (MAPK3/ERK1) plays a central role in cellular proliferation, inflammation, apoptosis, and survival signalling and has been implicated in oral squamous cell carcinoma (OSCC), periodontitis, oral lichen planus, and other chronic oral inflammatory diseases. The present study employed an integrated computational workflow combining machine learning (ML)-based quantitative structure–activity relationship (QSAR) modelling, molecular docking, density functional theory (DFT), molecular dynamics (MD) simulation, and MM-GBSA analysis to identify and characterise potent MAPK3 inhibitors. Methods: A curated dataset of 907 experimentally validated MAPK3 inhibitors was retrieved from the ChEMBL database and processed using molecular descriptors and Morgan fingerprints. Multiple ML algorithms were evaluated under scaffold-based validation, with Light Gradient Boosting Machine (LightGBM) demonstrating the best predictive performance. Results: The final model achieved strong classification capability with ROC-AUC values of 0.898 and 0.926. Feature importance analysis revealed that local structural motifs captured by fingerprint descriptors played dominant roles in MAPK3 inhibitory activity. The top-ranked compounds were subjected to molecular docking, where compounds 58324148 and 137531515 exhibited strong binding affinities of −11.9 and −11.0 kcal/mol, respectively. DFT calculations demonstrated favourable electronic properties with low HOMO–LUMO energy gaps, while MD simulations confirmed stable receptor–ligand interactions throughout 200 ns trajectories. MM-GBSA analysis further supported strong binding stability dominated by van der Waals interactions. Conclusions: Overall, the integrated computational framework successfully identified promising MAPK3 inhibitor candidates with potential therapeutic relevance for oral inflammatory and proliferative diseases.

Pharmaceuticals doi: 10.3390/ph19081308

Authors: Engi Algharably Ursula Gundert-Remy

Optimizing pharmacotherapy is a central goal of clinical pharmacology [...]

Pharmaceuticals doi: 10.3390/ph19081307

Authors: Maha Abdullah Alwaili Nawal Al-Hoshani Huda A. Alqahtani Rasha Alonaizan Khaled Alzhrani Tariq Aziz

Background/Objectives: Batai orthobunyavirus (BATV) is an emerging mosquito-borne zoonotic pathogen for which no licensed vaccine is currently available. The viral envelope glycoprotein plays an important role in viral attachment and host immune recognition, making it a potential target for rational vaccine design. Methods: In this study, an immunoinformatics-based framework was used to design and evaluate a multi-epitope vaccine candidate targeting the BATV envelope glycoprotein. Selected B-cell, cytotoxic T-lymphocyte (CTL), and helper T-lymphocyte (HTL) epitopes were assembled using appropriate linkers and a human β-defensin adjuvant. Population coverage and in silico immune simulations were conducted to evaluate the potential breadth and magnitude of immune response. Results: The final vaccine construct demonstrated favorable physicochemical characteristics, high predicted antigenicity (0.7959), and non-allergenic properties while maintaining favorable predicted structural characteristics and broad predicted population coverage (99.92%). Structural docking revealed a stable interaction between the vaccine construct and human TLR4, with a weighted docking score of −1194.8, suggesting favorable molecular recognition and receptor engagement. Normal Mode Analysis further supported the structural stability and conformational integrity of the vaccine receptor complex. Immune simulation predicted robust primary and secondary immune responses characterized by elevated IgM and IgG antibody production, sustained memory cell formation, and strong IFN-γ and IL-2 responses, indicating the potential to elicit balanced humoral and cellular immunity. Conclusions: This study presents a structurally optimized and validated multiepitope vaccine candidate against the emerging Batai orthobunyavirus. These computational findings identified a promising vaccine candidate for further investigation; however, its immunogenicity, safety, and protective efficacy before further vaccine development can be considered.

Pharmaceuticals doi: 10.3390/ph19081306

Authors: Md. Amdadul Huq Md. Ashikur Rahman Md. Rasel Rana Jong-Whi Park

The biosynthesis of copper and copper oxide nanoparticles (Cu/CuO-NPs) has attracted considerable interest due to its non-toxic, eco-friendly nature and wide-ranging applications, especially in nanomedicine and biomedical fields. Traditional nanoparticle production methods often involve toxic chemicals and generate harmful byproducts. In contrast, biological synthesis provides a cleaner, safer, more cost-effective, and sustainable alternative. Various biological sources, including plants, bacteria, fungi, and yeast, have been employed for the efficient and non-toxic production of Cu/CuO-NPs. These organisms contain diverse biomolecules such as enzymes, proteins, amino acids, vitamins, flavonoids, and alkaloids that function as reducing, capping, and stabilizing agents during nanoparticle formation. The biologically synthesized Cu/CuO-NPs are characterized using UV-VIS spectroscopy, Raman spectroscopy, TEM, SEM, EDX, XRD, TGA, XPS, FTIR, DLS, zeta potential analyzer, etc. Cu/CuO-NPs hold promise for applications in nanomedicine, primarily because of their strong antimicrobial and anticancer activities and potential use as disinfectants against infectious diseases. Various reports have suggested that the biologically synthesized Cu/CuO-NPs have exhibited significant antimicrobial and anticancer efficacies against pathogenic bacteria, fungi and viruses and various cancer cells. Due to their nanoscale dimensions and extensive surface area, Cu/CuO nanoparticles can readily infiltrate cell walls, disrupt membrane integrity, generate reactive oxygen species, and hinder both DNA replication and protein production, leading to cell death. The present review comprehensively describes the biological synthesis of Cu/CuO-NPs, their characterization techniques, and potential antibacterial, antifungal, antiviral, and anticancer applications. The modes of action for antibacterial, antifungal, antiviral, and anticancer properties have also been explored critically.

Pharmaceuticals doi: 10.3390/ph19081305

Authors: Francisco Arias-Aragón Carmen Mata-Martín Alba Sánchez-Redondo Jesús Novalbos Miguel Ángel Seguido Francisco Abad-Santos Mar Hernaez Mercè Brunet David Hansoe Heredero-Jung Alejandro de la Sota-Pérez María Isidoro-García Almudena Gil-Rodriguez Alba Barral-Raña Olalla Maroñas Gladys Guadalupe Olivera Pasquini Enrique G. Zucchet María José Herrero José Manuel Dodero-Anillo Alicia Alba Máñez María José Pedrosa-Martínez Adrián Llerena on behalf of the BioFRAM PGx Consortium

Background/Objectives: Pharmacogenetics (PGx) may improve drug safety and effectiveness, but its integration into routine care remains heterogeneous across the Spanish National Health System (NHS). This article presents BioFRAM PGx, a multicentre implementation framework that defines the methodological and operational basis for subsequent observational validation in cardiovascular and mental-health care settings. Methods: The framework was developed by healthcare centres from eight Spanish Autonomous Communities through a structured review of PharmGKB/ClinPGx clinical annotations, CPIC and DPWG guidelines, and national regulatory resources. Gene–drug pairs were prioritized according to clinical actionability, therapeutic relevance, applicability to the Spanish population, and analytical feasibility. BioFRAM PGx defines a panel of seven pharmacogenes and 35 drugs, standardized genotyping and phenotype-assignment procedures, pharmacovigilance and clinical variables, healthcare-resource measures, and centralized data management. The proposed non-interventional validation phase includes adults receiving at least one selected drug in hospital or primary-care settings, with a minimum six-month follow-up. PGx results are not returned to clinicians and do not modify treatment. Its primary objective is to assess the predictive value of PGx genotyping for adverse drug reactions (ADRs); secondary objectives include ADR incidence and implementation feasibility. Results: The main outputs are the proposed multicentre observational design, the prioritized gene–drug panel, harmonized analytical and pharmacovigilance workflows, standardized data domains, and a centralized data-management strategy. No patient-level clinical, implementation, or economic outcomes are presented. Conclusions: BioFRAM PGx provides a structured multicentre framework for harmonizing pharmacogenetic procedures across the Spanish NHS. Its clinical effectiveness, influence on prescribing, scalability, and economic outcomes remain to be assessed in subsequent studies.

Pharmaceuticals doi: 10.3390/ph19081304

Authors: Sara Shefa Jagoda Szwach Anna Karwowska Aleksandra Wojno Magdalena Łyko

Acne vulgaris is one of the most prevalent dermatological conditions, affecting up to 85% of individuals aged 11–30 years. Clascoterone cream is the first topical androgen receptor inhibitor approved by the U.S. Food and Drug Administration (FDA) for the treatment of acne vulgaris in patients aged 12 years and older, and the first agent in this class approved for use in both male and female patients. The aim of this narrative review is to summarize the current evidence regarding the efficacy, safety profile, and potential future applications of clascoterone in the treatment of acne vulgaris. A comprehensive literature search was conducted in PubMed/MEDLINE and ClinicalTrials.gov databases up to April 2026. Search terms included “clascoterone,” “cortexolone 17α-propionate,” and “androgen receptor inhibitor acne.” Eligible publications included randomized controlled trials, meta-analyses, open-label extension studies, retrospective analyses, case reports, and relevant review articles published in English. Clinical trial registries and regulatory agency announcements were also searched for ongoing studies and recent approval decisions. Phase 3 trials and meta-analyses demonstrate that twice-daily application of clascoterone 1% cream significantly reduces inflammatory and non-inflammatory lesion counts compared to vehicle, with efficacy sustained up to 12 months of treatment. The safety profile is favorable, with adverse events limited predominantly to mild and transient local skin reactions. Subgroup analyses have shown no significant sex-related differences in efficacy or safety, and the safety profile appears comparable between adolescents and adults. In an exploratory pooled subgroup analysis, lesion-count reductions were greater in adults than in adolescents, whereas the adjusted between-group comparison of IGA success was not statistically significant. Because no formal treatment-by-age interaction test was reported, these findings should be considered hypothesis-generating. Emerging but still incomplete data suggest potential applications in androgenetic alopecia and mild hidradenitis suppurativa. Clascoterone represents a novel addition to the therapeutic armamentarium for acne vulgaris, offering a unique mechanism of action with a favorable safety profile. Further research is warranted to establish its role in combination therapies and in other androgen-dependent dermatoses.

Pharmaceuticals doi: 10.3390/ph19081303

Authors: Nadezhda G. Berdnikova Dmitry V. Tsyganko Vladislav D. Vasiukov Vladimir S. Arnautov Aleksei V. Trofimov Valerii A. Menshov Evgeniy S. Melnikov Evgenia V. Shikh Susanna S. Sologova Elena A. Smolyarchuk Natalia B. Lazareva

Background/Objectives: Vancomycin remains a cornerstone antibiotic for Gram-positive infections, but its narrow therapeutic window and marked pharmacokinetic variability in critical patients complicate dosing. This study aimed to analyze survival and identify factors associated with mortality in intensive care unit (ICU) patients receiving vancomycin under therapeutic drug monitoring (TDM). Methods: A single-center, retrospective cohort study was conducted at City Clinical Hospital named after I.V. Davydovsky (Moscow, Russia) between December 2021 and January 2026 (IRB protocol VANCO-2021). A total of 190 adult patients from surgical, therapeutic, and cardiac ICUs receiving vancomycin for at least 3–5 days were analyzed; trough concentrations were determined by HPLC-MS/MS. Cox proportional hazards, spline Cox, and segmented (piecewise) Cox models were applied to identify mortality predictors, with bootstrap subsampling assessing threshold robustness. Results: Three independent predictors of mortality were identified: ln(minimum vancomycin concentration) (HR = 1.644, p = 0.028), age (HR = 1.019, p = 0.037), and serum creatinine (HR = 1.004, p < 0.001). A data-driven threshold trough concentration (Ctrough) of 25.5 µg/mL was identified, above which mortality increased substantially (63.0% vs. 26.4%; p < 0.05). The adjusted hazard ratio for Ctrough ≥ 25.5 µg/mL was approximately 1.88 (p = 0.054); bootstrap subsampling yielded an uncertainty interval of 23.2–33.6 µg/mL. Conclusions: Elevated minimum vancomycin concentrations exceeding 25.5 µg/mL are independently and statistically associated with increased mortality in ICU patients, likely reflecting altered drug exposure and illness severity rather than a direct causal effect. Systematic TDM is essential for maintaining vancomycin within the therapeutic range.

Pharmaceuticals doi: 10.3390/ph19081302

Authors: Xiaotong Chu Xiao Chen Yanxiang Yuan Yanfei Niu Wentao Zhou Yiming Cui Yongyue Pan Haifeng Liu Zhuoma Dongzhi Shan Huang Bin Li

Objective: Siweixizangmaoru decoction (SXD), a classical Tibetan prescription documented in the medical text Four Medical Tantras, has shown therapeutic activity in experimental rheumatoid arthritis (RA). However, its candidate active constituents and underlying mechanisms remain unclear. Methods: In this study, serum-absorbed constituents following SXD administration were profiled using ultra-high-performance liquid chromatography coupled with quadrupole-Orbitrap high-resolution mass spectrometry (UHPLC-Q-Orbitrap HRMS), and network pharmacology was employed to identify core targets and candidate active constituents of SXD against RA. Candidate active constituents were further screened using cell-based assays, and the selected constituents were quantified using HPLC fingerprint analysis. Both in vivo (CIA rat model) and in vitro (RAW264.7 pyroptosis model) systems were used to investigate the pharmacological effects and mechanisms of SXD. Results: A total of 11 absorbed constituents were identified in the serum of SXD-treated rats, and four preliminary candidate active constituents, including chebulagic acid, kaempferol, gentiopicroside, and berberine, were screened. The combined in vivo and in vitro results showed that SXD attenuated RA progression and reduced inflammatory cytokine levels in rat serum and cell culture supernatants. At the molecular level, SXD increased Pol β expression and reduced DNA damage markers, cytosolic dsDNA accumulation, and cGAS/STING-, NF-κB-, and NLRP3-associated signaling. Pol β knockdown partially reduced the effects of SXD. Conclusions: These findings support the partial involvement of Pol β-associated processes in the anti-arthritic effects of SXD.

Pharmaceuticals doi: 10.3390/ph19081301

Authors: Abdullah Hilmi Marangoz Burcu Delibaş Gamze Altun Arife Ahsen Kaplan Hala Mahgoub Hamour Abit Aktaş Sait Polat Süleyman Kaplan

Background/Objectives: Diabetes mellitus is known to exacerbate neural degeneration and impair regenerative capacity following injury. This study aimed to investigate the effects of Garcinia kola and curcumin on spinal cord morphology and neural structure in a diabetic sciatic nerve injury model. Methods: Spinal cord tissues were obtained from male Wistar albino rats subjected to sciatic nerve transection and diabetes induction in a previously established experimental model. Animals had been assigned to five experimental groups (n = 7 per group): control group, transected sciatic nerve group, transected sciatic nerve + diabetes mellitus (DM), transected sciatic nerve + diabetes mellitus + curcumin (DM + Cur), and transected sciatic nerve + diabetes mellitus + Garcinia kola (DM + GK). Curcumin (300 mg/kg/day) and Garcinia kola (200 mg/kg/day) were administered orally. All animals were sacrificed at three months post-injury. Cervical (C3–C5) spinal cord segments were processed for histological evaluation, and quantitative assessments were performed using unbiased stereological methods to estimate neuronal and structural parameters. Microscopic analyses were conducted to assess morphological alterations. Results: Diabetes combined with sciatic nerve transection induced marked structural alterations in the spinal cord, including reductions in neuronal density and disruption of tissue organization. The DM group showed a significantly lower number of motor neurons than the other groups; similarly, it showed a significant decrease in soma area compared to the Cont and Sham groups. Conclusions: Both the curcumin and Garcinia kola treatment groups exhibited partial preservation of spinal cord morphology. This study provides evidence that Garcinia kola and curcumin exert protective effects not only at the peripheral nerve level, but also in the spinal cord following diabetic nerve injury.

Pharmaceuticals doi: 10.3390/ph19081300

Authors: Ardak Jumagaziyeva Seitzhan Turganbay Anar Seisembekova Daniil Shepilov Zhanar Iskakbayeva Sabina Kenesheva Saltanat Jumabayeva Gaukhar Askhatkyzy Nurdaulet Temir Abdurashit Khamidulin Alexandr Ilin

Background: Antimicrobial resistance (AMR) is among the most pressing challenges in modern infectious medicine, driving progressive failure of standard antibacterial therapy and rising mortality from infections caused by multidrug-resistant (MDR) pathogens. Antibiotic potentiation through adjuvant compounds capable of restoring the activity of existing drugs represents a promising strategy to overcome resistance without developing fundamentally new antibacterial molecules. This study aimed to evaluate the antibiotic-potentiating activity of a dextrin/polyvinyl alcohol/iodine complex (D/PVA/I-1), developed at the Scientific Center for Anti-Infectious Drugs JSC (Almaty, Kazakhstan), against clinically relevant MDR reference strains. Methods: Nine reference strains, Staphylococcus aureus (ATCC 33591, BAA-39), Escherichia coli (ATCC BAA-196, BAA-2523), Klebsiella pneumoniae (ATCC BAA-2524, 700603), Acinetobacter baumannii (ATCC BAA-1790), Streptococcus pneumoniae (ATCC BAA-660), and Haemophilus influenzae (ATCC 33930), and two clinical isolates, P. aeruginosa SCAID PHRX1-2019 and E. coli SCAID WND1-2021, were tested. Antibiotic-potentiating activity was assessed by checkerboard assay with calculation of the fractional inhibitory concentration index (FICI); bactericidal kinetics were evaluated by time-kill analysis. The effect of D/PVA/I-1 on cytoplasmic membrane permeability was investigated using a crystal violet uptake assay. Results: Of 45 D/PVA/I-1–antibiotic combinations tested across nine antibiotics, synergy (FICI ≤ 0.5) was demonstrated in 44.4% of cases, partial synergy in 48.9%, and additive effects in 6.7%; no antagonistic interactions were detected. The most pronounced potentiating effect occurred against Gram-positive pathogens, particularly MRSA strains (66.7% synergistic combinations). Time-kill analysis confirmed suppression of the regrowth phenotype characteristic of MDR strains under monotherapy and restoration of bactericidal activity against antibiotics to which strains exhibited intrinsic resistance. D/PVA/I-1 induced a dose- and time-dependent increase in membrane permeability in both Gram-positive and Gram-negative organisms. Conclusions: D/PVA/I-1 is an effective broad-spectrum antibiotic potentiator and represents a promising basis for combination therapy regimens against MDR infections.

Pharmaceuticals doi: 10.3390/ph19081299

Authors: Carolina Holguín-Meraz Rita Elizabeth Martínez-Martínez Erasto Armando Zaragoza-Contreras Rubén Abraham Domínguez-Pérez Karla Lizette Tovar-Carrillo Alejandro Donohue-Cornejo Juan Carlos Cuevas-González Simón Yobanny Reyes-López Erika de Lourdes Silva-Benítez María Verónica Cuevas-González León Francisco Espinosa-Cristóbal

Background: Motor and intellectual disabilities (MIDs) belong to a restricted and vulnerable social group with health restrictions for maintaining and preserving adequate oral health. Silver nanoparticles (AgNPs) have appeared as an encouraging therapy due to their excellent antimicrobial effects; however, scientific information on the antimicrobial effectiveness of AgNPs against bacteria in patients with MIDs remains limited. This study aimed to evaluate the antimicrobial substantivity of AgNPs in oral bacteria from patients with and without MIDs. Methods: Fifty-four oral biofilm samples were obtained from 26 participants with MIDs and 28 participants without MIDs. The microbial inhibition activity and the antimicrobial substantivity of AgNPs were determined. The antibiofilm activity of the AgNPs was explored using scanning electron microscopy (SEM). Results: The AgNPs displayed uniform size distributions (9.8 ± 1.7 nm) and electrical charges in surface particles that limit particle clustering (−39.8 ± 3.3 mV). The AgNPs were more effective than chlorhexidine (CHX) in eliminating bacteria from oral biofilms derived from both MID and non-MID patients (p < 0.05); therefore, the antimicrobial substantivity of the AgNPs was statistically better than deionized water but lower than CHX at specific exposure times (p < 0.05). The antimicrobial effectiveness of AgNPs at even low concentrations was associated with the type of oral biofilm, the type of antimicrobial solution, and, in some cases, gender. SEM images indicate a clear alteration in oral biofilm structure when AgNPs were applied. Conclusions: AgNPs exhibited significant potential to limit oral bacterial proliferation from microbial biofilms while contributing to the maintenance of oral condition in individuals with MIDs.

Pharmaceuticals doi: 10.3390/ph19081298

Authors: Michał Janiak Katarzyna Mądra-Gackowska Lidia Wydeheft Marcin Gackowski

Background/Objectives: Oral small-molecule inhibitors of activated factor XI (FXIa) are intended to attenuate thrombosis with less disruption of hemostasis than established anticoagulants, but clinical outcomes have not consistently paralleled pharmacodynamic target engagement. This systematic review integrated molecular design, preclinical pharmacology, human pharmacokinetics/pharmacodynamics (PK/PD), and clinical outcomes to identify where translation is maintained or lost. Methods: The review followed PRISMA 2020 and PRISMA-S and was registered in PROSPERO (CRD420261356169). PubMed/MEDLINE, Scopus, and Web of Science Core Collection were searched from inception through 30 March 2026 and rerun on 11 August 2026, with a final publication-eligibility cut-off of 30 June 2026. Owing to heterogeneity, evidence was synthesized narratively. Results: Sixty-eight eligible reports were included, comprising seven randomized parent outcome trials and nine reports with in vivo thrombosis or bleeding experiments. In the OCEANIC-STROKE trial, ischemic stroke occurred in 6.2% with asundexian versus 8.4% with placebo (HR 0.74, 95% CI 0.65–0.84), while major bleeding was 1.9% versus 1.7% (HR 1.10, 95% CI 0.85–1.44). In OCEANIC-AF, stroke or systemic embolism occurred in 1.3% with asundexian versus 0.4% with apixaban (HR 3.79, 95% CI 2.46–5.83), despite less major bleeding (0.2% vs. 0.7%; HR 0.32, 95% CI 0.18–0.55). Conclusions: Oral small-molecule FXIa inhibition is pharmacologically coherent, but clinical benefit is indication-, comparator-, and background-therapy-dependent. PD markers confirm target engagement but are not validated efficacy surrogates; hard clinical outcomes remain decisive.

Pharmaceuticals doi: 10.3390/ph19081297

Authors: Zahira Tber Mohammed Loubidi Jabrane Jouha Ismail Hdoufane Mümin Alper Erdogan Luciano Saso Güliz Armagan Sabine Berteina-Raboin

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Pharmaceuticals doi: 10.3390/ph19081296

Authors: Lucyna Tomaszek Wioletta Mędrzycka-Dąbrowska Sandra Lange Sebastian Dąbrowski Mateusz Szczupak

Background: Intravenous magnesium sulfate has been proposed as an adjuvant analgesic in perioperative care; however, its effects on postoperative pain and related analgesic outcomes remain uncertain. Methods: A systematic search of the literature was conducted from 1 January 2015 through 19 January 2026 to identify randomized controlled trials comparing intravenous magnesium sulfate with placebo in adults undergoing general anesthesia. Pain intensity was harmonized to a 0–10 scale. Primary outcomes were early postoperative pain (0–6 h), pain at 12 h, and pain at 24 h. Secondary outcomes were postoperative opioid consumption, expressed as intravenous morphine equivalents, and time to first rescue analgesia. Random-effects meta-analyses were performed. Risk of bias was assessed using RoB 2, and certainty of evidence using GRADE. Results: A total of 24 randomized controlled trials were included in the qualitative synthesis, of which 22 provided data for quantitative meta-analysis, yielding 23 comparisons and a total of 1750 participants. Intravenous magnesium reduced early postoperative pain (mean difference [MD] −0.90, 95% confidence interval [CI] −1.33 to −0.47; I2 = 96.27%) and pain at 24 h (MD −0.59, 95% CI −1.00 to −0.18; I2 = 66.36%), but not pain at 12 h (MD −0.52, 95% CI −1.15 to 0.11; I2 = 77.38%). Magnesium also reduced postoperative opioid consumption (MD −6.28 mg morphine equivalents, 95% CI −8.94 to −3.61; I2 = 95.93%) and prolonged time to first rescue analgesia (MD 85.61 min, 95% CI 40.11 to 131.10; I2 = 79.37%). Certainty of evidence ranged from very low to moderate, with the highest certainty observed for pain at 24 h. Conclusions: Intravenous magnesium was associated with reduced postoperative pain, particularly at 24 h, and with opioid-sparing effects. However, substantial heterogeneity across outcomes and generally low-to-moderate certainty of evidence warrant cautious interpretation. Magnesium may be considered as part of multimodal analgesia in appropriately selected patients.

Pharmaceuticals doi: 10.3390/ph19081295

Authors: Yuma Iwashita Yoshihiro Uesawa

Background/Objectives: Transthyretin amyloidosis (ATTR) is a progressive disease caused by the dissociation of the transthyretin (TTR) tetramer, leading to amyloid fibril formation. Although pharmacological stabilizers have been developed, preventive strategies for wild-type ATTR (ATTRwt) have not been established. This study developed a computational model to predict TTR-binding activity from chemical structures and applied it to the exploratory prioritization of food-derived compounds. Methods: A machine learning model was constructed using TTR–8-anilino-1-naphthalenesulfonic acid displacement assay data from the Tox24 Challenge. An integrated dataset compiled from multiple literature sources was used to compare predicted TTR-binding activity with amyloid fibril formation inhibition, and the trained model was applied to the PhytoHub database. Results: The model achieved a root mean square error of 21.34 and a mean absolute error of 15.9 percentage points on an external test dataset, a performance comparable to that of the top-scoring models in the Tox24 Challenge. The predicted TTR-binding activity showed positive correlations with inhibition of amyloid fibril formation, Pearson’s r = 0.673 and Spearman’s ρ = 0.602. The model prioritized 39 compounds with high predicted binding activity, predominantly polyphenols. Conclusions: These results suggest that the proposed in silico method may be useful for the exploratory prioritization of food-derived compounds with potential relevance to TTR stabilization.

Pharmaceuticals doi: 10.3390/ph19081294

Authors: Renata Solis-Peña Vanessa Lizbeth Correa-Macias Hania Lizeth Peralez-Olguin Dalia Lizbeth Martínez-Medellín Andrea Melissa González-Rodríguez Lizeth Guadalupe Campos-Muzquiz Lissethe Palomo-Ligas Sendar Daniel Nery-Flores

The genus Dalea (Fabaceae) comprises approximately 188 recognized species distributed mainly in arid and semi-arid regions of the American continent, where several species have been traditionally used in folk medicine for the treatment of gastrointestinal disorders, infections, inflammation, and other ailments. In recent years, Dalea species have attracted considerable scientific interest due to their remarkable diversity of secondary metabolites, particularly prenylated flavonoids, isoflavonoids, rotenoids, chalcones, stilbenes, and terpenoid-rich essential oils. Among these compounds, prenylated and geranylated flavonoids represent some of the most characteristic metabolites of the genus and are strongly associated with its reported biological activities. This review summarizes current knowledge regarding the taxonomy, distribution, traditional uses, phytochemical profile, pharmacological activities, and toxicological aspects of the genus Dalea. Although only about 19 species (approximately 10% of currently recognized species) have been subjected to phytochemical investigation, substantial knowledge gaps remain within the genus. Special emphasis is placed on prenylated phenolic compounds because of their structural diversity and broad spectrum of biological activities, including antibacterial, antifungal, antiparasitic, anti-inflammatory, neuroprotective, anticancer, anti-tyrosinase, and xanthine-oxidase-inhibitory effects. Despite promising pharmacological findings, most available evidence remains limited to in vitro evaluations, with relatively few studies including in vivo validation or mechanistic investigations. Furthermore, important challenges remain regarding toxicity, pharmacokinetics, metabolomic characterization, and sustainable production of bioactive metabolites. Overall, the available evidence highlights Dalea as a valuable source of bioactive compounds with potential pharmaceutical applications and supports the need for further interdisciplinary research focused on drug discovery and development.

Pharmaceuticals doi: 10.3390/ph19081293

Authors: Lidia-Ioana Virchea Maria Totan Cecilia Georgescu Adina Frum Endre Máthé Monica Mironescu Bence Pecsenye Robert Nagy Oana Viorica Danci Maria-Lucia Mureșan Nastaca Alina Palade Felicia-Gabriela Gligor

Background/Objectives: Essential oils (EOs) are complex mixtures extracted from plants. The aim of this study was to evaluate the chemical composition, antioxidant and antimicrobial properties, as well as in vivo toxicity against Drosophila melanogaster of the EOs extracted from Achillea millefolium L. (AM), Mentha longifolia L. (ML), and Thymus serpyllum L. (TS) from the mountains in Sibiu County, Romania. Methods: EOs were extracted by hydrodistillation. Their phytocompounds were determined by Gas Chromatography–Mass Spectrometry. Antioxidant potential was evaluated by 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2′-azinobis [3-ethylbenzothiazoline-6-sulfonic acid]-diammonium salt (ABTS), and ferric-reducing antioxidant power (FRAP) assays. Antibacterial action was measured against eight pathogenic bacteria. An in vivo D. melanogaster diet-dependent viability assay was conducted to assess the toxicity of the EOs. Results: Chamazulene (49.72%), pulegone (36.20%), and thymol (42.11%) were the most abundant constituents of the AM, ML, and TS EOs, respectively. All tested EOs exerted antioxidant activity, with TS EO being the most efficient in the DPPH (98.19% inhibition) and ABTS (1.82 mmol Trolox equivalent (TE)/mL EO) scavenging assays, while AM EO performed best in the FRAP assay (66.70 μmol TE/mL EO). Against TS EO, Bacillus subtilis was the most sensitive, with an inhibition zone of 37 mm. A dose-dependent decrease in the viability of D. melanogaster L. was observed when their diet was supplemented with AM, ML, and TS EOs. Conclusions: The analyzed EOs demonstrated antioxidant and antimicrobial effects, suggesting their potential applications in various fields. However, due to their toxicity, an in-depth analysis is required before introducing such products in therapy.

Pharmaceuticals doi: 10.3390/ph19081292

Authors: Emily-Alice Russu Radu Ilinca Liliana Gabriela Popa Anca Silvia Dumitriu Stana Păunică Călin Giurcăneanu Cristina-Crenguța Albu

Background/Objectives: Rituximab has improved outcomes in pemphigus vulgaris (PV), but treatment response remains heterogeneous and no clinically validated biomarker currently supports reliable individual-level prediction. Oral lesions frequently represent the earliest manifestation of PV and contribute substantially to disease burden and longitudinal assessment. This perspective aimed to synthesize candidate biomarkers and organize them within a conceptual, task-specific artificial intelligence (AI) framework for future rituximab outcome modeling. Methods: A structured literature search and narrative synthesis evaluated genetic and pharmacogenomic, immunological and serological, cellular, clinical, and oral phenotypic variables potentially relevant to rituximab response. Variables were considered according to biological rationale, available evidence, temporal availability, and potential use in pretreatment prognosis or post-treatment response monitoring. No patient-level dataset was analyzed, and no model was trained or validated. Results: Candidate pharmacogenomic variables include FCGR3A rs396991, FCGR2A rs1801274, and IL6 rs1800795, although evidence derives mainly from autoimmune diseases other than PV. IL10, TNF, TNFRSF13B, and population-specific HLA variables remain exploratory. Anti-desmoglein autoantibodies, BAFF, cytokine profiles, CD19+ B-cell depletion, CD27+ memory B-cell repopulation, regulatory T-cell dynamics, natural killer cell phenotypes, and clinical and oral phenotypic variables may provide complementary information, but none currently demonstrates sufficient independent predictive validity. The proposed RTX-AI Response Framework distinguishes the Pretreatment Prediction Component from the Post-Treatment Response Monitoring Component and assigns no numerical weights, probabilities, thresholds, or treatment recommendations. Conclusions: The framework should be interpreted exclusively as a conceptual research architecture. Prospective multicenter data collection, standardized biomarker assessment, task-specific model development, calibration, external validation, and evaluation of clinical utility are required before potential clinical implementation.

Pharmaceuticals doi: 10.3390/ph19081291

Authors: Melek Karaaslan H. Kamuran İleri Özler Burçin Ergene Sevde Nur Biltekin Kaleli Neslihan Üstündağ Okur Simge Aykan H. Gülçin Saltan İşcan

Background/Objectives: Despite the widespread use of essential oils in aromatherapy, objective evidence supporting their neurophysiological effects remains limited. This study evaluated the effects of selected pharmacopoeial-quality essential oils (lavender, rosemary, lemon, and peppermint as well as a blend of these oils) on electroencephalographic (EEG) activity following inhalation and developed and characterized microemulsion formulations guided by exploratory EEG response patterns, with preliminary in vitro cytotoxicity assessment. Methods: EEG recordings were obtained from 28 healthy volunteers following inhalation of the essential oils. Neurophysiological responses were evaluated using exploratory paired comparisons between each oil and its corresponding grape-seed oil control, together with repeated-measures ANOVA of normalized EEG responses. Selected oils were formulated as microemulsions (M1-M3), which were characterized by their physicochemical properties. Cytotoxicity was assessed in HaCaT human keratinocytes using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Results: Exploratory paired comparisons suggested localized EEG changes, particularly in the alpha-1 frequency band. However, these effects did not remain statistically significant after correction for multiple comparisons, and repeated-measures ANOVA of normalized EEG responses did not demonstrate significant overall differences among oil conditions, although the alpha-1 band exhibited the most distinct response pattern. Microemulsions exhibited nanoscale droplet sizes and favorable physicochemical stability. The IC50 values of M1 and M2 were greater than 2000 µg/mL, whereas M3 had an IC50 of approximately 1220 µg/mL. Conclusions: Pharmacopoeial-quality essential oils produced exploratory, region-specific EEG response patterns, with the alpha-1 band showing the most consistent response profile. The developed microemulsion formulations demonstrated suitable physicochemical stability and generally low cytotoxicity, supporting their further evaluation as aromatherapy product prototypes.

Pharmaceuticals doi: 10.3390/ph19081290

Authors: Surabhi Gautam Swarandeep Singh Vidhi Thakkar Devyani Joshi Sanjeev Kumar

Therapeutic antibodies have transformed the management of infectious diseases, autoimmune disorders, and cancer, yet most approved agents continue to rely on a limited range of IgG-based Fc scaffolds. Consequently, the broader therapeutic potential of antibody isotypes, subclasses, allotypes, and host Fc receptor (FcR) genetics remains incompletely integrated into antibody design and clinical development. Emerging evidence indicates that these determinants profoundly influence effector function, pharmacokinetics, tissue distribution, immunogenicity, and therapeutic efficacy. This review provides a comprehensive, translational perspective on how antibody isotype selection, IgG subclass biology, allotypic variation, Fcγ receptor (FcγR) and neonatal Fc receptor (FcRn) genetics, and Fc engineering collectively shape therapeutic outcomes. Distinct from previous reviews that examine these factors individually, we integrate their roles across infectious diseases, autoimmunity, and cancer to identify common principles governing Fc-dependent immunity and therapeutic response. We further describe the clinical implications of Fc-mediated mechanisms, Fc glycoengineering, FcRn-targeted therapies, systems serology, and emerging non-IgG antibody platforms, highlighting both established applications and unresolved challenges. By unifying structural immunology, Fc pharmacology, host genetics, and disease biology, this review advances the concept of precision Fc pharmacology, in which antibody scaffold selection and Fc optimization are tailored to specific disease contexts and patient immune profiles. This framework provides a roadmap for the rational development of next-generation antibody therapeutics with improved efficacy, safety, and clinical precision.

Pharmaceuticals doi: 10.3390/ph19081289

Authors: Shun Lv Jian Hu Huan Chen Minyu Zhu Wei Jin Qiyin Liu Qianqian Zhang Yinghua Zhang Ying Li Zhengqi Dong

Background: Diabetic foot ulcers (DFUs) are chronic wounds characterized by persistent inflammation, oxidative stress, impaired angiogenesis, and defective extracellular matrix remodeling. Current therapeutic approaches remain insufficient for refractory diabetic wounds due to limited drug retention and inadequate regulation of the wound microenvironment. This study aimed to develop a centipede-derived protein fraction-loaded nanocapsule hybrid hydrogel for sustained bioactive delivery and diabetic wound repair. Methods: A bioactive protein fraction was isolated from processed medicinal centipede material and screened using cellular compatibility assays. The selected tropomyosin-containing protein fraction was incorporated into nanocapsules and subsequently integrated into a gallic acid-modified polyacrylamide hydrogel matrix. The physicochemical properties, antioxidant activity, rheological behavior, and protein release characteristics of the nanocapsule-hydrogel system were systematically evaluated. A streptozotocin-induced diabetic rat wound model was established to assess therapeutic efficacy through wound closure analysis, histological staining, and immunohistochemical evaluation. Results: The prepared nanocapsules exhibited a spherical morphology, nanoscale size distribution, and sustained protein delivery capability. The PAM-GA hydrogel demonstrated antioxidant activity, injectability, shear-thinning behavior, self-healing ability, and enhanced retention of protein release. In vivo experiments showed that the PT@NC@PAM-GA hydrogel significantly accelerated wound closure and promoted tissue regeneration, accompanied by enhanced angiogenesis, collagen deposition, and reduced inflammatory responses. Conclusions: The centipede-derived protein fraction-loaded nanocapsule hybrid hydrogel effectively integrates bioactive protein delivery with a multifunctional hydrogel matrix, providing a potential strategy for sustained treatment of diabetic foot ulcers.

Pharmaceuticals doi: 10.3390/ph19081288

Authors: Shuyi Xu Hai Hu Yifan Li Jiawei Zhang Lei Wang Pameila Paerhati Wenxin Bao Yanlin Bian Jianwei Zhu Mingyuan Wu

Background: The efficacy of chemotherapy in triple-negative breast cancer (TNBC) is limited by intrinsic resistance and the tumor microenvironment (TME). Accumulating evidence reveals a mechanistic connection between programmed death-ligand 1 (PD-L1) and c-x-c motif chemokine receptor 4 (CXCR4), which dominate stroma barriers, immune escape, and cancer metastasis. Earlier studies have shown that dual suppression of c-x-c motif ligand 12 (CXCL12)/CXCR4 and programmed cell death-1 (PD-1)/PD-L1 pathways regulates extracellular matrix (ECM) deposition, activation of cancer-associated fibroblasts (CAFs), and epithelial–mesenchymal transition (EMT) of pancreatic cancer cells. Methods: We combined BsNb PX4, a bispecific nanobody targeting PD-L1 and CXCR4, with paclitaxel or gemcitabine in multiple tumor cell lines and human peripheral blood mononuclear cell (hPBMC)-reconstituted xenograft mouse models. Antitumor activity was assessed by CCK-8, flow cytometry, and ELISA, and immune cell infiltration and TME remodeling were examined by immunofluorescence, immunohistochemistry, cytokine assays, and RNA-seq. Results: In MDA-MB-231 cells, BsNb PX4 synergistically enhanced paclitaxel-induced growth inhibition and apoptosis via G2/M cycle arrest. This combinatorial strategy profoundly remodeled tumor immunity by expanding CD8+ T cells and depleting Foxp3+ CD4+ regulatory T cells (Tregs), while concurrently restoring T-cell cytotoxicity and skewing the cytokine balance toward an antitumor state, with elevated IFN-γ and reduced TGF-β1. Notably, compared with paclitaxel monotherapy, the combination significantly elevated intratumoral CD8+ T-cell infiltration, decreased Treg abundance, and exerted robust inhibitory effects on tumor growth and metastasis in humanized TNBC xenografts. Conclusions: These findings reveal that dual blockade of PD-L1 and CXCR4 acts synergistically with chemotherapy by triggering tumor cell apoptotic effects and reversing the immunosuppressive microenvironment, thereby emerging as a promising therapeutic strategy for TNBC.

Pharmaceuticals doi: 10.3390/ph19081287

Authors: Maria Luiza Lima Nele Wild Eva Liebau Carsten Wrenger André Barateiro

Malaria remains a major global health problem, especially in low- and mid-income countries where Plasmodium parasites are commonly endemic. Despite the significant number of available drugs to treat the disease, some aspects, such as increasing resistance to frontline antimalarials, inefficient treatment of asymptomatic carriers and treatment inadequacy for certain vulnerable groups, are jeopardizing control measures, justifying the need for therapeutic targets driving the discovery of new compounds. In this context, glutathione transferases, which are essential for redox metabolism and detoxification of heme in the asexual blood stages of Plasmodium parasites, have emerged as promising targets more than two decades ago, but have been deprioritized due to significant gaps in research. This review aims to synthesize current biochemical, structural, genetic, and transcriptomic evidence on Plasmodium glutathione S-transferases and evaluate their role in parasite physiology across the life cycle while revising their potential for drug targeting using suggested guidelines for target candidate profiling and prioritization.

Pharmaceuticals doi: 10.3390/ph19081286

Authors: Shuang Pan Yumo Li Zhi Lin Zhe Lin He Lin Ying Lv

Hyperuricemia (HUA) is a key pathological basis for gout and uric acid (UA) nephropathy, and its increasing global prevalence presents a substantial public health challenge. Natural products (NPs) have emerged as a research hotspot due to their potential to reduce UA and favourable safety profile; however, comprehensive bibliometric analyses in this area remain limited. In this study, the English literature on NPs with anti-HUA activity published between 2004 and 2026 was retrieved from the Web of Science, Scopus, and PubMed, and analyzed using Bibliometrix (R package), VOSviewer, and CiteSpace. A total of 581 articles were identified, indicating a rapid increase in publication output across 53 countries, 870 institutions, 234 journals, and 3421 authors. China was found to lead the field, with Guangzhou University of Traditional Chinese Medicine contributing the highest number of publications. The Journal of Ethnopharmacology ranked first in publication volume, and Kong Ling-Dong emerged as the most prolific author. Major research foci included xanthine oxidase (XOD), UA metabolism, oxidative stress, inflammatory regulation, gut microbiota, and molecular docking. Perilla frutescens leaves and Terminalia chebula, together with six principal categories of bioactive compounds, demonstrated promising therapeutic potential. Overall, this study elucidates the developmental trends and research frontiers in the field, providing a valuable reference for future investigations.

Pharmaceuticals doi: 10.3390/ph19081285

Authors: Vicenta Albarral Ávila Anna Nardi-Ricart Aitor Caballero-Román Lara Martínez Pettina David Miñana-Galbis Montserrat Miñarro Carmona

Background/Objectives: Antimicrobial resistance is a critical threat to global public health. Eugenol is a bioactive compound with broad-spectrum antimicrobial activity that has attracted increasing interest as a naturally derived antimicrobial agent with potential complementary applications to conventional antibiotics, but its clinical application is severely limited by its high volatility, low water solubility and thermo-oxidative instability. The main objective of this study was to develop eugenol-loaded microparticles using a ternary biopolymer matrix, to characterise their main physicochemical properties, and to evaluate their in vitro antimicrobial efficacy against clinically relevant bacterial reference strains. Methods: The microparticles were formulated from an emulsion of maltodextrin, gum arabic and soy lecithin, and encapsulated using a spray-drying technique. Product recovery, particle morphology assessed by scanning electron microscopy (SEM), particle size distribution determined by laser diffraction, and encapsulation efficiency quantified by GC-FID were analysed. Subsequently, antimicrobial activity was evaluated by comparing the microparticles with free eugenol using agar well diffusion and broth microdilution assays to determine the minimum inhibitory concentration (MIC) against eight bacterial strains. Results: The spray-drying process achieved a product recovery of 61.88% and an encapsulation efficiency of 52.45%. The resulting microparticles exhibited a smooth, spherical morphology with diameters of less than 20 µm. In microbiological assays, microencapsulation significantly reduced MIC values by 4- to 16-fold compared with free eugenol for susceptible strains. The formulation exhibited potent activity against most of the Gram-positive and Gram-negative pathogens tested, except for Pseudomonas aeruginosa, which remained resistant to both formulations. Conclusions: The encapsulation of eugenol in this optimised biopolymer matrix substantially improved its antimicrobial efficacy against the tested bacterial strains. These findings highlight the potential of spray-dried eugenol microparticles as a promising antimicrobial formulation and provide a basis for their further development for topical applications. Further studies are warranted to evaluate their pharmaceutical performance and antimicrobial mechanisms.

Pharmaceuticals doi: 10.3390/ph19081284

Authors: Zheng Chao Yuchen Mei Hao Peng Guowen Li Xiaodong Hao Yuchuan Liu Guanglu Lyu Zhihua Wang Le Li

Background/Objectives: Immune checkpoint blockade targeting the PD-1/PD-L1 axis is a standard treatment for urothelial carcinoma (UC), but 20–30% of patients develop resistance. The underlying mechanisms, particularly regarding tumor microenvironment metabolic reprogramming, remain unclear. Methods: We performed single-cell RNA sequencing on pre-treatment tumors from eight pembrolizumab-treated UC patients (three responders, five non-responders). Bioinformatic analyses included cell clustering, pathway enrichment, and cell–cell communication, validated using public datasets. Functional experiments involved SGPL1 knockdown in MB49 cells and a syngeneic PD-1-resistant murine model (MB49-R5). Results: Responders exhibited a higher proportion of infiltrating T-cells. Unexpectedly, CD4+ subsets, rather than CD8+ cells, were numerically enriched in responders. However, all T-cell subsets in responders demonstrated enhanced oxidative phosphorylation-dominant metabolic activity. Enhanced macrophage–T-cell crosstalk was observed, with macrophages exhibiting M1-like polarization driven by CD4+ T-cell-derived IFN-γ and CD40L signaling. In contrast, non-responders displayed dominant tumor-derived MIF–CD74 signaling and M2 polarization. Integration of public datasets identified SGPL1 as a key metabolic regulator associated with poor prognosis and reduced immune activation. Functional experiments demonstrated that SGPL1 knockdown inhibited tumor proliferation and restored anti-PD-1 sensitivity, accompanied by increased T-cell infiltration. Conclusions: T-cell metabolic activation drives anti-PD-1 responses in UC, and CD4+ T-cell-mediated M1 polarization and tumor-intrinsic SGPL1 crucially shape therapeutic outcomes, highlighting SGPL1 as a candidate metabolic target to overcome PD-1 resistance.

Pharmaceuticals doi: 10.3390/ph19081283

Authors: Mingli Shen Qingping Shi Shuang Gao Beiyan Chen Jieru Han

Background/Objective: Cinnamaldehyde (CA) is a naturally occurring bioactive compound derived from the leaves, bark, roots, and flowers of the Chinese medicinal plant Cinnamomum cassia. It exhibits a broad spectrum of pharmacological properties, encompassing antioxidant, antibacterial, anti-diabetic, antifungal, and anticancer activities. Notably, it has shown potential therapeutic benefits in the management of type 2 diabetes mellitus (T2DM) and non-alcoholic fatty liver disease (NAFLD). Ochratoxin A (OTA), a common contaminant found in foods such as cereals, coffee, and raisins, is also present in traditional Chinese medicinal materials, including Astragalus and liquorice. T2DM and NAFLD share intertwined pathophysiological pathways, including insulin resistance, dyslipidaemia, chronic low-grade inflammation and oxidative stress, with insulin resistance serving as the common pathological hub for both conditions. Consequently, they frequently co-occur and exacerbate each other. OTA exerts dual-targeted toxicity to the pancreas and liver, which may synergistically drive the development of the comorbidity of T2DM and NAFLD. These two processes are mutually causal and together constitute the pathological basis of metabolic comorbidity. Methods: Network toxicology employs toxicological data, gene expression, and protein–protein interaction (PPI) networks to predict the targets of toxins, while network pharmacology, based on systems biology principles, reveals how drugs exert regulatory effects through multiple targets and pathways. In this study, we employed an integrated network toxicology and network pharmacology approach to jointly decipher the potential mechanisms by which CA intervenes in OTA-induced comorbid T2DM-NAFLD. First, a network toxicology approach was employed to preliminarily screen for core toxicological targets responsible for OTA’s pathogenicity. Subsequently, network pharmacology was used to identify potential targets of CA-mediated intervention in the disease. Finally, the common overlap among the CA intervention targets, OTA toxicity targets, and disease targets was defined as the final set of potential targets for CA-mediated intervention in OTA-induced T2DM-NAFLD comorbidity. A PPI network was constructed using the STRING database, and topological analysis was performed with Cytoscape. Core targets were selected using the median values of six parameters—betweenness centrality, closeness centrality, degree centrality, eigenvector centrality, LAC (local average connectivity) score, and network centrality—as cut-off thresholds, and the top 10 key genes were further identified using the cytoHubba plugin. Gene Ontology (GO) functional enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were conducted via the DAVID database, and the results were visualized on the CNSknowall platform. Lastly, molecular docking of the core targets was performed using the CB-DOCK2 platform to validate binding affinity. Results: Based on an integrated analysis of network toxicology, network pharmacology, and molecular docking, 10 key targets were systematically identified. These may serve as potential mediators of cinnamaldehyde in the treatment of OTA-induced T2DM-NAFLD comorbidity. Among these, six targets—albumin (ALB), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), interleukin-6 (IL-6), tumor necrosis factor (TNF), actin beta (ACTB), and estrogen receptor 1 (ESR1)—possess crystal structures amenable to molecular docking. KEGG enrichment analysis revealed that CA and OTA jointly participate in key pathological processes such as the cancer pathway, the lipid and atherosclerosis pathway, the advanced glycation end-products–receptor for advanced glycation end-products (AGE-RAGE) signaling pathway, the phosphatidylinositol 3-kinase–protein kinase B (PI3K-Akt) signaling pathway, the TNF signaling pathway, and the interleukin-17 (IL-17) signaling pathway. OTA exacerbates inflammatory responses, impairs insulin signaling, promotes hepatic steatosis, and disrupts systemic metabolic homeostasis, ultimately contributing to T2DM-NAFLD comorbidity. Conversely, cinnamaldehyde counteracts these pathological processes through multiple mechanisms, including antioxidant and anti-inflammatory effects as well as regulation of glucose and lipid metabolism, thereby restoring metabolic homeostasis. Conclusions: This study has preliminarily identified the toxicological targets of OTA and the potential intervention targets of CA, offering new avenues for preventing and intervening in OTA-induced metabolic toxicity. Furthermore, it provides a theoretical basis for CA as a potential multi-target therapeutic agent and presents novel insights worthy of further investigation into the prevention of T2DM-NAFLD comorbidity.

Pharmaceuticals doi: 10.3390/ph19081282

Authors: Miruna Valeria Moraru Oana Andreia Coman Aurelian Zugravu Smaranda Stoleru Cristina Isabel Viorica Ghiță Mihnea Costescu Elena Poenaru Adrian-Florentin Dragomir Aurelia Cristiana Barbu Dragos Constantin Lunca Andrei Stoian Mugurel Nader Jafal Clara Maria Stoleru Ion Fulga

Background: Benzodiazepines and non-benzodiazepine hypnotics (Z-drugs) act at the benzodiazepine binding site of GABAA receptors but differ in their receptor subtype selectivity and behavioural profiles. Diazepam acts as a non-selective positive allosteric modulator of benzodiazepine-sensitive GABAA receptors, whereas zolpidem and indiplon display preferential affinity for α1-containing receptor subtypes. Although these compounds have been investigated individually in experimental models of exploratory behaviour and cognition, comparative behavioural data obtained using the same experimental protocol remain limited. Methods: This study evaluated the behavioural effects of diazepam, zolpidem and indiplon in male Wistar rats using a two-trial Y-maze paradigm with a 24 h retention interval. Animals received one of three dose levels of each compound before the acquisition trial. Recall-phase behaviour was evaluated using conventional exploration measures together with time- and entry-based preference indices for the previously inaccessible arm. Primary analyses were performed separately for each compound using one-way ANOVA with appropriate post hoc comparisons. Complementary exploratory factorial analyses were subsequently conducted to examine overall behavioural patterns across drug and nominal dose categories. Results: Diazepam produced marked dose-dependent reductions in recall-phase exploration of the previously inaccessible arm and in both exploratory preference indices. In contrast, zolpidem and indiplon reduced exploratory activity during the acquisition phase without significantly affecting recall-phase exploratory behaviour. The complementary factorial analyses identified significant Drug × Nominal Dose Categories interactions across all recall-related endpoints, indicating that behavioural responses across the nominal dose categories differed among the three compound-specific experimental series. The strongest interaction effects were observed for Time Index % (F(6,72) = 10.522, p < 0.001, ηp2 = 0.467) and Entries Index % (F(6,72) = 6.870, p < 0.001, ηp2 = 0.364). Conclusions: Diazepam produced more pronounced dose-dependent alterations in recall-phase exploratory behaviour than zolpidem or indiplon in the experimental conditions employed. The complementary factorial analyses support the interpretation that the three compounds exhibited different behavioural patterns across the nominal dose categories evaluated, while these exploratory comparisons should be interpreted within the constraints of the study design. Overall, the findings contribute to the comparative behavioural characterization of benzodiazepine-site modulators in the two-trial Y-maze paradigm.

Pharmaceuticals doi: 10.3390/ph19081281

Authors: Siri Paulo Mafalda Laranjo Ana Margarida Abrantes Ricardo Jorge Teixo Diogo A. Fonseca João Mendes-Abreu José Pedro Figueiredo Maria Filomena Botelho Manuel Marques Ferreira Arménio Serra

Background/Objectives: Osteonecrosis of the jaw, a non-healing wound, is a side effect in patients treated with nitrogen-containing bisphosphonates (e.g., zoledronate) after some oral procedures. Considering zoledronate toxicity in surgical wounds and the capacity of calcium phosphate synthetic compounds to adsorb it, the aim of this work was to demonstrate and elucidate the mechanism underlying this adsorption. Methods: Four different calcium phosphate synthetic bone substitutes were used in this study: AdBone® BCP (0.5–1 mm), Guidor® easy-graft (0.5–1 mm), Maxresorb® (0.5–1 mm) and Maxresorb® (0.8–1.5 mm). The materials were incubated in aqueous zoledronate solutions. Samples of the solutions obtained after 1, 2, 3, 6, 24, 48, 72 and 120 h of conditioning were then analyzed by spectrophotometry to measure their absorbance and subsequently characterized by micro-elemental analysis. Results: All four materials demonstrated time-dependent zoledronate adsorption. AdBone® BCP achieved the most rapid and sustained sequestration, reaching 80% reduction at 24 h and stabilizing at 85% from 72 h onwards without re-release. Maxresorb® formulations showed comparable kinetics through 72 h (70–75%) but exhibited partial re-release at 120 h. Guidor® easy-graft showed negligible adsorption through the first 24 h, consistent with the physical barrier effect of its PLGA coating. Spectrophotometric findings were independently confirmed by elemental analysis. Conclusions: This work confirms zoledronate adsorption by calcium phosphate synthetic biomaterials. Among the tested materials, AdBone® BCP demonstrated the most favorable adsorption profile, with kinetics well matched to the critical window of alveolar wound healing. These findings support the use of biphasic calcium phosphate bone substitutes as a local strategy to reduce free zoledronate bioavailability at the surgical site, potentially lowering the risk of MRONJ in patients undergoing dentoalveolar procedures under bisphosphonate therapy.

Pharmaceuticals doi: 10.3390/ph19081280

Authors: Manyu Wang Yang Fu Peipei Yuan Yi Zeng Zihao Yang Shenggui Zhang Weisheng Feng Xiaoke Zheng

Background: Male infertility, largely driven by declining sperm quality, is a growing global concern. Radix Bupleuri Chinensis (Chaihu, CH)—a key component of traditional formulas such as Chaihu Shengjing Tang and Chaihu Shugan San—has been clinically used to enhance sperm quality. However, the specific role and mechanism of CH as a single entity in ameliorating spermatogenic dysfunction remain unclear. This study aimed to determine whether CH extract ameliorates spermatogenic dysfunction and whether GnRH/GnRHR signaling and taurine biosynthesis are involved. Methods: Male Kunming (KM) mice and GC-1 spermatogonial cells were used to establish experimental models. Histopathological and cellular changes were examined via hematoxylin and eosin (H&E) staining and immunofluorescence. Enzyme and hormone levels were quantified, while gene and protein expression were analyzed using quantitative real-time polymerase chain reaction, Western blotting, and high-content imaging. Results: CH extract mitigated DOX-induced spermatogenic dysfunction by activating the GnRHR signaling pathway, enhancing taurine biosynthesis, and reducing oxidative stress. In vitro, saikosaponin D alleviated DOX-induced injury in GC-1 cells and was associated with changes in GnRHR-related signaling and taurine-biosynthesis-related proteins, supporting its potential role as a candidate bioactive constituent of CH. Conclusions: CH extract alleviated DOX-induced spermatogenic impairment in mice. The in vivo cetrorelix experiments support the involvement of the GnRHR signaling pathway, whereas the concurrent changes in taurine, CDO1, and FMO1 are consistent with enhanced taurine biosynthesis. These findings provide experimental support for further investigation of CH as a potential intervention for chemotherapy-related reproductive injury.

Pharmaceuticals doi: 10.3390/ph19081279

Authors: Szymon Viscardi Anna Duda-Madej Paweł Krzyżek

Helicobacter pylori is one of the most common bacterial pathogens in humans and the primary etiological agent of chronic gastritis, peptic ulcer disease, and gastric cancer. Its ability to establish persistent gastric colonization relies on multiple virulence factors, including adhesins, urease, cytotoxins, motility, outer membrane vesicles, and biofilm formation, which collectively promote bacterial survival, chronic inflammation, and treatment failure. The increasing prevalence of antibiotic-resistant H. pylori strains has intensified the search for therapeutic strategies targeting both bacterial viability and virulence. Berberine (BBR), a natural isoquinoline alkaloid, has emerged as a promising candidate because of its antibacterial, anti-inflammatory, and antioxidant properties. Increasing evidence derived from native berberine, its derivatives, and berberine-based formulations indicates multifaceted anti-H. pylori activity, including direct antibacterial effects, inhibition of virulence determinants, and modulation of host inflammatory responses. This review summarizes current knowledge on the epidemiology and pathogenic mechanisms of H. pylori and provides a comprehensive overview of the available evidence regarding the anti-H. pylori pharmacological profile of BBR-based compounds. Particular attention is given to their effects on bacterial adhesion, motility, urease activity, efflux pump function, biofilm formation, and host inflammatory signaling pathways. The review also discusses findings from preclinical and clinical studies supporting BBR-based strategies as adjuncts to conventional eradication therapies. In addition, recent advances in nanotechnology-based drug delivery systems designed to overcome the poor oral bioavailability of BBR and improve its therapeutic efficacy against H. pylori are highlighted.

Pharmaceuticals doi: 10.3390/ph19081278

Authors: Nikoleta Đorđevski Ana Ćirić Uroš Gašić Marija Ivanov Alexia Delnatte Mikhael Bechelany Dina Tucović Jelena Kulaš Maja Čakić-Milošević Dejan Stojković

Background/Objectives: Verbena officinalis has long been used in traditional medicine for treating skin disorders, yet its potential in advanced wound-dressing systems remains insufficiently explored. This study aimed to characterize V. officinalis extracts obtained using different solvents, evaluate their antimicrobial, antibiofilm, cytotoxic, and wound-healing activities, and develop electrospun polycaprolactone–polyethylene glycol (PCL–PEG) nanofibers as a delivery platform for the most active extract. Methods: Extracts were chemically characterized by LC–MS and evaluated against clinical bacterial, yeast, and dermatophyte isolates associated with skin infections. Antibiofilm activity was assessed against Staphylococcus lugdunensis. The most active hydroalcoholic extract (EW7.5) was incorporated into PCL–PEG nanofibers (1–10%, w/w). Metabolite release was monitored by semi-quantitative LC–MS analysis. Cytotoxicity and wound-healing activity were evaluated using HaCaT keratinocytes, while in vivo efficacy was assessed in a full-thickness excisional wound model in Dark Agouti rats (five animals per group in two independent experiments). Results: EW7.5 exhibited the strongest antimicrobial activity, with MIC values of 0.06 mg/mL against all tested bacterial and Candida strains and 0.06–0.5 mg/mL against dermatophytes. It inhibited S. lugdunensis biofilm formation by 79.43% at MIC/4 and reduced pre-formed biofilm biomass by 64.98% at 2 × MBC. EW7.5 and EW7.5-loaded nanofibers showed no cytotoxicity up to 400 μg/mL. In the scratch assay, the free extract achieved 42.34 ± 1.58% wound closure, while nanofibers containing 10% EW7.5 achieved 25.87 ± 2.52%, compared to 7.73 ± 0.07% for the control. In vivo, EW7.5-loaded membranes significantly accelerated wound closure and the 10% formulation significantly improved extracellular matrix deposition and vascular remodeling compared to the control membrane. Conclusions: EW7.5-loaded PCL–PEG nanofibers combine antimicrobial, antibiofilm, biocompatible, and wound-healing properties, supporting their potential as multifunctional wound dressings. Nevertheless, further studies with mechanistic investigations are required to fully establish their therapeutic applicability.

Pharmaceuticals doi: 10.3390/ph19081277

Authors: Se-Eun Byeon Rengarajan Baskaran Young-Joon Park

Background/Objective: Niclosamide exhibits considerable potential for the treatment of drug-resistant cancers. However, its poor aqueous solubility substantially limits its oral bioavailability and therapeutic efficacy. To overcome this limitation, we developed a novel pharmaceutical supramolecular crystalline phase comprising niclosamide and nafamostat. Methods: In this study, we developed a niclosamide–nafamostat pharmaceutical supramolecular crystalline phase using a conventional solvent evaporation technique. Comprehensive solid-state characterization was carried out using X-ray powder diffraction (XRD), Fourier-transform infrared spectroscopy (FTIR), and differential scanning calorimetry (DSC). In vitro drug transport kinetics and barrier transport efficiency were evaluated across artificial transpermeable membranes using a diffusion cell setup. In vitro anticancer efficacy was systematically screened against human lung, breast, and pancreatic cancer cell lines. Finally, in vivo translation was established using female NOD/SCID mice tumor-bearing animal models; therapeutic efficacy and total tumor burden were evaluated. Results: Niclosamide–nafamostat cocrystal (NNC) was found to form a distinct crystalline phase, with characterization results supporting a unique crystal lattice stabilized through strong intermolecular hydrogen-bonding interactions and exhibiting high thermal purity. Crucially, the supramolecular crystalline phase considerably enhances membrane transport, yielding superior cumulative permeation and enhanced apparent permeability (Papp) values compared to those of pure niclosamide. In vitro evaluation across multiple cancer cell lines demonstrated a tenfold increase in antiproliferative potency compared to the parent compounds. Furthermore, in vivo studies revealed a twofold increase in tumor growth inhibition and a tenfold reduction in total tumor burden (p < 0.05). Conclusions: These findings demonstrate that the NNC supramolecular crystalline phase platform successfully optimizes membrane permeability and antitumor efficacy, highlighting its potential for treating advanced cancers.

Pharmaceuticals doi: 10.3390/ph19081276

Authors: Yong Huang Chengze Li Taichuang Li Jiuhui Zhao Xinyu Yang Maoqun Zhang Ying Liang

Background/Objectives: Antibody selection is critical for advancing pretargeted PET imaging toward clinical translation. This study directly compared two clinically approved anti-HER2 antibodies—trastuzumab and pertuzumab—within an identical pretargeting system, utilizing the specific chemical ligation between tetrazine and trans-cyclooctene (TCO) via the inverse electron-demand Diels–Alder (IEDDA) cycloaddition. Methods: In HER2-positive SKOV3 tumor-bearing mice, TCO-conjugated antibodies were administered 1–13 days prior to injection of a fluorine-18-labeled tetrazine probe ([18F]PEG12-Tz). Results: Both strategies enabled high-contrast imaging, overcoming the kinetic mismatch between slow-clearing antibodies and the short-lived 18F isotope. Distinct profiles were observed: pertuzumab-TCO yielded higher tumor uptake and a broader imaging window (3–13 days), whereas trastuzumab-TCO resulted in lower uptake and a narrower window (1–5 days). These differences are not affinity-driven, as both conjugates showed similar Kd values (~11–14 nM). Instead, pertuzumab-TCO’s superiority stems from a dual mechanism: (1) slower cellular internalization (65.1% vs. 94.0% at 60 min) preserves surface TCO for click reaction; and (2) markedly slower blood clearance (t1/2β = 285.9 h vs. 106.0 h) ensures sustained bioavailability and tumor extravasation. Conclusions: This head-to-head evaluation demonstrates that epitope-dictated internalization and clearance are decisive parameters for antibody selection, providing a mechanism-based framework for optimizing pretargeted imaging.

Pharmaceuticals doi: 10.3390/ph19081275

Authors: Huayan Li Xiying He Ting Cao Jinfeng Huang Bojun Chen Lijing Xu Yanxiao Yang Gang Li Lei Xiong

Depression is a leading cause of disability worldwide, and currently available antidepressants are limited by delayed therapeutic onset, inadequate efficacy in some patients, and adverse effects. G protein-coupled receptors (GPCRs), the largest family of membrane receptors in the central nervous system, regulate neurotransmission, neuroplasticity, neuroinflammation, stress responses, and reward processing, and are therefore important targets for antidepressant drug development. Natural products are a rich source of structurally diverse bioactive compounds, many of which show antidepressant-like effects through the modulation of GPCR-mediated signaling pathways. In this narrative review, we summarize the roles of major GPCR families implicated in depression and provide an updated overview of natural products that modulate these receptors. We particularly emphasize receptor-specific mechanisms, downstream signaling networks, and the pharmacological actions of representative natural compounds. We also highlight emerging concepts in GPCR biology, including receptor heteromerization, signaling bias, and allosteric modulation, that may create new opportunities for antidepressant discovery. Finally, we discuss current challenges related to target validation, pharmacokinetics, and clinical translation. Collectively, these insights support further investigation of natural product-derived GPCR modulators as potential leads for next-generation antidepressant development.

Pharmaceuticals doi: 10.3390/ph19081274

Authors: Marcin Zakrzewski Patrycja Bełdzińska Karolina Gackowska Aliaksandra Yurchak Marzena Jamrógiewicz Dariusz Wyrzykowski Katarzyna Bury Katarzyna Grzyb Grzegorz Gołuński Jacek Piosik

Background/Objectives: Cancer remains one of the leading causes of death worldwide. Although chemotherapy is widely used, it is associated with severe side effects, including myelosuppression and systemic toxicity. Nanoparticles have emerged as promising candidates for modulating drug activity. In this study, we investigated whether platinum nanoparticles (PtNPs) of various sizes interact with idarubicin (IDA), an anthracycline anticancer drug used primarily to treat acute leukaemia. Methods: Interactions between PtNPs and IDA were analysed using dynamic light scattering (DLS), atomic force microscopy (AFM), fluorescence spectroscopy, Fourier-transform infrared (FTIR) spectroscopy, and near-infrared (NIR) spectroscopy. Thermodynamic and thermal properties were assessed using isothermal titration calorimetry (ITC) and differential scanning calorimetry (DSC). Biological effects were evaluated using the Ames mutagenicity assay on the Salmonella enterica serovar Typhimurium TA98 strain and cytotoxicity assays on SK-BR-3 and MCF-7 cell lines. Results: DLS demonstrated changes in the hydrodynamic diameter of PtNPs following IDA addition, which were further supported by AFM imaging. PtNPs significantly quenched IDA fluorescence, indicating close molecular interactions, which were further supported by FTIR and NIR. ITC revealed that the interactions were endothermic, with enthalpy values ranging from 1.2 to 3.6 kcal/mol, and DSC demonstrated that the PtNP-IDA combination altered the melting temperature of IDA. Biological assays revealed that all examined PtNP sizes influenced IDA mutagenicity in the Salmonella enterica serovar Typhimurium TA98 strain. Furthermore, PtNPs modulated the cytotoxicity of IDA in SK-BR-3 and MCF-7 cell lines in a dose-dependent manner. Conclusions: These findings demonstrate that PtNPs interact with IDA and modulate its biological activity. However, in this study no non-cancerous cell lines were examined; therefore, the observed interactions and biological effects support further investigation of the PtNP-IDA combination in the context of nanoparticle-assisted anticancer therapy on a broader range of in vitro cell lines.

Pharmaceuticals doi: 10.3390/ph19081273

Authors: Umme Hani Mona Al Hamod Noura Al Hamood Yahya Alhamhoom Mohammed Ghazwani Fahad AlQahtani Helal A. Helal Riyaz Ali M. Osmani

Prostate cancer (PC) is one of the most common cancers in men globally and there is an urgent need for new immune-based approaches because many traditional therapies, including chemotherapy, radiotherapy, and anti-androgens, have limitations. Due to their distinct physicochemical and biological features, gold nanoparticles (AuNPs) are emerging as a potential nanoplatform for the development of strategies in prostate cancer therapy. These features include tunable size, shape, and surface plasmon resonance (SPR) and high surface-to-volume ratio, which result in enhanced drug loading, targeted delivery and improved bioavailability. In addition, AuNPs can also be functionalized for active targeting to promote selective accumulation in tumors while minimizing systemic toxicity. In addition, the intrinsic optical and photothermal properties of these nanoparticles allow them to serve for PTT, radiosensitization and multimodal imaging, i.e., CT (computed tomography) and photoacoustic imaging. These have shown potential in pre-clinical and clinical evaluation but face issues with long-term toxicity, biodistribution and large-scale manufacturing. In this review, we summarize the organizing features, functional properties, therapeutic activities and translational potentials of AuNPs in prostate cancer treatment, with emphasis on their contributions towards precision nanomedicine.

Pharmaceuticals doi: 10.3390/ph19081272

Authors: Husam Qanash Aisha M. H. Al-Rajhi Abdulrahman S. Bazaid Manar F. Alghassab Fahad Almarshadi Walid Alesefir Waleed Hakami Amro Duhduh Abdu Aldarhami

Background/Objectives: Delonix regia seeds contain phytochemicals with therapeutic potential, but their activity against Helicobacter pylori and biomedical properties remain incompletely characterized. This study aimed to characterize the phenolic and amino acid composition of D. regia seed extract (DRSE) and evaluate its anticancer, wound-healing, anti-inflammatory, anticoagulant, antibacterial, antibiofilm and urease-targeted docking activities. Methods: DRSE was analyzed by high-performance liquid chromatography (HPLC) and amino acid analysis. Biological effects were assessed using MTT cytotoxicity, scratch wound-healing, bovine serum albumin (BSA) denaturation, prothrombin time (PT) and partial thromboplastin time (PTT), antibacterial and crystal violet antibiofilm assays. Gallic acid and vanillin were docked against H. pylori urease (PDB ID: 1E9Y). Results: Gallic acid was the predominant phenolic compound (1417.90 µg/g), while aspartic and glutamic acids dominated the amino acid fraction. DRSE showed preferential cytotoxicity toward A431 carcinoma cells (IC50 = 108.97 ± 0.68 µg/mL) compared with HFB4 fibroblasts (IC50 = 318.56 ± 2.06 µg/mL), yielding a selectivity index of 2.92. Scratch closure was comparable to the control (81.71% versus 80.85%), although the migration rate increased to 16.12 µm. DRSE inhibited protein denaturation by 91.20% (IC50 = 6.39 ± 0.19 µg/mL) and prolonged PT and PTT to 27.37 and 90.50 s, respectively. It inhibited H. pylori with minimum inhibitory and bactericidal concentrations of 31.25 µg/mL and suppressed biofilm formation by 95.39%. Gallic acid and vanillin showed comparable urease docking scores of −4.72 and −4.71 kcal/mol. Conclusions: DRSE showed selective anticancer, anti-H. pylori, antibiofilm, anti-inflammatory, anticoagulant, and moderate pro-migratory activities. Mechanistic, safety and in vivo studies are warranted.

Pharmaceuticals doi: 10.3390/ph19081271

Authors: Nariman Nabil Hussein Sabit Jawaher Almulhim Borros Arneth Shaimaa Abdel-Ghany

Background/Objectives: This study engineered and evaluated a targeted, folate-functionalized chitosan nanoparticle (CS-FA NP) delivery system to enhance the therapeutic efficacy of standard quercetin and Coriandrum sativum seed extract against breast cancer. Methods: Phytochemical profiling confirmed a 14% crude yield for the methanolic extract, with gas chromatography–mass spectrometry (GC-MS) and high-performance liquid chromatography (HPLC) identifying quercetin as the principal bioactive agent. The synthesized CS-FA NPs exhibited a core size of 7–20 nm, an average hydrodynamic diameter of 150–160 nm, a stable zeta potential of −55 mV, and high encapsulation efficiencies (87.2% for quercetin and 80.5% for coriander). Kinetic assessments confirmed a biphasic, diffusion-controlled release matching Higuchi matrix kinetics. Anticancer activity was evaluated in vitro using MTT cytotoxicity, Annexin V-FITC/PI apoptosis analysis, RT-qPCR, and ex vivo rat aortic ring assays, followed by validation in a syngeneic 4T1 mammary tumor mouse model. Results: In vitro, folate-receptor-targeted quercetin nanoparticles (T4) demonstrated superior, selective cytotoxicity, particularly against triple-negative MDA-MB-231 cells, while sparing normal fibroblasts. Annexin V-FITC/PI apoptosis profiling and ex vivo aortic ring assays revealed profound, cell-line-dependent programmed cell death and up to 90% inhibition of microvessel sprout outgrowth. Mechanistically, RT-qPCR verified that nano-formulations induced complete transcriptional silencing of NANOG, MMP-1, VEGFA, TSPAN8, TWIST, EMMPRIN, and CDK1, alongside marked upregulation of P27KIP1 and P21CIP1. In vivo, these nano-formulations successfully improved tumor-associated pathological features, reduced aggressive tumor spindle-cell proliferation, and suppressed elevated serum CA15-3 and arginase biomarkers. Conclusions: Folate-functionalized chitosan nano-formulations significantly enhanced the anticancer efficacy of quercetin and Coriandrum sativum seed extract through improved targeted delivery, potent antiproliferative, anti-angiogenic, and pro-apoptotic activities, together with favorable modulation of multiple molecular pathways associated with breast cancer progression. These findings support their potential as promising targeted nanotherapeutic strategies for breast cancer treatment.