Search Results (68)

Alzheimer’s disease is a progressive neurodegenerative disorder marked by declining cognitive function. While early-stage treatment focuses on acetylcholinesterase (AChE) inhibition, butyrylcholinesterase (BChE) activity increases as the disease progresses, contributing to cholinergic deficits and neuroinflammation. This shift in enzyme dominance presents a compelling rationale for developing BChE-specific inhibitors as a potential therapeutic avenue. This study explores small, three-membered rings, scaffolds offering potential for interaction with the enzyme’s active site, as building blocks for novel BChE inhibitors. Employing a computational approach based on quantum–chemical multiligand simultaneous molecular docking, we virtually fitted these compounds into the BChE active site to predict binding affinity and key interactions. Our calculations extend beyond simple shape matching by incorporating accurate electronic properties, leading to more reliable predictions of binding strength and stability. The goal was not immediate identification of potent inhibitors, but a systematic assessment of how these rings interact with BChE. This foundational knowledge will inform the design and synthesis of larger, more complex molecules with enhanced binding affinity and selectivity, ultimately aiming to develop compounds to inhibit BChE activity and potentially slow Alzheimer’s progression. Full article

Histone deacetylases (HDACs) 1–3 are key regulators of gene expression and represent important therapeutic targets in cancer, neurodegenerative, and immune disorders. Many potent class I HDAC inhibitors display slow- and tight-binding kinetics, which profoundly influence their efficacy and pharmacodynamics. In particular, their dissociation rate (off-kinetic) is critical, since prolonged target engagement greatly influences drug efficacy in vivo. However, the off-kinetics of HDAC inhibitors are often overlooked in the early stages of drug development. Here, we investigated the dissociation kinetics of tucidinostat, trapoxin A, and TNG260 in comparison to the pan-HDAC inhibitor vorinostat. Using biochemical 100-fold jump dilution assays, NanoBRET assays, and cellular washout experiments, we characterized the dissociation of these compounds from purified proteins and in a cellular context. Tucidinostat showed moderately slow off-kinetics, while the clinical candidate TNG260 demonstrated pronounced tight-binding properties. Trapoxin A displayed remarkable discrepancies between assays, as it showed fast dissociation kinetics in the biochemical assay, but tight-binding properties in a cellular setting. These findings not only address the previously unexplored dissociation kinetics of two clinically relevant inhibitors, but also underscore the importance of comprehensive kinetic profiling of novel HDAC inhibitors in cellular models. Full article

Idiopathic pulmonary fibrosis (IPF) is a chronic and irreversible interstitial lung disease characterized by progressive scarring of the lungs. The available therapeutic strategies are limited and primarily focus on slowing disease progression rather than achieving fibrosis reversal. Cardamonin (CDN), a food-derived natural chalcone, has exhibited anti-fibrotic activity in liver and kidney fibrosis models; however, its role and underlying mechanism in IPF remain unelucidated. Herein, we integrated network pharmacology, machine learning, molecular simulations, and in vitro experiments. Network pharmacology identified 135 overlapping targets between CDN and IPF, which demonstrated a significant enrichment in the Phosphatidylinositol 3-Kinase/Protein Kinase B signaling pathway (PI3K/AKT). Machine learning further prioritized 6 core targets, with IGF1 emerging as a key candidate. Molecular docking revealed a favorable binding energy of −7.9 kcal/mol for the CDN-IGF1 complex. Subsequent 100 ns molecular dynamics simulations further confirmed its robust binding stability, yielding a mean binding free energy of −150.978 kcal/mol. In vitro, CDN significantly mitigated fibrosis in bleomycin (BLM)-challenged A549 cells, downregulating the expression of α-smooth muscle actin (α-SMA) and fibronectin. This effect was accompanied by a beneficial reversal of epithelial–mesenchymal transition (EMT), as indicated by increased E-cadherin levels and decreased vimentin expression. Mechanistically, CDN significantly suppressed the IGF1/PI3K/AKT axis; this inhibitory effect was partially reversed by exogenous IGF1 supplementation and further enhanced by the PI3K-specific inhibitor LY294002. This work provides the evidence that CDN alleviates BLM-induced pulmonary fibrosis by targeting the IGF1/PI3K/AKT-EMT axis. These findings lend support to a robust mechanistic basis for developing CDN as a potential therapeutic candidate for IPF. It should be noted that these conclusions are drawn from in vitro experiments using A549 cells, and further validation in primary alveolar epithelial cells and animal models is warranted to confirm their physiological relevance. Full article

Tyrosinase (EC 1.14.18.1) is the primary enzyme responsible for melanogenesis in mammals and enzymatic browning in food, creating a high demand for potent, safe inhibitors of this enzyme in the cosmetic, medical, and agricultural sectors. Conventional synthetic inhibitors often face limitations concerning their cytotoxicity and stability, necessitating the exploration of marine natural products (MNPs). Marine algae, comprising macroalgae (seaweeds) and microalgae (including cyanobacteria), represent an underexploited source of structurally diverse bioactives. Macroalgae, particularly brown species, yield complex phlorotannins, such as the non-competitive oligomer dieckol, which exhibits an IC50 of 2.16 µg/mL. Conversely, microalgae deliver high-potency, low-molecular-weight compounds, notably the synthesizable scytonemin monomer (ScyM) with an IC50 of 4.90 µM—significantly stronger than kojic acid. Mechanistic analysis, supported by molecular docking, reveals diverse modes of action, from the two-step slow binding of complex phlorotannins to the highly specific competitive binding of red algal bromophenols. Translational success requires the consistent application of green extraction techniques, such as Natural Deep Eutectic Solvents (NADESs), and advanced delivery systems, like Nanostructured Lipid Carriers (NLCs), to ensure the stability and bioavailability of these compounds for future cosmeceutical and medical applications. Full article

Alpha2-plasmin inhibitor (α2PI) has a heterogeneous structure due to proteolytic cleavages in the circulation. The C-terminally cleaved form loses the plasminogen binding site and is, therefore, a slow plasmin inhibitor (NPB-α2PI). As FXIII primarily crosslinks the plasminogen-binding intact form (PB-α2PI) to fibrin, the effect of NPB-α2PI on fibrinolysis has been less studied. Herein, we investigated the effect of C-terminal truncation. Total-, PB-, and NPB-α2PI antigen levels and α2PI incorporation were measured by ELISAs from samples of 80 healthy individuals. Clot lysis parameters of the same subjects were investigated using an in vitro clot lysis assay. α2PI incorporation into the clot was demonstrated by Western blotting. Clot lysis and clot structure were also analyzed using an α2PI-deficient plasma substituted with recombinant PB- and NPB-α2PI. Both plasma and clot-bound levels of total- and NPB-α2PI showed a significant positive correlation with clot lysis parameters. NPB-α2PI was detected in the clot due to non-covalent binding. Regardless of the type of binding, both forms affected the clot structure by increasing the thickness of the fibrin fibers and reducing the pore size. In conclusion, we found that NPB-α2PI can bind non-covalently to fibrin, and this binding contributes to changes in clot structure and inhibition of fibrinolysis. Full article

Background: Alopecia areata is an autoimmune disorder that causes hair loss in clumps about the size and shape of a quarter. The estimated prevalence of the disorder is approximately 1 in 1000 people, with a lifetime risk of approximately 2 percent. One of the systemic therapies for alopecia areata consists of the use of glucocorticoids or immunosuppressants. Methods: Baricitinib (BCT) is a Janus kinase (JAK) 1 and 2 selective inhibitor used as an immunosuppressant drug. In this study, three olive oil BCT formulations (Oil A, Oil B, and Oil C, which differ in their content in squalene, tocopherol, tyrosol, and hydroxytyrosol) have been developed for topical delivery. The formulations were physicochemically characterized and the in vitro drug release and ex vivo permeation through human skin tissues were assessed. Results: The results showed nearly identical viscosity across all three formulations, exhibiting Newtonian behavior. The mathematical modeling used to describe the drug release profiles was the one-site binding hyperbola for all formulations. Oil-based formulations showed a slow BCT penetration into human skin. Skin integrity remained intact during the experiments, with no signs of irritation or alterations observed. In addition, all the formulations proved their efficacy in vivo. Conclusions: Among the formulations, Oil A demonstrated the highest ability retention capacity (Q_r = 1875 ± 124.32 ng/cm²) in the skin, making it an excellent candidate for further investigation in the treatment of alopecia areata. Full article

Background/Objectives: Androgen receptor (AR) expression and signaling are critical for the progression of prostate cancer and have been the therapeutic focus of prostate cancer for over 50 years. While a variety of agents have been developed to target this axis, many of these fail due to the emergent expression of AR RNA splice variants, such as AR-V7, that can signal independently of ligand binding. Other therapies, such as vaccination against prostate-specific antigens, have achieved FDA approvals but have fallen short of being incorporated as standard-of-care therapies for advanced prostate cancer. This may be due to the elevated level of immunosuppression observed in prostate cancer, which remains largely refractory to immune checkpoint blockade. Methods: We developed a vaccine targeting AR-V7, a common isoform associated with treatment resistance, and demonstrated its ability to elicit AR-V7-specific immunity and enable anti-tumor responses against AR-V7+ cancers in subcutaneous tumor models. Results: Our studies also revealed that AR-V7 expression conferred an immune suppressive phenotype that was significant in a non-AR-dependent prostate cancer model. Notably, in this model, we found that vaccination in combination with enzalutamide, an AR antagonist, suppressed these aggressive immune suppressive cancers and resulted in enhanced survival in comparison to control vaccinated and enzalutamide-treated mice. While anti-PD-1 immune checkpoint inhibition (ICI) alone slowed tumor growth, the majority of vaccinated mice that received anti-PD-1 therapy showed complete tumor elimination. Conclusions: Collectively, these results validate the importance of AR signaling in prostate cancer immune suppression and suggest the potential of AR-V7-specific vaccines as therapeutic strategies against prostate cancer, offering significant protective and therapeutic anti-tumor responses, even in the presence of androgen signaling inhibitors. Full article

Prolonged exposure to HIV-1 transactivator of transcription (Tat) protein dysregulates monoamine transmission, a physiological change implicated as a key factor in promoting neurocognitive disorders among people living with HIV. We have demonstrated that in vivo expression of Tat in Tat transgenic mice decreases dopamine uptake through both dopamine transporter (DAT) and norepinephrine transporter (NET) in the prefrontal cortex. Further, our novel allosteric inhibitor of monoamine transporters, SRI-32743, has been shown to attenuate Tat-inhibited dopamine transport through DAT and alleviates Tat-potentiated cognitive impairments. The current study reports the pharmacological profiles of SRI-32743 in basal and Tat-induced inhibition of human NET (hNET) function. SRI-32743 exhibited less affinity for hNET binding than desipramine, a classical NET inhibitor, but displayed similar potency for inhibiting hDAT and hNET activity. SRI-32743 concentration-dependently increased hNET affinity for [³H]DA uptake but preserved the V_max of dopamine transport. SRI-32743 slowed the cocaine-mediated dissociation of [³H]Nisoxetine binding and reduced both [³H]DA and [³H]MPP+ efflux but did not affect d-amphetamine-mediated [³H]DA release through hNET. Finally, we determined that SRI-32743 attenuated a recombinant Tat_1–86-induced decrease in [³H]DA uptake via hNET. Our findings demonstrated that SRI-32743 allosterically disrupts the recombinant Tat_1–86–hNET interaction, suggesting a potential treatment for HIV-infected individuals with concurrent cocaine abuse. Full article

SHP2, a pivotal component downstream of both receptor and non-receptor tyrosine kinases, has been underscored in the progression of various human cancers and neurodevelopmental disorders. Allosteric inhibitors have been proposed to regulate its autoinhibition. However, oncogenic mutations, such as E76K, convert SHP2 into its open state, wherein the catalytic cleft becomes fully exposed to its ligands. This study elucidates the dynamic properties of SHP2 structures across different states, with a focus on the effects of oncogenic mutation on two known binding sites of allosteric inhibitors. Through extensive modeling and simulations, we further identified an alternative allosteric binding pocket in solution structures. Additional analysis provides insights into the dynamics and stability of the potential site. In addition, multi-tier screening was deployed to identify potential binders targeting the potential site. Our efforts to identify a new allosteric site contribute to community-wide initiatives developing therapies using multiple allosteric inhibitors to target distinct pockets on SHP2, in the hope of potentially inhibiting or slowing tumor growth associated with SHP2. Full article

The kinetics and mechanism of drug binding to its target are critical to pharmacological efficacy. A high throughput (HTS) screen often results in hundreds of hits, of which usually only simple IC₅₀ values are determined during reconfirmation. However, kinetic parameters such as residence time for reversible inhibitors and the k_inact/K_I ratio, which is the critical measure for evaluating covalent inactivators, are early predictive measures to assess the chances of success of the hits in the clinic. Using the promising cancer target human histone deacetylase 8 as an example, we present a robust method that calculates concentration-dependent apparent rate constants for the inhibition or inactivation of HDAC8 from dose–response curves recorded after different pre-incubation times. With these data, hit compounds can be classified according to their mechanism of action, and the relevant kinetic parameters can be calculated in a highly parallel fashion. HDAC8 inhibitors with known modes of action were correctly assigned to their mechanism, and the binding mechanisms of some hits from an internal HDAC8 screening campaign were newly determined. The oxonitriles SVE04 and SVE27 were classified as fast reversible HDAC8 inhibitors with moderate time-constant IC₅₀ values of 4.2 and 2.6 µM, respectively. The hit compound TJ-19-24 and SAH03 behave like slow two-step inactivators or reversible inhibitors, with a very low reverse isomerization rate. Full article

This study aimed to enhance the stability of the Rotigotine (ROT) patch using polymers as crystal inhibitors. Three polymers (Poloxamer 188, Soluplus, TPGS) were selected as crystal inhibitors to formulate ROT patches with varying drug loadings (20%, 40%, 60%, and 80%, w/w). SEM and XRD analysis revealed that the Soluplus and Soluplus-TPGS groups with a high concentration (80%, w/w) of ROT could be stored at room temperature for at least 90 days without crystallization. Moreover, the crystallization nucleation time and growth rate were utilized to assess the ability of Poloxamer 188, Soluplus, and TPGS to hinder the formation of ROT crystals and slow down its crystallization rate. Molecular docking results elucidated the intermolecular forces between ROT and different polymers, revealing their mechanisms for crystal inhibition. The ROT-Soluplus-TPGS combination exhibited the lowest binding free energy (−5.3 kcal/mol), indicating the highest binding stability, thereby effectively reducing crystal precipitation. In vitro skin permeation studies demonstrated that ROT patches containing crystal inhibitors exhibited promising transdermal effects. With increasing ROT concentration, the cumulative drug permeation substantially increased, while the lag time was notably reduced. This study offers novel insights for the development of ROT patches. Full article

PLK1 is found to be highly expressed in various types of cancers, but the development of inhibitors for it has been slow. Most inhibitors are still in clinical stages, and many lack the necessary selectivity and anti-tumor effects. This study aimed to create new inhibitors for the PLK1-PBD by focusing on the PBD binding domain, which has the potential for greater selectivity. A 3D QSAR model was developed using a dataset of 112 compounds to evaluate 500 molecules. ADMET prediction was then used to select three molecules with strong drug-like characteristics. Scaffold hopping was employed to reconstruct 98 new compounds with improved drug-like properties and increased activity. Molecular docking was used to compare the efficient compound abbapolin, confirming the high-activity status of [(14S)-14-hydroxy-14-(pyridin-2-yl)tetradecyl]ammonium,[(14S)-15-(2-furyl)-14-hydroxypentadecyl]ammonium and [(14S)-14-hydroxy-14-phenyltetradecyl]ammonium. Molecular dynamics simulations and MMPBSA were conducted to evaluate the stability of the compounds in the presence of proteins. An in-depth analysis of [(14S)-15-(2-furyl)-14-hydroxypentadecyl]ammonium and [(14S)-14-hydroxy-14-phenyltetradecyl]ammonium identified them as potential candidates for PLK1 inhibitors. Full article

Monoamine transporters, including dopamine, norepinephrine, and serotonin transporters (DAT, NET, and SERT, respectively), are important therapeutic targets due to their essential roles in the brain. To overcome the slow action of selective monoamine reuptake inhibitors, dual- or triple-acting inhibitors have been developed. Here, to examine whether combination treatments of selective reuptake inhibitors have synergistic effects, the pharmacological properties of DAT, NET, and SERT were investigated using the selective inhibitors of each transporter, which are vanoxerine, nisoxetine, and fluoxetine, respectively. Potencies were determined via fluorescence-based substrate uptake assays in the absence and presence of other inhibitors to test the multi-drug effects on individual transporters, resulting in antagonistic effects on DAT. In detail, fluoxetine resulted in a 1.6-fold increased IC₅₀ value of vanoxerine for DAT, and nisoxetine produced a more drastic increase in the IC₅₀ value by six folds. Furthermore, the effects of different inhibitors, specifically monovalent ions, were tested on DAT inhibition by vanoxerine. Interestingly, these ions also reduced vanoxerine potency in a similar manner. The homology models of DAT suggested a potential secondary inhibitor binding site that affects inhibition in an allosteric manner. These findings imply that the use of combination therapy with monoamine reuptake inhibitors should be approached cautiously, as antagonistic effects may occur. Full article

Replication protein A (RPA) is the major single-stranded DNA (ssDNA) binding protein that is essential for DNA replication and processing of DNA double-strand breaks (DSBs) by homology-directed repair pathways. Recently, small molecule inhibitors have been developed targeting the RPA70 subunit and preventing RPA interactions with ssDNA and various DNA repair proteins. The rationale of this development is the potential utility of such compounds as cancer therapeutics, owing to their ability to inhibit DNA replication that sustains tumor growth. Among these compounds, (1Z)-1-[(2-hydroxyanilino) methylidene] naphthalen-2-one (HAMNO) has been more extensively studied and its efficacy against tumor growth was shown to arise from the associated DNA replication stress. Here, we study the effects of HAMNO on cells exposed to ionizing radiation (IR), focusing on the effects on the DNA damage response and the processing of DSBs and explore its potential as a radiosensitizer. We show that HAMNO by itself slows down the progression of cells through the cell cycle by dramatically decreasing DNA synthesis. Notably, HAMNO also attenuates the progression of G2-phase cells into mitosis by a mechanism that remains to be elucidated. Furthermore, HAMNO increases the fraction of chromatin-bound RPA in S-phase but not in G2-phase cells and suppresses DSB repair by homologous recombination. Despite these marked effects on the cell cycle and the DNA damage response, radiosensitization could neither be detected in exponentially growing cultures, nor in cultures enriched in G2-phase cells. Our results complement existing data on RPA inhibitors, specifically HAMNO, and suggest that their antitumor activity by replication stress induction may not extend to radiosensitization. However, it may render cells more vulnerable to other forms of DNA damaging agents through synthetically lethal interactions, which requires further investigation. Full article

Introduction: Upregulation of complement system factors are reported to be increased in polycystic ovary syndrome (PCOS) and may be due to obesity and insulin resistance rather than inherently due to PCOS. We directly compared complement factors from an obese, insulin-resistant PCOS population to a nonobese, non-insulin-resistant PCOS population in a proteomic analysis to investigate this. Methods: Plasma was collected from 234 women (137 with PCOS and 97 controls) from a biobank cohort and compared to a nonobese, non-insulin-resistant population (24 with PCOS and 24 controls). Slow off-rate modified aptamer (SOMA) scan plasma protein measurement was undertaken for the following complement system proteins: C1q, C1r, C2, C3, C3a, iC3b, C3b, C3d, C3adesArg, C4, C4a, C4b, C5, C5a, C5b-6 complex, C8, properdin, factor B, factor D, factor H, factor I, Mannose-binding protein C (MBL), complement decay-accelerating factor (DAF) and complement factor H-related protein 5 (CFHR5). Results: The alternative pathway of the complement system was overexpressed in both obese and nonobese PCOS, with increased C3 (p < 0.05) and properdin (p < 0.01); additionally, factor B increased in obese PCOS (p < 0.01). For inhibitors of this pathway, factor I was increased (p < 0.01) in both slim and obese PCOS, with an increase in CFHR5 and factor H in obese PCOS (p < 0.01). Complement factors iC3b, C3d and C5a, associated with an enhanced B cell response and inflammatory cytokine release, were increased in both slim and obese PCOS (p < 0.05). C3a and its product, C3adesArg, were both significantly elevated in nonobese PCOS (<0.01) but not altered in obese PCOS. Hyperandrogenemia correlated positively with properdin and iC3b in obese PCOS (p < 0.05) but not in nonobese PCOS. There was no association with insulin resistance. BMI correlated positively in both groups with factor B, factor H and C5a. Additionally, in obese PCOS, BMI correlated with C3d, factor D, factor I, CFHR5 and C5a (p < 0.05), and in nonobese PCOS, BMI correlated with properdin, iC3b, C3, C3adesArg, C3a, C4, C5, C5a and C1q. In obese controls, BMI correlated with C3, C3desArg, C3a, C3d, C4, factor I, factor B, C5a and C5, whilst in nonobese controls, BMI only correlated negatively with C1q. Comparison of nonobese and obese PCOS showed that properdin, C3b, iC3b, C4A, factor D, factor H and MBL differed. Conclusion: The upregulation of the alternative complement pathway was seen in nonobese PCOS and was further exacerbated in obese PCOS, indicating that this is an inherent feature of the pathophysiology of PCOS that is worsened by obesity and is reflected in the differences between the nonobese and obese PCOS phenotypes. However, the increase in the complement proteins associated with activation was counterbalanced by upregulation of complement inhibitors; this was evident in both PCOS groups, suggesting that insults, such as a cardiovascular event or infection, that cause activation of complement pathways may be amplified in PCOS. Full article