Targeting Protein-protein Interactions Research Articles

Enhanced motivated behavior mediated by pharmacological targeting of the FGF14/Nav1.6 complex in nucleus accumbens neurons

Protein/protein interactions (PPI) play crucial roles in neuronal functions. Yet, their potential as drug targets for brain disorders remains underexplored. The fibroblast growth factor 14 (FGF14)/voltage-gated Na+ channel 1.6 (Nav1.6) complex regulates excitability of medium spiny neurons (MSN) of the nucleus accumbens (NAc), a central hub of reward circuitry that controls motivated behaviors. Here, we identified compound 1028 (IUPAC: ethyl 3-(2-(3-(hydroxymethyl)-1H-indol-1-yl)acetamido)benzoate), a brain-permeable small molecule that targets FGF14R117, a critical residue located within a druggable pocket at the FGF14/Nav1.6 PPI interface. We found that 1028 modulates FGF14/Nav1.6 complex assembly and depolarizes the voltage-dependence of Nav1.6 channel inactivation with nanomolar potency by modulating the intramolecular interaction between the III-IV linker and C-terminal domain of the Nav1.6 channel. Consistent with the compound’s effects on Nav1.6 channel inactivation, 1028 enhances MSN excitability ex vivo and accumbal neuron firing rate in vivo in murine models. Systemic administration of 1028 maintains behavioral motivation preferentially during motivationally deficient conditions in murine models. These behavioral effects were abrogated by in vivo gene silencing of Fgf14 in the NAc and were accompanied by a selective reduction in accumbal dopamine levels during reward consumption in murine models. These findings underscore the potential to selectively regulate complex behaviors associated with neuropsychiatric disorders through targeting of PPIs in neurons.

Peptide design to control protein-protein interactions.

Targeting of protein-protein interactions has become of huge interest in every aspect of medicinal and biological sciences. The control of protein interactions selectively offers the opportunity to control biological processes while limiting off target effects. This interest has massively increased with the development of cryo-EM and protein structure prediction with tools such as RosettaFold and AlphaFold. When designing molecules to control protein interactions, either inhibition or stabilisation, a starting point is commonly peptide design. This tutorial review describes that process, highlighting the selection of an initial sequence with and without structural information. Subsequently, methods for how the sequence can be analysed for key residues and how this information can be used to optimise the ligand efficiency are highlighted. Finally a discussion on how peptides can be further modified to increase their affinity and cell permeability, improving their drug-like properties, is presented.

Targeting protein-protein interactions in drug discovery: Modulators approved or in clinical trials for cancer treatment.

Protein-protein interactions (PPIs) form complex cellular networks fundamental to many key biological processes, including signal transduction, cell proliferation and DNA repair. In consequence, their perturbation is often associated with many human diseases. Targeting PPIs offers a promising approach in drug discovery and ongoing advancements in this field hold the potential to provide highly specific therapies for a wide range of complex diseases. Despite the development of PPI modulators is challenging, advances in the genetic, proteomic and computational level have facilitated their discovery and optimization. Focusing on anticancer drugs, in the last years several PPI modulators have entered clinical trials and venetoclax, which targets Bcl-2 family proteins, has been approved for treating different types of leukemia. This review discusses the clinical development status of drugs modulating several PPIs, such as MDM2-4/p53, Hsp90/Hsp90, Hsp90/CDC37, c-Myc/Max, KRAS/SOS1, CCR5/CCL5, CCR2/CCL2 or Smac/XIAP, in cancer drug discovery.

Exploring potential drug targets for SLE through Mendelian randomization and network pharmacology.

Systemic lupus erythematosus (SLE) is a complex and incurable autoimmune disease, so several drug remission for SLE symptoms have been developed and used at present. However, treatment varies by patient and disease activity, and existing medications for SLE were far from satisfactory. Novel drug targets to be found for SLE therapy are still needed. Mendelian randomization (MR), an observational study way, was performed to explore potential drug targets for SLE using protein quantitative trait loci (pQTL) from recently published genome-wide association studies (GWAS) of cerebrospinal fluid (CSF) and plasma proteins, which obtained genetic instruments for 154 CSF proteins of 971 participants, and 734 plasma proteins of 23591 participants. Bidirectional Mendelian randomization analysis, colocalization analysis, and phenotype scanning were performed to find key proteins for SLE. In addition, external data verification was implemented to further consolidate the Mendelian randomization findings. Candidate proteins as targets to find drugs and discuss the druggability. Finally, Network pharmacology and molecular docking methods were used to verify the effects of Voclosporin and Cyclosporine on SLE targets. Protein-protein interaction (PPI) and core target analysis of candidate drugs and SLE overlapping targets were performed to identify potential hub targets and interactions. The affinity between drug targets and SLE targets was confirmed by molecular docking. In the preliminary analysis, we identified four key proteins as possible drug targets in CSF and plasma proteins, included ICAM-1(P = 4.62E-05, OR = 0.90(0.86, 0.95)), sICAM-1(P = 4.62E-05, OR = 0.49(0.35, 0.69)), FCG2B (P = 7.63E-11, OR = 0.57(0.48, 0.67)), PPP3CA; PPP3R1 (P = 5.47E-07, OR = 0.66(0.57, 0.78)). Among them, ICAM1 was detected in both CSF and plasma proteins. By excluding reverse causality, confounding factors, and linkage disequilibrium (LD), we identified PPP3CA; PPP3R1 as novel drug targets for SLE, including Voclosporin and Cyclosporine. Finally, the Drugbank database shows that novel drugs contain 33 targets for treating SLE. PPI suggested that SIRT1, ACE, PTGS2, and BACE1 were pivotal targets for SLE treatment. In addition, the molecular docking showed that the bioactive molecules of Voclosporin and Cyclosporine had a good affinity with the target of SLE. Our integrative analysis suggested that levels of circulating PPP3CA; PPP3R1 had causal effects on SLE risk and served as potential treatment targets. Moreover, this study provides new evidence for Voclosporin as an SLE treatment through Mendelian randomization and Network pharmacology, and warrants further clinical investigation.

Guanidinium-Stapled Helical Peptides for Targeting Protein-Protein Interactions.

Peptide stapling has emerged as a versatile approach in drug discovery to reinforce secondary structure elements especially α-helices and improve properties of linear bioactive peptides. Inspired by the prevalence of arginine in protein-protein and protein-DNA interfaces, we investigated guanidinium-stapling as a means to constrain helical peptides. Guanidinium stapling was readily achieved on solid support, utilizing two orthogonally protected lysine or unatural α-amino acid residues with an amino function. This method allows for easy modulation of the nature and size of the staple as well as helix propensity. Evaluating a set of guanidinium-stapled peptides for their interaction with different protein targets identified several binders with increased target affinity. X-ray structure determination of four complexes revealed that all stapled peptides adopt a helical conformation upon protein binding. Notably, the disubstituted guanidinium generally exhibits a distinct cis/trans conformation and, in one instance, retains a conserved hydrogen bond with the protein surface. By identifying, for the first time, the guanidinium moiety as an effective helical peptide stapling group, this research significantly expands the repertoire of α-helix stapling techniques for the creation of useful protein mimics.

A split intein and split luciferase-coupled system for detecting protein-protein interactions.

Elucidation of protein-protein interactions (PPIs) represents one of the most important methods in biomedical research. Recently, PPIs have started to be exploited for drug discovery purposes and have thus attracted much attention from both the academic and pharmaceutical sectors. We previously developed a sensitive method, Split Intein-Mediated Protein Ligation (SIMPL), for detecting binary PPIs via irreversible splicing of the interacting proteins being investigated. Here, we incorporated tripart nanoluciferase (tNLuc) into the system, providing a luminescence signal which, in conjunction with homogenous liquid phase operation, improves the quantifiability and operability of the assay. Using a reference PPI set, we demonstrated an improvement in both sensitivity and specificity over the original SIMPL assay. Moreover, we designed the new SIMPL-tNLuc ('SIMPL2') platform with an inherent modularity allowing for flexible measurement of molecular modulators of target PPIs, including inhibitors, molecular glues and PROTACs. Our results demonstrate that SIMPL2 is a sensitive, cost- and labor-effective tool suitable for high-throughput screening (HTS) in both PPI mapping and drug discovery applications.

MiR-1, miR-133a, miR-29b and skeletal muscle fibrosis in chronic limb-threatening ischaemia.

Chronic limb-threatening ischaemia (CLTI), the most severe manifestation of peripheral arterial disease (PAD), is associated with a poor prognosis and high amputation rates. Despite novel therapeutic approaches being investigated, no significant clinical benefits have been observed yet. Understanding the molecular pathways of skeletal muscle dysfunction in CLTI is crucial for designing successful treatments. This study aimed to identify miRNAs dysregulated in muscle biopsies from PAD cohorts. Using MIcroRNA ENrichment TURned NETwork (MIENTURNET) on a publicly accessible RNA-sequencing dataset of PAD cohorts, we identified a list of miRNAs that were over-represented among the upregulated differentially expressed genes (DEGs) in CLTI. Next, we validated the altered expression of these miRNAs and their targets in mice with hindlimb ischaemia (HLI). Our results showed a significant downregulation of miR-1, miR-133a, and miR-29b levels in the ischaemic limbs versus the contralateral non-ischaemic limb. A miRNA target protein-protein interaction network identified extracellular matrix components, including collagen-1a1, -3a1, and -4a1, fibronectin-1, fibrin-1, matrix metalloproteinase-2 and -14, and Sparc, which were upregulated in the ischaemic muscle of mice. This is the first study to identify miR-1, miR-133a, and miR-29b as potential contributors to fibrosis and vascular pathology in CLTI muscle, which supports their potential as novel therapeutic agents for this condition.

Integrative Investigation of Flavonoids Targeting YBX1 Protein-Protein Interaction Network in Breast Cancer: From Computational Analysis to Experimental Validation.

Y-box-binding protein 1 (YBX1) is a multifunctional oncoprotein with its nuclear localization contributing to chemo-resistance in breast cancer. Through its interactions with various proteins and lncRNAs, YBX1 promotes cancer cell migration, invasion, and metastasis. Despite its significant role in cancer progression, studies on YBX1's protein-protein interactions (PPIs) remain limited. Flavonoids are natural compounds with anticancer properties that inhibit metastasis, modulate immunity, and induce apoptosis, with minimal systemic toxicity, making them strong candidates for cancer therapy. Targeting PPIs offers a promising approach for cancer therapy and flavonoids, with their anticancer properties, may modulate these interactions. Our study focused on the YBX1 PPI network, specifically targeting HSPA1A, IGF2BP1, MECP2, G3BP1, EWSR1, PURA, and SYNCRIP. We selected four flavonoids Quercetin, Fisetin, Rutin, and Myricitrin based on literature and conducted 26 docking sessions. Further ADMET analysis indicated Quercetin and Fisetin as more favorable for drug-likeness parameters than Rutin and Myricitrin, which was underscored by MD simulation data. In vitro studies showed that Quercetin and Fisetin downregulated YBX1 expression in a dose-dependent manner (50μM to 150μM) in MCF-7 cells. Our study provides a preliminary understanding of YBX1 PPI and the potential of flavonoids to disrupt these interactions. This study investigates the potential of flavonoids to target YBX1 PPIs, providing insights into novel therapeutic strategies for YBX1-driven cancers.

Exploration of the Active Components and Mechanism of Jiegeng (Platycodonis Radix) in the Treatment of Influenza Virus Pneumonia Through Network Pharmacology Analysis and Experimental Verification.

This study aimed to explore the pathogenesis of platycodin D and luteolin, which are both active components in Jiegeng (Platycodonis Radix), in the treatment of influenza virus pneumonia through network pharmacology analysis combined with experimental verification. The bioactive components of Jiegeng (Platycodonis Radix) were screened by TCMSP and literature mining, and the results were standardized via the UniProt database. The action targets for the disease were identified from databases including OMIM, GeneCards, TTD, DisGeNET, and PharmGKB. Then, the visualized key target regulatory network and protein-protein interaction (PPI) network for the active components were established using Cytoscape3.7.1 software. The findings were illustrated through Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses. The intervention concentrations of platycodin D and luteolin were screened by the CCK8 method, and the important signaling pathways of platycodin D and luteolin for treating influenza virus pneumonia were verified by RT-qPCR and Western blot tests. From data mining, 89 common drug-disease targets were screened out, and five major active components of Jiegeng (Platycodonis Radix), including platycodin D and luteolin, were obtained. Besides, 11 therapeutic targets including IL-17, IL-6, TNF-α, JUN, and MAKP1 were identified by PPI network analysis. GO and KEGG enrichment analyses showed that the pathways most related to the mechanisms of Jiegeng (Platycodonis Radix) against influenza virus pneumonia included the TNF and IL-17 signaling pathways and apoptosis. Invitro experiments demonstrated that the model group exhibited a notable elevation in mRNA levels of IL-6, IL-17, TNF-α, JUN, MAPK1, and the IL-17/-acting protein ratio, as compared to the control group (p < 0.05). In contrast to the model group, the IL-6, IL-17, TNF-α, JUN, MAPK1 mRNA expression levels, and the IL-17 protein ratio in both the platycodin D group and luteolin group were considerably decreased (p < 0.05). Combined with network pharmacology and experimental verification, this study revealed that platycodin D and luteolin in Jiegeng (Platycodonis Radix) may treat influenza virus pneumonia by regulating inflammation through the IL-17 signaling pathway.

Molecular mechanisms of Buqing granule for the treatment of diabetic retinopathy: Network pharmacology analysis and experimental validation.

Diabetic retinopathy (DR) is a common microvascular complication of diabetes mellitus. Its blindness rate is high; therefore, finding a reasonable and safe treatment plan to prevent and control DR is crucial. Currently, there are abundant and diverse research results on the treatment of DR by Chinese medicine Traditional Chinese medicine compounds are potentially advantageous for DR prevention and treatment because of its safe and effective therapeutic effects. To investigate the effects of Buqing granule (BQKL) on DR and its mechanism from a systemic perspective and at the molecular level by combining network pharmacology and in vivo experiments. This study collected information on the drug targets of BQKL and the therapeutic targets of DR for intersecting target gene analysis and protein-protein interactions (PPI), identified various biological pathways related to DR treatment by BQKL through Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses, and preliminarily validated the screened core targets by molecular docking. Furthermore, we constructed a diabetic rat model with a high-fat and high-sugar diet and intraperitoneal streptozotocin injection, and administered the appropriate drugs for 12 weeks after the model was successfully induced. Body mass and fasting blood glucose and lipid levels were measured, and pathological changes in retinal tissue were detected by hematoxylin and eosin staining. ELISA was used to detect the oxidative stress index expression in serum and retinal tissue, and immunohistochemistry, real-time quantitative reverse transcription PCR, and western blotting were used to verify the changes in the expression of core targets. Six potential therapeutic targets of BQKL for DR treatment, including Caspase-3, c-Jun, TP53, AKT1, MAPK1, and MAPK3, were screened using PPI. Enrichment analysis indicated that the MAPK signaling pathway might be the core target pathway of BQKL in DR treatment. Molecular docking prediction indicated that BQKL stably bound to these core targets. In vivo experiments have shown that compared with those in the Control group, rats in the Model group had statistically significant (P < 0.05) severe retinal histopathological damage; elevated blood glucose, lipid, and malondialdehyde (MDA) levels; increased Caspase-3, c-Jun, and TP53 protein expression; and reduced superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) levels, ganglion cell number, AKT1, MAPK1, and MAPK3 protein expression. Compared with the Model group, BQKL group had reduced histopathological retinal damage and the expression of blood glucose and lipids, MDA level, Caspase-3, c-Jun and TP53 proteins were reduced, while the expression of SOD, GSH-Px level, the number of ganglion cells, AKT1, MAPK1, and MAPK3 proteins were elevated. These differences were statistically significant (P < 0.05). BQKL can delay DR onset and progression by attenuating oxidative stress and inflammatory responses and regulating Caspase-3, c-Jun, TP53, AKT1, MAPK1, and MAPK3 proteins in the MAPK signaling pathway mediates these alterations.

Polymers for Disrupting Protein-Protein Interactions: Where Are We and Where Should We Be?

Protein-protein interactions (PPIs) are central to the cellular signaling and regulatory networks that underlie many physiological and pathophysiological processes. It is challenging to target PPIs using traditional small molecule or peptide-based approaches due to the frequent lack of well-defined binding pockets at the large and flat PPI interfaces. Synthetic polymers offer an opportunity to circumvent these challenges by providing unparalleled flexibility in tuning their physiochemical properties to achieve the desired binding properties. In this review, we summarize the current state of the field pertaining to polymer-protein interactions in solution, highlighting various polyelectrolyte systems, their tunable parameters, and their characterization. We provide an outlook on how these architectures can be improved by incorporating sequence control, foldability, and machine learning to mimic proteins at every structural level. Advances in these directions will enable the design of more specific protein-binding polymers and provide an effective strategy for targeting dynamic proteins, such as intrinsically disordered proteins.

Discovery of Inhibitors Targeting Protein-Protein Interaction between Bacterial RNA Polymerase and NusG as Novel Antimicrobials.

Bacterial RNA polymerase (RNAP), the core enzyme responsible for bacterial transcription, requires the NusG factor for efficient transcription elongation and termination. As the primary binding site for NusG, the RNAP clamp-helix (CH) domain represents a potential protein-protein interaction (PPI) target for novel antimicrobial agent design and discovery. In this study, we designed a pharmacophore model based on the essential amino acids of the CH for binding to NusG, such as R270, R278, and R281 (Escherichia coli numbering), and identified a hit compound with mild antimicrobial activity. Subsequent rational design and synthesis of this hit compound led to improved antimicrobial activity against Streptococcus pneumoniae, with the minimum inhibitory concentration (MIC) reduced from 128 to 1 μg/mL. Additional characterization of the antimicrobial activity, inhibitory activity against RNAP-NusG interaction, and cell-based transcription and fluorescent assays of the optimized compounds demonstrated their potential for further lead optimization.

A Novel Confocal Scanning Protein-Protein Interaction Assay (PPI-CONA) Reveals Exceptional Selectivity and Specificity of CC0651, a Small Molecule Binding Enhancer of the Weak Interaction between the E2 Ubiquitin-Conjugating Enzyme CDC34A and Ubiquitin.

Protein-protein interactions (PPIs) are some of the most challenging target classes in drug discovery. Highly sensitive detection techniques are required for the identification of chemical modulators of PPIs. Here, we introduce PPI confocal nanoscanning (PPI-CONA), a miniaturized, microbead based high-resolution fluorescence imaging assay. We demonstrate the capabilities of PPI-CONA by detecting low affinity ternary complex formation between the human CDC34A ubiquitin-conjugating (E2) enzyme, ubiquitin, and CC0651, a small molecule enhancer of the CDC34A-ubiquitin interaction. We further exemplify PPI-CONA with an E2 enzyme binding study on CC0651 and a CDC34A binding specificity study of a series of CC0651 analogues. Our results indicate that CC0651 is highly selective toward CDC34A. We further demonstrate how PPI-CONA can be applied to screening very low affinity interactions. PPI-CONA holds potential for high-throughput screening for modulators of PPI targets and characterization of their affinity, specificity, and selectivity.

The immunotherapy mechanism of Hedyotis Diffusae Herba in treating liver cancer: a study based on network pharmacology, bioinformatics, and experimental validation.

Liver cancer is a malignant tumor that develops on or inside the liver. Hedyotis diffusa Willd (HDW) plays a significant role in anti-tumor activities; however, its mechanism against liver cancer remains unclear. This study aims to evaluate the immunotherapeutic mechanism of HDW in treating liver cancer through network pharmacology, bioinformatics analysis, and experimental validation. Network pharmacology was utilized to identify the active components and potential targets of HDW from the TCMSP database. A potential target protein-protein interaction (PPI) network was constructed using the STRING database, followed by function and pathway enrichment analysis of the targets using GO and KEGG methods. In addition, the key targets for HDW against liver cancer were identified using five different algorithms in Cytoscape. The TCGA and HPA databases were used to assess the mRNA and protein expression of core target genes in normal liver and liver cancer tissues and their relationship with overall survival in liver cancer, as well as their role in immune infiltration. Molecular docking between the core components of HDW and the core targets was performed using PyMOL software. The effects of HDW on the proliferation and apoptosis of liver cancer cells were examined using MTT and flow cytometry. The regulatory effects of the core component quercetin on core targets were validated using RT-qPCR and Western blot. A total of 163 potential targets were identified by searching for intersections among 7 types of active components and all potential and liver cancer targets. PPI network analysis revealed the core targets IL6 and TNF. GO enrichment analysis involved 2089 biological processes, 76 cellular components, and 196 molecular functions. KEGG enrichment analysis suggested that the anti-cancer effects of HDW might be mediated by the AGE-RAGE signaling pathway, IL-17 signaling pathway, TNF signaling pathway, PI3K-Akt signaling pathway, and NF-κB signaling pathway. Database validation of key targets showed that mRNA and protein expression results for the IL6 gene were contradictory, while those for the TNF gene were consistent, both being underexpressed in liver cancer. Importantly, the expression of IL6 and TNF was related to the infiltration of 24 types of immune cells, with the highest correlation with macrophages. Molecular docking showed that IL6 and TNF had high binding stability with quercetin, with binding energies of - 7.4 and - 6.0kJ∙mol-1, respectively. Experimental validation showed that quercetin inhibited liver cancer cell proliferation and promoted apoptosis in a dose-dependent manner, with protein results indicating that quercetin downregulated the mRNA and protein expression of IL6 and TNF, and upregulated key proteins in the AGE-RAGE signaling pathway, AGEs, and RAGE. This study comprehensively elucidates the activity, potential targets, and molecular mechanisms of HDW against liver cancer, providing a promising strategy for the scientific basis and treatment mechanism of traditional Chinese medicine in treating liver cancer.

Graph Association Analyses for Early Drug Discovery

We demonstrate MedHunter, a system for assisting the early stage of drug development. MedHunter builds a biomedical knowledge graph DDKG by integrating data from eleven biochemical libraries and data banks, and aligning entities from different data sources by means of heterogeneous entity resolution. It identifies drug-disease associations and protein-protein interactions in DDKG by employing graph association rules (GARs). GARs use graph patterns to extract relevant entities and embed ML models as predicates. MedHunter discovers GARs from DDKG and incrementally enriches DDKG with external data; it cleans DDKG with a special form of GARs. We demonstrate MedHunter for its (a) interfaces, (b) data enrichment/cleaning, and (c) applications in target identification, drug-drug interaction and protein-protein interaction.

Impact of the near-physiological temperature on the in vitro maturation of bovine oocytes: A comparative proteomic approach

In vivo, the temperature inside preovulatory follicles of cows is approximately 1 °C lower than rectal temperature. However, standard bovine oocyte in vitro maturation (IVM) protocols use 38.5 °C based on rectal temperature. This study evaluated the effect of reducing IVM temperature to 37.5 °C on the proteomic profile of oocytes compared to the routine 38.5 °C. Nuclear maturation rate and cumulus cell (CC) expansion (30 COCs per group, 21 replicates) were assessed by observing the first polar body and using a subjective scoring method (0–4). Total nitrite concentrations in the culture medium were measured using the Griess method. Differential proteomics was performed using LC-MS/MS on pooled oocyte samples (500 matured oocytes per group, three replicates), followed by gene ontology enrichment, protein-protein interaction, and putative miRNA target analyses. No significant differences were observed between the groups in nuclear maturation, CC expansion, or nitrite concentration (P > 0.05). A total of 806 proteins were identified, with 7 up-regulated and 12 down-regulated in the treatment group compared to the control. Additionally, 12 proteins were unique to the control group, and 8 were unique to the treatment group. IVM at 37.5 °C resulted in the upregulation of proteins involved in protein folding and GTP binding, and the downregulation of enzymes with oxidoreductase activity and proteins involved in cytoskeletal fiber formation. Furthermore, 43 bovine miRNAs potentially regulating these genes (DES, HMOX2, KRT75, FARSA, IDH2, CARHSP1) were identified. We conclude that IVM of bovine oocytes at 37.5 °C induces significant proteomic changes without impacting nuclear maturation, cumulus cell expansion, or nitrite concentration in the IVM medium.

Combining Network Pharmacology and Experimental Verification to Ascertain the Mechanism of Action of Asparagus officinalis Against the Brain Damage Caused by Fluorosis.

Asparagus officinalis (ASP) has antioxidation, anti-inflammatory, antiaging, and immune system-enhancing effects. We explored the preventive and therapeutic consequences of ASP on the brain damage elicited by fluorosis through network pharmacology and in vivo experimental validation. We ascertained the pharmaceutically active ingredients and drug targets of ASP from the Traditional Chinese Medicine Systems Pharmacology database, predicted the disease targets of fluorosis-induced brain injury using GeneCards and Online Mendelian Inheritance in Man databases, obtained target protein-protein interaction networks in the Search Tool for the Retrieval of Interacting Genes/Proteins database, used Cytoscape to obtain key targets and active ingredients, and conducted enrichment analyses of key targets in the Database for Annotation, Visualization and Integrated Discovery. Enrichment analyses showed that "mitogen-activated protein kinase" (MAPK), "phosphoinositide 3-kinase/protein kinase B" (PI3K-Akt), "nuclear factor-kappa B" (NF-κB), and the "neurotrophin signaling pathway" were the most enriched biological processes and signaling pathways. ASP could alleviate fluorosis-based injury, improve brain-tissue damage, increase urinary fluoride content, and improve oxidation levels and inflammatory-factor levels in the body. ASP could also reduce dental fluorosis, bone damage, fluoride concentrations in blood and bone, and accumulation of lipid peroxide. Upon ASP treatment, expression of silent information regulator (SIRT)1, brain-derived neurotrophic factor (BDNF), tropomyosin receptor kinase B (TrkB), MAPK, NF-κB, PI3K, Akt, and B-cell lymphoma-2 in rat brain tissue increased gradually, whereas that of Bax, caspase-3, and p53 decreased gradually. We demonstrated that ASP could regulate the brain damage caused by fluorosis through the SIRT1/BDNF/TrkB signaling pathway, and reported the possible part played by ASP in preventing and treating fluorosis.

Targeting protein-protein interactions for therapeutic intervention in Parkinson's disease to delay progression.

Background: Parkinson's disease (PD) is a neurodegenerative condition with few treatments to slow or stop development. Protein-protein interactions (PPIs) are intriguing therapeutic targets. This research examined the safety and effectiveness of a new Parkinson's disease medication targeting PPIs. Methods: After enrollment, 60 individuals were randomly assigned to the treatment and control groups. MDS-UPDRS Part III score change from baseline to week 12 was the main outcome measure. Secondary outcome measures were the Hoehn and Yahr scale, NMSS, and Montreal Cognitive Assessment. Study-wide adverse events were tracked. Results: Compared to the control group, the treatment group exhibited a substantial improvement in MDS-UPDRS Part III score (p &lt; 0.001). Additionally, the therapy group showed substantial improvements in Hoehn and Yahr stage, NMSS score, and MoCA score compared to the control group (p &lt; 0.001). No significant adverse effects were documented with the experimental medication. Conclusion: This research suggests that targeting PPIs may treat Parkinson's disease. No harmful side effects were detected with the experimental medication, which improved motor and non-motor symptoms in PD patients. These results require more study to determine the long-term safety and effectiveness of targeting PPIs in Parkinson's disease.

Mechanism of ergosterol peroxide on MCF-7 breast cancer cells based on network pharmacology and in vitro experiments

This study investigated the effects of ergosterol peroxide(EP) on the proliferation and apoptosis of MCF-7 breast cancer cells, explored its possible mechanisms of action, and verified the effects and mechanisms by in vitro experiments. Network pharmaco-logy was used to screen the target proteins of EP and construct target networks and protein-protein interaction(PPI) networks to predict the potential target proteins and related pathways involved in EP anti-breast cancer effects. The MTT assay was performed to measure the inhibitory effect of EP on MCF-7 cell proliferation, and the colony formation assay was used to assess the cell cloning ability. Flow cytometry and laser confocal microscopy were employed to evaluate cell apoptosis, mitochondrial membrane potential and reactive oxygen species(ROS) levels. Western blot analysis was conducted to examine the expression levels of B-cell lymphoma 2(Bcl-2), Bcl-2-associated X protein(Bax), cytochrome C(Cyt C), caspase-7, cleaved caspase-7, phosphatidylinositol 3-kinase(PI3K), and se-rine/threonine kinase B(AKT) in MCF-7 cells treated with EP. The results of network pharmacology prediction yielded 173 common targets between EP and breast cancer; the results of Kyoto Encyclopedia of Genes and Genomes(KEGG) enrichment analysis showed that EP treatment for breast cancer mainly affected the signaling pathways such as cancer pathway, PI3K-AKT signaling pathway, cellular senescence signaling pathway, and viral carcinogenesis pathway; and the MTT assay results showed that the viability of MCF-7 cells in the EP group was significantly lower than that in the control group, exhibiting a time-and concentration-dependent trend, and EP can inhibit colony formation of MCF-7 breast cancer cells. Treatment with 10, 20, and 40 μmol·L~(-1) EP for 24 h resulted in a significant increase in the total apoptosis rate of MCF-7 cells, a significant decrease in mitochondrial membrane potential, and a significant increase in ROS levels. In addition, treatment with EP led to an upregulation of Cyt C, Bax, and cleaved caspase-7 protein expression, and a downregulation of p-PI3K, p-AKT, and Bcl-2 protein expression in MCF-7 cells. Studies have shown that EP inhibits MCF-7 breast cancer cell proliferation and reduces colony formation by a mechanism that may be related to the PI3K-AKT pathway mediating the mitochondrial apoptotic pathway.

Research on the mechanism of Ursolic acid for treating Parkinson's disease by network pharmacology and experimental verification

The objective of this study was to investigate the potential targets and mechanisms of UA in the treatment of PD. The efficacy of UA in PD was assessed through network pharmacology, molecular docking, and experimental methods. Common target protein-protein interaction (PPI) networks were constructed and visualized using Cytoscape. As a result, 9 key genes, namely CASP3, IL6, IL1B, PTGS2, CREB1, TNF, MAPK3, JUN, and CASP8, were selected. Molecular docking simulations were performed using Discovery Studio 2019 to validate the correlation between UA and the core targets. The results demonstrated a favorable binding affinity between UA and CASP8, IL1B, CASP3, TNF, MAPK3 and IL6. In vivo studies showed UA ameliorated motor dysfunction, and UA can significantly increase the protein expression of tyrosine hydroxylase (TH) in PD mice model. In addition, in vitro experiments confirmed that UA effectively reduced the protein expression of CASP8, CASP3 and MAPK3 in PD cell models and suppressed the gene expression of TNF-α, IL-6, and IL-1β. These findings indicate that the therapeutic effects of UA on PD could be due to its influence on various targets within both the apoptosis and neuroinflammatory signaling pathways. Consequently, this study provides a methodological and theoretical foundation for further elucidating the pharmacological mechanism of UA.