(Homo-)harringtonine prevents endothelial inflammation through IRF-1 dependent downregulation of VCAM1 mRNA expression and inhibition of cell adhesion molecule protein biosynthesis.

The plant alkaloid homoharringtonine (HHT) is a Food and Drug Administration (FDA)-approved drug for the treatment of hematologic malignancies. In addition to its well-established antitumor activity, accumulating evidence attributes anti-inflammatory effects to HHT, which have mainly been studied in leukocytes to date. However, a potential influence of HHT on inflammatory activation processes in endothelial cells, which are a key feature of inflammation and a prerequisite for the leukocyte-endothelial cell interaction and leukocyte extravasation, remains poorly understood. In this study, the anti-inflammatory potential of HHT and its derivative harringtonine (HT) on the TNF-induced leukocyte-endothelial cell interaction was assessed, and the underlying mechanistic basis of these effects was elucidated. HHT affected inflammation in vivo in a murine peritonitis model by reducing leukocyte infiltration and proinflammatory cytokine expression as well as ameliorating abdominal pain behavior. In vitro, HT and HHT impaired the leukocyte-endothelial cell interaction by decreasing the expression of the endothelial cell adhesion molecules intracellular adhesion molecule -1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). This effect was mediated by a bipartite mechanism. While HHT did not affect the prominent TNF-induced pro-inflammatory NF-ĸB signaling cascade, the compound downregulated the VCAM1 mRNA expression in an IRF-1-dependent manner and diminished active ICAM1 mRNA translation as determined by polysome profiling. This study highlights HHT as an anti-inflammatory compound that efficiently hampers the leukocyte-endothelial cell interaction by targeting endothelial activation processes.

HPLC- and NMR-Based Chemical Profiling, Wound-Healing Potential, Anti-Inflammatory and Antibacterial Activities of Satureja pilosa (Lamiaceae), a Neglected Medicinal-Aromatic Herb.

Satureja pilosa Velen. (Lamiaceae) is a perennial and melliferous aromatic-medicinal subshrub which is range-restricted in adjacent parts of Greece and Bulgaria and locally in Italy, known in Northern Greece as wild oregano ("agriorigani") and traditionally collected from the wild for culinary purposes. Since the ethnopharmacological data and modern biological activities of Satureja spp. suggest promising applications in skin conditions, the present study aimed to investigate the hitherto unknown phenolic content of cultivated S. pilosa and its potential biological activities, focusing mainly on wound-healing and anti-inflammatory effects. An HPLC-PDA-MS-targeted phytochemical investigation, along with NMR, allowed for the isolation and characterization of the main constituents, resulting in 18 compounds. Representative extracts and purified compounds were tested for wound-healing activity on NIH/3T3 fibroblasts. The butanol extract exhibited a significantly higher cell migration rate (73.4%) compared to aqueous (50.6%) and methanolic (49.6%) ones, enhancing the cell migration more rapidly at both concentration levels, whilst rosmarinic acid was the most potent among the isolated compounds, with a migration rate of 64.0% at the concentration level of 10-5 mg/mL, followed by 3,4-dihydrophenyllactic acid (54.7%). Moreover, potential effects on endothelial activation processes were explored, including the leukocyte-endothelial cell interaction during inflammatory processes and the migratory capacity during angiogenic actions, since these processes are commonly associated with skin diseases. Finally, extracts and purified compounds demonstrated weak antibacterial potential against two important pathogens (Staphylococcus aureus and Pseudomonas aeruginosa), suggesting that further investigation is warrented.

Venomics of the milos viper (Macrovipera schweizeri) unveils patterns of venom composition and exochemistry across blunt-nosed viper venoms.

Introduction: Snakebite is a neglected tropical disease and a globally important driver of death and morbidity. Vipers of the genus Macrovipera (Viperidae: Viperinae) are among the snakes of higher medical importance in the Old World. Despite the medical relevance of Macrovipera venoms, the knowledge regarding them is heterogeneously distributed with virtually all works conducted so far focusing on subspecies of Macrovipera lebetinus, while other species within the genus are largely overlooked. Here we present the first proteomic evaluation of the venom from the Greek endemic Milos viper (Macrovipera schweizeri). In line with clinical symptoms typically elicited by Macrovipera envenomations, Milos viper venom primarily comprises coagulotoxic and cytotoxic protein families, such as metalloproteinases (svMP) and serine proteases (svSP). Methods: We conducted comparative bioactivity assays on venoms from M. schweizeri and the M. lebetinus subspecies M. lebetinus cernovi, M. lebetinus obtusa, and M. lebetinus turanica, and showed that they all exhibit similarities in levels of cytotoxicity proteolytic activity, and inhibition of prokaryotic growth. Lastly, we compared Macrovipera venom profiles by 1D-SDS-PAGE and RP-HPLC, as well as our proteomic data with previously published Macrovipera venom proteomes. Results and discussion: The analyzes performed to reveal that a general venom profile seems to be conserved across blunt-nosed vipers, and that, M. schweizeri envenomations, similarly to those caused by other blunt-nosed vipers, are able to cause significant tissue damage. The present work represents an important starting point for the development of comparative studies across the full taxonomic range of the genus Macrovipera and can potentially help optimize the treatment of envenomations caused by M. schweizeri.

The Microtubule-Targeting Agent Pretubulysin Impairs the Inflammatory Response in Endothelial Cells by a JNK-Dependent Deregulation of the Histone Acetyltransferase Brd4.

The anti-inflammatory effects of depolymerizing microtubule-targeting agents on leukocytes are known for a long time, but the potential involvement of the vascular endothelium and the underlying mechanistic basis is still largely unclear. Using the recently synthesized depolymerizing microtubule-targeting agent pretubulysin, we investigated the anti-inflammatory potential of pretubulysin and other microtubule-targeting agents with respect to the TNF-induced leukocyte adhesion cascade in endothelial cells, to improve our understanding of the underlying biomolecular background. We found that treatment with pretubulysin reduces inflammation in vivo and in vitro via inhibition of the TNF-induced adhesion of leukocytes to the vascular endothelium by down-regulation of the pro-inflammatory cell adhesion molecules ICAM-1 and VCAM-1 in a JNK-dependent manner. The underlying mechanism includes JNK-induced deregulation and degradation of the histone acetyltransferase Bromodomain-containing protein 4. This study shows that depolymerizing microtubule-targeting agents, in addition to their established effects on leukocytes, also significantly decrease the inflammatory activation of vascular endothelial cells. These effects are not based on altered pro-inflammatory signaling cascades, but require deregulation of the capability of cells to enter constructive transcription for some genes, setting a baseline for further research on the prominent anti-inflammatory effects of depolymerizing microtubule-targeting agents.

Integrating Siderophore Substructures in Thiol-Based Metallo-ß-Lactamase Inhibitors.

Metallo beta lactamases (MBLs) are among the most problematic resistance mechanisms of multidrug-resistant Gram-negative pathogens due to their broad substrate spectrum and lack of approved inhibitors. In this study, we propose the integration of catechol substructures into the design of thiol-based MBL inhibitors, aiming at mimicking bacterial siderophores for the active uptake by the iron acquisition system of bacteria. We synthesised two catechol-containing MBL inhibitors, as well as their dimethoxy counterparts, and tested them for in vitro inhibitory activity against NDM-1, VIM-1, and IMP-7. We demonstrated that the most potent catechol-containing MBL inhibitor is able to bind Fe3+ ions. Finally, we could show that this compound restores the antibiotic activity of imipenem in NDM-1-expressing K. pneumoniae, while leaving HUVEC cells completely unaffected. Thus, siderophore-containing MBL inhibitors might be a valuable strategy to overcome bacterial MBL-mediated resistance to beta lactam antibiotics.

The Pharmacological Potential of Novel Melittin Variants from the Honeybee and Solitary Bees against Inflammation and Cancer.

The venom of honeybees is composed of numerous peptides and proteins and has been used for decades as an anti-inflammatory and anti-cancer agent in traditional medicine. However, the bioactivity of specific biomolecular components has been evaluated for the predominant constituent, melittin. So far, only a few melittin-like peptides from solitary bee species have been investigated, and the molecular mechanisms of bee venoms as therapeutic agents remain largely unknown. Here, the preclinical pharmacological activities of known and proteo-transcriptomically discovered new melittin variants from the honeybee and more ancestral variants from phylogenetically older solitary bees were explored in the context of cancer and inflammation. We studied the effects of melittin peptides on cytotoxicity, second messenger release, and inflammatory markers using primary human cells, non-cancer, and cancerous cell lines. Melittin and some of its variants showed cytotoxic effects, induced Ca2+ signaling and inhibited cAMP production, and prevented LPS-induced NO synthesis but did not affect the IP3 signaling and pro-inflammatory activation of endothelial cells. Compared to the originally-described melittin, some phylogenetically more ancestral variants from solitary bees offer potential therapeutic modalities in modulating the in vitro inflammatory processes, and hindering cancer cell viability/proliferation, including aggressive breast cancers, and are worth further investigation.

Atropopeptides are a Novel Family of Ribosomally Synthesized and Posttranslationally Modified Peptides with a Complex Molecular Shape.

Biomacromolecules are known to feature complex three-dimensional shapes that are essential for their function. Among natural products, ambiguous molecular shapes are a rare phenomenon. The hexapeptide tryptorubin A can adopt one of two unusual atropisomeric configurations. Initially hypothesized to be a non-ribosomal peptide, we show that tryptorubin A is the first characterized member of a new family of ribosomally synthesized and posttranslationally modified peptides (RiPPs) that we named atropopeptides. The sole modifying enzyme encoded in the gene cluster, a cytochrome P450 monooxygenase, is responsible for the atropospecific formation of one carbon-carbon and two carbon-nitrogen bonds. The characterization of two additional atropopeptide biosynthetic pathways revealed a two-step maturation process. Atropopeptides promote pro-angiogenic cell functions as indicated by an increase in endothelial cell proliferation and undirected migration. Our study expands the biochemical space of RiPP-modifying enzymes and paves the way towards the chemoenzymatic utilization of atropopeptide-modifying P450s.

Detailed Characterization of Small Extracellular Vesicles from Different Cell Types Based on Tetraspanin Composition by ExoView R100 Platform.

Small extracellular vesicles (sEV) hold enormous potential as biomarkers, drug carriers, and therapeutic agents. However, due to previous limitations in the phenotypic characterization of sEV at the single vesicle level, knowledge of cell type-specific sEV signatures remains sparse. With the introduction of next-generation sEV analysis devices, such as the single-particle interferometric reflectance imaging sensor (SP-IRIS)-based ExoView R100 platform, single sEV analyses are now possible. While the tetraspanins CD9, CD63, and CD81 were generally considered pan-sEV markers, it became clear that sEV of different cell types contain several combinations and amounts of these proteins on their surfaces. To gain better insight into the complexity and heterogeneity of sEV, we used the ExoView R100 platform to analyze the CD9/CD63/CD81 phenotype of sEV released by different cell types at a single sEV level. We demonstrated that these surface markers are sufficient to distinguish cell-type-specific sEV phenotypes. Furthermore, we recognized that tetraspanin composition in some sEV populations does not follow a random pattern. Notably, the tetraspanin distribution of sEV derived from mesenchymal stem cells (MSCs) alters depending on cell culture conditions. Overall, our data provide an overview of the cell-specific characteristics of sEV populations, which will increase the understanding of sEV physiology and improve the development of new sEV-based therapeutic approaches.

Designing a Small Fluorescent Inhibitor to Investigate Soluble Epoxide Hydrolase Engagement in Living Cells.

Soluble epoxide hydrolase (sEH) is a promising target for a number of inflammation-related diseases. In addition, inhibition of sEH has been shown to reduce neuroinflammation, which plays a critical role in the development of central nervous system (CNS) diseases such as Alzheimer's disease. In this study, we present the rational design of a small fluorescent sEH inhibitor. Starting from the clinical candidate GSK2256294A, we replaced the triazine moiety with the 4-chloro-7-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl) fluorophore. The resulting fluorescent sEH inhibitor displayed excellent potency in an in vitro enzyme activity assay (IC50 < 2 nM). The developed inhibitor is applicable in a NanoBRET-based assay system suitable for studying sEH target engagement in living cells. Furthermore, the inhibitor can be used to visualize sEH in sEH-transfected HEK293 cells and in primary mouse astrocytes by fluorescence microscopy.

Human 5-lipoxygenase regulates transcription by association to euchromatin.

Human 5-lipoxygenase (5-LO) is the key enzyme of leukotriene biosynthesis, mostly expressed in leukocytes and thus a crucial component of the innate immune system. In this study, we show that 5-LO, besides its canonical function as an arachidonic acid metabolizing enzyme, is a regulator of gene expression associated with euchromatin. By Crispr-Cas9-mediated 5-LO knockout (KO) in MonoMac6 (MM6) cells and subsequent RNA-Seq analysis, we identified 5-LO regulated genes which could be clustered to immune/defense response, cell adhesion, transcription and growth/developmental processes. Analysis of differentially expressed genes identified cyclooxygenase-2 (COX2, PTGS2) and kynureninase (KYNU) as strongly regulated 5-LO target genes. 5-LO knockout affected MM6 cell adhesion and tryptophan metabolism via inhibition of the degradation of the immunoregulator kynurenine. By subsequent FAIRE-Seq and 5-LO ChIP-Seq analyses, we found an association of 5-LO with euchromatin, with prominent 5-LO binding to promoter regions in actively transcribed genes. By enrichment analysis of the ChIP-Seq results, we identified potential 5-LO interaction partners. Furthermore, 5-LO ChIP-Seq peaks resemble patterns of H3K27ac histone marks, suggesting that 5-LO recruitment mainly takes place at acetylated histones. In summary, we demonstrate a noncanonical function of 5-LO as transcriptional regulator in monocytic cells.

The protein biosynthesis inhibitor vioprolide A evokes anti-angiogenic and pro-survival actions by targeting NOP14 and decreasing VEGF receptor 2- and TAZ-signaling.

Angiogenesis contributes to the progression of several diseases including cancer or age-related macular degeneration and is crucially driven by pathologically hyperactive endothelial cells (ECs). Targeting angiogenic processes in ECs thus represents a promising strategy to treat these conditions. Vioprolide A (vioA) is a myxobacterial cyclic depsipeptide that targets the nucleolar protein 14 (NOP14) and possesses strong anti-cancer and anti-inflammatory actions. Here, we present evidence that vioA promotes anti-angiogenic actions in vivo and in ECs in vitro. VioA reduced the choroidal neovascularization after laser-induced photocoagulation in mice in vivo, the sprouting of choroidal explant cultures ex vivo and key angiogenic features of ECs in vitro. Mechanistically, vioA decreased VEGFR2 protein levels and phosphorylation leading to impaired downstream pro-angiogenic signaling. Concurrently, vioA influenced TAZ signaling by diminishing its nuclear translocation and protein level, resulting in a reduced expression of pro-angiogenic target genes and dynamic cytoskeletal remodeling. Surprisingly, vioA induced pro-survival signaling in ECs by activating Akt and inhibiting p53-dependent apoptosis. Knockdown of the cellular target NOP14 further revealed a partial involvement in the anti-angiogenic and pro-survival actions of vioA. Taken together, our study introduces vioA as an interesting anti-angiogenic compound that warrants further investigations in preclinical studies.

Ginger Constituent 6-Shogaol Inhibits Inflammation- and Angiogenesis-Related Cell Functions in Primary Human Endothelial Cells.

Rhizomes from Zingiber officinale Roscoe are traditionally used for the treatment of a plethora of pathophysiological conditions such as diarrhea, nausea, or rheumatoid arthritis. While 6-gingerol is the pungent principle in fresh ginger, in dried rhizomes, 6-gingerol is dehydrated to 6-shogaol. 6-Shogaol has been demonstrated to exhibit anticancer, antioxidative, and anti-inflammatory actions more effectively than 6-gingerol due to the presence of an electrophilic Michael acceptor moiety. In vitro, 6-shogaol exhibits anti-inflammatory actions in a variety of cell types, including leukocytes. Our study focused on the effects of 6-shogaol on activated endothelial cells. We found that 6-shogaol significantly reduced the adhesion of leukocytes onto lipopolysaccharide (LPS)-activated human umbilical vein endothelial cells (HUVECs), resulting in a significantly reduced transmigration of THP-1 cells through an endothelial cell monolayer. Analyzing the mediators of endothelial cell-leukocyte interactions, we found that 30 µM of 6-shogaol blocked the LPS-triggered mRNA and protein expression of cell adhesion molecules. In concert with this, our study demonstrates that the LPS-induced nuclear factor κB (NFκB) promoter activity was significantly reduced upon treatment with 6-shogaol. Interestingly, the nuclear translocation of p65 was slightly decreased, and protein levels of the LPS receptor Toll-like receptor 4 remained unimpaired. Analyzing the impact of 6-shogaol on angiogenesis-related cell functions in vitro, we found that 6-shogaol attenuated the proliferation as well as the directed and undirected migration of HUVECs. Of note, 6-shogaol also strongly reduced the chemotactic migration of endothelial cells in the direction of a serum gradient. Moreover, 30 µM of 6-shogaol blocked the formation of vascular endothelial growth factor (VEGF)-induced endothelial sprouts from HUVEC spheroids and from murine aortic rings. Importantly, this study shows for the first time that 6-shogaol exhibits a vascular-disruptive impact on angiogenic sprouts from murine aortae. Our study demonstrates that the main bioactive ingredient in dried ginger, 6-shogaol, exhibits beneficial characteristics as an inhibitor of inflammation- and angiogenesis-related processes in vascular endothelial cells.

Lecanoric acid mediates anti-proliferative effects by an M phase arrest in colon cancer cells.

Lichen extracts containing, among other compounds, depsides such as evernic acid, atranorin, and lecanoric acid possess anti-proliferative effects. We aimed to identify lichen metabolites that are responsible for the observed anti-proliferative effects. We performed cytotoxicity, cell colony, cell cycle and apoptosis assays in various cell lines or primary immune cells. We analyzed several cell cycle proteins and apoptosis-related proteins to gain insights into the underlying mechanism. All depsides reduced the viability of the tested cell lines (HCT-116, HEK293T, HeLa, NIH3T3, RAW246.7) in a cell line-dependent manner with lecanoric acid being the most effective. Atranorin did not influence the cell cycle or colony formation in HCT-116 cells, but induced apoptosis in HCT-116 cells. Evernic acid showed no anti-proliferative effects. Lecanoric acid inhibited cell colony formation already at 0.03 µg/ml in HCT-116 cells and induced a G2 cell cycle block in several cell lines. Moreover, lecanoric acid arrested the cell cycle, presumably in the M phase, since expression of cyclin B1 and phosphorylated histone H3 was upregulated, whereas the inactive cyclin-dependent kinase 1 (CDK1) was reduced in HCT-116 cells. Most importantly, cell death induced by lecanoric acid was more prominent in cancer cells than in primary human immune and endothelial cells. In conclusion, lecanoric acid seems to mediate its anti-proliferative effects via arrest of cells in the M phase. Our data suggest lecanoric acid may be a potential new candidate for anti-cancer therapy, because it has anti-proliferative effects on cancer cell lines, and does not affect primary immune cells.

Nitroxoline and its derivatives are potent inhibitors of metallo-ß-lactamases.

Carbapenemases such as metallo-ß-lactamases (MBLs) are spreading among Gram-negative bacterial pathogens. Infections due to these multidrug-resistant bacteria constitute a major global health challenge. Therapeutic strategies against carbapenemase producing bacteria include ß-lactamase inhibitor combinations. Nitroxoline is a broad-spectrum antibiotic with restricted indication for urinary tract infections. In this study, we report on nitroxoline as an inhibitor of MBLs. We investigate the structure-activity relationships of nitroxoline derivatives considering in vitro MBL inhibitory potency in a fluorescence based assay using purified recombinant MBLs, NDM-1 and VIM-1. We investigated the most potent nitroxoline derivative in combination with imipenem against clinical isolates as well as transformants producing MBL by broth microdilution and time-kill kinetics. Our findings demonstrate that nitroxoline derivatives are potent MBL inhibitors and in combination with imipenem overcome MBL-mediated carbapenem resistance.

The natural product vioprolide A exerts anti-inflammatory actions through inhibition of its cellular target NOP14 and downregulation of importin-dependent NF-ĸB p65 nuclear translocation.

Chronic inflammation is characterized by persisting leukocyte infiltration of the affected tissue, which is enabled by activated endothelial cells (ECs). Chronic inflammatory diseases remain a major pharmacotherapeutic challenge, and thus the search for novel drugs and drug targets is an ongoing demand. We have identified the natural product vioprolide A (vioA) to exert anti-inflammatory actions in vivo and in ECs in vitro through inhibition of its cellular target nucleolar protein 14 (NOP14). VioA attenuated the infiltration of microglia and macrophages during laser-induced murine choroidal neovascularization and the leukocyte trafficking through the vascular endothelium in the murine cremaster muscle. Mechanistic studies revealed that vioA downregulates EC adhesion molecules and the tumor necrosis factor receptor (TNFR) 1 by decreasing the de novo protein synthesis in ECs. Most importantly, we found that inhibition of importin-dependent NF-ĸB p65 nuclear translocation is a crucial part of the action of vioA leading to reduced NF-ĸB promotor activity and inflammatory gene expression. Knockdown experiments revealed a causal link between the cellular target NOP14 and the anti-inflammatory action of vioA, classifying the natural product as unique drug lead for anti-inflammatory therapeutics.

Discovery of Irbesartan Derivatives as BLT2 Agonists by Virtual Screening.

Leuktriene B4 receptor 2 (BLT2) is a G-protein coupled receptor modulation of which is discussed to be a therapeutic option for healing of intestinal lesions. In this work, new BLT2 agonists were identified by a virtual screening of a repurposing library and in vitro assay of the most promising compounds. Irbesartan, an approved type-1 angiotensin II receptor (AT1) antagonist, was identified as a moderate BLT2 agonist. An initial SAR study on the irbesartan scaffold was performed resulting in the discovery of a new potent BLT2 agonist (8f, EC50 = 67.6 nM). Irbesartan and 8f were shown to promote proliferation of epithelial colon cells, an effect which was reversible by a BLT2 antagonist.

Identification of novel anti-cancer agents by the synthesis and cellular screening of a noscapine-based library.

Noscapine is a natural product first isolated from the opium poppy (Papaver somniferum L.) with anticancer properties. In this work, we report the synthesis and cellular screening of a noscapine-based library. A library of novel noscapine derivatives was synthesized with modifications in the isoquinoline and phthalide scaffolds. The so generated library, consisting of fifty-seven derivatives of the natural product noscapine, was tested against MDA-MB-231 breast cancer cells in a cellular proliferation assay (with a Z' > 0.7). The screening resulted in the identification of two novel noscapine derivatives as inhibitors of MDA cell growth with IC50 values of 5 µM and 1.5 µM, respectively. Both hit molecules have a five-fold and seventeen-fold higher potency, compared with that of lead compound noscapine (IC50 26 µM). The identified active derivatives retain the tubulin-binding ability of noscapine. Further testing of both hit molecules, alongside the natural product against additional cancer cell lines (HepG2, HeLa and PC3 cells) confirmed our initial findings. Both molecules have improved anti-proliferative properties when compared to the initial natural product, noscapine.

Benefits of Ginger and Its Constituent 6-Shogaol in Inhibiting Inflammatory Processes.

Ginger (Zingiber officinale Roscoe) is widely used as medicinal plant. According to the Committee on Herbal Medicinal Products (HMPC), dried powdered ginger rhizome can be applied for the prevention of nausea and vomiting in motion sickness (well-established use). Beyond this, a plethora of pre-clinical studies demonstrated anti-cancer, anti-oxidative, or anti-inflammatory actions. 6-Shogaol is formed from 6-gingerol by dehydration and represents one of the main bioactive principles in dried ginger rhizomes. 6-Shogaol is characterized by a Michael acceptor moiety being reactive with nucleophiles. This review intends to compile important findings on the actions of 6-shogaol as an anti-inflammatory compound: in vivo, 6-shogaol inhibited leukocyte infiltration into inflamed tissue accompanied with reduction of edema swelling. In vitro and in vivo, 6-shogaol reduced inflammatory mediator systems such as COX-2 or iNOS, affected NFκB and MAPK signaling, and increased levels of cytoprotective HO-1. Interestingly, certain in vitro studies provided deeper mechanistic insights demonstrating the involvement of PPAR-γ, JNK/Nrf2, p38/HO-1, and NFκB in the anti-inflammatory actions of the compound. Although these studies provide promising evidence that 6-shogaol can be classified as an anti-inflammatory substance, the exact mechanism of action remains to be elucidated. Moreover, conclusive clinical data for anti-inflammatory actions of 6-shogaol are largely lacking.

Propranolol Activates the Orphan Nuclear Receptor TLX to Counteract Proliferation and Migration of Glioblastoma Cells.

The ligand-sensing transcription factor tailless homologue (TLX, NR2E1) is an essential regulator of neuronal stem cell homeostasis with appealing therapeutic potential in neurodegenerative diseases and central nervous system tumors. However, knowledge on TLX ligands is scarce, providing an obstacle to target validation and medicinal chemistry. To discover TLX ligands, we have profiled a drug fragment collection for TLX modulation and identified several structurally diverse agonists and inverse agonists of the nuclear receptor. Propranolol evolved as the strongest TLX agonist and promoted TLX-regulated gene expression in human glioblastoma cells. Structure-activity relationship elucidation of propranolol as a TLX ligand yielded a structurally related negative control compound. In functional cellular experiments, we observed an ability of propranolol to counteract glioblastoma cell proliferation and migration, while the negative control had no effect. Our results provide a collection of TLX modulators as initial chemical tools and set of lead compounds and support therapeutic potential of TLX modulation in glioblastoma.