Discovery and properties of novel analogues of the aphid pheromones nepetalactone and nepetalactol.

BACKGROUND: Pheromones have unique advantages for pest control. Current aphid pheromone research focuses on alarm and sex pheromones. However, practical applications are limited so far, as (E)-ß-farnesene has only been investigated to a small extent as an alarm pheromone and only male aphids are targeted by sex pheromones. Previous literature reports electrophysiological responses and repellent behavior of asexual aphids to nepetalactone (1B), therefore our objective was to modify nepetalactone's structure to identify key fragments responsible for repellent effects, as guidance for subsequent modifications and further investigation. RESULTS: In this study, seven derivatives were designed and synthesized based on nepetalactol (1A) and nepetalactone (1B) as lead compounds. Free-choice tests, conducted using cowpea aphids (Aphis craccivora), revealed that the lactone moiety was crucial for the repellent activity, and the removal of the carbonyl group eliminated the repelling effect. Compound (±)1I, an analogue of nepetalactone (1B), demonstrated a significantly higher repellent value than nepetalactone (1B) at three different concentrations, and even at 0.1 mg/mL it maintained a considerable repellent effect (26.5%). Electrostatic potential and density functional theory calculations supported the importance of the carbonyl group for the repellent effects. CONCLUSION: The newly discovered para-pheromone (±)1I shows improved repellent effects and potential for development as a novel biological control agent. Based on our innovative findings, analogues with improved efficacy and properties can be designed and prepared. Our research contributes to understanding the effects of structural modifications on pheromone activity and properties, which is crucial for exploring novel pheromone-based products for crop protection. © 2024 Society of Chemical Industry.

ZIM, a Norbornene Derived from 4-Aminoantipyrine, Induces DNA Damage and Cell Death but in Association Reduces the Effect of Commercial Chemotherapeutics.

Cancer incidence is increasing, and the drugs are not very selective. These drugs cause adverse effects, and the cells become resistant. Therefore, new drugs are needed. Here, we evaluated the effects of ZIM, a candidate for chemotherapy, and 4-AA alone and in association with commercial chemotherapeutic agents. Subsequently, the results of ZIM and 4-AA were compared. Male Swiss mice were treated with doses of 12, 24, or 48 mg/kg ZIM or 4-AA alone or in association with cisplatin (6 mg/kg), doxorubicin (16 mg/kg), and cyclophosphamide (100 mg/kg). Biometric parameters, DNA damage (comet and micronuclei), cell death, and splenic phagocytosis were evaluated. DNA docking was also performed to confirm the possible interactions of ZIM and 4-AA with DNA. 4-AA has been shown to have low genotoxic potential, increase the frequency of cell death, and activate phagocytosis. ZIM causes genomic and chromosomal damage in addition to causing cell death and activating phagocytosis. In association with chemotherapeutical agents, both 4-AA and ZIM have a chemopreventive effect and, therefore, reduce the frequency of DNA damage, cell death, and splenic phagocytosis. The association of 4-AA and ZIM with commercial chemotherapeutic agents increased the frequency of lymphocytes compared to chemotherapeutic agents alone. Molecular docking demonstrated that ZIM has more affinity for DNA than 4-AA and its precursors (1 and 2). This was confirmed by the lower interaction energy of the complex (-119.83 kcal/mol). ZIM can break the DNA molecule and, therefore, its chemotherapeutic effect can be related to DNA damage. It is considered that ZIM has chemotherapeutic potential. However, it should not be used in combination with cisplatin, doxorubicin, and cyclophosphamide as it reduces the effects of these drugs.

Comprehensive evaluation of novel fungicide benzovindiflupyr at the enantiomeric level: Bioactivity, toxicity, mechanism, and dissipation behavior.

Racemates in the environment can lead to inaccurate risk assessment. To obtain the enantiomeric level information of benzovindiflupyr for accurate risk assessment, the absolute configuration of benzovindiflupyr was first confirmed, and the enantioseparation method was developed by supercritical fluid chromatography tandem mass spectrometry. The enantioselectivity for bioactivity and toxicity was investigated, and the mechanism was explored by molecular docking and detecting succinate dehydrogenase (SDH) activity and content of succinate acid. 1S,4R-(-)-benzovindiflupyr was identified as the most active against the six targeted phytopathogens, which showed higher 1.7-54.5 times than 1R,4S-(+)-benzovindiflupyr. Additionally, 1S,4R-(-)-benzovindiflupyr (LD50: 21.54 µg L-1) was 103.7 times more toxic than 1R,4S-(+)-benzovindiflupyr against Daphnia magna. 1S,4R-(-)-benzovindiflupyr had a stronger affinity for SDH and significantly inhibited SDH activity, resulting in an increase in succinate acid in the tricarboxylic acid cycle, while its downstream products, fumaric and L-malic acid were significantly reduced. Moreover, the dissipation behavior of benzovindiflupyr on three vegetables was evaluated. 1S,4R-(-)-benzovindiflupyr was preferentially degraded in tomato, but opposite in leaves. The enantioselectivity in pepper and cucumber leaves was the same as in tomato, while there was no enantioselectivity in pepper and cucumber. The study provides a basis for accurate risk assessment and the development of high-effective and low-risk fungicides.

Synthesis, Docking Studies and Pharmacological Evaluation of Serotoninergic Ligands Containing a 5-Norbornene-2-Carboxamide Nucleus.

A new series of 5-norbornene-2-carboxamide derivatives was prepared and their affinities to the 5-HT1A, 5-HT2A, and 5-HT2C receptors were evaluated and compared to a previously synthesized series of derivatives characterized by exo-N-hydroxy-5-norbornene-2,3-dicarboximidenucleus, in order to identify selective ligands for the above-mentioned subtype receptors. Arylpiperazines represents one of the most important classes of 5-HT1AR ligands, and recent research concerning new derivatives has been focused on the modification of one or more portions of such pharmacophore. The combination of structural elements (heterocyclic nucleus, propyl chain and 4-substituted piperazine), known to be critical to the affinity to 5-HT1A receptors, and the proper selection of substituents led to compounds with high specificity and affinity towards serotoninergic receptors. The most active compounds were selected for further in vivo assays to determine their functional activity. Finally, to rationalize the obtained results, molecular docking studies were performed. The results of the pharmacological studies showed that Norbo-4 and Norbo-18 were the most active and promising derivatives for the serotonin receptor considered in this study.

In Silico and Ex Vivo Studies on the Spasmolytic Activities of Fenchone Using Isolated Guinea Pig Trachea.

Fenchone is a bicyclic monoterpene found in a variety of aromatic plants, including Foeniculum vulgare and Peumus boldus, and is used in the management of airways disorders. This study aimed to explore the bronchodilator effect of fenchone using guinea pig tracheal muscles as an ex vivo model and in silico studies. A concentration-mediated tracheal relaxant effect of fenchone was evaluated using isolated guinea pig trachea mounted in an organ bath provided with physiological conditions. Sustained contractions were achieved using low K+ (25 mM), high K+ (80 mM), and carbamylcholine (CCh; 1 µM), and fenchone inhibitory concentration-response curves (CRCs) were obtained against these contractions. Fenchone selectively inhibited with higher potency contractions evoked by low K+ compared to high K+ with resultant EC50 values of 0.62 mg/mL (0.58-0.72; n = 5) and 6.44 mg/mL (5.86-7.32; n = 5), respectively. Verapamil (VRP) inhibited both low and high K+ contractions at similar concentrations. Pre-incubation of the tracheal tissues with K+ channel blockers such as glibenclamide (Gb), 4-aminopyridine (4-AP), and tetraethylammonium (TEA) significantly shifted the inhibitory CRCs of fenchone to the right towards higher doses. Fenchone also inhibited CCh-mediated contractions at comparable potency to its effect against high K+ [6.28 mg/mL (5.88-6.42, n = 4); CCh] and [6.44 mg/mL (5.86-7.32; n = 5); high K+]. A similar pattern was obtained with papaverine (PPV), a phosphodiesterase (PDE), and Ca2+ inhibitor which inhibited both CCh and high K+ at similar concentrations [10.46 µM (9.82-11.22, n = 4); CCh] and [10.28 µM (9.18-11.36; n = 5); high K+]. However, verapamil, a standard Ca2+ channel blocker, showed selectively higher potency against high K+ compared to CCh-mediated contractions with respective EC50 values of 0.84 mg/mL (0.82-0.96; n = 5) 14.46 mg/mL (12.24-16.38, n = 4). The PDE-inhibitory action of fenchone was further confirmed when its pre-incubation at 3 and 5 mg/mL potentiated and shifted the isoprenaline inhibitory CRCs towards the left, similar to papaverine, whereas the Ca2+ inhibitory-like action of fenchone pretreated tracheal tissues were authenticated by the rightward shift of Ca2+ CRCs with suppression of maximum response, similar to verapamil, a standard Ca2+ channel blocker. Fenchone showed a spasmolytic effect in isolated trachea mediated predominantly by K+ channel activation followed by dual inhibition of PDE and Ca2+ channels. Further in silico molecular docking studies provided the insight for binding of fenchone with Ca2+ channel (-5.3 kcal/mol) and K+ channel (-5.7), which also endorsed the idea of dual inhibition.

Fenchone Derivatives as a Novel Class of CB2 Selective Ligands: Design, Synthesis, X-ray Structure and Therapeutic Potential.

A series of novel cannabinoid-type derivatives were synthesized by the coupling of (1S,4R)-(+) and (1R,4S)-(-)-fenchones with various resorcinols/phenols. The fenchone-resorcinol derivatives were fluorinated using Selectfluor and demethylated using sodium ethanethiolate in dimethylformamide (DMF). The absolute configurations of four compounds were determined by X-ray single crystal diffraction. The fenchone-resorcinol analogs possessed high affinity and selectivity for the CB2 cannabinoid receptor. One of the analogues synthesized, 2-(2',6'-dimethoxy-4'-(2″-methyloctan-2″-yl)phenyl)-1,3,3-trimethylbicyclo[2.2.1]heptan-2-ol (1d), had a high affinity (Ki = 3.51 nM) and selectivity for the human CB2 receptor (hCB2). In the [35S]GTPγS binding assay, our lead compound was found to be a highly potent and efficacious hCB2 receptor agonist (EC50 = 2.59 nM, E(max) = 89.6%). Two of the fenchone derivatives were found to possess anti-inflammatory and analgesic properties. Molecular-modeling studies elucidated the binding interactions of 1d within the CB2 binding site.

Molecular and functional characterization of a conserved odorant receptor from Aedes albopictus.

BACKGROUND: The Asian tiger mosquito Aedes albopictus is a competent vector of several viral arboviruses including yellow fever, dengue fever, and chikungunya. Several vital mosquito behaviors (e.g., feeding, host-seeking, mating, and oviposition) are primarily dependent on the olfactory system for semiochemicals detection and discrimination. However, the limited number of studies hampers our understanding of the relationships between the Ae. albopictus olfactory system and the complex chemical world. METHODS: We performed RT-qPCR assay on antennae of Ae. albopictus mosquitoes of different sexes, ages and physiological states, and found odorant receptor 11 (AalbOr11) enriched in non-blood-fed female mosquitoes. Then, we examined the odorant preference with a panel of physiologically and behaviorally relevant odorants in Xenopus oocytes. RESULTS: The results indicated that AalbOr11 could be activated by ten aromatics, seven terpenes, six heterocyclics, and three alcohols. Furthermore, using post-RNA interference (RNAi) hand-in-cage assay, we found that reducing the transcript level of AalbOr11 affected the repellency activity mediated by (+)-fenchone at a lower concentration (0.01% v/v). CONCLUSIONS: Using in vitro functional characterization, we found that AalbOr11 was a broadly tuned receptor. Moreover, we found that AalbOr11 shared a conserved odorant reception profile with homologous Anopheles gambiae Or11. In addition, RNAi and bioassay suggested that AablOr11 might be one of the receptors mediating (+)-fenchone repellency activity. Our study attempted to link odor-induced behaviors to odorant reception and may lay the foundation for identifying active semiochemicals for monitoring or controlling mosquito populations.

Nitrobenzoxadiazole derivatives of the rat selective toxicant norbormide as fluorescent probes for live cell imaging.

Norbormide [5-(α-hydroxy-α-2-pyridylbenzyl)-7-(α-2-pyridylbenzylidene)-5-norbornene-2,3-dicarboximide] (NRB, 1), an existing but infrequently used rodenticide, is known to be uniquely toxic to rats, but relatively harmless to other rodents and mammals. As a vasoactive agent, NRB induces a species-specific vasocontractile effect that is restricted to the peripheral arteries of the rat. Despite the precise mechanisms behind this phenomenon having yet to be fully clarified, it is postulated that the molecular target of NRB could be located within the plasma membrane of rat peripheral artery myocytes (e.g. rat caudal artery myocytes). As such, the primary objective of this study was to develop a fluorescently labelled derivative of NRB to investigate its subcellular distribution/localization in both NRB-sensitive (freshly isolated rat caudal artery myocytes, FIRCAMs) and NRB-insensitive (human hepatic stellate, LX2) cells. Of the examples prepared, lead structure endo-NRB-NBD-bPA subsequently demonstrated retention of the parent toxicant's pharmacological profile (in terms of its ability to induce both a vasocontractile response in rat caudal artery rings in vitro, and a lethal end-point in rats in vivo). Endo-NRB-NBD-bPA was also shown to be significantly less permeable (an integral feature in the design of fluorescent probes targeting cell-surface receptors) to both LX2 cells and FIRCAMs. Disappointingly, no fluorescence could be observed on the plasma membrane of FIRCAMs stained with endo-NRB-NBD-bPA.

D609 inhibition of phosphatidylcholine-specific phospholipase C attenuates prolonged insulin stimulation-mediated GLUT4 downregulation in 3T3-L1 adipocytes.

Glucose uptake is stimulated by insulin via stimulation of glucose transporter 4 (GLUT4) translocation to the plasma membrane from intracellular compartments in adipose tissue and muscles. Insulin stimulation for prolonged periods depletes GLUT4 protein, particularly in highly insulin-responsive GLUT4 storage vesicles. This depletion mainly occurs via H2O2-mediated retromer inhibition. However, the post-receptor mechanism of insulin activation of oxidative stress remains unknown. Here, we show that phosphatidylcholine-specific phospholipase C (PC-PLC) plays an important role in insulin-mediated downregulation of GLUT4. In the study, 3T3-L1 adipocytes were exposed to a PC-PLC inhibitor, tricyclodecan-9-yl-xanthogenate (D609), for 30 min prior to the stimulation with 500 nM insulin for 4 h, weakening the depletion of GLUT4. D609 also prevents insulin-driven H2O2 generation in 3T3-L1 adipocytes. Exogenous PC-PLC and its product, phosphocholine (PCho), also caused GLUT4 depletion and promoted H2O2 generation in 3T3-L1 adipocytes. Furthermore, insulin-mediated the increase in the cellular membrane PC-PLC activity was observed in Amplex Red assays. These results suggested that PC-PLC plays an important role in insulin-mediated downregulation of GLUT4 and that PCho may serve as a signaling molecule.

Amyloid Beta-Peptide 25-35 (Aß25-35) Induces Cytotoxicity via Multiple Mechanisms: Roles of the Inhibition of Glucosylceramide Synthase by Aß25-35 and Its Protection by D609.

Sphingolipids (SLs), such as ceramide, glucosylceramide (GlcCer), and sphingomyelin, play important roles in the normal development/functions of the brain and peripheral tissues. Disruption of SL homeostasis in cells/organelles, specifically up-regulation of ceramide, is involved in multiple diseases including Alzheimer's disease (AD). One of the pathological features of AD is aggregates of amyloid beta (Aß) peptides, and SLs regulate both the formation/aggregation of Aß and Aß-induced cellular responses. Up-regulation of ceramide levels via de novo and salvage synthesis pathways is reported in Aß-treated cells and brains with AD; however, the effects of Aß on ceramide decomposition pathways have not been elucidated. Thus, we investigated the effects of the 25-35-amino acid Aß peptide (Aß25-35), the fundamental cytotoxic domain of Aß, on SL metabolism in cells treated with the fluorescent nitrobenzo-2-oxa-1,3-diazole-labeled C6-ceramide (NBD-ceramide). Aß25-35 treatment reduced the formation of NBD-GlcCer mediated by GlcCer synthase (GCS) without affecting the formation of NBD-sphingomyelin or NBD-ceramide-1-phosphate, and reduced cell viability. Aß25-35-induced responses decreased in cells treated with D609, a putative inhibitor of sphingomyelin synthases. Aß25-35-induced cytotoxicity significantly increased in GCS-knockout cells and pharmacological inhibition of GCS alone demonstrated cytotoxicity. Our study revealed that Aß25-35-induced cytotoxicity is at least partially mediated by the inhibition of GCS activity.

Metabolic adaptations to hypoxia in the neonatal mouse forebrain can occur independently of the transporters SLC7A5 and SLC3A2.

Neonatal encephalopathy due to hypoxia-ischemia is associated with adverse neurodevelopmental effects. The involvement of branched chain amino acids (BCAAs) in this is largely unexplored. Transport of BCAAs at the plasma membrane is facilitated by SLC7A5/SLC3A2, which increase with hypoxia. We hypothesized that hypoxia would alter BCAA transport and metabolism in the neonatal brain. We investigated this using an organotypic forebrain slice culture model with, the SLC7A5/SLC3A2 inhibitor, 2-Amino-2-norbornanecarboxylic acid (BCH) under normoxic or hypoxic conditions. We subsequently analysed the metabolome and candidate gene expression. Hypoxia was associated with increased expression of SLC7A5 and SLC3A2 and an increased tissue abundance of BCAAs. Incubation of slices with 13C-leucine confirmed that this was due to increased cellular uptake. BCH had little effect on metabolite abundance under normoxic or hypoxic conditions. This suggests hypoxia drives increased cellular uptake of BCAAs in the neonatal mouse forebrain, and membrane mediated transport through SLC7A5 and SLC3A2 is not essential for this process. This indicates mechanisms exist to generate the compounds required to maintain essential metabolism in the absence of external nutrient supply. Moreover, excess BCAAs have been associated with developmental delay, providing an unexplored mechanism of hypoxia mediated pathogenesis in the developing forebrain.

New class of hantaan virus inhibitors based on conjugation of the isoindole fragment to (+)-camphor or (-)-fenchone hydrazonesv.

This work presents the design and synthesis of camphor, fenchone, and norcamphor N-acylhydrazone derivatives as a new class of inhibitors of the Hantaan virus, which causes haemorrhagic fever with renal syndrome (HFRS). A cytopathic model was developed for testing chemotherapeutics against the Hantaan virus, strain 76-118. In addition, a study of the antiviral activity was carried out using a pseudoviral system. It was found that the hit compound possesses significant activity (IC50 = 7.6 ± 2 µM) along with low toxicity (CC50 > 1000 µM). Using molecular docking procedures, the binding with Hantavirus nucleoprotein was evaluated and the correlation between the structure of the synthesised compounds and the antiviral activity was established.

Chemical composition and synergistic effect of three Moroccan lavender EOs with ciprofloxacin against foodborne bacteria: a promising approach to modulate antimicrobial resistance.

The aim of this study was to determine the chemical profile of the essential oils (EOs) of three Moroccan lavender species (Lavandula pedunculata, LP; Lavandula angustifolia, LA; and Lavandula maroccana, LM) and to investigate, for the first time, the synergistic effect of the optimal mixture of the EOs with conventional antibiotic ciprofloxacin against three pathogenic foodborne bacteria. Gas chromatography/mass spectrometry analysis showed that eucalyptol (39·05%), camphor (24·21%) and borneol (8·29%) were the dominant compounds of LA-EO. LP-EO was characterized by the abundance of camphor (74·51%) and fenchone (27·06%), whereas carvacrol (42·08%), camphor (17·95%) and fenchone (12·05%) were the main constituents of LM-EO. EOs alone or combined showed a remarkable antimicrobial activity against the tested bacteria with minimum inhibitory concentrations (MICs) ranging from 3·53 to 15·96 mg ml-1 . The optimal mixture, calculated using a mixture design, corresponded to 19% LA, 38% LP and 43% LM. All combination of the EOs and the best EO mixture with ciprofloxacin exhibited a total synergism with fractional inhibitory concentration index values ranging from 0·27 to 0·37. The best EO mixture showed the highest gain of 128-fold, especially against Salmonella spp., more than that found testing the EOs separately. These findings should be taken into consideration for a possible application in the pharmaceutical and food industries.

Genetic polymorphisms associated to SDHI fungicides resistance in selected Aspergillus flavus strains and relation with aflatoxin production.

Aspergillus flavus is a common and ubiquitous fungal species able to colonize several agricultural commodities, in both pre- and post-harvest conditions. This species represents a very harmful plant pathogen for its ability to synthesize aflatoxin B1, responsible for human primary hepatocellular carcinoma and classified as a group I (human carcinogenic) by the International Agency for Research on Cancer. Several approaches have been proposed to control A. flavus development and related aflatoxin production in field and storage conditions. The Succinate Dehydrogenase Inhibitor (SDHI) fungicide boscalid has been shown to control A. flavus growth and aflatoxin contamination both in vitro and in field experiments. However, this compound is classified as medium-high risk fungicide for triggering fungal resistance and, indeed, resistant strains can occur on crops treated with boscalid. In this paper, we selected laboratory A. flavus strains resistant to boscalid grown on agar medium containing 50 mg/L of boscalid. In order to investigate the molecular mechanism responsible for the resistant phenotype, specific primer pairs were designed to amplify the whole SdhB, SdhC and SdhD genes. By amino acid sequence analysis, two point mutations, Tyrosine replacing Histidine at codon 249 of SdhB (H249Y) and Arginine replacing Glycine at codon 91 of SdhC (G91R), were identified. The effect of SDHI boscalid and isopyrazam on mycelial growth and conidial germination was evaluated. Both resistant genotypes showed high resistance (MIC and EC50 > 1000 mg/L) to boscalid. A positive cross-resistance was found between boscalid and isopyrazam. Specific sub-lethal doses of both fungicides (0.5 mg/L of boscalid and 0.01 mg/L of isopyrazam) interfered with the mechanisms associated to pigmentation of colonies. In particular, fungal colonies appeared depigmented lacking the typical A. flavus green colour shown on un-amended fungicide medium. A strict correlation between lack of pigmentation and increasing aflatoxin production was also observed.

Functional role of phosphatidylcholine-specific phospholipase C in regulating leukotriene synthesis and degranulation in human eosinophils.

Phosphatidylinositol-specific phospholipase C (PI-PLC) and cytosolic phospholipase A2 (cPLA2) regulate both eosinophil degranulation and leukotriene (LT) synthesis via PI-PLC-mediated calcium influx and cPLA2 activation. Phosphatidylcholine-specific phospholipase C (PC-PLC) likely plays a key role in cellular signaling, including the eosinophilic allergic inflammatory response. This study examined the role of PC-PLC in eosinophil LT synthesis and degranulation using tricyclodecan-9-yl-xanthogenate (D609), a PC-specific PLC inhibitor. D609 inhibited N-formyl-met-leu-phe + cytochalasin B (fMLP/B)-induced arachidonic acid (AA) release and leukotriene C4 (LTC4) secretion. However, at concentrations that blocked both AA release and LTC4 secretion, D609 had no significant inhibitory effect on stimulated cPLA2 activity. D609 also partially blocked fMLP/B-induced calcium influx, indicating that inhibition of AA release and LTC4 secretion by D609 is due to inhibition of calcium-mediated cPLA2 translocation to intracellular membranes, not inhibition of cPLA2 activity. In addition, D609 inhibited fMLP/B-stimulated eosinophil peroxidase release, indicating that PC-PLC regulates fMLP/B-induced eosinophil degranulation by increasing the intracellular calcium concentration ([Ca2+]i). Overall, our results showed that PC-PLC is critical for fMLP/B-stimulated eosinophil LT synthesis and degranulation. In addition, degranulation requires calcium influx, while PC-PLC regulates LTC4 synthesis through calcium-mediated cPLA2 activation.

Structure optimization of positive allosteric modulators of GABAB receptors led to the unexpected discovery of antagonists/potential negative allosteric modulators.

Positive allosteric modulators (PAMs) of GABAB receptor represent an interesting alternative to receptor agonists such as baclofen, as they act on the receptor in a more physiological way and thus are devoid of the side effects typically exerted by the agonists. Based on our interest in the identification of new GABAB receptor PAMs, we followed a merging approach to design new chemotypes starting from selected active compounds, such as GS39783, rac-BHFF, and BHF177, and we ended up with the synthesis of four different classes of compounds. The new compounds were tested alone or in the presence of 10 µM GABA using [35S]GTPγS binding assay to assess their functionality at the receptor. Unexpectedly, a number of them significantly inhibited GABA-stimulated GTPγS binding thus revealing a functional switch with respect to the prototype molecules. Further studies on selected compounds will clarify if they act as negative modulators of the receptor or, instead, as antagonists at the orthosteric binding site.

Open and rearranged norbornane derived polycyclic cage molecules as potential neuroprotective agents through attenuation of MPP+- and calcium overload-induced excitotoxicity in neuroblastoma SH-SY5Y cells.

The neuroprotective effects of closed polycyclic cage molecules such as NGP1-01, memantine and amantadine have been extensively explored. These effects are mostly linked to the antagonism of the N-methyl-d-aspartate (NMDA) receptor- and the blockage of voltage gated calcium channels (VGCC). The synthesis of structurally related open and rearranged cage derivatives has been studied in depth. However, very little is known on their neuroprotective effects. In this study, a series of open and rearranged polycyclic cage molecules containing a norbornane derived scaffold were synthesised and evaluated for cytotoxicity, neuroprotection and calcium blocking effects via the NMDA receptor and VGCC on neuroblastoma cells at a 10 µM concentration. All compounds showed negligible cytotoxicity and were able to significantly attenuate MPP+-induced neurotoxicity between 26.07 ± 12.50% to 48.42 ± 0.76%, with compound 14 showing the best neuroprotective effect. In comparison to known NMDA receptor antagonists, all compounds demonstrated moderate to excellent calcium blocking effects of 26.50 ± 2.28 to 72.95 ± 3.38%. Docking studies suggest that these compounds are able to show significant NMDA receptor channel blocking ability since they bind in a comparable manner to the crystallographic pose of MK-801 inside the NMDAR ion channel. Some compounds were also able to attenuate calcium influx through VGCC channels between 21.28 ± 3.69% to 50.34 ± 7.67%. Compound 4 and 15 showed the highest inhibition of calcium influx at the VGCC and NMDA receptor, respectively. The compounds exhibiting good cytotoxicity-, neuroprotective- and calcium blocking profiles could potentially act as neuroprotective agents to clinically benefit people suffering from neurodegenerative disorders.

Hemocompatible LAT1-inhibitor can induce apoptosis in cancer cells without affecting brain amino acid homeostasis.

Increased amounts of amino acids are essential for cancer cells to support their sustained growth and survival. Therefore, inhibitors of amino acid transporters, such as L-type amino acid transporter 1 (LAT1) have been developed. In this study, a previously reported LAT1-inhibitor (KMH-233) was studied for its hemocompatibility and toxicity towards human umbilical vein endothelial cells (HUVEC) and human aortic smooth muscle cells (AoSMCs). Furthermore, the cytotoxic effects against human breast adenocarcinoma cells (MCF-7) and its ability to affect mammalian (or mechanistic) target of rapamycin (mTOR) and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling were evaluated. Moreover, the effects of this inhibitor to modulate LAT1 function on the cell surface and the brain amino acid homeostasis were evaluated after intraperitoneal (i.p.) administration of LAT1-inhibitor (23 µmol/kg) in mice. The results showed that LAT1-inhibitor (KMH-233) is hemocompatible at concentrations below 25 µM and it does not affect coagulation in plasma. However, it can reduce the total protein amount of mTOR and NF-κB, resulting in increased apoptosis in LAT1-expressing cancer cells. Most importantly, the inhibitor did not affect mouse brain levels of L-Leu, L-Tyr or L-Trp or modulate the function of LAT1 on the MCF-7 cell surface. Therefore, this inhibitor can be considered as a safe but effective anti-cancer agent. However, due to the compensative mechanism of cancer cells for their increased amino acid demand, this compound is most effective inducing apoptosis when used in combinations with other chemotherapeutics, such as protease inhibitor, bestatin, as demonstrated in this study.

D609 protects retinal pigmented epithelium as a potential therapy for age-related macular degeneration.

Accumulated oxidative damage may lead to irreversible retinal pigmented epithelium (RPE) cell death, which is considered to be the primary cause of dry age-related macular degeneration (AMD), leading to blindness in the elderly. However, an effective therapy for this disease is lacking. Here, we described a robust high-content screening procedure with a library of 814 protective compounds and found that D609 strongly protected RPE cells from sodium iodate (SI)-induced oxidative cell death and prolonged their healthy survival. D609 effectively attenuated excessive reactive oxygen species (ROS) and prevented severe mitochondrial loss due to oxidative stress in the RPE cells. Surprisingly, the potent antioxidative effects of D609 were not achieved through its own reducibility but were primarily dependent on its ability to increase the expression of metallothionein. The injection of this small water-soluble molecule also showed an explicit protective effect of the RPE layer in an SI-induced AMD mouse model. These findings suggested that D609 could serve as a novel antioxidative protector of RPE cells both in vitro and in vivo and unveiled a novel antioxidative mechanism of D609, which may ultimately have clinical applications for the treatment of AMD.

Regulation of Melanogenesis by the Amino Acid Transporter SLC7A5.

Integration of chromatin immunoprecipitation-sequencing and microarray data enabled us to identify previously unreported MITF-target genes, among which the amino acid transporter SLC7A5 is also included. We reported that small interfering RNA-mediated SLC7A5 knockdown decreased pigmentation in B16F10 cells but neither affected morphology nor dendricity. Treatment with the SLC7A5 inhibitors 2-amino-2-norbornanecarboxylic acid (BCH) or JPH203 also decreased melanin synthesis in B16F10 cells. Our findings indicated that BCH was as potent as reference depigmenting agent, kojic acid, but acted through a different pathway not affecting tyrosinase activity. BCH also decreased pigmentation in human MNT1 melanoma cells or normal human melanocytes. Finally, we tested BCH on a more physiological model, using reconstructed human epidermis and confirmed a strong inhibition of pigmentation, revealing the clinical potential of SLC7A5 inhibition and positioning BCH as a depigmenting agent suitable for cosmetic or dermatological intervention in hyperpigmentation diseases.