Aurantoside L, a New Tetramic Acid Glycoside with Anti-Leishmanial Activity Isolated from the Marine Sponge Siliquariaspongia japonica.

A new tetramic acid glycoside, aurantoside L (1), was isolated from the sponge Siliquariaspongia japonica collected at Tsushima Is., Nagasaki Prefecture, Japan. The structure of aurantoside L (1) composed of a tetramic acid bearing a chlorinated polyene system and a trisaccharide part was elucidated using spectral analysis. Aurantoside L (1) showed anti-parasitic activity against L. amazonensis with an IC50 value of 0.74 µM.

Metabolic Investigation and Auxiliary Enzyme Modelization of the Pyrrocidine Pathway Allow Rationalization of Paracyclophane-Decahydrofluorene Formation.

Fungal paracyclophane-decahydrofluorene-containing natural products are complex polycyclic metabolites derived from similar hybrid PKS-NRPS pathways. Herein we studied the biosynthesis of pyrrocidines, one representative of this family, by gene inactivation in the producer Sarocladium zeae coupled to thorough metabolic analysis and molecular modeling of key enzymes. We characterized nine pyrrocidines and analogues as well as in mutants a variety of accumulating metabolites with new structures including rare cis-decalin, cytochalasan, and fused 6/15/5 macrocycles. This diversity highlights the extraordinary plasticity of the pyrrocidine biosynthetic gene cluster. From accumulating metabolites, we delineated the scenario of pyrrocidine biosynthesis. The ring A of the decahydrofluorene is installed by PrcB, a membrane-bound cyclizing isomerase, on a PKS-NRPS-derived pyrrolidone precursor. Docking experiments in PrcB allowed us to characterize the active site suggesting a mechanism triggered by arginine-mediated deprotonation at the terminal methyl of the substrate. Next, two integral membrane proteins, PrcD and PrcE, each predicted as a four-helix bundle, perform hydroxylation of the pyrrolidone ring and paracyclophane formation, respectively. Modelization of PrcE highlights a topological homology with vitamin K oxido-reductase and the presence of a disulfide bond. Our results suggest a previously unsuspected coupling mechanism via a transient loss of aromaticity of tyrosine residue to form the strained paracyclophane motif. Finally, the lipocalin-like protein PrcX drives the exo-cycloaddition yielding ring B and C of the decahydrofluorene to afford pyrrocidine A, which is transformed by a reductase PrcI to form pyrrocidine B. These insights will greatly facilitate the microbial production of pyrrocidine analogues by synthetic biology.

Design, Synthesis, and Herbicidal Evaluation of Pyrrolidinone-Containing 2-Phenylpyridine Derivatives as Novel Protoporphyrinogen Oxidase Inhibitors.

In this work, a series of pyrrolidinone-containing 2-phenylpyridine derivatives were synthesized and evaluated as novel protoporphyrinogen IX oxidase (PPO, EC 1.3.3.4) inhibitors for herbicide development. At 150 g ai/ha, compounds 4d, 4f, and 4l can inhibit the grassy weeds of Echinochloa crus-galli (EC), Digitaria sanguinalis (DS), and Lolium perenne (LP) with a range of 60 to 90%. Remarkably, at 9.375 g ai/ha, these compounds showed 100% inhibition effects against broadleaf weeds of Amaranthus retroflexus (AR) and Abutilon theophrasti (AT), which were comparable to the performance of the commercial herbicides flumioxazin (FLU) and saflufenacil (SAF) and better than that of acifluorfen (ACI). Molecular docking analyses revealed significant hydrogen bonding and π-π stacking interactions between compounds 4d and 4l with Arg98, Asn67, and Phe392, respectively. Additionally, representative compounds were chosen for in vivo assessment of PPO inhibitory activity, with compounds 4d, 4f, and 4l demonstrating excellent inhibitory effects. Notably, compounds 4d and 4l induced the accumulation of reactive oxygen species (ROS) and a reduction in the chlorophyll (Chl) content. Consequently, compounds 4d, 4f, and 4l are promising lead candidates for the development of novel PPO herbicides.

Aniracetam: An Evidence-Based Model for Preventing the Accumulation of Amyloid-ß Plaques in Alzheimer's Disease.

 Alzheimer's disease is the leading cause of dementia in the world. It affects 6 million people in the United States and 50 million people worldwide. Alzheimer's disease is characterized by the accumulation of amyloid-ß plaques (Aß), an increase in tau protein neurofibrillary tangles, and a loss of synapses. Since the 1990s, removing and reducing Aß has been the focus of Alzheimer's treatment and prevention research. The accumulation of Aß can lead to oxidative stress, inflammation, neurotoxicity, and eventually apoptosis. These insults impair signaling systems in the brain, potentially leading to memory loss and cognitive decline. Aniracetam is a safe, effective, cognitive-enhancing drug that improves memory in both human and animal studies. Aniracetam may prevent the production and accumulation of Aß by increasing α-secretase activity through two distinct pathways: 1) increasing brain derived neurotrophic factor expression and 2) positively modulating metabotropic glutamate receptors. This is the first paper to propose an evidence-based model for aniracetam reducing the accumulation and production of Aß.

Psychiatric assessment prior to and after switch from levetiracetam to brivaracetam.

PURPOSE: Brivaracetam is often used as an alternative to levetiracetam in patients with epilepsy (PWE) encountering efficacy issues or adverse events with levetiracetam. This study evaluated the psychological status of PWE who were switched from levetiracetam to brivaracetam due to psychiatric tolerability concerns in comparison to those who remained on levetiracetam. METHODS: We used various psychological assessments including the Symptom Checklist SCL-90-R, the Beck Depression Inventory-II, and the adverse event profile. Eligible participants completed the questionnaires at baseline and again 8 days later. Psychological changes were assessed using standard statistical methods to show differences between a group that immediately switched from levetiracetam to brivaracetam and another group with unchanged levetiracetam. RESULTS: Between May 2020 and May 2021, 63 patients participated in the study, of whom 34 switched from levetiracetam to brivaracetam. At baseline, participants who switched to brivaracetam had fewer antiseizure medications but experienced more monthly seizures. Baseline scores for anxiety (p = 0.020) and psychoticism (p = 0.046) on SCL-90-R in PWE switched to brivaracetam were higher than in the remaining group. In the subsequent assessment, all psychological scores were reduced and were no longer significantly different between both groups. Using multiple regression, initial treatment with a single antiseizure medication and male gender emerged as predictors of psychological improvement. CONCLUSION: Our study found no increased risk of adverse events or psychiatric symptoms after switching from levetiracetam to brivaracetam. Though statistically non-significant, a trend towards improved psychiatric outcomes in the switch group warrants further investigation in future trials with stronger designs for enhanced statistical power.

Water-compatible cross-linked pyrrolidone acrylate pressure-sensitive adhesives with persistent adhesion for transdermal delivery: Synergistic effect of hydrogen bonding and electrostatic force.

Poor skin adhesion and mechanical properties are common problems of pressure-sensitive adhesive (PSA) in transdermal drug delivery system (TDDS). Its poor water compatibility also causes the patch to fall off after sweating or soaking in the application site. To solve this problem, poly (2-Ethylhexyl acrylate-co-N-Vinyl-2-pyrrolidone-co-N-(2-Hydroxyethyl)acrylamide) (PENH), a cross-linked pyrrolidone polyacrylate PSA, was designed to improve the adhesion and water resistance of PSA through electrostatic force and hydrogen bonding system. The structure of PENH was characterized by 1H NMR, FTIR, DSC, and other methods. The mechanism was studied by FTIR, rheological test, and molecular simulation. The results showed that the PENH patch could adhere to human skin for more than 10 days without cold flow, and it could still adhere after sweating or water contact. In contrast, the commercial PSA Duro-Tak® 87-4098 and Duro-Tak® 87-2852 fell off completely on the 3rd and 6th day, respectively, and Duro-Tak® 87-2510 showed a significant dark ring on the second day. Mechanism studies have shown that the hydrogen bond formed by 2-ethylhexyl acrylate (2-EHA), N-vinyl-2-pyrrolidinone (NVP), and N-(2-Hydroxyethyl)acrylamide (HEAA) enhances cohesion, the interaction with skin improves skin adhesion, and the electrostatic interaction with water or drug molecules enhances the ability of water absorption and drug loading. Due to the synergistic effect of hydrogen bonds and electrostatic force, PENH can maintain high cohesion after drug loading or water absorption. PENH provides a choice for the development of water-compatible patches with long-lasting adhesion. STATEMENT OF SIGNIFICANCE: Based on the synergistic effect of hydrogen bonding and electrostatic force, a hydrogen-bonded, cross-linked pyrrolidone acrylate pressure-sensitive adhesive for transdermal drug delivery was designed and synthesized, which has high adhesion and cohesive strength and is non-irritating to the skin. The patch can be applied on the skin surface continuously for more than 10 days without the phenomenon of "dark ring", and the patch can remain adherent after the patient sweats or bathes. This provides a good strategy for choosing a matrix for patches that require prolonged administration.

Brivaracetam exposure-response predictions in pediatric patients from age 1 month: Extrapolation of levetiracetam adult-pediatric scaling to brivaracetam.

BACKGROUND: An adult population pharmacokinetic/pharmacodynamic (PK/PD) model for the antiseizure medication (ASM) brivaracetam (BRV) was previously extended to children aged 4-16 years by using a pediatric BRV population PK model. Effects were scaled using information from a combined adult-pediatric PK/PD model of a related ASM, levetiracetam (LEV). OBJECTIVE: To scale an existing adult population PK/PD model for BRV to children aged 1 month to < 4 years using information from a combined adult-pediatric PK/PD model for LEV, and to predict the effective dose of BRV in children aged 1 month to < 4 years using the adult BRV PK/PD model modified for the basal seizure rate in children. MATERIAL AND METHODS: An existing adult population PK/PD model for BRV was scaled to children aged from 1 month to < 4 years using information from a combined adult-pediatric PK/PD model for LEV, an ASM binding to the same target protein as BRV. An existing adult-pediatric PK/PD model for LEV was extended using data from UCB study N01009 (NCT00175890) to include children as young as 1 month of age. The BRV population PK model was updated with data up to 180 days after first administration from BRV pediatric studies N01263 (NCT00422422) and N01266 (NCT01364597). PK and PD simulations for BRV were performed for a range of mg/kg doses to predict BRV effect in pediatric participants, and to provide dosing recommendations. RESULTS: The extended adult-pediatric LEV PK/PD model was able to describe the adult and pediatric data using the same PD model parameters in adults and children and supported the extension of the adult BRV PK/PD model to pediatric patients aged 1 month to < 4 years. Simulations predicted exposures similar to adults receiving BRV 100 mg twice daily (b.i.d.), when using 3 mg/kg b.i.d. for weight < 10 kg, 2.5 mg/kg b.i.d. for weight ≥ 10 kg and < 20 kg, and 2 mg/kg b.i.d. for weight ≥ 20 kg in children aged 1 month to < 4 years. PK/PD simulations show that maximum BRV response is expected to occur with 2-3 mg/kg b.i.d. dosing of BRV in children aged 1 month to < 4 years, with an effective dose of 1 mg/kg b.i.d. for some participants. CONCLUSION: Development of an adult-pediatric BRV PK/PD model allowed characterization of the exposure-response relationship of BRV in children aged 1 to < 4 years, providing a maximal dose allowance based on weight.

Mutagenetic analysis of the biosynthetic pathway of tetramate bripiodionen bearing 3-(2H-pyran-2-ylidene)pyrrolidine-2,4-dione skeleton.

BACKGROUND: Natural tetramates are a family of hybrid polyketides bearing tetramic acid (pyrrolidine-2,4-dione) moiety exhibiting a broad range of bioactivities. Biosynthesis of tetramates in microorganisms is normally directed by hybrid polyketide synthase (PKS) and nonribosomal peptide synthetase (NRPS) machineries, which form the tetramic acid ring by recruiting trans- or cis-acting thioesterase-like Dieckmann cyclase in bacteria. There are a group of tetramates with unique skeleton of 3-(2H-pyran-2-ylidene)pyrrolidine-2,4-dione, which remain to be investigated for their biosynthetic logics. RESULTS: Herein, the tetramate type compounds bripiodionen (BPD) and its new analog, featuring the rare skeleton of 3-(2H-pyran-2-ylidene)pyrrolidine-2,4-dione, were discovered from the sponge symbiotic bacterial Streptomyces reniochalinae LHW50302. Gene deletion and mutant complementation revealed the production of BPDs being correlated with a PKS-NRPS biosynthetic gene cluster (BGC), in which a Dieckmann cyclase gene bpdE was identified by sit-directed mutations. According to bioinformatic analysis, the tetramic acid moiety of BPDs should be formed on an atypical NRPS module constituted by two discrete proteins, including the C (condensation)-A (adenylation)-T (thiolation) domains of BpdC and the A-T domains of BpdD. Further site-directed mutagenetic analysis confirmed the natural silence of the A domain in BpdC and the functional necessities of the two T domains, therefore suggesting that an unusual aminoacyl transthiolation should occur between the T domains of two NRPS subunits. Additionally, characterization of a LuxR type regulator gene led to seven- to eight-fold increasement of BPDs production. The study presents the first biosynthesis case of the natural molecule with 3-(2H-pyran-2-ylidene)pyrrolidine-2,4-dione skeleton. Genomic mining using BpdD as probe reveals that the aminoacyl transthiolation between separate NRPS subunits should occur in a certain population of NRPSs in nature.

Identification of Lydicamycins as Main Antioomycete Compounds from the Biocontrol Agent Streptomyces sp. NEAU-S7GS2.

Oomycetes are well-known phytopathogens that seriously threaten many important crops worldwide. In this study, the endophytic actinobacterium Streptomyces sp. NEAU-S7GS2 demonstrated significant antagonistic activity against Phytophthora and Pythium and showed a potent biocontrol effect on suppression of soybean phytophthora root rot and pepper phytophthora blight. Two compounds were subsequently isolated as the main active components by bioassay-guided fractionation and identified as lydicamycins A and B. These two compounds showed high antioomycete activity against Phytophthora and Pythium with EC50 values of 0.73-2.67 µg/mL, which are equal to or lower than those of commercialized drug metalaxyl. In vivo bioassay using detached leaves demonstrated that lydicamycin A had a better control efficiency against soybean phytophthora root rot than metalaxyl. Taken together, these results suggest that the biocontrol agent Streptomyces sp. NEAU-S7GS2 and lydicamycins have the potential to be developed as promising pesticides to control diseases caused by oomycetes.

The Effect of 4-(Dimethylamino)phenyl-5-oxopyrrolidines on Breast and Pancreatic Cancer Cell Colony Formation, Migration, and Growth of Tumor Spheroids.

A series of hydrazones, azoles, and azines bearing a 4-dimethylaminophenyl-5-oxopyrrolidine scaffold was synthesized. Their cytotoxic effect against human pancreatic carcinoma Panc-1 and triple-negative breast cancer MDA-MB-231 cell lines was established by MTT assay. Pyrrolidinone derivatives 3c and 3d, with incorporated 5-chloro and 5-methylbenzimidazole fragments; hydrazone 5k bearing a 5-nitrothien-2-yl substitution; and hydrazone 5l with a naphth-1-yl fragment in the structure significantly decreased the viability of both cancer cell lines. Compounds 3c and 5k showed the highest selectivity, especially against the MDA-MB-231 cancer cell line. The EC50 values of the most active compound 5k against the MDA-MB231 cell line was 7.3 ± 0.4 µM, which were slightly higher against the Panc-1 cell line (10.2 ± 2.6 µM). Four selected pyrrolidone derivatives showed relatively high activity in a clonogenic assay. Compound 5k was the most active in both cell cultures, and it completely disturbed MDA-MB-231 cell colony growth at 1 and 2 µM and showed a strong effect on Panc-1 cell colony formation, especially at 2 µM. The compounds did not show an inhibitory effect on cell line migration by the 'wound-healing' assay. Compound 3d most efficiently inhibited the growth of Panc-1 spheroids and reduced cell viability in MDA-MB-231 spheroids. Considering these different activities in biological assays, the selected pyrrolidinone derivatives could be further tested to better understand the structure-activity relationship and their mechanism of action.

Efficacy and Tolerability of Adjunctive Brivaracetam in Patients with Focal-Onset Seizures on Specific Concomitant Antiseizure Medications: Pooled Analysis of Double-Blind, Placebo-Controlled Trials.

INTRODUCTION: This article aimed to assess the efficacy and tolerability of adjunctive brivaracetam (BRV) in adults with focal-onset seizures on specific concomitant antiseizure medications (ASMs) taken as part of their treatment regimen. METHODS: This was a post hoc analysis of pooled data from double-blind, placebo-controlled trials (N01252/NCT00490035, N01253/NCT00464269, and N01358/NCT01261325) in patients with uncontrolled focal-onset seizures randomized to BRV (50-200 mg/day) or placebo on the most common concomitant ASMs at trial initiation. RESULTS: Nine concomitant ASMs were analyzed: carbamazepine (CBZ), lamotrigine (LTG), valproate (VPA), oxcarbazepine (OXC), topiramate (TPM), phenytoin (PHT), lacosamide (LCM), clobazam (CLB), and phenobarbital (PHB). Reduction over placebo in focal-onset seizure frequency per 28 days with BRV ranged from 11.7% (concomitant OXC) to 33.5% (concomitant PHB). The median percentage reduction from baseline in focal-onset seizure frequency per 28 days ranged from 25.5% to 42.8% in patients on BRV (placebo 4.4-21.2%); 50% responder rates ranged from 31.9% to 44.9% in patients on BRV (placebo 11.4-25.2%). In patients on BRV, seizure freedom ranged from 1.4% (concomitant PHT) to 12.5% (concomitant LCM); seizure freedom ranged from 0% to 1.2% in patients on placebo. All efficacy endpoints analyzed were consistently numerically higher in patients on BRV versus placebo. The overall incidence of treatment-emergent adverse events (TEAEs) was generally similar across subgroups by specific concomitant ASMs in patients on BRV (range 60.8-74.5%) or placebo (range 53.8-66.7%). Drug-related TEAEs were numerically higher across all subgroups by concomitant ASM in patients on BRV (range 35.2-48.3%) versus placebo (range 23.9-37.1%). Discontinuations due to TEAEs ranged from 2.9% to 13.3% in patients on BRV and was 0-5.7% for patients taking placebo across subgroups. CONCLUSION: BRV was efficacious and well tolerated regardless of the specific concomitant ASMs used as part of their treatment regimen. These data show that in patients with focal-onset seizures, BRV provides additional efficacy to a broad range of ASMs.

The combination of pleconaril, rupintrivir, and remdesivir efficiently inhibits enterovirus infections in vitro, delaying the development of drug-resistant virus variants.

Enteroviruses are a significant global health concern, causing a spectrum of diseases from the common cold to more severe conditions like hand-foot-and-mouth disease, meningitis, myocarditis, pancreatitis, and poliomyelitis. Current treatment options for these infections are limited, underscoring the urgent need for effective therapeutic strategies. To find better treatment option we analyzed toxicity and efficacy of 12 known broad-spectrum anti-enterovirals both individually and in combinations against different enteroviruses in vitro. We identified several novel, synergistic two-drug and three-drug combinations that demonstrated significant inhibition of enterovirus infections in vitro. Specifically, the triple-drug combination of pleconaril, rupintrivir, and remdesivir exhibited remarkable efficacy against echovirus (EV) 1, EV6, EV11, and coxsackievirus (CV) B5, in human lung epithelial A549 cells. This combination surpassed the effectiveness of single-agent or dual-drug treatments, as evidenced by its ability to protect A549 cells from EV1-induced cytotoxicity across seven passages. Additionally, this triple-drug cocktail showed potent antiviral activity against EV-A71 in human intestinal organoids. Thus, our findings highlight the therapeutic potential of the pleconaril-rupintrivir-remdesivir combination as a broad-spectrum treatment option against a range of enterovirus infections. The study also paves the way towards development of strategic antiviral drug combinations with virus family coverage and high-resistance barriers.

Targeted Metabolic Investigation of Ligandrol and Analytical Methods Validation for Its Main Long-term Metabolite.

Prompted by the need for related analytical reference material in the frame of the fight against doping in sports, synthetic efforts towards the main long-term bishydroxylated metabolite (LGD-LTM1) of the nonsteroidal selective androgen receptor modulator (SARM) ligandrol have produced related derivatives that were exploited for a targeted metabolite analysis of urine samples obtained in the course of previous excretion studies of this SARM. Further clarifying ligandrol's metabolic profile, the availability of synthetic reference material permitted the structural elucidation of a previously reported pyrrolidinone-type metabolite and revealed its potential analytical utility as an additional long-term marker. Moreover, synthetic reference material enabled the comparison and validation of liquid chromatography coupled with mass spectrometry (LC-MS)-based and gas chromatography coupled with mass spectrometry (GC-MS)-based detection and identification methods focusing on the LGD-LTM1 marker.

Investigation, scaffold hopping of novel donepezil-based compounds as anti-Alzhiemer's agents: synthesis, in-silico and pharmacological evaluations.

Alzheimer's disease (AD) is a multifaceted neurodegenerative condition. The pathogenesis of AD is highly intricate and the disease is apparent in the aged population ~ 50-70 years old. Even after > 100 years of research, the root origin of AD and its pathogenesis is unclear, complex and multifaceted. Herein, we have designed and synthesized 9 novel molecules with three different heterocyclic scaffolds namely pyrrolidone-2-one, quinoline & indoline-2-one to imitate and explore the novel chemical space around donepezil. The synthesized molecules were evaluated for their potential as anti-Alzheimer's agents through in-vitro and in-vivo studies in appropriate animal models. To further understand their interaction with acetylcholinesterase enzyme (AChE), extra-precision docking, and molecular dynamics simulation studies were carried out. As the number of compounds was limited to thoroughly explore the structure-activity relationship, atom-based 3D-quantitative structure-activity relationships (QSAR) studies were carried out to get more insights. All the designed compounds were found to inhibit AChE with IC50 in the micromolar range. From pyrrolidone-2-one series, 6-chloro-N-(1-(1-(3,4-dimethoxybenzyl)-2-oxopyrrolidin-3-yl)piperidin-4-yl)pyridine-3-sulfonamide (9), 2-(1-benzylpiperidin-4-yl)-6,7-dimethoxy-4-(4-methoxyphenyl)quinoline (18) from quinoline series and N-(1-benzylpiperidin-4-yl)-2-(2-oxoindolin-3-yl)acetamide (23) from indolin-2-one series inhibited AChE with an IC50 value of 0.01 µM. Based on other biochemical studies like lipid peroxidation, reduced glutathione, superoxide dismutase, catalase, nitrite, and behavioural studies (Morris water maze), compound 9 was found to be a potent AChE inhibitor which can be further explored as a lead molecule to design more potent and effective anti-Alzheimer's agents.

The impact of brivaracetam on cognitive processes and anxiety in various experimental models.

BACKGROUND: Memory deficits and anxiety symptoms are undesirable effects that occur in epilepsy patients. They may be associated with the pathophysiology of the disease but also with anticonvulsant therapy. Brivaracetam (BRV) is one of the newest antiseizure drugs. It acts as a ligand for synaptic vesicle glycoprotein 2A (SV2A), which may play a significant role in cognitive processes. Although BRV has a favorable safety profile, its central side effects remain unclear. Hence, this study aimed to evaluate the effect of BRV on various types of memory and anxiety in rats. METHODS: BRV was given to adult male Wistar rats (n = 80) via gastric tube as a single dose (6 mg/kg or 20 mg/kg) or chronically (6 mg/kg). The effect of the drug on spatial memory was evaluated in the Morris water maze (MWM), fear-learning by passive avoidance (PA), and recognition memory with novel object recognition (NOR). The elevated plus maze (EPM) was used to assess anxiety-like behaviors. RESULTS: The impact of BRV on memory is dose-dependent and mainly high doses may alter retrieval memory and fear-learning. Sub-chronic administration also impaired retrieval and spatial memory in animals. Moreover, chronic BRV may increase anxiety levels in rats but did not affect recognition memory. CONCLUSIONS: BRV may cause transient memory deficits as well as anxiety disturbances. However, the results are varied and depend on the type of memory, used dose, and duration of administration.

Thiolutin, a selective NLRP3 inflammasome inhibitor, attenuates cyclophosphamide-induced impairment of sperm and fertility in mice.

OBJECTIVE: The activation of the NLRP3 inflammasome has been implicated in male infertility. Our study aimed to investigate the therapeutic role of Thiolutin (THL), an inhibitor of the NLRP3 inflammasome, on oligoasthenospermia (OA) and to elucidate its mechanisms. MATERIALS AND METHODS: Semen from 50 OA and 20 healthy males were analyzed to assess the sperm quality and levels of inflammatory markers. Their correlation was determined using Pearson's correlation coefficient. The BALB/c mice were intraperitoneal injected by cyclophosphamide at 60 mg/kg/day for five days to induce OA, followed by a two-week treatment with THL or L-carnitine. Reproductive organ size and H&E staining were determined to observe the organ and seminiferous tubule morphology. ELISA and western blotting were utilized to measure sex hormone levels, inflammatory markers, and NLRP3 inflammasome levels. Furthermore, male and female mice were co-housed to observe pregnancy success rates. RESULTS: OA patients exhibited a decrease in sperm density and motility compared to healthy individuals, along with elevated levels of IL-1ß, IL-18 and NLRP3 inflammasome. In vivo, THL ameliorated OA-induced atrophy of reproductive organs, hormonal imbalance, and improved sperm density, motility, spermatogenesis and pregnancy success rates with negligible adverse effects on weight or liver-kidney function. THL also demonstrated to be able to inhibit the activation of NLRP3 inflammasome and associated proteins in OA mice. DISCUSSION: THL can improve sperm quality and hormonal balance in OA mice through the inhibition of NLRP3 inflammasome activation. Thus, THL holds promising potential as a therapeutic agent for OA.

Thiolutin has complex effects in vivo but is a direct inhibitor of RNA polymerase II in vitro.

Thiolutin is a natural product transcription inhibitor with an unresolved mode of action. Thiolutin and the related dithiolopyrrolone holomycin chelate Zn2+ and previous studies have concluded that RNA Polymerase II (Pol II) inhibition in vivo is indirect. Here, we present chemicogenetic and biochemical approaches to investigate thiolutin's mode of action in Saccharomyces cerevisiae. We identify mutants that alter sensitivity to thiolutin. We provide genetic evidence that thiolutin causes oxidation of thioredoxins in vivo and that thiolutin both induces oxidative stress and interacts functionally with multiple metals including Mn2+ and Cu2+, and not just Zn2+. Finally, we show direct inhibition of RNA polymerase II (Pol II) transcription initiation by thiolutin in vitro in support of classical studies that thiolutin can directly inhibit transcription in vitro. Inhibition requires both Mn2+ and appropriate reduction of thiolutin as excess DTT abrogates its effects. Pause prone, defective elongation can be observed in vitro if inhibition is bypassed. Thiolutin effects on Pol II occupancy in vivo are widespread but major effects are consistent with prior observations for Tor pathway inhibition and stress induction, suggesting that thiolutin use in vivo should be restricted to studies on its modes of action and not as an experimental tool.

Brivaracetam as add-on therapy in children with developmental epileptic encephalopathies: A study of 42 patients.

OBJECTIVE: Here we present a multicenter series of patients with developmental epileptic encephalopathies (DEE) who were treated with brivaracetam (BRV) as add-on therapy. METHODS: Medical records of 42 patients with DEE treated with add-on BRV seen at four pediatric neurology centers in Argentina between January 2021 and July 2023 were retrospectively analyzed. RESULTS: We included 42 patients (26 males, 16 females) with a mean age of 7 years (SD, ± 3.8; median, 9; range, 2-16). The children had different types of childhood-onset treatment-resistant DEEs and received BRV as add-on therapy for a mean period of 2 years (SD, ± 1.3 years; median, 1.5 years; range, 0.5-3 years). Thirty-three patients received levetiracetam (LEV) before the introduction of BRV. In nine patients, BRV was started without prior LEV because of behavioral disturbances. Three patients (9.5 %) became seizure free and 26/42 patients (62.1 %) had a greater than 50 % decrease in seizures after a mean follow-up of 21 months. Ten patients (23.8 %) had a 25-50 % seizure reduction, while seizure frequency remained unchanged in two (4.7 %) and increased in one patient (2.4 %). The interictal EEG abnormalities improved in all the responders. Adverse effects, consisting of drowsiness, irritability, and decreased appetite, were observed in seven patients (16.6 %), but did not lead to treatment discontinuation. CONCLUSION: Brivaracetam was found to be effective, safe, and well tolerated in children with DEE. In patients on LEV with behavioral disturbances, BRV may be tried. BRV may also be given without a previous trial with LEV in patients with behavioral problems.

Synaptic Terminal Density Early in the Course of Schizophrenia: An In Vivo UCB-J Positron Emission Tomographic Imaging Study of SV2A.

BACKGROUND: The synaptic hypothesis is an influential theory of the pathoetiology of schizophrenia (SCZ), which is supported by the finding that there is lower uptake of the synaptic terminal density marker [11C]UCB-J in patients with chronic SCZ than in control participants. However, it is unclear whether these differences are present early in the illness. To address this, we investigated [11C]UCB-J volume of distribution (VT) in antipsychotic-naïve/free patients with SCZ who were recruited from first-episode services compared with healthy volunteers. METHODS: Forty-two volunteers (SCZ n = 21, healthy volunteers n = 21) underwent [11C]UCB-J positron emission tomography to index [11C]UCB-J VT and distribution volume ratio in the anterior cingulate, frontal, and dorsolateral prefrontal cortices; the temporal, parietal and occipital lobes; and the hippocampus, thalamus, and amygdala. Symptom severity was assessed in the SCZ group using the Positive and Negative Syndrome Scale. RESULTS: We found no significant effects of group on [11C]UCB-J VT or distribution volume ratio in most regions of interest (effect sizes from d = 0.0-0.7, p > .05), with two exceptions: we found lower distribution volume ratio in the temporal lobe (d = 0.7, uncorrected p < .05) and lower VT/fp in the anterior cingulate cortex in patients (d = 0.7, uncorrected p < .05). The Positive and Negative Syndrome Scale total score was negatively associated with [11C]UCB-J VT in the hippocampus in the SCZ group (r = -0.48, p = .03). CONCLUSIONS: These findings indicate that large differences in synaptic terminal density are not present early in SCZ, although there may be more subtle effects. When taken together with previous evidence of lower [11C]UCB-J VT in patients with chronic illness, this may indicate synaptic density changes during the course of SCZ.