Aryl amino acetamides prevent Plasmodium falciparum ring development via targeting the lipid-transfer protein PfSTART1.

With resistance to most antimalarials increasing, it is imperative that new drugs are developed. We previously identified an aryl acetamide compound, MMV006833 (M-833), that inhibited the ring-stage development of newly invaded merozoites. Here, we select parasites resistant to M-833 and identify mutations in the START lipid transfer protein (PF3D7_0104200, PfSTART1). Introducing PfSTART1 mutations into wildtype parasites reproduces resistance to M-833 as well as to more potent analogues. PfSTART1 binding to the analogues is validated using organic solvent-based Proteome Integral Solubility Alteration (Solvent PISA) assays. Imaging of invading merozoites shows the inhibitors prevent the development of ring-stage parasites potentially by inhibiting the expansion of the encasing parasitophorous vacuole membrane. The PfSTART1-targeting compounds also block transmission to mosquitoes and with multiple stages of the parasite's lifecycle being affected, PfSTART1 represents a drug target with a new mechanism of action.

In vitro and in silico analyses reveal the toxicity of metolachlor to grass carp hepatocytes and the antagonism of melatonin.

Due to the widespread use of metolachlor (MET), the accumulation of MET and its metabolites in the environment has brought serious health problems to aquatic organisms. At present, the toxicity of MET on the physiological metabolism of aquatic animals mainly focused on the role of enzymes. There is still a lack of research on the molecular mechanisms of MET hepatotoxicity, especially on antagonizing MET toxicity. Therefore, this study focuses on grass carp hepatocytes (L8824 cells) closely related to toxin accumulation. By establishing a MET exposed L8824 cells model, it is determined that MET exposure induces pyrolytic inflammation of L8824 cells. Subsequent mechanistic studies found that MET exposure induces pyroptosis in L8824 cells through mitochondrial dysfunction, and siCaspase-1 inhibits the MET induced ROS production, suggesting a regulation of ROS-NLRP3- Caspase-1 pyroptotic inflammation cycling center in MET induced injury to L8824 cells. Molecular docking revealed a strong binding energy between melatonin (MT) and Caspase-1. Finally, a model of L8824 cells with MT intervention in MET exposure was established. MT can antagonize the pyroptosis induced by MET exposure in L8824 cells by targeting Caspase-1, thereby restoring mitochondrial function and inhibiting the ROS-pyroptosis cycle. This study discovered targets and mechanisms of MT regulating pyroptosis in MET exposed-L8824 cells, and the results are helpful to provide new targets for the design of MET antidotes.

2-(Methyl(phenyl)amino)-N-(phenyloxyphenyl)acetamide structural motif representing a framework for selective SIRT2 inhibition.

The mammalian cytoplasmic protein SIRT2, a class III histone deacetylase family member, possesses NAD+-dependent lysine deacetylase/deacylase activity. Dysregulation of SIRT2 has been implicated in the pathogenesis of several diseases, including neurological and metabolic disorders and cancer; thus, SIRT2 emerges as a potential therapeutic target. Herein, we identified a series of diaryl acetamides (ST61-ST90) by the structural optimization of our hit STH2, followed by enhanced SIRT2 inhibitory potency and selectivity. Among them, ST72, ST85, and ST88 selectively inhibited SIRT2 with IC50 values of 9.97, 5.74, and 8.92 µM, respectively. Finally, the entire study was accompanied by in silico prediction of binding modes of docked compounds and the stability of SIRT2-ligand complexes. We hope our findings will provide substantial information for designing selective inhibitors of SIRT2.

Novel Acetamide-Based HO-1 Inhibitor Counteracts Glioblastoma Progression by Interfering with the Hypoxic-Angiogenic Pathway.

Glioblastoma multiforme (GBM) represents the deadliest tumor among brain cancers. It is a solid tumor characterized by uncontrolled cell proliferation generating the hypoxic niches in the cancer core. By inducing the transcription of hypoxic inducible factor (HIF), hypoxia triggers many signaling cascades responsible for cancer progression and aggressiveness, including enhanced expression of vascular endothelial growth factor (VEGF) or antioxidant enzymes, such as heme oxygenase-1 (HO-1). The present work aimed to investigate the link between HO-1 expression and the hypoxic microenvironment of GBM by culturing two human glioblastoma cell lines (U87MG and A172) in the presence of a hypoxic mimetic agent, deferoxamine (DFX). By targeting hypoxia-induced HO-1, we have tested the effect of a novel acetamide-based HO-1 inhibitor (VP18/58) on GBM progression. Results have demonstrated that hypoxic conditions induced upregulation and nuclear expression of HO-1 in a cell-dependent manner related to malignant phenotype. Moreover, our data demonstrated that the HO-1 inhibitor counteracted GBM progression by modulating the HIFα/HO-1/VEGF signaling cascade in cancer cells bearing more malignant phenotypes.

Synthesis of new N-(5,6-methylenedioxybenzothiazole-2-yl)-2-[(substituted)thio/piperazine]acetamide/propanamide derivatives and evaluation of their AChE, BChE, and BACE-1 inhibitory activities.

In this study, the synthesis of N-(5,6-methylenedioxybenzothiazole-2-yl)-2-[(substituted)thio/piperazine]acetamide/propanamide derivatives (3a-3k) and to investigate their acetylcholinesterase (AChE), butyrylcholinesterase (BChE) and ß-secretase 1 (BACE-1) inhibition activity were aimed. Mass, 1H NMR, and 13C NMR spectra were utilized to determine the structure of the synthesized compounds. Compounds 3b, 3c, 3f, and 3j showed AChE inhibitory activity which compound 3c (IC50 = 0.030 ± 0.001 µM) showed AChE inhibitory activity as high as the reference drug donepezil (IC50 = 0.0201 ± 0.0010 µM). Conversely, none of the compounds showed BChE activity. Compounds 3c and 3j showed the highest BACE-1 inhibitory activity and IC50 value was found as 0.119 ± 0.004 µM for compound 3j whereas IC50 value was 0.110 ± 0.005 µM for donepezil, which is one of the reference substance. Molecular docking studies have been carried out using the data retrieved from the server of the Protein Data Bank (PDBID: 4EY7 and 2ZJM). Using in silico approach behavior active compounds (3c and 3j) and their binding modes clarified.

Design, synthesis, and biological evaluation of chalcone acetamide derivatives against triple negative breast cancer.

Chalcones are chemical scaffolds found in natural products, particularly in plants, and are considered for structural diversity in medicinal chemistry for drug development. Herein, we designed and synthesised novel acetamide derivatives of chalcone, characterizing them using 1H NMR, 13C NMR, HRMS, and IR spectroscopic methods. These derivatives were then screened against human cancer cells for cytotoxicity using the SRB assay. Among the tested derivatives, 7g, with a pyrrolidine group, exhibited better cell growth inhibition activity against triple-negative breast cancer (TNBC) cells. Further assays, including SRB, colony formation, and fluorescent dye-based microscopic analysis, confirmed that 7g significantly inhibited MDA-MB-231 cell proliferation. Furthermore, 7g promoted apoptosis by upregulating cellular reactive oxygen species (ROS) levels and disrupting mitochondrial membrane potential (MMP). Elevated expression of pro-apoptotic proteins (Bax and caspase-3) and a higher Bax/Bcl-2 ratio with downregulation of anti-apoptotic (Bcl-2) protein levels were observed in TNBC cells. The above results suggest that 7g can promote cellular death through apoptotic mechanisms in TNBC cells.

High-dose Agomelatine Combined with Haloperidol Decanoate Improves Cognition, Downregulates MT2, Upregulates D5, and Maintains Krüppel-like Factor 9 But Alters Cardiac Electrophysiology.

Haloperidol decanoate (HD) has been implicated in cognitive impairment. Agomelatine (AGO) has been claimed to improve cognition. We aimed at investigating the effects of HD + low- or high-dose AGO on cognition, verifying the melatonergic/dopaminergic to the cholinergic hypothesis of cognition and exploring relevant cardiovascular issues in adult male Wistar albino rats. HD + high-dose AGO prolonged the step-through latency by +61.47% (P < 0.0001), increased the time spent in bright light by +439.49% (P < 0.0001), reduced the time spent in dim light by -66.25% (P < 0.0001), and increased the percent of alternations by +71.25% (P < 0.0001), despite the reductions in brain acetylcholine level by -10.67% (P < 0.0001). Neurodegeneration was minimal, while the mean power frequency of the source wave was reduced by -23.39% (P < 0.05). Concurrently, the relative expression of brain melatonin type 2 receptors was reduced by -18.75% (P < 0.05), against increased expressions of dopamine type 5 receptors by +22.22% (P < 0.0001) and angiopoietin-like 4 by +119.18% (P < 0.0001). Meanwhile, electrocardiogram (ECG) demonstrated inverted P wave, reduced P wave duration by -36.15% (P < 0.0001) and PR interval by -19.91% (P < 0.0001), prolonged RR interval by +27.97% (P < 0.05), increased R wave amplitude by +523.15% (P < 0.0001), and a depressed ST segment and inverted T wave. In rats administered AGO, HD, or HD+ low-dose AGO, Alzheimer's disease (AD)-like neuropathologic features were more evident, accompanied by extensive ECG and neurochemical alterations. HD + high-dose AGO enhances cognition but alters cardiac electrophysiology. SIGNIFICANCE STATEMENT: Given the issue of cognitive impairment associated with HD and the claimed cognitive-enhancing activity of AGO, combined high-dose AGO with HD improved cognition of adult male rats, who exhibited minimal neurodegenerative changes. HD+ high-dose AGO was relatively safe regarding triggering epileptogenesis, while it altered cardiac electrophysiology. In the presence of low acetylcholine, the melatonergic/dopaminergic hypothesis, added to angiopoietin-like 4 and Krüppel-like factor 9, could offer some clue, thus offering novel targets for pharmacologic manipulation of cognition.

Synthesis of novel indol-3-acetamido analogues as potent anticancer agents, biological evaluation and molecular modeling studies.

Cannabinoids bind to cannabinoid receptors CB1 and CB2 and their antitumoral activity has been reported against some various cancer cell lines. Some synthetic cannabinoids possessing indole rings such as JWH-015 and JWH-133 particularly bind to the cannabinoid CB2 receptor and it was reported that they inhibit the proliferation and growth of various cancer cells without their psychoactive effects. However, the pharmacological action mechanisms of the cannabinoids are completely unknown. In this study, we report the synthesis of some new cannabinoidic novel indoles and evaluate their anticancer activity on various cancerous and normal cell lines (U87, RPMI 8226, HL60 and L929) using several cellular and molecular assays including MTT assay, real-time q-PCR, scratch assay, DAPI assay, Annexin V-PE/7AAD staining, caspase3/7 activity tests. Our findings indicated that compounds 7, 10, 13, 16, and 17 could reduce cell viability effectively. Compound 17 markedly increased proapoptotic genes (BAX, BAD, and BIM), tumor suppressor gene (p53) expression levels as well as the BAX/BCL-2 ratio in U87 cells. In addition, 17 inhibited cell migration. Based on these results, 17 was chosen for determining the mechanism of cell death in U87 cells. DAPI and Annexin V-7AAD staining results showed that 17 induced apoptosis, moreover activated caspase 3/7 significantly. Hence, compound 17, was selected as a lead compound for further pharmacomodulation. To rationalize the observed biological activities of 17, our study also included a comprehensive analysis using molecular docking and MD simulations. This integrative approach revealed that 17 fits tightly into the active site of the CB2 receptor and is involved in key interactions that may be responsible for its anti-proliferative effects.

MPoMA protects against lung epithelial cell injury via p65 degradation.

Numerous cases of lung injury caused by viral infection were reported during the coronavirus disease-19 pandemic. While there have been significant efforts to develop drugs that block viral infection and spread, the development of drugs to reduce or reverse lung injury has been a lower priority. This study aimed to identify compounds from a library of compounds that prevent viral infection that could reduce and prevent lung epithelial cell damage. We investigated the cytotoxicity of the compounds, their activity in inhibiting viral spike protein binding to cells, and their activity in reducing IL-8 production in lung epithelial cells damaged by amodiaquine (AQ). We identified N-(4-(4-methoxyphenoxy)-3-methylphenyl)-N-methylacetamide (MPoMA) as a non-cytotoxic inhibitor against viral infection and AQ-induced cell damage. MPoMA inhibited the expression of IL-8, IL-6, IL-1ß, and fibronectin induced by AQ and protected against AQ-induced morphological changes. However, MPoMA did not affect basal IL-8 expression in lung epithelial cells in the absence of AQ. Further mechanistic analysis confirmed that MPoMA selectively promoted the proteasomal degradation of inflammatory mediator p65, thereby reducing intracellular p65 expression and p65-mediated inflammatory responses. MPoMA exerted potent anti-inflammatory and protective functions in epithelial cells against LPS-induced acute lung injury in vivo. These findings suggest that MPoMA may have beneficial effects in suppressing viral infection and preventing lung epithelial cell damage through the degradation of p65 and inhibition of the production of inflammatory cytokines.

Microwave-Assisted Atom Transfer Radical Cyclization in the Synthesis of 3,3-Dichloro-γ- and δ-Lactams from N-Alkenyl-Tethered Trichloroacetamides Catalyzed by RuCl2(PPh3)3 and Their Cytotoxic Evaluation.

An expeditious synthesis of γ- and δ-lactams from tethered alkenyl trichloroacetamides in the presence of 5% of RuCl2(PPh3)3 is reported. In this investigation we have demonstrated that microwave activation significantly enhances reaction rates, leading to the formation of the corresponding lactams in yields ranging from good to excellent. Thus, we have been able to prepare a wide range of lactams, including indole and morphan bicyclic scaffolds, where the corresponding reactions were completely diastereoselective. This process was successfully extended to α,α-dichloroamides without affecting either their yield or their diastereoselectivity. Some of the lactams prepared in this work were evaluated for their hemolytic and cytotoxic responses. All compounds were found to be non-hemolytic at the tested concentration, indicating their safety profile in terms of blood cell integrity. Meanwhile, they exhibited interesting cytotoxicity responses that depend on both their lactam structure and cell line. Among the molecules tested, γ-lactam 2a exhibited the lowest IC50 values (100-250 µg/mL) as a function of its cell line, with promising selectivity against squamous carcinoma cells (A431) in comparison with fibroblasts (3T3 cell line).

A Potential Multitarget Insect Growth Regulator Candidate: Design, Synthesis, and Biological Activity of Novel Acetamido Derivatives Containing Hexacyclic Pyrazole Carboxamides.

Insect growth regulators (IGRs) are important green insecticides that disrupt normal growth and development in insects to reduce the harm caused by pests to crops. The ecdysone receptor (EcR) and three chitinases OfChtI, OfChtII, and OfChi-h are closely associated with the molting stage of insects. Thus, they are considered promising targets for the development of novel insecticides such as IGRs. Our previous work identified a dual-target compound 6j, which could act simultaneously on both EcR and OfChtI. In the present study, 6j was first found to have inhibitory activities against OfChtII and OfChi-h, too. Subsequently, taking 6j as a lead compound, 19 novel acetamido derivatives were rationally designed and synthesized by introducing an acetamido moiety into the amide bridge based on the flexibility of the binding cavities of 6j with EcR and three chitinases. Then, their insecticidal activities against Plutella xylostella (P. xylostella), Ostrinia furnacalis (O. furnacalis), and Spodoptera frugiperda (S. frugiperda) were carried out. The bioassay results revealed that most of these acetamido derivatives possessed moderate to good larvicidal activities against three lepidopteran pests. Especially, compound I-17 displayed excellent insecticidal activities against P. xylostella (LC50, 93.32 mg/L), O. furnacalis (LC50, 114.79 mg/L), and S. frugiperda (86.1% mortality at 500 mg/L), significantly better than that of 6j. In addition, further protein validation and molecular docking demonstrated that I-17 could act simultaneously on EcR (17.7% binding activity at 8 mg/L), OfChtI (69.2% inhibitory rate at 50 µM), OfChtII (71.5% inhibitory rate at 50 µM), and OfChi-h (73.9% inhibitory rate at 50 µM), indicating that I-17 is a potential lead candidate for novel multitarget IGRs. This work provides a promising starting point for the development of novel types of IGRs as pest management agents.

Protein quality control systems in the endoplasmic reticulum and the cytosol coordinately prevent alachlor-induced proteotoxic stress in Saccharomyces cerevisiae.

Alachlor, a widely used chloroacetanilide herbicide for controlling annual grasses in crops, has been reported to rapidly trigger protein denaturation and aggregation in the eukaryotic model organism Saccharomyces cerevisiae. Therefore, this study aimed to uncover cellular mechanisms involved in preventing alachlor-induced proteotoxicity. The findings reveal that the ubiquitin-proteasome system (UPS) plays a crucial role in eliminating alachlor-denatured proteins by tagging them with polyubiquitin for subsequent proteasomal degradation. Exposure to alachlor rapidly induced an inhibition of proteasome activity by 90 % within 30 min. The molecular docking analysis suggests that this inhibition likely results from the binding of alachlor to ß subunits within the catalytic core of the proteasome. Notably, our data suggest that nascent proteins in the endoplasmic reticulum (ER) are the primary targets of alachlor. Consequently, the unfolded protein response (UPR), responsible for coping with aberrant proteins in the ER, becomes activated within 1 h of alachlor treatment, leading to the splicing of HAC1 mRNA into the active transcription activator Hac1p and the upregulation of UPR gene expression. These findings underscore the critical roles of the protein quality control systems UPS and UPR in mitigating alachlor-induced proteotoxicity by degrading alachlor-denatured proteins and enhancing the protein folding capacity of the ER.

Evaluation of novel 2-(quinoline-2-ylthio)acetamide derivatives linked to diphenyl-imidazole as α-glucosidase inhibitors: Insights from in silico, in vitro, and in vivo studies on their anti-diabetic properties.

The inhibition of the α-glucosidase enzyme is crucial for targeting type 2 diabetes mellitus (DM). This study introduces a series of synthetic analogs based on thiomethylacetamide-quinoline derivatives linked to diphenyl-imidazole as highly potential α-glucosidase inhibitors. Twenty derivatives were synthesized and screened in vitro against α-glucosidase, revealing IC50 values ranging from 0.18 ± 0.00 to 2.10 ± 0.07 µM, in comparison to the positive control, acarbose. Among these derivatives, compound 10c (IC50 = 0.180 µM) demonstrated the highest potency and revealed a competitive inhibitory mechanism in kinetic studies (Ki = 0.15 µM). Docking and molecular dynamic evaluations elucidated the binding mode of 10c with the active site residues of the α-glucosidase enzyme. Moreover, in vivo assessments on a rat model of DM affirmed the anti-diabetic efficacy of 10c, evidenced by reduced fasting and overall blood glucose levels. The histopathological evaluation enhanced pancreatic islet architecture and hepatocytes in liver sections. In conclusion, novel 2-(quinoline-2-ylthio)acetamide derivatives as potent α-glucosidase inhibitors were developed. Compound 10c emerged as a promising candidate for diabetes management, warranting further investigation for potential clinical applications and mechanistic insights.

Activity refinement of aryl amino acetamides that target the P. falciparum STAR-related lipid transfer 1 protein.

Malaria is a devastating disease that causes significant morbidity worldwide. The development of new antimalarial chemotypes is urgently needed because of the emergence of resistance to frontline therapies. Independent phenotypic screening campaigns against the Plasmodium asexual parasite, including our own, identified the aryl amino acetamide hit scaffold. In a prior study, we identified the STAR-related lipid transfer protein (PfSTART1) as the molecular target of this antimalarial chemotype. In this study, we combined structural elements from the different aryl acetamide hit subtypes and explored the structure-activity relationship. It was shown that the inclusion of an endocyclic nitrogen, to generate the tool compound WJM-715, improved aqueous solubility and modestly improved metabolic stability in rat hepatocytes. Metabolic stability in human liver microsomes remains a challenge for future development of the aryl acetamide class, which was underscored by modest systemic exposure and a short half-life in mice. The optimized aryl acetamide analogs were cross resistant to parasites with mutations in PfSTART1, but not to other drug-resistant mutations, and showed potent binding to recombinant PfSTART1 by biophysical analysis, further supporting PfSTART1 as the likely molecular target. The optimized aryl acetamide analogue, WJM-715 will be a useful tool for further investigating the druggability of PfSTART1 across the lifecycle of the malaria parasite.

2-(3-Indolyl)acetamides and their oxazoline analogues: Anticancer SAR study.

We previously studied 2-aryl-2-(3-indolyl)acetohydroxamates as potential agents against melanoma. These compounds were ineffective in a mouse melanoma xenograft model, most likely due to unfavorable metabolic properties, specifically due to glucuronidation of the N-hydroxyl of the hydoxamic moiety. In the present work, we prepared a series of analogues, 2-aryl-2-(3-indolyl)acetamides and their oxazoline derivatives, which do not contain the N-hydroxyl group. We investigated the structure-activity relationship in both series of compounds and found that the 2-naphthyl is a preferred group at C-2 of the indole in the amide series, whereas the tetralin moiety is favorable in the same location in the oxazoline series. Overall, three compounds in the amide series have GI50 values as low as 0.2-0.3 µM and the results clearly indicate that the N-hydroxyl group is not necessary for high potency in vitro.

Molecular Docking, Synthesis, and Characterization of Furanyl-Pyrazolyl Acetamide and 2,4-Thiazolidinyl-Furan-3-Carboxamide Derivatives as Neuroinflammatory Protective Agents.

Microglia are key immune cells in the brain that maintain homeostasis and defend against immune threats. Targeting the dysfunctional microglia is one of the most promising approaches to inhibit neuroinflammation. In the current study, a diverse series of molecular hybrids were designed and screened through molecular docking against two neuroinflammatory targets, namely HMGB1 (2LY4) and HMGB1 Box A (4QR9) proteins. Based on the outcomes of docking scores fifteen compounds; ten furanyl-pyrazolyl acetamides 11(a-j), and five 2,4-thiazolidinyl-furan-3-carboxamide 15(a-e) derivatives were selected for further synthesis, followed by biological evaluation. The selected compounds, 11(a-j) and 15(a-e) were successfully synthesized with moderate to good yields, and structures were confirmed by IR, NMR, and mass spectra. The in-vitro cytotoxicity was evaluated on microglial cells namely BV-2, N-9, HMO6, leukemic HAP1, and human fibroblast cells. Further western-blot analysis revealed that 11h, 11f, 11c, 11j, 15d, 15c, 15e, and 15b compounds significantly suppressed anti-inflammatory markers such as TNF-α, IL-1, IL-6, and Bcl-2. All derivatives were moderate in potency compared to reference doxorubicin and could potentially act as novel anti-neuroinflammatory agents. This study can act as a beacon for further research in the application of furan-pyrazole and furan-2,4-thiazolidinediones as lead moieties for anti-neuroinflammatory and related diseases.

Design and Synthesis of 2-Phenylindolizine Acetamides: Molecular Docking, in Vitro Antimicrobial and Anticancer Activity Evaluation.

In the present work, we synthesized a small library of 2-phenylindolizine acetamide derivatives 7a-i and studied their biological activity. The synthesis was accomplished starting with easily available starting material phenacyl bromide 1 proceeding through the key intermediate 6-methyl-7-nitro-2-phenylindolizine 4. All the compounds 7a-i were characterized using spectroscopy viz., 1H-NMR, 13C NMR, FTIR, and mass spectrometry. Interestingly, 2-phenylindolizine scaffolds 7c, 7f and 7g revealed a remarkable antibacterial activity against relevant organisms S. aureus, E. coli, S. pneumoniae, P. aeruginosa. The target compounds 7e and 7h showed excellent anticancer activity against Colo-205 and MDA-MB-231 cell lines with IC50 values of 68.62, 62.91, 54.23 and 46.34 µM respectively. Additionally, all the 2-phenylindolizine acetamide derivatives 7a-i were subjected to molecular docking prediction by Autodock 4.2. Compounds 7a, 7f and 7c exhibited very good hydrogen bonding amino acid interactions Asp83 (2.23 Å), Asp83 (2.08 Å), His74 (2.05 Å), His76 (1.71 Å), Ser80 (1.05 Å) with active site of Topoisomerase-IV from S. pneumoniae (4KPE). Further, the compounds 7a-i have revealed acceptable ranges for drug-likeliness properties upon evaluation using SwissADME for ADMET and physiochemical properties.

Development of sperm cryopreservation protocol for patin buah, Pangasius nasutus.

The development of sperm cryopreservation for Pangasius nasutus is necessary in order to serve the growing demand of this species through artificial fertilization and the preservation of valuable strains of male broodstocks. In the present study, the basic protocol of sperm cryopreservation for P. nasutus was established by identifying the optimal conditions for optimum cryoprotectant, toxicity of cryoprotectants, extenders, freezing condition and dilution ratio. Methanol (MeOH) at 10% concentration had the best post-thaw motility (26.3 ± 0.9%) and curvilinear velocity (VCL) compared to dimethyl acetamide and dimethyl sulfoxide. MeOH was the least toxic cryoprotectant; sperm suspended in 5 and 10% MeOH maintained motility up to 50 min. No significant differences were detected between the three types of extenders tested (0.9% sodium chloride, Calcium-free Hanks' Balance salt solution and ringer solution). P. nasutus sperm had a narrow range of optimal cooling rate. Significantly higher post-thaw motility was identified when cooling at 9.23 °C min-1, obtained by freezing at height of 14 cm above liquid nitrogen vapor for 7 min, showing lower cooling rate is suitable for this species. However, when cooling below and above the optimal cooling rate, post-thaw motility dropped drastically. There were no significant differences among the dilution ratios investigated, indicating the volume of cryodiluent at all tested ratios (1:9, 1:19 and 1:49) was sufficient for the protection of cells during the cryopreservation process. The development of the protocol for cryopreserved P. nasutus sperm will assist artificial seed production and provide an important tool for genetic and breeding research.

Genome-wide study of glutathione transferases and their regulation in flufenacet susceptible and resistant black-grass (Alopecurus myosuroides Huds.).

BACKGROUND: Glutathione transferases (GSTs) are enzymes with a wide range of functions, including herbicide detoxification. Up-regulation of GSTs and their detoxification activity enables the grass weed black-grass (Alopecurus myosuroides Huds.) to metabolize the very-long-chain fatty acid synthesis inhibitor flufenacet and other herbicides leading to multiple herbicide resistance. However, the genomic organization and regulation of GSTs genes is still poorly understood. RESULTS: In this genome-wide study the location and expression of 115 GSTs were investigated using a recently published black-grass genome. Particularly, the most abundant GSTs of class tau and phi were typically clustered and often followed similar expression patterns but possessed divergent upstream regulatory regions. Similarities were found in the promoters of the most up-regulated GSTs, which are located next to each other in a cluster. The binding motif of the E2F/DP transcription factor complex in the promoter of an up-regulated GST was identical in susceptible and resistant plants, however, adjacent sequences differed. This led to a stronger binding of proteins to the motif of the susceptible plant, indicating repressor activity. CONCLUSIONS: This study constitutes the first analysis dealing with the genomic investigation of GST genes found in black-grass and their transcriptional regulation. It highlights the complexity of the evolution of GSTs in black-grass, their duplication and divergence over time. The large number of GSTs allows weeds to detoxify a broad spectrum of herbicides. Ultimately, more research is needed to fully elucidate the regulatory mechanisms of GST expression. © 2024 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Discovery of N-(1-(6-Oxo-1,6-dihydropyrimidine)-pyrazole) Acetamide Derivatives as Novel Noncovalent DprE1 Inhibitors against Mycobacterium tuberculosis.

Decaprenylphosphoryl-ß-d-ribose oxidase (DprE1) is a promising target for treating tuberculosis (TB). Currently, most novel DprE1 inhibitors are discovered through high-throughput screening, while computer-aided drug design (CADD) strategies are expected to promote the discovery process. In this study, with the aid of structure-based virtual screening and computationally guided design, a series of novel scaffold N-(1-(6-oxo-1,6-dihydropyrimidine)-pyrazole) acetamide derivatives with significant antimycobacterial activities were identified. Among them, compounds LK-60 and LK-75 are capable of effectively suppressing the proliferation of Mtb with MICMtb values of 0.78-1.56 µM, comparable with isoniazid and much superior to the phase II candidate TBA-7371 (MICMtb = 12.5 µM). LK-60 is also the most active DprE1 inhibitor derived from CADD so far. Further studies confirmed their high affinity to DprE1, good safety profiles to gut microbiota and human cells, and synergy effects with either rifampicin or ethambutol, indicating their broad potential for clinical applications.