Isoglutaminyl Cyclase Overexpression Enhances KYSE30 Cancer Cell Proliferation and Migration via the MAPK Signaling Pathway.

To understand how upregulated isoglutaminyl cyclase (isoQC) is involved in the initiation of diseases such as cancer, we developed a human KYSE30 carcinoma cell model in which isoQC was stably overexpressed. GO and KEGG analysis of the DEGs (228) and DEPs (254) respectively implicated isoQC on the proliferation invasion and metastasis of cells and suggested that isoQC might participate in the regulation of MAPK, RAS, circadian rhythm, and related pathways. At the functional level, isoQC-overexpressing KYSE30 cells showed enhanced proliferation, migration, and invasion capacity. Next, we decided to study the precise effect of isoQC overexpression on JNK, p-JNK, AKT, p-AKT, ERK, p-ERK, and PER2, as RNA levels of these proteins are significantly correlated with signal levels indicated in RNA-Seq analysis, and these candidates are the top correlated DEPs enriched in RT-qPCR analysis. We saw that only p-ERK expression was inhibited, while PER2 was increased. These phenotypes were inhibited upon exposure to PER2 inhibitor KL044, which allowed for the restoration of p-ERK levels. These data support upregulated isoQC being able to promote cancer cell proliferation and migration in vitro, likely by helping to regulate the MAPK and RAS signaling pathways, and the circadian protein PER2 might be a potential mediator.

Structure-activity relationship of dual inhibitors containing maleimide and imidazole motifs against glutaminyl cyclase and glycogen synthase kinase-3ß.

Alzheimer's disease (AD) is a major cause of dementia and one of the most common chronic diseases affecting the aging population. Because AD is considered a public health priority, there is a critical need to discover novel and effective agents for the treatment of this condition. In view of the known contribution of up-regulated glutaminyl cyclase (QC) and glycogen synthase kinase-3ß (GSK-3ß) to the initiation of AD, we previously evaluated a series of dual inhibitors containing maleimide and imidazole motifs as potential anti-AD agents. Here, we assessed another series of hybrids containing maleimide and imidazole motifs to gain an in-depth understanding of the structure-activity relationship (SAR). Based on the primary screening, the introduction of 5-methyl imidazole at one side of the molecule did not enhance the QC-specific inhibitory activity of these hybrids (2, IC50 = 1.22 µM), although the potency was increased by 2' substitution on the maleimide motif at the other side of the molecule. Interestingly, compounds containing 5-methyl imidazole exhibited stronger GSK-3ß-specific inhibitory activity (2, IC50 = 0.0021 µM), and the electron-withdrawing group and 2' and 3' substitution were favorable. Further investigation of substitutions on the maleimide motif in compounds 14-35 revealed that QC-specific inhibition in the presence of piperidine was improved by introduction of a methoxy group (R2). Increasing the linker length and introduction of a methoxy group (R2) also increased the GSK-3ß-specific inhibitory potency. These findings were further confirmed by molecular docking analysis of 33 and 24 with QC and GSK-3ß. Overall, these hybrids exhibited enhanced inhibitory potency against both QC and GSK-3ß, highlighting an important strategy for improving the potency of hybrids as dual-targeting anti-AD agents.

Discovery of potential scaffolds for glutaminyl cyclase inhibitors: Virtual screening, synthesis, and evaluation.

Glutaminyl cyclase (QC) plays a crucial role in the early stages of Alzheimer's disease (AD), thus inhibition of QC may be a promising strategy for the treatment of early AD. Therefore, QC inhibitors with novel chemical scaffolds may contribute to the development of additional anti-AD agents. We conducted a virtual screening of 3 million compounds from the Chemdiv and Enamine databases, to discover potential scaffolds for QC inhibitors. Three scaffolds, 120974, 147706, and 141449, were selected from this structure-based virtual screening through a combination of pharmacophore modeling, a receptor-ligand pharmacophore model, and the GALAHAD model, and furtherly filtered by chelation with zinc ion and docking properties. Consequently, three compounds, 1, 2, and 3, were designed and synthesized based on these three scaffolds, respectively. The IC50 of compounds 1 and 3 against QC were 14.19 ± 4.21 and 4.34 ± 0.35 µM, respectively. Our results indicate that the new scaffolds selected using a virtual screening process exhibit potential as novel QC inhibitors.

Cytotoxic Compounds from Belamcanda chinensis (L.) DC Induced Apoptosis in Triple-Negative Breast Cancer Cells.

Four compounds (1, 5, 7, and 8) were first isolated from the genus Belamcanda Adans. nom. conserv., and six known compounds (2-4, 6, 9, and 10) were isolated from the rhizome of Belamcanda chinensis (L.) DC. Their structures were confirmed by spectroscopic data. Herein, compounds 1-10 were rhapontigenin, trans-resveratrol, 5,7,4'-trihydroxy-6,3',5'-trimethoxy-isoflavone, irisflorentin, 6-hydroxybiochannin A, iridin S, pinoresinol, 31-norsysloartanol, isoiridogermanal, and iristectorene B, respectively. All compounds were evaluated for their antiproliferative effects against five tumor cell lines (BT549, 4T1, MCF7, MDA-MB-231, and MDA-MB-468). Among them, compound 9 (an iridal-type triterpenoid) showed the highest activity against 4T1 and MDA-MB-468 cells. Further studies displayed that compound 9 inhibited cell metastasis, induced cells cycle arrest in the G1 phase, exhibited significant mitochondrial damage in 4T1 and MDA-MB-468 cells including excess reactive oxygen species, decreased mitochondrial membrane potential, and induced 4T1 and MDA-MB-468 cell apoptosis for the first time. In summary, these findings demonstrate that compound 9 exerts promising potential for triple-negative breast cancer treatment and deserves further evaluation.

Isolation and Identification of Chemical Compounds from Agaricus blazei Murrill and Their In Vitro Antifungal Activities.

This study explores the antifungal properties of Agaricus blazei Murrill, a valuable medicinal and edible fungus. Six compounds (1-6) were first isolated from A. blazei using various isolation techniques and identified using spectroscopic methods. These compounds include linoleic acid, 1,1'-oxybis(2,4-di-tert-butylbenzene), glycerol monolinoleate, volemolide (17R)-17-methylincisterol, (24s)-ergosta-7-en-3-ol, and dibutyl phthalate. This study also assesses the antifungal activities of these compounds against Trichophyton mentagrophology, Trichophyton rubrum, Candida albicans, and Cryptococcus neoformans. The results demonstrate varied sensitivities against these pathogenic fungi, with compound 2 showing significant inhibition against T. mentagrophology, compound 3 showing significant inhibition against T. rubrum, and compound 6 showing significant inhibition against C. albicans. This study underscores the medicinal potential of A. blazei as an antifungal agent and sheds light on its valuable research implications.

METTL7B contributes to the malignant progression of glioblastoma by inhibiting EGR1 expression.

Glioblastoma (GBM), a predominant central nervous system (CNS) malignancy, is correlated with high mortality and severe morbidity. Mammalian methyltransferase-like 7B (METTL7B) as a methyltransferase has been identified to participate in cancer progression. However, its function in GBM is elusive. Accordingly, we aimed to explore the effect of METTL7B on GBM. The expression of METTL7B and EGR2 in GBM patients and GBM cells were detected by qPCR, western blots and immunohistochemical staining. Cell viability was assessed by CCK-8 assays. Cell proliferation was determined by EdU, colony formation, and tumor sphere formation assays. METTL7B shRNA was injected into the Balb/c nude mice. The size and weight of isolated tumor was measured. And the expression levels of Ki67, METTL7B and EGR1 were examined by immunohistochemical staining. METTL7B was significantly elevated, while EGR1 was downregulated in clinical GBM tissues. METTL7B upregulation was associated with the low overall survival of GBM patients. Moreover, METTL7B depletion remarkably attenuated GBM cell proliferation. Mechanistically, METTL7B overexpression inhibited EGR1 expression in GBM cells. EGR1 knockdown rescued the inhibitory effect of METTL7B depletion on GBM cell proliferation. Meanwhile, METTL7B depletion arrested more GBM cells at the G0/G1, but fewer cells at the S phase, which EGR1 knockdown reversed these effects. Furthermore, tumorigenicity analysis revealed that METTL7B promotes tumor growth of GBM cells in vivo. METTL7B contributes to the malignant progression of GBM by inhibiting EGR1 expression. METTL7B and EGR1 may be utilized as the treatment targets for GBM therapy.

Regulation of ferroptosis by bioactive phytochemicals: Implications for medical nutritional therapy.

Ferroptosis is an iron- and lipotoxicity-dependent regulated cell death that has been implicated in various diseases, such as cancer, neurodegeneration and stroke. The biosynthesis of phospholipids, coenzyme Q10, and glutathione, and the metabolism of iron, amino acids and polyunsaturated fatty acid, are tightly associated with cellular sensitivity to ferroptosis. Up to now, only limited drugs targeting ferroptosis have been documented and exploring novel effective ferroptosis-modulating compound is needed. Natural bioactive products are conventional resources for drug discovery, and some of them have been clinically used against cancers and neurodegenerative diseases as dietary supplements or pharmaceutic agents. Notably, increasing evidence demonstrates that natural compounds, such as saponins, flavonoids and isothiocyanates, can either induce or inhibit ferroptosis, further expanding their therapeutic potentials. In this review, we highlight current advances of the emerging molecular mechanisms and disease relevance of ferroptosis. We also systematically summarize the regulatory effects of natural phytochemicals on ferroptosis, and clearly indicate that saponins, terpenoids and alkaloids induce ROS- and ferritinophagy-dependent ferroptosis, whereas flavonoids and polyphenols modulate iron metabolism and nuclear factor erythroid 2-related factor 2 (NRF2) signaling to inhibit ferroptosis. Finally, we explore their clinical applications in ferroptosis-related diseases, which may facilitate the development of their dietary usages as nutraceuticals.

Peroxide- and transition metal-free electrochemical synthesis of α,ß-epoxy ketones.

A novel electrochemical method for the synthesis of α,ß-epoxy ketones is reported. With KI as the redox mediator, methyl ketones reacted with aldehydes under peroxide- and transition metal-free electrolytic conditions and afforded α,ß-epoxy ketones in one pot (36 examples, 52-90% yield). This safe and environmental-friendly method has a broad substrate scope and can readily provide a variety of α,ß-epoxy ketones in gram-scales for evaluation of their anti-cancer activities.

A sensitive LC-MS assay using derivatization with boron trifluoride to quantify curcuminoids in biological samples.

A procedure is described to measure curcumin (C), demethoxycurcumin (DMC), bisdemethoxycurcumin (BDMC), tetrahydrocurcumim (TC) and their glucuronidated metabolites (CG, DMCG, and BDMCG) in plasma, brain, liver and tumor samples. The procedure involves converting the analytes to their boron difluoride derivatives and analyzing them by combined liquid chromatography coupled to an ion trap mass spectrometer operating in the negative ion MSn scan mode. The method has superb limits of detection of 0.01 nM for all curcuminoids and 0.5 nM for TC and the glucuroniated metabolites, and several representative chromatograms of biological samples containing these analytes are provided. In addition, the pharmacokinetic profile of these compounds in one human who daily consumed an over-the-counter curcuminoid product shows the peak and changes in circulating concentrations achieved by this mode of administration.

Glutaminyl cyclase inhibitor contributes to the regulation of HSP70, HSP90, actin, and ribosome on gene and protein levels in vitro.

Because of the crucial roles of upregulated glutaminyl cyclase (QC) in the initiation and development of Alzheimer's disease (AD), QC inhibitors are supposed as disease-modifying agents for the treatment of AD. And reported compounds encourage this hypothesis greatly based on the remarkable anti-AD effects in vivo. To illustrate the mechanism in detail, the actions of a selected QC inhibitor (23) were assessed firstly in a cell system here. It was demonstrated that QC activities and the generation of pyroglutamate-modified ß-amyloids in PC12 cells were both inhibited obviously after the treatment of 23. A total of 13 and 15 genes were up- and downregulated significantly in treated cells by RNA-sequencing analysis. Quantitative real-time polymerase chain reaction, enzyme-linked immunosorbent assay, WB, and immunofluorescence analysis supported the effects of 23 on the transcriptome of PC12 cells consequently. The expressions of chaperones, heat shock proteins (HSP) 70, and 90, were upreglutated, while gene expression of actin and the level of encoded protein were reduced significantly in PC12 cells with the treatment. Furthermore, the regulations of ribosome were observed after the treatment. These results indicate the potency of 23 to improve the translation, expression and folding regulation of proteins and affect the multivalent cross-linking of cytoskeletal protein and other proteins subsequently in the cell system and might contribute to the understanding of the mechanism of QC inhibitor as potential anti-AD agents.

Inhibitory effect of flavonoids on human glutaminyl cyclase.

Glutaminyl cyclase (QC) plays an important role in the pathogenesis of Alzheimer's disease (AD) and can be a potential target for the development of novel anti-AD agents. However, the study of QC inhibitors are still less. Here, phenol-4' (R1-), C5-OH (R2-) and C7-OH (R3-) modified apigenin derivatives were synthesized as a new class of human QC (hQC) inhibitors. The efficacy investigation of these compounds was performed by spectrophotometric assessment and the structure-activity relationship (SAR) was evaluated. Molecular docking was also carried out to analyze the binding mode of the synthesized flavonoid to the active site of hQC.

Syntheses of tanshinone anhydrides and their suppression on oxidized LDL uptake in macrophages and foam cell formation.

We synthesized eight tanshinone anhydrides and the alcoholytic derivatives through a mild oxygen-insertion under Pd/C catalytic hydrogenation conditions. The suppressive effects of the anhydrides on the oxidized low-density lipoprotein (oxLDL) uptake and the oxLDL-induced macrophage-derived foam cell formation were studied. Our results revealed that both anhydrides 1a and 2a could significantly suppress the oxLDL uptake in macrophages and the foam cell formation at micromolar level, which might be partially attributed to their inhibition of oxLDL-induced LOX-1 expression in macrophages.

Development of lactoferrin-coated multifunctional copolymer micelles to cross the blood-brain barrier.

The blood-brain barrier (BBB) prevents pathogens and toxins in the bloodstream from reaching the brain, but also inhibits the delivery of agents intended to treat central nervous system disorders, such as Alzheimer's disease (AD). In this study, we prepared and evaluated a novel nano-delivery vehicle system composed of lactoferrin-conjugated (Lf-PIC@Se) micelles. We used a COOH-PEG-PAsp-PV@Se synthesis-based method to prepare the micelles, which involved self-assembly followed by EDC-NHS coupling. Using glutaminyl cyclase inhibitor 8 as a model encapsulated chemical, Lf-PIC@Se micelles achieved a good loading capacity. In vitro analysis demonstrated that Lf-PIC@Se/8 micelles were stable in both neutral and acidic pH solutions in the presence or absence of H2O2, and confirmed their biosafety and compatibility in PC12 and bEND.3 cells. Notably, the cell uptake of Lf-PIC@Se/C6 micelles was much higher than that of PIC@Se micelles, and occurred through LfR-mediated endocytosis. The presence of Se meant that Lf-PIC@Se micelles acted as ROS scavengers in PC12 cells under H2O2-induced oxidative stress, which inhibited oxidative damage and increased mitochondrial membrane potential. Hemolysis assays further demonstrated that Lf-PIC@Se represent a biocompatible carrier. Finally, in vivo experiments in mice suggested that Lf-PIC@Se micelles successfully crossed the BBB, confirming their potential as vehicles for drug delivery when treating AD and other central nervous system disorders.

Chitosan-based oral nanoparticles as an efficient platform for kidney-targeted drug delivery in the treatment of renal fibrosis.

There is increasingly keen interest in developing orally delivered targeted drugs, especially for diseases that require long-term medication. Hence, we manufactured nanoparticles derived from methoxypolyethylene glycol-chitosan (PCS) to enhance the oral delivery and kidney-targeted distribution of salvianolic acid B (SalB), a naturally occurring renoprotective and anti-fibrotic compound, as a model drug for the treatment of renal fibrosis. Orally administered SalB-loaded PCS nanoparticles (SalB-PCS-NPs) maintained good stability in the gastrointestinal environment, improved mucus-penetrating capacity, and enhanced transmembrane transport through a Caco-2 cell monolayer. The relative oral bioavailability of SalB-PCS-NPs to free SalB and SalB-loaded chitosan nanoparticles (SalB-CS-NPs) was 367.0 % and 206.2 %, respectively. The structural integrity of SalB-PCS-NPs after crossing the intestinal barrier was also validated by Förster resonance energy transfer (FRET) in vitro and in vivo. Fluorescein isothiocyanate (FITC)-labeled SalB-PCS-NPs showed higher kidney accumulation than free FITC and FITC-labeled SalB-CS-NPs (4.6-fold and 2.1-fold, respectively). Significant improvements in kidney function, extracellular matrix accumulation, and pathological changes were observed in a unilateral ureter obstruction mouse model of renal fibrosis after once daily oral treatment with SalB-PCS-NPs for 14 days. Thus, oral administration of SalB-PCS-NPs represents a promising new strategy for kidney-targeted drug delivery.

Design, synthesis and anti-AD effects of dual inhibitor targeting glutaminyl cyclase/GSK-3ß.

Alzheimer's disease (AD), multifactorial disease, is recognized as one of the most common forms of dementia, and the efficacy of anti-AD drugs is limited clinically. Up-regulated glutaminyl cyclase (QC) and glycogen synthase kinase-3ß (GSK-3ß) have been identified as two critical elements involved in AD recently. Here, a series of novel chemicals containing maleimide and imidazole motif were designed and synthesized as dual inhibitors targeting QC and GSK-3ß. Based on primary screening, compound 2 (2.26 µM), 5 (2.37 µM), 8 (1.34 µM), 21 (2.44 µM), 25 (0.36 µM), 27 (1.76 µM), 28 (1.04 µM), 33 (2.08 µM) and 34 (2.33 µM) exhibited notable human QC (hQC) inhibitory potency, while compound 1 (0.014 µM), 7 (0.04 µM), 8 (0.057 µM), 19 (0.034 µM), 24 (0.014 µM), 32 (0.032 µM), 38 (0.051 µM), 39 (0.044 µM), 44 (0.048 µM), 47 (0.011 µM), 49 (0.021 µM) and so on showed remarkable GSK-3ß inhibitory activities. And as expected, these chemicals possessed significant inhibitory potency on both hQC and GSK-3ß, such as compound 1 (2.80 and 0.014 µM), 8 (1.34 and 0.057 µM), 25 (0.36 and 0.15 µM), 27 (1.76 and 0.069 µM), 28 (1.04 and 0.090 µM), 33 (2.08 and 0.19 µM), 34 (2.33 and 0.11 µM), 35 (2.55 and 0.14 µM), 36 (2.34 and 0.11 µM), etc. Subsequent in vivo studies demonstrated that compound 8 attenuated cognitive deficits and decreased the anxiety-like behavior in 3 × Tg-AD mice. The treatment decreased both pE-Aß and Aß accumulation by inhibiting the activity of QC, and decreased the hyperphosphorylation of Tau by reducing the levels of GSK-3ß in the brains of AD mice. Results obtained in this research suggested that these novel compounds could be supposed as potential anti-AD agents targeting QC and GSK-3ß.

Near-infrared AIE-active phosphorescent iridium(III) complex for mitochondria-targeted photodynamic therapy.

Mitochondria-targeted photodynamic therapy (PDT) has recently been recognized as a promising strategy for effective cancer treatment. In this work, a mitochondria-targeted near-infrared (NIR) aggregation-induced emission (AIE)-active phosphorescent Ir(III) complex (Ir1) is reported with highly favourable mitochondria-targeted bioimaging and cancer PDT properties. Complex Ir1 has strong absorption in the visible light region (∼500 nm) and can effectively produce singlet oxygen (1O2) under green light (525 nm) irradiation. It preferentially accumulates in the mitochondria of human breast cancer MDA-MB-231 cells as revealed by colocalization analysis. Complex Ir1 displays high phototoxicity toward human breast cancer MDA-MB-231 cells and mouse breast cancer 4T1 cells. Complex Ir1 induces reactive oxygen species (ROS) production, mitochondrial dysfunction, and endoplasmic reticulum (ER) stress in MDA-MB-231 cells upon photoirradiation, leading to apoptotic cell death. The favorable PDT performance of Ir1in vivo has been further demonstrated in tumour-bearing mice. Together, the results suggest that Ir1 is a promising photosensitizer for mitochondria-targeted imaging and cancer phototherapy.

A novel chemosensor based on rhodamine derivative for colorimetric and fluorometric detection of Cu2+ in aqueous solution.

A new rhodamine-based reversible chemosensor (2) was synthesized, which exhibits high sensitivity and selectivity for Cu(2+) but no significant response toward other competitive metal ions in aqueous solution. Upon the addition of Cu(2+), the spirolactam ring of 2 was opened and the solution color changed from colorless to red. Strangely, an unexpected fluorescence quenching was observed, which is contrary to the fluorescence turn-on of the most rhodamine-based chemosensors. The likely novel sensing mechanism has been proposed.

Essential oil components from Asarum sieboldii Miquel are toxic to the house dust mite Dermatophagoides farinae.

In our effort to develop novel plant-derived acaricides, we examined the contact and fumigant toxicity of Asarum heterotropoides (Asarum sieboldii Miquel) essential oil constituents to the house dust mite Dermatophagoides farinae (Acari: Pyroglyphidae). Ten constituents, including methyl eugenol (relative amount 42.18 %), were identified by gas chromatography-mass spectroscopy (GC-MS) in the A. sieboldii Miq. essential oil. In contact toxicity tests, methyl eugenol (4.2 µg/cm(2), 24 h LD50) was most toxic to D. farinae, followed by benzyl benzoate (9.1 µg/cm(2)), A. sieboldii Miq. essential oil (37.7 µg/cm(2)), and dibutyl phthalate (DBP 57.9 µg/cm(2)). The potency of methyl eugenol and A. sieboldii Miq. essential oil was higher than benzyl benzoate and DBP, with mortalities of 100, 100, 94.6, and 13.2 %, respectively, after 2.5 h of exposure. In the vapor phase mortality bioassay, methyl eugenol and A. sieboldii Miq. essential oil resulted in 100 % mortality in closed containers after 24-h exposure, but only 4.7 and 7.9 %, respectively, in open containers, indicating that the toxicity in these tests was largely due to the vapor phase. Methyl eugenol and A. sieboldii Miq. essential oil merit further study as potential D. farinae control compounds.

Acaricidal activity of DHEMH, derived from patchouli oil, against house dust mite, Dermatophagoides farinae.

This study characterized the acaricidal activity of constituents of patchouli oil extracted from (Pogostemon cablin (Blanco) BENTH) against the house dust mite, Dermatophagoides farinae. A new compound, 2-(1,3-dihydroxy-but-2-enylidene)-6-methyl-3-oxo-heptanoic acid (DHEMH), was isolated from patchouli oil and characterized by (1)H-NMR, (13)C-NMR, LC-MS and elemental analysis (EA). This active component was identified as the hydrolysate of pogostone. Fifteen other constituents found in patchouli oil were also identified by GC-MS, including patchouli alcohol and pogostone. LD(50) studies carried out over 24 h using contact toxicity tests identified DHEMH as the most toxic compound to D. farinae (2.04 µg/cm(2)), followed by patchouli oil (6.11 µg/cm(2)), benzyl benzoate (BP) (9.31 µg/cm(2)) and dibutyl phthalate (DBP) (58.52 µg/cm(2)). In vapor phase toxicity tests, all of these compounds were more effective in closed than open containers, indicating that the most efficient mode of delivery for these compounds is the vapor phase. These results indicate that DHEMH and patchouli oil merit further study as potential agents for the control of D. farinae.

The Leaves of Cyclocarya paliurus: A Functional Tea with Preventive and Therapeutic Potential of Type 2 Diabetes.

Type 2 diabetes mellitus (T2DM) has become a universal and chronic global public health concern and causes multiple complex complications. In order to meet the rapidly growing demand for T2DM treatment, increased research has been focused on hypoglycemic drugs. Cyclocarya paliurus (Batal.) Iljinsk is the only living species of the genus Cyclocarya Iljinskaja, whose leaves have been extensively used as a functional tea to treat obesity and diabetes in China. An enormous amount of very recent pharmacological research on the leaves of C. paliurus has demonstrated that they carry out numerous biological activities, such as hypoglycemic, anti-inflammatory, and intestinal microbiota regulation. Multiple in vitro and in vivo studies have also shown that the extracts of C. paliurus leaves are innocuous and safe. This study aims to provide an up-to-date review of the botany, traditional uses, phytochemistry, pharmacological effects against diabetes, toxicology, and clinical studies of C. paliurus leaves, in hopes of promoting a better understanding of their role in the prevention and treatment of T2DM.