Synthesis of novel 4-substituted isatin Schiff base derivatives as potential autophagy inducers and evaluation of their antitumour activity.

Induction of autophagic death in cancer cells is one of the promising strategies for the development of anti-cancer therapeutics. In the present study, we designed and synthesized a series of isatin Schiff base derivatives containing thioether structures. After discovering the highly active target compound H13 (IC50 = 4.83 µM) based on in vitro antiproliferation, we also found it had a high safety against normal cells HEK293 with CC50 of 69.01 µM, indicating a sufficient therapeutic window. In addition, to provide reference for subsequent studies, a model was successfully constructed by Sybyl software. Preliminary mechanistic studies suggested that H13-induced apoptosis may be closely related to ROS accumulation and mitochondrial dysfunction. Subsequent studies revealed that H13 inhibited cell proliferation by inducing cellular autophagy mainly through blocking signal of the PI3K/AKT/mTOR pathway. Altogether, these results suggested that H13 was potentially valuable as a lead compound.

A Novel Tryptanthrin Derivative D6 Induces Apoptosis and DNA Damage in Non-small-cell Lung Cancer Cells Through Regulating the EGFR Pathway.

BACKGROUND: Non-small-cell lung cancer is a prevalent malignancy associated with significant morbidity and mortality rates. Tryptanthrin and its derivatives have exhibited potent antitumor activity. OBJECTIVE: This study aims to investigate the inhibitory effect of a novel synthesized tryptanthrin derivative D6 on proliferation and the possible mechanism of human non-small cell lung cancer cell lines (A549) in vitro. METHODS: In this study, MTT assay, cell migration, colony formation assay, cell cycle analysis, cell apoptosis, JC- 1 staining assay, reactive oxygen species analysis, proteomics, western blotting, high content screening and absorption titrations analysis were performed. RESULTS: We found that D6 inhibited both the proliferation and migration, induced cell cycle arrest in the G2/M phase, increased levels of ROS, decreased mitochondrial membrane potential, and promoted apoptosis in A549 cells. Further mechanistic studies found that D6 reduced EGFR expression in A549 cells and inhibited the EGFR pathway by decreasing phosphorylation levels of EGFR, Stat3, AKT and Erk1/2. Moreover, DNA damage induced by D6 involved an increase in p53/MDM2 ratio and concentration-dependent accumulation of micronuclei. CONCLUSION: D6 demonstrated significant antitumor activity against A549 cells by inhibiting the EGFR signaling pathway, inducing DNA damage, and subsequently leading to oxidative stress, apoptosis, and cell cycle arrest. Our findings suggest that D6 exhibits potential as an NSCLC drug, owing to its attributes such as antiproliferative activity and ability to induce apoptosis by attenuating the EGFR-mediated signaling pathway.

Discovery of tryptanthrin analogues bearing F and piperazine moieties as novel phytopathogenic antibacterial and antiviral agents.

BACKGROUND: Plant bacterial infections and plant viruses seriously affect the yield and quality of crops. Based on the various activities of tryptanthrin, a series of tryptanthrin analogues bearing F and piperazine moieties were designed, synthesized, and evaluated for their biological activities against three plant bacteria and tobacco mosaic virus (TMV). RESULTS: Bioassay results indicated that compounds 6a-6l displayed excellent antibacterial activities in vitro and 6a-6c and 6g exhibited better antiviral activities against TMV than commercial ribavirin. In particular, 6b showed the most effect on Xanthomonas oryzae pv. oryzae (Xoo) with a half-maximal effective concentration (EC50 ) of 1.26 µg mL-1 , compared with the commercial pesticide bismerthiazol (BT; EC50 = 34.3 µg mL-1 ) and thiodiazole copper (TC; EC50 = 73.3 µg mL-1 ). Meanwhile, 6a also had the best antiviral activity at 500 µg mL-1 for curative, protection, and inactivation purposes, compared with ribavirin in vivo. CONCLUSION: Compound 6b could cause changes in bacterial morphology, induce the accumulation of reactive oxygen species, promote apoptosis of bacterial cells, inhibit the formation of biofilm, and block the growth of Xoo cells. Proteomic analysis revealed major differences in the bacterial secretory system pathways T2SS and T6SS, which inhibited membrane transport. Molecular docking revealed that 6a and 6g could interact with TMV coat protein preventing virus assembly. These results suggest that tryptanthrin analogues bearing F and piperazine moieties could be promising candidate agents for antibacterial and antiviral use in agricultural production. © 2023 Society of Chemical Industry.

Design, Synthesis, and Mechanism of Novel 9-Aliphatic Amine Tryptanthrin Derivatives against Phytopathogenic Bacteria.

Taking inspiration from the use of natural product-derived bactericide candidates in drug discovery, a series of novel 9-aliphatic amine tryptanthrin derivatives were designed, synthesized, and evaluated for their biological activity against three plant bacteria. The majority of these compounds exhibited excellent antibacterial activity in vitro. Compound 7c exhibited a significantly superior bacteriostatic effect against Xanthomonas axonopodis pv Citri (Xac), Xanthomonas oryzae pv Oryzae (Xoo), and Pseudomonas syringae pv Actinidiae (Psa) with final corrected EC50 values of 0.769, 1.29, and 15.5 µg/mL, respectively, compared to the commercial pesticide thiodiazole copper which had EC50 values of 58.8, 70.9, and 91.9 µg/mL. Preliminary mechanism studies have demonstrated that 7c is capable of altering bacterial morphology, inducing reactive oxygen species accumulation, promoting bacterial cell apoptosis, inhibiting normal cell growth, and affecting cell membrane permeability. Moreover, in vivo experiments have substantiated the effectiveness of 7c as a therapeutic and defensive agent against the citrus canker. The proteomic analysis has unveiled that the major disparities are located within the bacterial secretion system pathway, which hinders membrane transportation. These discoveries imply that 7c could be an auspicious prototype for developing antiphytopathogenic bacterial agents.

Novel 2-Amino-1,4-Naphthoquinone Derivatives Induce A549 Cell Death through Autophagy.

A series of 1,4-naphthoquinone derivatives containing were synthesized as anti-cancer agents and the crystal structure of compound 5a was confirmed by X-ray diffraction. In addition, the inhibitory activities against four cancer cell lines (HepG2, A549, K562, and PC-3) were tested, respectively, and compound 5i showed significant cytotoxicity on the A549 cell line with the IC50 of 6.15 µM. Surprisingly, in the following preliminary biological experiments, we found that compound 5i induced autophagy by promoting the recycling of EGFR and signal transduction in the A549 cell, resulting in the activation of the EGFR signal pathway. The potential binding pattern between compound 5i and EGFR tyrosine kinase (PDB ID: 1M17) was also identified by molecular docking. Our research paves the way for further studies and the development of novel and powerful anti-cancer drugs.

Discovery and Mechanism of Azatryptanthrin Derivatives as Novel Anti-Phytopathogenic Bacterial Agents for Potent Bactericide Candidates.

The natural alkaloids of tryptanthrin and their derivatives have a wide range of biological activities. In this research, four series of azatryptanthrin derivatives containing 4-aza/3-aza/2-aza/1-aza tryptanthrin were prepared by condensation cyclization reaction against plant pathogens to develop a new natural product-based bacterial pesticide. Compound 4Aza-8 displayed a remarkable growth inhibitory effect on pathogenic bacteria of Xanthomonas axonopodis pv. citri (Xac), Xanthomonas oryzae pv. Oryzae (Xoo), and Pseudomonas syringae pv. actinidiae (Psa) with the final corrected EC50 values of 0.312, 1.91, and 18.0 µg/mL, respectively, which were greatly superior than that of tryptanthrin (Tryp). Moreover, 4Aza-8 also showed effective therapeutic and protective activities in vivo on citrus canker. Further mechanism studies on Xac elucidated that compound 4Aza-8 was able to affect the growth curve of Xac and the formation of biofilm, cause severe shrinkage in bacterial morphology, increase reactive oxygen species levels, and induce apoptosis in bacterial cells. Quantitative analysis of differential protein profiles found that the major differences were mainly concentrated on the endometrial protein in the bacterial secretion system pathway, which blocked the membrane transport and affected the transfer of DNA to the host cell. In summary, these research results suggest that 4Aza-8 represents a promising anti-phytopathogenic-bacteria agent, which is worth being further investigated as a bactericide candidate.

Discovery of Tryptanthrin and Its Derivatives and Its Activities against NSCLC In Vitro via Both Apoptosis and Autophagy Pathways.

In this study, a series of novel tryptanthrin derivatives were synthesized and their inhibitory activities against selected human cancer cell lines, namely, lung (A549), chronic myeloid leukemia (K562), prostate (PC3), and live (HepG2), were evaluated using a methyl thiazolyl tetrazolium colorimetric (MTT) assay. Among the tested compounds, compound C1 exhibited a promising inhibitory effect on the A549 cell line with an IC50 value of 0.55 ± 0.33 µM. The observation of the cell morphological result showed that treatment with C1 could significantly inhibit the migration of A549 cells through the cell migration assay. Moreover, after treatment with C1, the A549 cells exhibited a typical apoptotic morphology and obvious autophagy. In addition, the detection of apoptosis and the mitochondrial membrane potential indicated that C1 induced A549 cell apoptosis via modulating the levels of Bcl2 family members and disrupted the mitochondrial membrane potential. Compound C1 also suppressed the expression of cyclin D1 and increased the expression of p21 in the A549 cells, inducing cell cycle arrest in the G2/M phase in a dose dependent manner. The further mechanism study found that C1 markedly increased the transformation from LC3-I to LC3-II. Taken together, our results suggest that C1 is capable of inhibiting the proliferation of non-small cell lung cancer (NSCLC) cells, inducing cell apoptosis, and triggering autophagy.

Synthesis and biological assessment of indole derivatives containing penta-heterocycles scaffold as novel anticancer agents towards A549 and K562 cells.

Herein, a new series of 2-chloro-N-(5-(2-oxoindolin-3-yl)-4H-pyrazol-3-yl) acetamide derivatives containing 1,3,4-thiadiazole (10a-i) and 4H-1,2,4-triazol-4-amine (11a-r) moiety was designed, synthesised as novel anticancer agents. The antiproliferative activity values indicated that compound 10 b stood as the most potent derivative with IC50 values of 12.0 nM and 10 nM against A549 and K562 cells, respectively. Mechanism investigation and docking studies of 10 b indicated that it possessed good apoptosis characteristic and dose-dependent growth arrest of A549 and K562 cells, blocked cell cycle into G2/M phase. Interestingly, 10 b suppressed the growth of A549 and K562 cells via modulation of EGFR and p53-MDM2 mediated pathway.

Toehold-mediated ligation-free rolling circle amplification enables sensitive and rapid imaging of messenger RNAs in situ in cells.

In situ analysis of tumor-related messenger RNAs (mRNAs) is significant in identifying cancer cells at the genetic level in the early stage. Rolling circle amplification (RCA)-based methods are primary tools for in situ mRNA assay, however, the necessary ligation reaction not only shows low ligation efficiency, but also greatly prolongs the assay time that increases the risk of cells losing and mRNAs leakage. In this work, we propose a novel toehold-mediated ligation-free RCA (TMLFRCA) on a designed structure-switchable dumbbell-shaped probe (SDP). Target mRNA can specifically activate SDP from its circular form by toehold strand displacement, thereby initiates in situ RCA for mRNA imaging with the help of a short DNA primer. For the proof-of-concept demonstration, the TK1 mRNA was sensitively detected by TMLFRCA in less than 3.5 h with a limit of detection (LOD) of 0.39 fM (corresponds to 2.39×108copiesL-1), and significantly improved specificity capable for distinguishing single base difference. The sensitivity of the TMLFRCA for TK1 mRNA in situ assay is ∼29-fold and ∼7-fold higher than that of FISH and ligase-assisted RCA method, respectively, which enables the TMLFRCA method capability of highly sensitive and specific distinction mRNA expression levels between cancer cells and normal cells. We believe this TMLFRCA strategy would be of great value in both basic research and clinical diagnosis.

Polymerization retardation isothermal amplification strategy enables the sensitive and facile investigation of the flanking sequence preference of ten-eleven translocation 2 protein.

Active DNA demethylation process critically relies on the intrinsic properties of ten-eleven translocation proteins (Tets), particularly the flanking sequence preference. Challenged by the fact that the proximate bases to the 5-methylcytosine (5mC) are multitudinous and their influence on the Tets/DNA interplay is minute, the current methodologies are very limited in terms of cost, sensitivity and efficiency. Herein, we propose a polymerization retardation isothermal amplification (PRIA) strategy that enables sensitive and fast study of the flanking sequence preference of Tet. By arranging DNA polymerase to repetitively pass DNA strands through an isothermal replication-scission amplification reaction, the tiny difference in the Tet/DNA interplay can be consecutively accumulated and amplified. Low amount sample (80 ng) even multiple samples can be simultaneously analyzed within 10 h on an easily accessible laboratory real-time quantitative PCR instrument. For a proof-of-concept study, the binding preference (PB) of Tet2 for XmCGX, (X = C, G, T, A) was analyzed by PRIA and computational analysis, showing an order of AmCGT > TmCGA ≈ GmCGC > CmCGG. Furthermore, the binding and oxidation preference in Tet/DNA interplay process was individually considered. By comparative evaluation of the total flanking sequence preference (PT) and the PB, for the first time, we revealed that Tet2 has a lower oxidation preference (PO) to proximal flanking bases and the main contributor to PT of Tet2 is PB. The PRIA strategy, due to its reliable, cost-effective, high efficiency and low sample input, would hopefully advance epigenetic researches and other relative studies.

Matrine enhances the efficacy of adriamycin chemotherapy in osteosarcoma cells by the STAT3 pathway.

Matrine and adriamycin have been extensively considered to be effective in anticancer therapies. However, the role of matrine in the antitumor activity of adriamycin against human osteosarcoma (OS) remains elusive. The aim of this study was to investigate the effect of matrine in OS chemotherapy of adriamycin. In the study, we found that matrine promoted the inhibitory effect of adriamycin against OS cell proliferation and growth. Wound healing and transwell assays showed that the inhibitory effect of adriamycin against migration and invasion of OS cells was significantly enhanced by matrine. For the underlying mechanism investigation, we showed that adriamycin reduced the protein level of PCNA, MMP-9, phosphorylated STAT3, and survivin, which was further intensified by the application of matrine. These results show that matrine could promote the therapeutic efficacy of adriamycin against human OS.

Metastable Dumbbell Probe-Based Hybridization Chain Reaction for Sensitive and Accurate Imaging of Intracellular-Specific MicroRNAs In Situ in Living Cells.

Sensitive and accurate imaging of intracellular-specific microRNAs (miRNAs) in situ in living cells is seriously challenged by the susceptibility of nucleic acid probes and the low dynamics of the hybridization reaction in cellular environments. Herein, we engineer a set of new metastable dumbbell probes (M xDPs) to overcome these key limitations by concurrently boosting transfection, antidigestibility, assembly dynamics, and nanostructural uniformity. The M xDPs can maintain their stability up to 16 h in living cells and produce uniform and dense DNA nanostructures rapidly (<2 h) and specifically from a hybridization chain reaction (HCR). A sharp signal from the cascade accumulation of fluorescence resonance energy transfer (FRET) further minimizes the effect of system fluctuations. The M xDPs-based HCR (M xDPHCR) method showed identical performance in the analysis of miR-27a in cell lysate and buffer condition and obtained a limit of detection down to 3.2 pM (corresponding to 160 amol per 50 µL), which is 44-fold lower than on conventional hairpin probes. The M xDPHCR method clearly distinguished normal cells from tumor cells and provided more accurate quantitative information on the intracellular-specific miRNAs. The strategy would offer a powerful tool for visualizing and localizing desired nucleic acids in living cells.

Real-Time Sensing of TET2-Mediated DNA Demethylation In Vitro by Metal-Organic Framework-Based Oxygen Sensor for Mechanism Analysis and Stem-Cell Behavior Prediction.

Active DNA demethylation, mediated by O2-dependent ten-eleven translocation (TET) enzymes, has essential roles in regulating gene expression. TET kinetics assay is vital for revealing mechanisms of demethylation process. Here, by a metal-organic framework (MOF)-based optical O2 sensor, we present the first demonstration on real-time TET2 kinetics assay in vitro. A series of luminescent Cu(I) dialkyl-1,2,4-triazolate MOFs were synthesized, which were noble-metal-free and able to intuitively response to dissolved O2 in a wide range from cellular hypoxia (≤15 µM) to ambient condition (∼257 µM). By further immobilization of the MOFs onto transparent silicon rubber (MOF@SR) to construct O2 film sensors, and real-time monitoring of O2 consumption on MOF@SR over the reaction time, the complete TET2-mediated 5-methylcytosine (5mC) oxidation process were achieved. The method overcomes the limitations of the current off-line methods by considerably shortening the analytical time from 0.5-18 h to 10 min, and remarkably reducing the relative standard deviation from 10%-68% to 0.68%-4.2%. As a result, the Michaelis-Menten constant ( Km) values of TET2 for 5mC and O2 in ascorbic acid-free (AA-) condition were precisely evaluated to be 24 ± 1 and 43.8 ± 0.3 µM, respectively. By comparative study on AA-containing (AA+) conditions, and further establishing kinetics models, the stem-cell behavior of TETs was successfully predicted, and the effects of key factors (AA, O2, Fe2+) on TETs were revealed, which were fully verified in mouse embryonic stem (mES) cells. The method is promising in wide application in kinetics analysis and cell behavior prediction of other important O2-related enzymes.

Short-probe-based duplex-specific nuclease signal amplification strategy enables imaging of endogenous microRNAs in living cells with ultrahigh specificity.

Specific nucleic acids amplification at a constant and mild temperature is important for imaging assay of endogenous microRNAs (miRNAs) in living cells. Duplex-specific nuclease (DSN) is attractive in one-step isothermal assay of miRNA; however, its intrinsic limitations of low amplification specificity and high reaction temperature greatly restrict the application scope. Herein, we present a short-probe-based DSN signal amplification (spDSNSA) strategy enabling analysis of miRNAs at body temperature with significantly high specificity. From systematic investigation of amplification reaction on different types of DNA probes, we revealed that the annealing rate between probe and target miRNA greatly affects the dynamics of amplification process. By simply shortening the length of DNA probe, the spDSNSA remarkably improved specificity without loss of amplification efficiency at 37 °C. As a proof-of-concept, let-7a was sensitively detected by spDSNSA with a limit of detection down to 30 p.M., and a specificity 102 ‒ 104 folds higher than those of traditional DSNSA methods. The analysis of the let-7a in the lysates of A549 human lung cancer cells and BEAS-2B human lung normal bronchial epithelial cells exhibited well agreement with rt-qPCR method. Furthermore, the endogenous let-7a in A549 and BEAS-2B living cells was clearly imaged without damaging the original morphology of cells. The method provide a facile idea for extension of DNS related signal amplification strategies in the application in living cells and POCTs, and would pose a great impact on the development of simple and rapid molecular diagnostic applications for short oligonucleotides.

[Treatment of anterior canal benign paroxysmal positional vertigo by Yacovino repositioning maneuver].

OBJECTIVE: To evaluate the efficacy of Yacovino repositioning maneuver in patients with anterior semicircular canal benign paroxysmal positional vertigo (ASC-BPPV). METHOD: Nine patients were diagnosed as ASC-BPPV from January 2013 to October 2014. All the patients were performed with Yacovino repositioning maneuver and the effective rate were evaluated by Dix-Hallpike tests. RESULT: Among the nine ASC-BPPV patients, 2 cases were successfully controlled by the first maneuver, 2 cases by the second time, and the nystagmus of 1 case was disappeared after 1 months' follow-up. The remaining 3 cases were respectively followed up till 7,8, 12 months with consistent positional downbeat nystagmus. CONCLUSION: Being a relative low incidence disease, of ASC-BPPV also has low effective rate after Yacovino repositioning maneuver.

[The correlations between varying tinnitus severity and anxiety and depression in non-acute tinnitus patients].

OBJECTIVE: To investigate the correlation betwen varying degrees of non-acute tinnitus and anxiety and depression. METHOD: Seventy-seven outpatients with non-acute tinnitus as their in chief complaint were submitted to Tinnitus Handicap Inventory(THI), Self-Rating Anxiety Scale(SAS), and Self-Rating Depression Scale (SDS). RESULT: THI and its three subscales were found to have significant correlations with SAS and SDS. The group (THI ≥ 38) had more anxiety and depression than the mild (THI < 38). Significant correlations were also observed between THI, SAS and SDS in the group with THI ≥ 38, in contrast with the group of THI < 38. CONCLUSION: Patients with THI ≥ 38 suffered from severe anxiety and depression than the mild. Doctors should pay more attention to these patients, especially their psychological disorders.

Broadband conversion of visible light to near-infrared emission by Ce3+, Yb3+-codoped yttrium aluminum garnet.

We show that Ce(3+) can be an efficient sensitizer for Yb(3+) in the host lattice of yttrium aluminum garnet (YAG). With blue-light excitation to induce the 4f-->5d transition of Ce(3+), characteristic near-IR emission of Yb(3+) due to transition of (2)F(5/2)-->(2)F(7/2) peaking at 1030 nm is generated as a result of energy transfer from Ce(3+) to Yb(3+). The result of spectral evolution with temperature indicates that the efficiency of energy transfer is enhanced owing to thermal effect. This evidence implies that the phonon-assisted process participates in the downconversion of YAG: Ce(3+), Yb(3+).

Infrared broadband emission of bismuth-doped barium-aluminum-borate glasses.

We report near infrared broadband emission of bismuth-doped barium-aluminum-borate glasses. The broadband emission covers 1.3microm window in optical telecommunication systems. And it possesses wide full width at half maximum (FWHM) of ~200nm and long lifetime as long as 350micros. The luminescent properties are quite sensitive to glass compositions and excitation wavelengths. Based on energy matching conditions, we suggest that the infrared emission may be ascribed to 3P1? 3P0 transition of Bi+. The broad infrared emission characteristics of this material indicate that it might be a promising candidate for broadband optical fiber amplifiers and tunable lasers.