Blocking senescence and tolerogenic function of dendritic cells induced by γδ Treg cells enhances tumor-specific immunity for cancer immunotherapy.

BACKGROUND: Regulatory T (Treg) cells are a key component in maintaining the suppressive tumor microenvironment and immune suppression in different types of cancers. A precise understanding of the molecular mechanisms used by Treg cells for immune suppression is critical for the development of effective strategies for cancer immunotherapy. METHODS: Senescence development and tolerogenic functions of dendritic cells (DCs) induced by breast cancer tumor-derived γδ Treg cells were fully characterized using real-time PCR, flow cytometry, western blot, and functional assays. Loss-of-function strategies with pharmacological inhibitor and/or neutralizing antibody were used to identify the potential molecule(s) and pathway(s) involved in DC senescence and dysfunction induced by Treg cells. Impaired tumor antigen HER2-specific recognition and immune response of senescent DCs induced by γδ Treg cells were explored in vitro and in vivo in humanized mouse models. In addition, the DC-based HER2 tumor vaccine immunotherapy in breast cancer models was performed to explore the enhanced antitumor immunity via prevention of DC senescence through blockages of STAT3 and programmed death-ligand 1 (PD-L1) signaling. RESULTS: We showed that tumor-derived γδ Treg cells promote the development of senescence in DCs with tolerogenic functions in breast cancer. Senescent DCs induced by γδ Treg cells suppress Th1 and Th17 cell differentiation but promote the development of Treg cells. In addition, we demonstrated that PD-L1 and STAT3 signaling pathways are critical and involved in senescence induction in DCs mediated by tumor-derived γδ Treg cells. Importantly, our complementary in vivo studies further demonstrated that blockages of PD-L1 and/or STAT3 signaling can prevent γδ Treg-induced senescence and reverse tolerogenic functions in DCs, resulting in enhanced HER2 tumor-specific immune responses and immunotherapy efficacy in human breast cancer models. CONCLUSIONS: These studies not only dissect the suppressive mechanism mediated by tumor-derived γδ Treg cells on DCs in the tumor microenvironment but also provide novel strategies to prevent senescence and dysfunction in DCs and enhance antitumor efficacy mediated by tumor-specific T cells for cancer immunotherapy.

TMEM97/Sigma 2 Receptor Increases Estrogen Receptor α Activity in Promoting Breast Cancer Cell Growth.

Aberrant estrogen receptor (ER) signaling is a major driver of breast tumor growth and progression. Sigma 2 receptor has long been implicated in breast carcinogenesis based on pharmacological studies, but its molecular identity had been elusive until TMEM97 was identified as the receptor. Herein, we report that the TMEM97/sigma 2 receptor is highly expressed in ER-positive breast tumors and its expression is strongly correlated with ERs and progesterone receptors (PRs) but not with HER2 status. High expression levels of TMEM97 are associated with reduced overall survival of patients. Breast cancer cells with increased expression of TMEM97 had a growth advantage over the control cells under both nutrition-limiting and sufficient conditions, while the knockdown of TMEM97 expression reduced tumor cell proliferations. When compared to their vector control cells, MCF7 and T47D cells with increased TMEM97 expression presented increased resistance to tamoxifen treatment and also grew better under estrogen-depleted conditions. The TMEM97/sigma 2 receptor enhanced the ERα transcriptional activities and increased the expression of genes responsive to estrogen treatment. Increased TMEM97 also stimulated the mTOR/S6K1 signaling pathways in the MCF7 and T47D cells. The increased level of active, phosphorylated ERα, and the enhanced resistance to tamoxifen treatment with increased TMEM97, could be blocked by an mTOR inhibitor. The knockdown of TMEM97 expression reduced the ERα and mTOR/S6K1 signaling activities, rendering the cells with an increased sensitivity to tamoxifen. The observations suggest that the TMEM97/sigma 2 receptor is a novel regulator of ERα activities in breast tumor cell growth.

Blockades of effector T cell senescence and exhaustion synergistically enhance antitumor immunity and immunotherapy.

BACKGROUND: Current immunotherapies still have limited successful rates among cancers. It is now recognized that T cell functional state in the tumor microenvironment (TME) is a key determinant for effective antitumor immunity and immunotherapy. In addition to exhaustion, cellular senescence in tumor-infiltrating T cells (TILs) has recently been identified as an important T cell dysfunctional state induced by various malignant tumors. Therefore, a better understanding of the molecular mechanism responsible for T cell senescence in the TME and development of novel strategies to prevent effector T cell senescence are urgently needed for cancer immunotherapy. METHODS: Senescent T cell populations in the TMEs in mouse lung cancer, breast cancer, and melanoma tumor models were evaluated. Furthermore, T cell senescence induced by mouse tumor and regulatory T (Treg) cells in vitro was determined with multiple markers and assays, including real-time PCR, flow cytometry, and histochemistry staining. Loss-of-function strategies with pharmacological inhibitors and the knockout mouse model were used to identify the potential molecules and pathways involved in T cell senescence. In addition, melanoma mouse tumor immunotherapy models were performed to explore the synergistical efficacy of antitumor immunity via prevention of tumor-specific T cell senescence combined with anti-programmed death-ligand 1 (anti-PD-L1) checkpoint blockade therapy. RESULTS: We report that both mouse malignant tumor cells and Treg cells can induce responder T cell senescence, similar as shown in human Treg and tumor cells. Accumulated senescent T cells also exist in the TME in tumor models of lung cancer, breast cancer and melanoma. Induction of ataxia-telangiectasia mutated protein (ATM)-associated DNA damage is the cause for T cell senescence induced by both mouse tumor cells and Treg cells, which is also regulated by mitogen-activated protein kinase (MAPK) signaling. Furthermore, blockages of ATM-associated DNA damage and/or MAPK signaling pathways in T cells can prevent T cell senescence mediated by tumor cells and Treg cells in vitro and enhance antitumor immunity and immunotherapy in vivo in adoptive transfer T cell therapy melanoma models. Importantly, prevention of tumor-specific T cell senescence via ATM and/or MAPK signaling inhibition combined with anti-PD-L1 checkpoint blockade can synergistically enhance antitumor immunity and immunotherapy in vivo. CONCLUSIONS: These studies prove the novel concept that targeting both effector T cell senescence and exhaustion is an effective strategy and can synergistically enhance cancer immunotherapy.

Exploration of the H2O2 Oxidation Process and Characteristic Evaluation of Humic Acids from Two Typical Lignites.

To study the effect of H2O2 on the content and properties of humic acids (HAs) in lignites, the experimental conditions including oxidation time, H2O2 concentration, and the solid-liquid ratio were investigated. Under the optimum oxidation conditions, the contents of HAs of YL and HB lignite were 45.4 and 40.9%, respectively. The HAs extracted from oxidized and raw lignites were characterized and compared. The results showed that the HAs extracted from oxidized lignites contain more total acidic groups, carboxyl groups, and aliphatic carbon than that in HAs extracted from raw lignites, and their hydrophilic-hydrophobic index value is higher and thermooxidative stability is better than those in HAs extracted from raw lignites. In addition, the composition of polycyclic aromatic hydrocarbons and fluorophore types in HAs extracted from oxidized lignites are similar to the HAs extracted from raw lignites. The results indicated that the oxidation operation can increase the content of HAs in lignites, and simultaneously increase the content of oxygen-containing functional groups and biological activity of HAs, which provided a reference for the subsequent application of HAs.

Megestrol acetate is a specific inducer of CYP3A4 mediated by human pregnane X receptor.

PURPOSE: Megestrol acetate is a synthetic progestogen used to treat some cancers and cancer-associated cachexia, but its potential interactions with other drugs are not well known. This study aims to determine the regulation of drug metabolizing enzymes by megestrol acetate. METHODS: Primary human hepatocytes were treated and analyzed by PCR array to identify genes involved in drug metabolism that are impacted by megestrol acetate. P450 3A4 (CYP3A4) reporter gene assay and HPLC analyses of nifedipine metabolites were used to determine CYP3A4 gene expression and activities. Competitive ligand binding assay was used to determine the affinity of megestrol acetate toward human pregnane x receptor (hPXR). Electrophoretic mobility shift assay and mammalian two hybrid assay were used to determine the mechanism of megestrol to activate hPXR. RESULTS: The levels and activities of CYP3A4 were significantly induced (> 4-folds) by megestrol acetate in human hepatocytes and HepG2 cells. Megestrol treatment induced CYP3A4 through the activation of hPXR, a ligand-activated transcription factor that plays a role in drug metabolism and transport. Other tested nuclear receptors showed no response. The mechanism studies showed that megestrol activated hPXR by binding to the ligand binding domain (LBD) of hPXR and increasing the recruitment of the cofactors such as steroid receptor cofactor (SRC-1). CONCLUSION: The results suggest that megestrol acetate is a specific inducer of CYP3A4 mediated by hPXR and therefore has the potential to cause drug interactions, especially in the co-administration with drugs that are substrates of CYP3A4.

Structural Characterization and Molecular Simulation of Baoqing Lignite.

The molecular structure of Baoqing lignite was analyzed by ultimate analysis, Fourier transform infrared spectroscopy, X-ray diffraction spectroscopy, 13C solid-state nuclear magnetic resonance, and X-ray photoelectron spectroscopy. The results revealed that the aromaticity of Baoqing lignite is 27.64%, and the aromatic structure mainly contains benzene and naphthalene. The aliphatic structure consists of alkyl side chains and cycloalkyl. Oxygen atoms are present in phenol, ether, carbonyl, and carboxyl groups; nitrogen atoms are chiefly in pyridine and pyrrole; sulfur atoms mainly exist in sulfoxide sulfur. The molecular structure model of Baoqing lignite was constructed based on experimental data, and the molecular formula is C184H199O50N2S. The molecular configuration was optimized by adopting the M06-2X basis set in the framework of density functional theory. Moreover, the simulated FTIR spectrum was in good agreement with the experimental spectra, proving the accuracy of the molecular structure. The molecular model of Baoqing lignite contains a majority of aliphatic structures and aromatic rings with a poor condensation degree. Moreover, the aromatic layers irregularly arrange in space.

Production of biosilica nanoparticles from biomass power plant fly ash.

When it comes to the combustion of biomass, per ton of solid biofuel will generate 70 kg ash on average. Additionally, these ashes have a high specific surface area, especially fly ash, which may adsorb harmful substances and damage to human health. This work was aimed to reutilize biomass power plant fly ash to produce silica material, to reduce the hazard of ash landfill for environment. The ash underwent acid leaching with 1.5 M HCl after proper heating pre-treatment. Then, 2 M NaOH was direct to react with residue to obtain sodium silicate. Finally, acid titration of solution was used to precipitate silica. The results showed that the amorphous silica has been produced from fly ash successfully with the purity from 44.41% to 93.63% and yield of 20.45%, and the optimal calcination conditions for amorphous transformation of silica in fly ash were temperature of 611 °C with time of 5 h and the minimum crystallinity was 17.41%, modeled with response surface methodology. Spectroscopy analysis revealed that the three-dimensional network silica was hydroxylated to form the linear structure. Thermal analysis indicated that the decomposition of silanol groups tend to be stable at 400 °C, but the ash was decomposing up to 1000 °C. Morphological analysis demonstrated that BET surface area ranged from 24 m2/g to 115 m2/g, agglomerate particle size from 380.9 nm to 178.8 nm, when the ash was conversion to spherical silica. Consequently, it is possible to turn blend biomass fly ash into amorphous silica nanoparticles.

A comparative study on the structural features of humic acids extracted from lignites using comprehensive spectral analyses.

The lignite reserves of Zhaotong and Mile in China are abundant and lignite utilizations are limited, however, humic acids (HAs) extracted from lignites play a significant role in many fields including agriculture, environmental protection and so on. Herein, the structures of HAs extracted from Zhaotong and Mile lignites (denoted as ZLHA and MLHA, respectively) were characterized and compared to each other using comprehensive spectral analyses. As a result, the UV-Vis spectrum analyses of HAs indicated that the molecular weight of MLHA is larger than that of ZLHA. Cross polarization magic angle spinning 13C NMR, which is rarely used to analyze the structures of HAs using fitting peaks, and FT-IR spectrum analyses indicated that both the aromaticity and the oxygen-containing group contents of ZLHA are higher than those of MLHA, and the HAs' aromaticity could be confirmed by the results of the X-ray diffraction patterns. Additionally, the main existing forms of the elements in the HAs were obtained from X-ray photoelectron spectrum analyses, which are not commonly used for HA analyses. In this work, the utilization of comprehensive spectral analyses was an effective method to study the structural features of ZLHA and MLHA and it could provide a basic reference for the applications of ZLHA and MLHA.

Re-emergence of a genotype VIII virulent Newcastle disease virus isolated from Chinese game fowl after 13 years.

Circulation of dominant genotypes VI and VII of Newcastle disease virus (NDV) is causing significant economic losses to the poultry industry in China. However, reports of Newcastle disease (ND) outbreaks caused by genotype VIII strains of NDV are rare. In this study, a virulent genotype VIII strain of NDV, designated GXGB2011, was isolated from a vaccinated game fowl flock showing clinic signs of infection in Pinxiang county, Guangxi, China. The whole genome of the isolate was completely sequenced and was found to be comprised of 15,192 nucleotides (nt), encoding the six structural proteins in the order of 3'-NP-P-M-F-HN-L-5'. The pattern of cleavage site 112 RRQKR↓F117 in the fusion (F) protein and the intracerebral pathogenicity index (ICPI) value of 1.5 showed that the strain GXGB2011 was a velogenic NDV. The results of the challenge experiment with the 5-week-old SPF chickens showed that the strain was highly pathogenic with 100% morbidity and mortality of the challenged birds. Based on the detection of virus in different organs of the infected birds, the highest viral load in caecal tonsils was observed and viral levels in immune organs were higher than those in the respiratory organs. Bayesian reconstruction of complete genomes based on the sequences of 66 NDV reference strains showed that the strain belonged to the genotype VIII of NDV. Phylogenetic analysis showed that the strain was more closely related to the foreign strains gamefowl/U.S.(CA)/24225/98, 1ITTY94060 and IT-147/94 rather than to the first domestic strains of the emergence genotype VIII in Qinghai, China during 1979-1985. In summary, the results of the study demonstrated the re-emergence of a highly pathogenic virulent isolate of genotype VIII of NDV. These results indicate the risk that this genotype VIII of NDV may spread to commercial chickens from game fowl.

Moderate levels of dietary arachidonic acid reduced lipid accumulation and tended to inhibit cell cycle progression in the liver of Japanese seabass Lateolabrax japonicus.

To investigate the physiological roles of dietary arachidonic acid (ARA) in fish, a feeding trial with Japanese seabass was conducted, followed by a hepatic transcriptome assay. Six experimental diets differing basically in ARA level (0.05%, 0.22%, 0.37%, 0.60%, 1.38% and 2.32% of dry matter) were used in the feeding trial. Liver samples from fish fed diets with 0.05% and 0.37% ARA were subjected to transcriptomic assay, generating a total of 139 differently expressed unigenes, which were primarily enriched in lipid metabolism and cell cycle-related signaling pathways. Then, qRT-PCR validation on lipid metabolism and cell cycle-related genes as well as corresponding enzyme-linked immunosorbent assay of selected proteins were conducted with liver samples from all six groups. Moderated ARA levels reduced lipogenesis and stimulated ß-oxidation concurrently, but high ARA levels seemed to affect lipid metabolism in complicated ways. Both gene expression and protein concentration of cell cycle-related proteins were decreased by moderate levels of dietary ARA. The lipid content and fatty acid composition in fish confirmed the transcription and protein concentration results related to lipid metabolism. In conclusion, moderate levels of dietary ARA (0.37% and 0.60%) reduced lipid accumulation and tended to inhibit cell cycle progression in the liver of Japanese seabass.

Molecular characterization of two novel sub-sublineages of pigeon paramyxovirus type 1 in China.

Pigeon paramyxovirus type 1 (PPMV-1) infection is enzootic in pigeon flocks and poses a potential risk to the poultry industry in China. To gain insight into the biological characteristics and transmission routes of circulating PPMV-1 in pigeons, 13 PPMV-1 isolates from domestic pigeons isolated during 2011-2015 in Guangxi province, China, were characterized using a pathogenicity assessment and phylogenetic analysis. All PPMV-1 isolates were mesogenic or lentogenic strains and had a mean death time (MDT) in 9-day-old SPF chicken embryos and a intracerebral pathogenicity index (ICPI) values of 54-154 h and 0.00-0.90, respectively. Analysis of the F and HN gene sequences of the PPMV-1 isolates and the Newcastle Disease (ND) vaccine strain La Sota, revealed that the nucleotide sequence similarity of the F and HN genes were all < 85% between the PPMV-1 isolates and La Sota, significantly lower than those > 98% among the PPMV-1 isolates. The amino acids sequence of the F protein at the cleavage site of the 13 PPMV-1 isolates was 112RRQKR↓F117, characteristic of virulent Newcastle disease virus (NDV). All 13 isolates were classified as sublineage 4b by phylogenetic analysis and evolutionary distances, based on the F gene sequences. It was also found that the 13 isolates were divided into two novel sub-groups of sublineage 4b, sub-sublineages 4biig and 4biih. Since these two novel sub-sublineages had two different geographic sources, we speculated that they represent two different transmission routes of PPMV-1 in China. Phylogenetic analysis of these isolates will help to elucidate the sources of the transmission and evolution of PPMV-1 and may help to control PPMV-1 infection in the pigeon industry in China.

Effects of different dietary DHA:EPA ratios on gonadal steroidogenesis in the marine teleost, tongue sole (Cynoglossus semilaevis).

The present study was conducted to investigate the effects of dietary DHA and EPA on gonadal steroidogenesis in mature females and males, with a feeding trial on tongue sole, a typical marine teleost with sexual dimorphism. Three experimental diets differing basically in DHA:EPA ratio, that is, 0·68 (diet D:E-0·68), 1·09 (D:E-1·09) and 2·05 (D:E-2·05), were randomly assigned to nine tanks of 3-year-old tongue sole (ten females and fifteen males in each tank). The feeding trail lasted for 90 d before and during the spawning season. Fish were reared in a flowing seawater system and fed to apparent satiation twice daily. Compared with diet D:E-0·68, diet D:E-1·09 significantly enhanced the oestradiol production in females, whereas diet D:E-2·05 significantly enhanced the testosterone production in males. In ovaries, diet D:E-1·09 induced highest mRNA expression of follicle-stimulating hormone receptor (FSHR), steroidogenic acute regulatory protein, 17α-hydroxylase (P450c17) and 3ß-hydroxysteroid dehydrogenase (3ß-HSD). In testes, diet 2·05 resulted in highest mRNA expression of FSHR, cholesterol side-chain cleavage enzyme, P450c17 and 3ß-HSD. Fatty acid profiles in fish tissues reflected closely those of diets. Female fish had more gonadal EPA content but less DHA content than male fish, whereas there was a reverse observation in liver. In conclusion, the dietary DHA:EPA ratio, possibly combined with the dietary EPA:arachidonic acid ratio, differentially regulated sex steroid hormone synthesis in mature female and male tongue soles. Females seemed to require more EPA but less DHA for the gonadal steroidogenesis than males. The results are beneficial to sex-specific nutritive strategies in domestic teleost.

Cloning and characterization of fatty acid-binding proteins (fabps) from Japanese seabass (Lateolabraxjaponicus) liver, and their gene expressions in response to dietary arachidonic acid (ARA).

In the present study, putative cDNA of five fabp isoforms, i.e., fabp1, fabp2, fabp3, fabp4, and fabp7, was cloned and characterized from the liver of Japanese seabass (Lateolabrax japonicus), and their expression in response to diets with different arachidonic acid (ARA) levels (0.05%, 0.22%, 0.37%, 0.60%, 1.38% and 2.32% of dry matter) was investigated following a feeding trial. The Japanese seabass fabps showed high identity to their orthologs in other fish species and mammals. However, a specific fabp of Japanese seabass showed much lower identity to other Japanese seabass fabps. fabp1 has high expressions in liver and intestine, whereas fabp2 is mainly expressed in the gastrointestinal tract. The highest expression level of fabp3, fabp4, and fabp7 was observed in muscle, eye, and liver respectively. Different tissue expression patterns of fabp2, fabp4, and fabp7 between Japanese seabass and other teleost may indicate specific evolutionary Fabp functions in Japanese seabass. Moderate levels of dietary ARA (0.37-0.60%) enhanced the gene expressions of fabp1 in liver and intestine, fabp2 in intestine, and fabp3 in intestine, whereas excess dietary ARA levels (1.38-2.323%) were ineffective. The highest level of dietary ARA (2.32%) increased only the expression of fabp3 in muscle compared to the control diet. Gene expressions of fabp3 and fabp7 in liver, and fabp4 in liver, intestine, and muscle were not significantly influenced by dietary ARA. To our knowledge, this is the first study investigating the regulation of fabp expressions by dietary ARA.

Transcriptomic analysis of the effects of Toll-like receptor 4 and its ligands on the gene expression network of hepatic stellate cells.

BACKGROUND: Intact Toll-like receptor 4 (TLR4) has been identified in hepatic stellate cells (HSCs), the primary fibrogenic cell type in liver. Here, we investigated the impact of TLR4 signaling on the gene expression network of HSCs by comparing the transcriptomic changes between wild-type (JS1) and TLR4 knockout (JS2) murine HSCs in response to two TLR4 ligands, lipopolysacchride (LPS), or high-mobility group box 1 (HMGB1). RESULTS: Whole mouse genome microarray was performed for gene expression analysis. Gene interaction and co-expression networks were built on the basis of ontology and pathway analysis by Kyoto Encyclopedia of Genes and Genomes (KEGG). Gene expression profiles are markedly different between Wild type (JS1) and TLR4 knockout (JS2) HSCs under basal conditions or following stimulation with LPS or HMGB1. The differentially expressed genes between TLR4 intact and null HSCs were enriched in signaling pathways including p53, mTOR, NOD-like receptor, Jak-STAT, chemokine, focal adhesion with some shared downstream kinases, and transcriptional factors. Venn analysis revealed that TLR4-dependent, LPS-responsive genes were clustered into pathways including Toll-like receptor and PI3K-Akt, whereas TLR4-dependent, HMGB1-responsive genes were clustered into pathways including metabolism and phagosome signaling. Genes differentially expressed that were categorized to be TLR4-dependent and both LPS- and HMGB1-responsive were enriched in cell cycle, ubiquitin mediated proteolysis, and mitogen-activated protein kinase (MAPK) signaling pathways. CONCLUSIONS: TLR4 mediates complex gene expression alterations in HSCs. The affected pathways regulate a wide spectrum of HSC functions, including inflammation, fibrogenesis, and chemotaxis, as well as cell growth and metabolism. There are common and divergent regulatory signaling downstream of LPS and HMGB1 stimulation via TLR4 on HSCs. These findings emphasize the complex cascades downstream of TLR4 in HSCs that could influence their cellular biology and function.

Calcium-calmodulin dependent protein kinase I from Macrobrachium nipponense: cDNA cloning and involvement in molting.

Calcium-calmodulin dependent protein kinase I is a component of a calmodulin-dependent protein kinase cascade and involved in many physiological processes. The full-length cDNA of calcium-calmodulin dependent protein kinase I (MnCaMKI) was cloned from the freshwater prawn Macrobrachium nipponense and its expression pattern during the molt cycle and after eyestalk ablation is described. The full-length cDNA of MnCaMKI is 3,262 bp in length and has an open reading frame (ORF) of 1,038 bp, encoding a 345 amino acid protein. The expression of MnCaMKI in three examined tissues was upregulated in the premolt stage of the molt cycle. Its expression was induced after eyestalk ablation (ESA): the highest expression level was reached 1 day after ESA in hepatopancreas, and 3 days after ESA in muscle. By dsRNA-mediated RNA interference assay, expression of MnCaMKI and ecydone receptor gene (MnEcR) was significantly decreased in prawns treated by injection of dsMnCaMKI, while expression of these two genes was also significantly decreased in prawns treated by injection of dsMnEcR, demonstrating a close correlation between the expression of these two genes. These results suggest that CaMKI in M. nipponense is involved in molting.

EZH2 regulates cancer cell migration through repressing TIMP-3 in non-small cell lung cancer.

Histone methylations play important roles in human cancer metastasis. Enhancer of zeste homolog 2 (EZH2) is a key component of the polycomb repressor complex 2, which is responsible for histone H3K27 methylation. EZH2 is overexpressed in lung cancer and epigenetically silences tumor suppressor genes. Here, we showed that EZH2 was up-regulated in lung cancer and had a positive correlation with pathologic stage, nodal involvement in lung cancer patients. Moreover, overexpression of EZH2 was correlated with reduced tissue inhibitor of metalloproteinase-3 (TIMP-3) expression, which was shown to be negatively associated with tumor metastasis. Of note, overall survival time of patients with high EZH2/low TIMP-3 expression was significantly shorter than that of patients with low EZH2/high TIMP-3 (P = 0.031). RNA interfering and pharmacologic inhibition of EZH2 reduced histone H3 lysine 27 tri-methylation level and increased TIMP-3 expression level. Knockdown of EZH2 by siRNA significantly reduced A549 cancer cell migration. In contrast, reduction of TIMP-3 in A549 cells partially rescued EZH2 deficiency-induced loss of cell migration capacity. Taken together, our findings indicate that EZH2 accelerates cancer cell migration, in part, via the repression of TIMP-3 expression, suggesting a potential mechanism by which EZH2 promotes lung cancer progression and metastasis.

Carmichaeline A: a new C20-diterpenoid alkaloid from Aconitum carmichaeli.

The roots of Aconitum carmichaeli Debx. are known for their medicinal value. A new C20-diterpenoid alkaloid designated as carmichaeline A (1) has been isolated, along with eight known diterpenoid alkaloids from the roots of the plant. Their structures were elucidated on the basis of spectroscopic data interpretation.

Study on the interactions between diketo-acid inhibitors and prototype foamy virus integrase-DNA complex via molecular docking and comparative molecular dynamics simulation methods.

Human immunodeficiency virus type 1 (HIV-1) integrase (IN) is an important drug target for anti-acquired immune deficiency disease (AIDS) treatment and diketo-acid (DKA) inhibitors are potent and selective inhibitors of HIV-1 IN. Due to lack of three-dimensional structures including detail interactions between HIV-1 IN and its substrate viral DNA, the drug design and screening platform remains incompleteness and deficient. In addition, the action mechanism of DKA inhibitors with HIV-1 IN is not well understood. In view of the high homology between the structure of prototype foamy virus (PFV) IN and that of HIV-1 IN, we used PFV IN as a surrogate model for HIV-1 IN to investigate the inhibitory mechanism of raltegravir (RLV) and the binding modes with a series of DKA inhibitors. Firstly, molecular dynamics simulations of PFV IN, IN-RLV, IN-DNA, and IN-DNA-RLV systems were performed for 10 ns each. The interactions and inhibitory mechanism of RLV to PFV IN were explored through overall dynamics behaviors, catalytic loop conformation distribution, and hydrogen bond network analysis. The results show that the coordinated interactions of RLV with IN and viral DNA slightly reduce the flexibility of catalytic loop region of IN, and remarkably restrict the mobility of the CA end of viral DNA, which may lead to the partial loss of the inhibitory activity of IN. Then, we docked a series of DKA inhibitors into PFV IN-DNA receptor and obtained the IN-DNA-inhibitor complexes. The docking results between PFV IN-DNA and DKA inhibitors agree well with the corresponding complex of HIV-1 IN, which proves the dependability of PFV IN-DNA used for the anti-AIDS drug screening. Our study may help to make clear some theoretical questions and to design anti-AIDS drug based on the structure of IN.

1,4-Bis(1H-benzimidazol-1-yl)benzene.

In the title compound, C(20)H(14)N(4), the dihedral angles between the central benzene ring and the pendant benzimidazole ring systems are 46.60 (15) and 47.89 (16)°. The dihedral angle between the benzimidazole ring systems is 85.62 (12)° and the N atoms lie to the same side of the mol-ecule. In the crystal, mol-ecules are linked by C-H⋯N inter-actions and weak aromatic π-π stacking [shortest centroid-centroid separation = 3.770 (2) Å] is observed.

(1ß,2α,3α,7α,11α,13ß)-1,3,7,11-Tetra-acet-oxy-2,13-bis-(benz-yloxy)-21-methyl-19,21-secohetisan-19-al hemi-hydrate.

In the crystal structure of the title compound, C(43)H(46)NO(13)·0.5H(2)O, the mol-ecule assumes a U-shaped conformation, the terminal benzene rings being approximately parallel and partially overlapped with each other. The mol-ecule contains eight alicyclic and heterocyclic rings. The cyclo-hexane rings adopt chair conformations, the other three six-membered carbocyclic rings form a bicyclo-[2.2.2]octane system with a boat conformation for each six-membered ring, the six-membered heterocyclic ring has a chair conformation and both of the five-membered rings have envelope conformations. The solvent water mol-ecule links with the organic mol-ecule via classic O-H⋯O and weak C-H⋯O hydrogen bonding in the crystal structure.