Synthesis and biological evaluation of esculetin derivatives as potential anti-HBV agents.

Previous in vivo and in vitro studies revealed that esculetin (Fig. 1) has anti-hepatitis B virus (anti-HBV) activity as well as a protective effect on liver damage caused by duck hepatitis B virus. We designed and synthesized a series of esculetin derivatives, introduced side chains containing various amino groups into site 7 of the parent structure, and synthesized C-4 and C-8 substituted derivatives with the goal of investigating their anti-HBV activities. In vitro anti-HBV activity was performed against HepG2.2.15 cells by using Enzyme-Linked Immunosorbent Assay(ELISA) kit and cytotoxicity was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay with lamivudine as the positive control. The results demonstrated that several compounds showed moderate anti-HBV activity, while the introduction of morpholine groups could significantly inhibit the expression of hepatitis B e antigen (HBeAg) and the introduction of the 2-methylimidazole group could significantly inhibit the expression of Hepatitis B surface antigen (HBsAg). Among all tested compounds, compound 4a demonstrated the best anti-HBeAg activity (IC50 = 15.8 ± 4.2 µM), while compound 6d demonstrated the best anti-HBsAg activity (IC50 = 21.4 ± 2.8 µM). Compounds 6b and 6c showed moderate anti-HBV activity and HBsAg inhibition. Compounds 4b showed moderate anti-HBV activity and an inhibitory effect on HBeAg. In addition, compounds 4a, 4c, 4d, 6b, 6c and 6d showed improved metabolic stability. This study provides useful guidance for the discovery of anti-HBV drugs, which merits further investigation.

Two new isoquinolines from Corydalis saxicola.

Two new isoquinolines (1 and 3), along with 4 known isoquinolines were obtained from the ethanol extract of Corydalis saxicola Bunting. Their structures were elucidated based on detailed spectroscopic data (NMR, HR-ESIMS) and comparison with literature data. The absolute configurations of the new compounds were assigned by comparing computed electronic circular dichroism (ECD). The anti-inflammatory effects of the isolates were assessed by inhibiting NO production in LPS-induced RAW264.7 macrophage cells, and the results showed that compounds 1-6 exhibited anti-inflammatory activities, with IC50 values ranged from 44.24 ± 1.16 to 69.00 ± 5.41 µM.

Design and synthesis of a ratiometric photoacoustic imaging probe activated by selenol for visual monitoring of pathological progression of autoimmune hepatitis.

Photoacoustic (PA) imaging with both the high contrast of optical imaging and the high spatial resolution of ultrasound imaging has been regarded as a robust biomedical imaging technique. Autoimmune hepatitis (AIH) is the second largest liver inflammatory disease after viral hepatitis, but its pathogenesis is not fully understood probably due to the lack of an effective in vivo monitoring approach. In this work, an innovative selenol-activated ratiometric PA imaging probe APSel was developed for visual monitoring of pathological progress of AIH. Selenols including selenocysteine (Sec, the major form of Se-containing species in vivo) have been demonstrated to have an effective antioxidant role in inflammation. The reaction of APSel with selenol results in a blue shift of the PA spectrum peak from 860 nm to 690 nm, which enables the ratiometric PA imaging. The APSel probe displays high sensitivity and selectivity to Sec and other selenols. The APSel probe was then employed for ratiometric PA imaging of selenol in cells, and for monitoring the development of AIH in a murine model by tracking the changes of selenol level. The results revealed that the level of selenol was closely correlated with the development of AIH. The proposed APSel, as the first example of a selenol-responsive PA imaging probe, provides a new tool and approach to study and diagnose AIH diseases.

Copper(I)-catalyzed [4 + 2] cycloaddition of aza-ortho-quinone methides with bicyclic alkenes.

A variety of tetrahydroquinoline-fused bicycles bearing multiple stereocenters are prepared in good yields with high diastereoselectivity through Cu2O-catalyzed [4 + 2] cycloaddition of aza-ortho-quinone methides (ao-QMs) with bicyclic alkenes. Mechanistic studies reveal that the Cu(i) catalyst not only promotes the formation of ao-QMs through a radical process by single electron transfer but also accelerates [4 + 2] cycloaddition. The reaction was easily performed on gram scale and the obtained tetrahydroquinoline-fused bicycles can be converted to diverse tetrahydroquinoline scaffolds.

Mitochondrial-Targeted and Near-Infrared Fluorescence Probe for Bioimaging and Evaluating Monoamine Oxidase A Activity in Hepatic Fibrosis.

Monoamine oxidase A (MAO-A) is a promising diagnostic marker for cancer, depression, Parkinson's disease, and liver disease. The fluorescence detection of MAO-A in living animals is of extreme importance for the early diagnosis of related diseases. However, the development of specific and mitochondrial-targeted and near-infrared (NIR) fluorescence MAO-A probes is still inadequate. Here, we designed and synthesized four NIR fluorescence probes containing a dihydroxanthene (DH) skeleton to detect MAO-A in complex biological systems. The specificity of our representative probe DHMP2 displays a 31-fold fluorescence turn-on in vitro, and it can effectively accumulate in the mitochondria and specifically detect the endogenous MAO-A concentrations in PC-3 and SH-SY5Y cell lines. Furthermore, the probe DHMP2 can be used to visualize the endogenous MAO-A activity in zebrafish and tumor-bearing mice. More importantly, it is the first time that the MAO-A activity of hepatic fibrosis tissues is detected through the probe DHMP2. The present study shows that the synthesized DHMP2 might serve as a potential tool for monitoring MAO-A activity in vivo and diagnosing related diseases.

Coumarinolignoids and lignanoids from the stems and leaves of Sapium discolor.

Two new coumarinolignoids, sapiumins D (1) and E (2), a new lignanoid, lariciresinol 9'-benzoate (3), together with six known coumarinolignoids (4-9) and eight known lignanoids (10-17), were isolated from the stems and leaves of Sapium discolor. The structures of the isolated compounds were elucidated by extensive spectroscopic methods, including NMR, MS, and single crystal X-ray diffraction experiments. Compounds 5, 10, 11, and 13 significantly inhibited nitric oxide production in lipopolysaccharide-induced BV-2 microglial cells, with IC50 values in the range of 2.13-11.37 µM.

Phthalazino[1,2-b]quinazolinones as p53 Activators: Cell Cycle Arrest, Apoptotic Response and Bak-Bcl-xl Complex Reorganization in Bladder Cancer Cells.

p53 inactivation is a clinically defined characteristic for cancer treatment-nonresponsiveness. It is therefore highly desirable to develop anticancer agents by restoring p53 function.1 Herein the synthesized phthalazino[1,2-b]quinazolinones were discovered as p53 activators in bladder cancer cells. 10-Bromo-5-(2-dimethylamino-ethylamino)phthalazino[1,2-b]quinazolin-8-one (5da) was identified as the most promising candidate in view of both its anticancer activity and mechanisms of action. 5da exhibited strong anticancer activity on a broad range of cancer cell lines and significantly reduced tumor growth in xenograft models at doses as low as 6 mg/kg. Furthermore, 5da caused cell cycle arrest at S/G2 phase, induced apoptosis, changed cell size, and led to cell death by increasing the proportion of sub-G1 cells. Molecular mechanism studies suggested that accumulation of phospho-p53 in mitochondria after 5da treatment resulted in conformational activation of Bak, thereby evoking cell apoptosis, finally leading to irreversible cancer cell inhibition. Our present studies furnish new insights into the molecular interactions and anticancer mechanisms of phospho-p53-dependent quinazolinone compound.

Tandem C-N Bond Formation through Condensation and Metal-Free N-Arylation: Protocol for Synthesizing Diverse Functionalized Quinoxalines.

Diverse functionalized quinoxalines were synthesized in good yields from arylamines and readily available ß-keto oximes through condensation and metal-free N-arylation. The reaction was compatible with various functional groups, such as halides, cyano, and esters. A mechanism was proposed based on the experimental results. These quinoxalines were easily obtained on a gram scale and converted to various useful scaffolds. Compound LASSBio-1022 was prepared in 83% yield in two steps.

Anti-inflammatory polyphenol constituents derived from Cissus pteroclada Hayata.

A new bergenin derivative, bergenin-11-O-α-d-galactopyranoside (compound 1), together with seven known polyphenolic compounds, were isolated from the stem of Cissus pteroclada Hayata. The structures of the 8 compounds were elucidated by spectroscopic methods, including extensive 1D and 2D NMR techniques. Moreover, the in vitro anti-inflammatory effects of compounds (1-8) in LPS-stimulated murine macrophage RAW 264.7 cells were also investigated. Our results revealed that compound 1 inhibited the production of pro-inflammatory mediators NO and PGE2 and the expression of NF-κB, TNF-α, IL-1ß, iNOS and COX-2.

Synthesis, Fluorescence Properties, and Antiproliferative Potential of Several 3-Oxo-3H-benzo[f]chromene-2-carboxylic Acid Derivatives.

In this study, two series of 3-oxo-3H-benzo[f]chromene-2-carboxylic acid derivatives (compounds 5a-i and 6a-g) were synthesized. Their in vitro proliferation inhibitory activities against the A549 and NCI-H460 human non-small cell lung cancer (NSCLC) cell lines were evaluated. Their photophysical properties were measured. Among these target compounds, 5e exhibited the strongest antiproliferative activity by inducing apoptosis, arresting cell cycle, and elevating intracellular reactive oxygen species (ROS) level, suggesting that it may be a potent antitumor agent. In addition, compound 6g with very low cytotoxicity, demonstrated excellent fluorescence properties, which could be used as an effective fluorescence probe for biological imaging.

Purification, antioxidant and hepatoprotective activities of polysaccharide from Cissus pteroclada Hayata.

Polysaccharide of Cissus pteroclada Hayata (CPHP) was extracted and purified. Three major fractions (CPHP I, CPHP II-1 and CPHP II-2) from the CPHP were purified by column chromatography and investigated for their monosaccharide compositions, scavenging radical effects and hepatoprotective activities in vitro. The results showed that glucose and galactose were the main monosaccharides of three polysaccharide fractions, CPHP II-1 and CPHP II-2 were acidic polysaccharide fractions which contained glucuronic acid and galacturonic acid. Antioxidant activity determination suggested that CPHP I and CPHP II-1 had a higher scavenging effects on DPPH, superoxide radical, hydroxyl radical and ABTS radical. And the results of antioxidant test in vitro showed that CPHP II-2 could significantly increase (P<0.01) the activities of SOD and GSH-Px and decreased MDA level in human hepatocyte cell line (HL7702 cell), which indicating that CPHP II-2 possessed good hepatoprotective activity.

Distinct novel quinazolinone exhibits selective inhibition in MGC-803 cancer cells by dictating mutant p53 function.

The mutant p53 proteins and their corresponding cellular response can be manipulated by novel quinazolinone derivatives 4-8 (a-i) in p53 mutant cancer cells. Of the two most potent compounds, 4a exhibited promising broad-spectrum anti-cancer effects, whereas 6c showed selective and exclusive inhibition activity in p53 mutant cancer cell lines but low toxicity to wild-type p53 cancer cell A375 and normal lung fibroblast WI-38 cells. Furthermore, 6c exhibited a more sophisticated mechanism for cell-destructive response by causing S/G2 phase arrest effect and cell size reduction. Compared with the cellular response of 6b and genetic background of cell lines studied, p53 mutation was found to be the key factor and main target for 6c evoked cell-destructive response. Molecular mechanism studies indicated that p53 phosphorylation and acetylation dual-targeting inhibitor 6c exerted anti-cancer activities with a special mechanism in evoking cell apoptosis by arresting mutant p53 function to trigger the deregulation of Cdk2 caused Bim-mediated apoptosis. To the best of our knowledge, 6c is the first quinazolinone derivative to dictate mutant p53 function for apoptotic cell death.

Design, synthesis and biological evaluation of novel 1-hydroxyl-3-aminoalkoxy xanthone derivatives as potent anticancer agents.

A series of novel 1-hydroxyl-3-aminoalkoxy xanthone derivatives were designed, synthesized and evaluated for in vitro anticancer activity against four selected human cancer cell lines (nasopharyngeal neoplasm CNE, liver cancer BEL-7402, gastric cancer MGC-803, lung adenocarcinoma A549). Most of the synthesized compounds exhibit effective cytotoxic activity against the four tested cancer cell lines with the IC50 values at micromolar concentration level. Some preliminary structure-activity relationships were also discussed. In this series of derivatives, compound 3g shows excellent broad spectrum anticancer activity with IC50 values ranging from 3.57 to 20.07 µM. The in vitro anticancer activity effect and action mechanism of compound 3g on human gastric carcinoma MGC-803 cell were further investigated. The results showed that compound 3g exhibits dose- and time-dependent anticancer effects on MGC-803 cells through apoptosis, which might be associated with its decreasing intracellular calcium and the mitochondrial membrane potential.

2,4-Di-bromo-1,3-dihy-droxy-9H-xanthen-9-one.

The title compound, C13H6Br2O4, derived from xanthone, a fundamental structural framework of active ingredients in many medicinal plants, and was synthesized by bromination of 1,3-di-hydroxyxanthen-9-one with N-bromo-succinimide. The mol-ecular conformation is essentially planar, the dihedral angle between the benzene rings being 1.1 (4)°. This conformation is favorable for the formation of an intra-molecular O-H⋯O hydrogen bond between a hy-droxy group and the xanthone carbonyl group. In the crystal, mol-ecules are associated into chains along the b-axis direction via C=O⋯H-O hydrogen bonds involving the other hy-droxy group.

Synthesis and antitumor activities of novel thiourea α-aminophosphonates from dehydroabietic acid.

A series of novel thiourea α-aminophosphonate derivatives containing DHA structure was designed and synthesized as antitumor agents. Their inhibitory activities against the NCI-H460 (lung), A549 (lung adenocarcinoma), HepG2 (liver) and SKOV3 (ovarian) human cancer cell lines were estimated using MTT assay in vitro. The screening results revealed that many compounds exhibited moderate to high levels of antitumor activities against the tested cancer cell lines and that most demonstrated more potent inhibitory activities compared with the commercial anticancer drug 5-fluorouracil. The mechanism of compound 5f was preliminarily investigated by acridine orange/ethidium bromide staining, Hoechst 33258 staining, JC-1 mitochondrial membrane potential staining, TUNEL assay, DNA ladder assay and flow cytometry, which indicated that the compound can induce cell apoptosis in A549 cells.

Human serum albumin-based design of a diflunisal prodrug.

The cyclooxygenase-2 inhibitor, diflunisal, is used in the clinic for its anti-inflammatory activity. About 99% of a dose of diflunisal is unavailable for reaction with the target enzyme, because diflunisal strongly binds to human serum albumin (HSA). To reduce the binding affinity of diflunisal to albumin, we designed and synthesized the prodrug acetyldiflunisal. The crystal structure of HSA complexed with fatty acid and acetyldiflunisal revealed that acetyldiflunisal binds to the IIA subdomain and that upon binding, it acetylates lysine 199. Mass spectrometry confirmed that acetyldiflunisal acetylates Lys199. The acetylated albumin had twofold weaker binding affinity for diflunisal as demonstrated by fluorescence quenching. Reduced binding affinity means that diflunisal is more easily released from acetylated albumin into the circulation. Therefore, lower doses of acetyldiflunisal compared to diflunisal will be required. Taken together, our results not only provide a template for design of HSA-based prodrugs, but also pave the way toward more effective use of diflunisal in the clinic.

[Study on steroidal and triterpenoid constituents from Cissus pteroclada].

OBJECTIVE: To study the chemical constituents of ethnical drug Cissus pteroclada. METHODS: Silica gel column chromatography was employed to separate the constituents from EtOAc extraction of Cissus pteroclada and their structures were identified by physicochemical properties as well as spectrum analysis. RESULTS: Five steroidal compounds and 2 triterpenoid constituents were obtained. Their structures were identified as: stigmasterol (1), stigmasterol acetate (2), stigmasta-5, 22-dien-3-O-beta-D-glucopyranoside (3), beta-sitosterol (4), daucousterol (5), taraxerone (6), oleanolic acid (7). CONCLUSION: All the compounds are isolated from this plant for the first time except for compound 4 and 5. Compounds 1 - 3 are obtained from this genus for the first time.

Tanshinone IIA activates calcium-dependent apoptosis signaling pathway in human hepatoma cells.

Tanshinone IIA (Tan IIA), a natural product from herb Salvia miltiorrhiza Bunge, has potential anti-tumor activity. The aim of this study was to pinpoint the molecular mechanisms underlying Tan IIA-induced cancer cell apoptosis. Human hepatoma BEL-7402 cells treated with Tan IIA underwent assessment with MTT assay for cell viability, 10-day culture for colony formation, flow cytometry and fluorescence microscopy for apoptosis and cell cycle analysis. Changes in intracellular [Ca(2+)] and mitochondrial membrane potential (∆ψ) reflected the calcium-dependent apoptosis pathway. RT-PCR was used to detect gene expression of Bad and metallothionein 1A (MT 1A). Cytotoxicity of Tan IIA was tested in human amniotic mesenchymal stem cells (HAMCs). Tan IIA exhibited dose-dependent and time-dependent anticancer effects on BEL-7402 cells through apoptosis and G(0)/G(1) arrest. Cells treated with Tan IIA increased their intracellular calcium, decreased their mitochondrial membrane potential and induced Bad and MT 1A mRNA expression. No adverse effects of Tan IIA were found in HAMCs. In conclusion, these results indicate that Tan IIA-induced cancer cell apoptosis acts via activation of calcium-dependent apoptosis signaling pathways and upregulation of MT 1A expression.

[Studies on the chemical constituents of Rubus parvifolius].

OBJECTIVE: To study the chemical constituents of Rubus parvifoliu. METHODS: The constituents were isolated by column chromatography and their structures were elucidated through spectroscopic analysis such as 1H-NMR, 13C-NMR, FT-IR, et al. RESULTS: Seven compounds were isolated from the roots of Rubus parvifolius L., they were identified as p-sitosterol (I), lauric acid (II), O-nitrophenol (III), beta-daucosterol (IV), euscaphic acid (V), camelliagenin A (VI) and(+) -catech in (VII). CONCLUSION: Compounds III and VII are isolated from the plant for the first time.

Hybrids of oxoisoaporphine-tacrine congeners: novel acetylcholinesterase and acetylcholinesterase-induced ß-amyloid aggregation inhibitors.

A series of dual binding site acetylcholinesterase (AChE) inhibitors have been designed, synthesized, and tested for their ability to inhibit AChE, butyrylcholinesterase (BChE), AChE-induced and self-induced ß-amyloid (Aß) aggregation. The new hybrids consist of a unit of 1-azabenzanthrone and a tacrine or its congener, connected through an oligomethylene linker containing an amine group at variable position. These hybrids exhibit high AChE inhibitory activity with IC(50) values in the nanomolar range in most cases. Moreover, five out of the 12 hybrids of this series, particularly those bearing a tetrahydroacridine moiety, exhibit a significant in vitro inhibitory activity toward the AChE-induced and self-induced Aß aggregation, which makes them promising anti-Alzheimer drug candidates.