DFT investigation of the DDQ-catalytic mechanism for constructing C-O bonds.

In this study, we investigated the photo-catalytic mechanisms for the construction of C-O bonds from arenes (benzene, 2',6'-dimethyl-[1,1'-biphenyl]-2-carboxylic acid, or 2,4-dichloro-1-fluorobenzene), catalyzed by 2,3-dichloro-5,6-dicyano-p-benzoquinone (DDQ). All the structures for the Gibbs free surfaces were calculated at the M06-2X-D3/ma-def2-SVP level in the SMD solvent model. Also, TDDFT calculations of DDQ were performed at the PBE1PBE-D3/ma-def2-SVP level in the SMD solvent model. The computational results indicated that DDQ, serving as a photo-catalyst, would be excited under visible light of 450 nm, aligning well with experimental observations as reflected in the UV-vis spectrum. Gibbs free energy surface analyses of the three reactions suggested that the path involving 3DDQ* activating the reactant (-COOH, H2O, or CH3OH) is favorable. Additionally, the role of O2 was investigated, revealing that it could facilitate the recycling of DDQ by lowering the energy barrier for the conversion of the DDQH˙ radical (not DDQH2) into DDQ. The use of ρhole and ρele can reveal the photo-catalytic reaction and charge transfer processes, while localized orbital locator isosurfaces and electron spin density isosurface graphs were employed to analyze structures and elucidate the single electron distribution. These computational results offer valuable insights into the studied interactions and related processes, shedding light on the mechanisms governing C-O bond formation from arenes catalyzed by DDQ.

Discovery of (aza)indole derivatives as novel respiratory syncytial virus fusion inhibitors.

A new class of indole derivatives (3) have been identified as potent RSV fusion inhibitors. SAR exploration revealed that 5-Cl and the sulfonyl side chain of the indole scaffold are crucial for anti-RSV activity. Further optimization led to the discovery of a cyclic sulfone (8i) with 2 nM anti-RSV activity and a much improved PK profile compared to the non-cyclic sulfone counterpart.

Discovery of Ziresovir as a Potent, Selective, and Orally Bioavailable Respiratory Syncytial Virus Fusion Protein Inhibitor.

Ziresovir (RO-0529, AK0529) is reported here for the first time as a promising respiratory syncytial virus (RSV) fusion (F) protein inhibitor that currently is in phase 2 clinical trials. This article describes the process of RO-0529 as a potent, selective, and orally bioavailable RSV F protein inhibitor and highlights the in vitro and in vivo anti-RSV activities and pharmacokinetics in animal species. RO-0529 demonstrates single-digit nM EC50 potency against laboratory strains, as well as clinical isolates of RSV in cellular assays, and more than one log viral load reduction in BALB/c mouse model of RSV viral infection. RO-0529 was proven to be a specific RSV F protein inhibitor by identification of drug resistant mutations of D486N, D489V, and D489Y in RSV F protein and the inhibition of RSV F protein-induced cell-cell fusion in cellular assays.

Discovery of Benzoazepinequinoline (BAQ) Derivatives as Novel, Potent, Orally Bioavailable Respiratory Syncytial Virus Fusion Inhibitors.

A novel benzoazepinequnoline (BAQ) series was discovered as RSV fusion inhibitors. BAQ series originated from compound 2, a hit from similarity-based virtual screening. In SAR exploration, benzoazepine allowed modifications in the head moiety. Benzylic sulfonyl on benzoazepine and 6-Me on quinoline were crucial for good anti-RSV activity. Although the basic amine in the head portion was crucial for anti-RSV activity, the attenuated basicity was required to reduce Vss. Introducing oxetane to the head portion led to discovery of compound 1, which demonstrated single-digit nM anti-RSV activity against different RSV strains, reasonable oral exposure in plasma, and 78-fold higher exposure in lung. Compound 1 also displayed 1 log viral reduction in a female BALB/c mice RSV model by b.i.d. oral dosing at 12.5 mg/kg. A single resistant mutant at L138F in fusion protein proved compound 1 to be a RSV fusion inhibitor.

OsERF101, an ERF family transcription factor, regulates drought stress response in reproductive tissues.

KEY MESSAGE: An ERF transcription factor OsERF101 is predominantly expressed in rice reproductive tissues and plays an important role in improving rice seed setting rate under drought stress. Drought reduces grain yield due to the cumulative damage effects to plant vegetative and reproductive developmental processes. However, the genes involved in these processes are still not completely understood. In this study, we identified a gene named OsERF101 as an important positive regulator in the adaptive responses to dehydration stress during the reproductive and vegetative stages. This gene encodes a member of APETALA2/Ethylene-Responsive Element Binding Protein (AP2/EREBP) family. OsERF101 was predominantly expressed in flowers, particularly in the tapetum and microspores under normal growth conditions. It was induced by drought, PEG6000 and abscisic acid (ABA) in leaves. During the vegetative stage, OsERF101-overexpression plants were more resistant to osmotic stress caused by PEG6000 compared to the control plants. They also had higher survival and seed setting rates than wild type when subjected to reproductive-stage drought stress. Further physiological analysis revealed that the pollen fertility was improved in the overexpression lines, while the knockout mutant and RNAi lines showed reduced pollen fertility and compromised drought tolerance during the reproductive stage. The increased proline content and peroxidase activity in OsERF101-overexpression plants might contribute to the improved drought-tolerance of plants. In addition, OsERF101-overexpression plants displayed ABA susceptible phenotype, in which the expression levels of ABA-responsive genes RD22, LEA3, and PODs were up-regulated. Taken together, our results indicate that OsERF101 is a gene that regulates dehydration responses during the vegetative and reproductive stages.

Discovery of methylsulfonyl indazoles as potent and orally active respiratory syncytial Virus(RSV) fusion inhibitors.

Recently we described a novel class of imidazopyridine compounds that showed exceptional anti-RSV potency in cell culture. However, unfavorable pharmacokinetic (PK) properties and glutathione (GSH) adduct liabilities impeded their further development. In a bid to address the PK and early safety concerns, a small compound library consisting of dozens of scaffold-hopping analogues was designed and synthesized for RSV CPE assay screening, which led to the identification of a new chemical starting point: methylsulfonyl indole compound 8. In this paper, we report the discovery and optimization of a series of methylsulfonyl indazoles as potent RSV fusion inhibitors. In particular, compound 47 was orally efficacious in a RSV mouse model, with 1.6 log unit viral load reduction at 25 mg/kg BID upon oral dosing. The results may have broad implications for the design of new RSV fusion inhibitors, and demonstrate the potential for developing novel therapies for RSV infection.

Discovery of Piperazinylquinoline Derivatives as Novel Respiratory Syncytial Virus Fusion Inhibitors.

A novel series of piperazinylquinoline derivatives were discovered as respiratory syncytial virus (RSV) fusion inhibitors by the ligand-based screening approach. Among 3,000 hits, 1-amino-3-[[2-(4-phenyl-1-piperidyl)-4-quinolyl]amino]propan-2-ol (7) was proven to be active against the RSV long (A) strain. The anti-RSV activity was improved by converting piperidine to benzylcarbonyl substituted piperazine. The basic side chain was also found to be crucial for anti-RSV activity. The selected analogues, 45 and 50, demonstrated anti-RSV activities up to EC50 = 0.028 µM and 0.033 µM, respectively. A direct anti-RSV effect was confirmed by a plaque reduction assay and a fusion inhibition assay. Both 45 and 50 showed promising DMPK properties with good oral bioavailability, and could potentially lead to novel therapeutic agents targeting the RSV fusion process.

Discovery of imidazopyridine derivatives as highly potent respiratory syncytial virus fusion inhibitors.

A series of imidazolepyridine derivatives were designed and synthesized according to the established docking studies. The imidazopyridine derivatives were found to have good potency and physical-chemical properties. Several highly potent compounds such as 8ji, 8jl, and 8jm were identified with single nanomolar activities. The most potent compound 8jm showed an IC50 of 3 nM, lower microsome clearance and no CYP inhibition. The profile of 8jm appeared to be superior to BMS433771, and supported further optimization.

[Effect of NaCl stress on physiological characteristics of Physalis alkekengi var. franchetii test-tube plantlet].

OBJECTIVE: To investigate the effect of NaCl stress on the growth and physiological characteristics of Physalis alkekengi var. franchetii, and to discuss the mechanism of salt tolerance. METHODS: The test-tube plantlet was cultured on the stress medium with NaCl (0.2%~1.0%), the content of proline, soluble sugar, soluble protein, chlorophyll and malondialdthyde (MDA), and membrane relative permeability, as well as the activities of superoxide dismutase (SOD), catalase (CAT) and peroxidase (POD) were determined and analyzed. RESULTS: The average height, stem diameter, root length, number of root and chlorophyll content gradually decreased at salinity less than or equal 0.6% NaCl, then decreased rapidly at salinity greater than or equal 0.8% NaCl, while the root formation was significantly inhibited. The content of free proline and SOD activities always increased with NaCl concentration increasing (0.2%~ 1.0%). Nevertheless, the contents of soluble sugar, soluble protein and the activities of CAT and POD increased firstly and then decreased, but the contents of soluble sugar and soluble protein were higher than those of the control group, whereas the content of MDA and membrane relative permeability accumulated slowly at salinity less than or equal 0.6% NaCl, then increased rapidly at salinity greater than 0.8% NaCl. CONCLUSION: The test-tube plantlet has an adaptability to salty environment at salinity less than or equal 0.6% NaCl, and the test-tube plantlet can alleviate the salty injury by increasing the contents of osmotic regulation substances, such as soluble sugar and soluble protein, decreasing the osmotic potential, and increasing the activities of antioxidant enzymes.

[Effects of different salt-alkaline stress on seed germination and seedling physiological characteristics of Codonopsis pilosula].

OBJECTIVE: To improve salt resistance of Codonopsis pilosula seeds and seedlings under different salt-alkaline stress, the seed germination and physiological characteristics of seedlings were studied. METHODS: Several physiological indexes of Codonopsis pilosula under different salt-alkaline stress were measured, such as the germination vigor, germination rate, relative germination rate and recovery germination rate,as well as others indexes like the contents of chlorophyll, soluble protein, water and MDA and the activities of POD and SOD. RESULTS: The inhibitory effects of three salts on seed germination were Na2 CO3 > NaCl + Na2 CO3 > NaCl with an increased salt concentration. The contents of chlorophyll, soluble protein and water were found significant different (P < 0.05) from the negative control group, along with the decreased of these indexes. However, positive correlation existed between MDA content and salt treatment concentration. In addition, the change trend of SOD and POD activity were similar. They were increased firstly, and then decreased as the stress intensity extended. CONCLUSION: The seeds and seedlings under different salt-alkaline stress were inhibited, with different inhibitory extent as Na2CO3 > NaCl + Na2 CO3 > NaCl.

[Study on the optimum combinations of plant hormone in callus culture of Thermopsis lanceolata].

OBJECTIVE: To explore the effects of different hormone combinations on callus induction of Thermopsis lanceolata. METHODS: The effects of different hormone concentrations and ratio on callus induction and subculture multiplication of Thermopsis lanceolata were investigated by orthogonal design methods. RESULTS: The optimal medium of callus induction for hypocotyls explants was MS + 2, 4-D (0.5 mg/L) + BA (1.0 mg/L) + NAA (1.0 mg/L) + IAA (0.5 mg/L). The best medium of callus subculture multiplication for hypocotyl explants was MS with 2,4-D (0.1 mg/L) + BA (0.1 mg/L) + NAA (0.2 mg/L) + IAA (0.5 mg/L) . In the process of callus subculture multiplication, the effect extent of four kinds of hormones for hypocotyl explants was BA > 2,4-D > IAA > NAA. The growth curve of callus culture optimal hormone combination was "S" curve. CONCLUSION: The rapid callus culture system for Thermopsis lanceolata is established for the first time. It provides a new way for protecting and utilizing Thermopsis lanceolata species resources through the modern biotechnology.

[Specific anti-tumor effect of immune competent cell induced by HSP-70 of HeLa cells in vitro].

BACKGROUND & OBJECTIVE: Recent studies indicated that heat shock proteins (HSPs) are associated with antigenic peptides and participate in the antitumor T cell response in animals. This study was designed to investigate the mechanism of anti-tumor effect of immune competent cells (ICC) induced by HSP-70 of HeLa (cervical carcinoma) cells in vitro. METHODS: The normal peripheral blood mononuclear cells(PBMCs) stimulated with HSP-70 of HeLa cells in vitro. T cell phenotypes were analyzed before and after stimulated by PBMCs. HSP-70 primed the immune competent cells (ICCs) from PBMCs were tested for cytotoxic activity against HeLa cell. RESULTS: CD3+ cell was 85.73 +/- 1.44%, up from 57.68 +/- 1.46% before the amplification(P < 0.002). CD8+ cell was 48.06 +/- 1.66%, up from 23.56 +/- 1.86% before the amplification(P < 0.002). The ICCs induced by HSP-70 were shown significant cytotoxic activity (82.69 +/- 1.97%, 62.11 +/- 1.61%) against HeLa cells and cervical carcinoma cells, and the cytotoxic activity (31.05 +/- 2.09%) of the ICC against HeLa cells could be blocked by anti-HSP-70 antibody. CONCLUSION: The ICC can be induced selectively by PBMCs with HSP-70 of HeLa cells, and has specific cytotoxic activity against HeLa cell.