Uncommon Epoxyquinols Pyrrolocytosporin A and Cytosporin E2 from the Fungus Eutypella sp. D-1.

Two uncommon epoxyquinols, pyrrolocytosporin A (1) and cytosporin E2 (2), along with the known cytosporin Y1 (3), were isolated from the solid defined medium of the Arctic-derived fungus Eutypella sp. D-1. Their structures were established through comprehensive analyses of spectroscopic and electronic circular dichroism data. Structurally, compound 1 represented the first nitrogen-containing epoxyquinol characterized by a pyrrole fused cytosporin framework, while compound 2 contained an uncommon cyclic carbonate functionality. The antibacterial, immunosuppressive, anti-inflammatory, and cytotoxic activities of all compounds were evaluated. Among the three metabolites, only compound 1 exhibited inhibitory effects on nitric oxide production induced by lipopolysaccharide with an IC50 value of 6.55 µM. Additionally, only compound 2 displayed inhibitory activity against ConA-induced T-cell proliferation with an IC50 value of 9.85 µM.

Cytosporin Derivatives from Arctic-Derived Fungus Eutypella sp. D-1 via the OSMAC Approach.

A chemical investigation of the Arctic-derived fungus Eutypella sp. D-1 based on the OSMAC (one strain many compounds) approach resulted in the isolation of five cytosporin polyketides (compounds 1-3 and 11-12) from rice medium and eight cytosporins (compounds 2 and 4-11) from solid defined medium. The structures of the seven new compounds, eutypelleudesmane A (1), cytosporin Y (2), cytosporin Z (3), cytosporin Y1 (4), cytosporin Y2 (5), cytosporin Y3 (6), and cytosporin E1 (7), were elucidated by analyzing their detailed spectroscopic data. Structurally, cytosporin Y1 (4) may be a key intermediate in the biosynthesis of the isolated cytosporins, rather than an end product. Compound 1 contained a unique skeleton formed by the ester linkage of two moieties, cytosporin F (12) and the eudesmane-type sesquiterpene dihydroalanto glycol. Additionally, the occurrence of cyclic carbonate moieties in compounds 6 and 7 was found to be rare in nature. The antibacterial, immunosuppressive, and cytotoxic activities of all compounds derived from Eutypella sp. D-1 were evaluated. Unfortunately, only compounds 3, 6, 8, and 10-11 displayed immunosuppressive activity, with inhibitory rates of 62.9%, 59.5%, 67.8%, 55.8%, and 68.7%, respectively, at a concentration of 5 µg/mL.

Alterations of oral microbiota in patients with obstructive sleep apnea-hypopnea syndrome treated with continuous positive airway pressure: a pilot study.

PURPOSE: Obstructive sleep apnea-hypopnea syndrome (OSAHS) is an independent risk factor for cardiovascular diseases, including hypertension. In our previous study, it was demonstrated that oral microbiota alteration in patients with OSAHS, particularly in the genera Aggregatibacter and Porphyromonas, may influence the development of hypertension. Continuous positive airway pressure (CPAP) is the main therapy for OSAHS and OSAHS-associated hypertension. However, the role of oral microbiota post CPAP treatment remains unknown. METHODS: We conducted 16S rDNA pyrosequencing and bioinformatic analyses to compare the bacterial composition of oral specimens from patients with OSAHS before and after overnight CPAP treatment. RESULTS: This approach enabled a relatively comprehensive description of oral microbiota, with decreases in Gemella and increases in Staphylococcus, f_Lachnospiraceae, Parabacteroides, and f_Ruminococcaceae after CPAP treatment. CONCLUSION: Alteration of oral microbiota may shed new insight on the underlying pathogenesis of OSAHS-associated hypertension.

Palitantin derivatives from the Antarctic fungus Geomyces sp. 3-1.

Two new polyketides, palitantins B and C (1 and 2), along with one known related compound (+)-palitantin (3) were obtained from the culture of the Antarctic fungus Geomyces sp. 3-1. The structures of the new compounds were elucidated by detailed analysis of HRESIMS, NMR, CD, and ECD data. Compound 3 showed potent PTP1B inhibitory activity with an IC50 value of 7.9 µM (ursolic acid as positive control, IC50 = 8.3 µM).

Eutypellacytosporins A-D, Meroterpenoids from the Arctic Fungus Eutypella sp. D-1.

The Arctic fungus Eutypella sp. D-1, previously found to produce a variety of cytotoxic cyclopropyl-fused and cyclobutyl-fused pimarane diterpenoids when grown in the defined medium, was induced to produce unusual metabolites by growing on solid rice medium. A chemical investigation on the rice medium extract led to the isolation of four new meroterpenoids, eutypellacytosporins A-D (1-4), along with the known biogenetically related compound cytosporin D (5). The structures of the new compounds were elucidated by their detailed spectroscopic analysis and modified Mosher's method. Compounds 1-4 may be formed by the 12,32-ester linkage of two moieties, cytosporin D (5) and decipienolide A or B. All isolated compounds, except 5, showed weak cytotoxicity against DU145, SW1990, Huh7, and PANC-1 cell lines with IC50 values ranging from 4.9 to 17.1 µM.

Bioactive metabolites from the Arctic fungus Nectria sp. B-13.

Two new cyclohexanone derivatives, nectriatones A-B (1-2), and one new natural product, nectriatone C (3), together with three known phenolic sesquiterpene derivatives (4-6), were isolated from the culture of Nectria sp. B-13 obtained from high-latitude soil of the Arctic. The structures of all compounds were unambiguously elucidated by extensive spectroscopic analysis, as well as by comparison with the literature. These compounds were evaluated in cytotoxic and antibacterial activities. Compounds 1-6 showed cytotoxicities against SW1990, HCT-116, MCF-7, and K562 cells, with IC50 values in the range of 0.43 to 42.64 µM. Only compound 4 exhibited antibacterial activity against Escherichisa coli, Bacillus subtilis, and Staphylococcus aureus (MIC 4.0, 2.0, and 4.0 µg/ml, respectively).

Libertellenones O-S and Eutypellenones A and B, Pimarane Diterpene Derivatives from the Arctic Fungus Eutypella sp. D-1.

Seven new pimarane-type diterpene derivatives, libertellenones O-S (1-5) and eutypellenones A and B (6 and 7), together with two known compounds (8 and 9), were isolated from the culture of Eutypella sp. D-1 obtained from high-latitude soil of the Arctic. Their structures were elucidated from spectroscopic data, as well as experimental and calculated electronic circular dichroism (ECD) analysis. Structurally, compounds 1-5 possess a cyclopropyl-fused pimarane diterpene moiety, whereas compounds 6 and 7 share an unusual cyclobutyl-fused pimarane diterpene skeleton. Compounds 1-9 exhibited cytotoxicities against HeLa, MCF-7, HCT-116, PANC-1, and SW1990 cells, with IC50 values in the range of 0.3 to 29.4 µM. Compounds 6 and 7 could dose-dependently inhibit the activity of NF-κB and exhibited significantly inhibitory effects on nitric oxide production induced by lipopolysaccharide.

Eutypellenoids A⁻C, New Pimarane Diterpenes from the Arctic Fungus Eutypella sp. D-1.

Three new pimarane diterpenes, eutypellenoids A⁻C (1⁻3), together with a known compound, eutypenoid C (4), were isolated from the culture extract of Eutypella sp. D-1 derived from the Arctic region. Compounds 1⁻3 possessed an uncommon tetrahydrofuran-fused pimarane diterpene skeleton. The structures of all compounds were determined by detailed spectroscopic analysis, electronic circular dichroism (ECD) analysis, as well as a comparison with the literature data. Antibacterial, antifungal, and cytotoxic activities of these compounds were evaluated. Compound 2 displayed antibacterial activity against Staphylococcus aureus and Escherichia coli with MIC values of 8 and 8 µg/mL, respectively. Additionally, compound 2 showed antifungal activity against Candidaparapsilosis, Candida albicans, Candida glabrata, and Candida tropicalis with MIC values of 8, 8, 16, and 32 µg/mL, respectively. Furthermore, compound 2 exhibited moderate cytotoxic activity against HCT-116 cell line with IC50 value of 3.7 µM.

Furanone derivative and sesquiterpene from Antarctic marine-derived fungus Penicillium sp. S-1-18.

A new furanone derivative, butanolide A (1), and a new sesquiterpene, guignarderemophilane F (2), together with six known compounds, were isolated from the fungus Penicillium sp. S-1-18 derived from Antarctic marine. The new structures were determined by spectroscopic studies such as 1D- and 2D-NMR and MS analyses. The absolute configuration of 1 was determined by the modified Mosher's method, while the absolute configuration of 2 was determined by calculated ECD spectroscopy. The isolated secondary metabolites were evaluated for their protein tyrosine phosphatase 1B (PTP1B) inhibitory activity. Compound 1 showed moderate inhibitory activity against PTP1B with IC50 value of 27.4 µM.

Biotransformation of total coumarins of Radix Glehniae by Lecanicillium attenuatum W-1-9.

The biotransformation of total coumarins of Radix Glehniae by Lecanicillium attenuatum W-1-9 yielded three new products, lecaniside A (1), lecaniside B (2), and lecaniside C (3). The chemical structures of these metabolites were elucidated based on extensive spectral data, including 2D NMR and HRMS. The hydrogenation, dealkylation, glycosylation, and O-methylation reactions of these metabolites were observed in the present study. In the in vitro assays, compound 1 displayed a little PTP1B inhibitory activity.

Marine Antibody-Drug Conjugates: Design Strategies and Research Progress.

Antibody-drug conjugates (ADCs), constructed with monoclonal antibodies (mAbs), linkers, and natural cytotoxins, are innovative drugs developed for oncotherapy. Owing to the distinctive advantages of both chemotherapy drugs and antibody drugs, ADCs have obtained enormous success during the past several years. The development of highly specific antibodies, novel marine toxins' applications, and innovative linker technologies all accelerate the rapid R&D of ADCs. Meanwhile, some challenges remain to be solved for future ADCs. For instance, varieties of site-specific conjugation have been proposed for solving the inhomogeneity of DARs (Drug Antibody Ratios). In this review, the usages of various natural toxins, especially marine cytotoxins, and the development strategies for ADCs in the past decade are summarized. Representative ADCs with marine cytotoxins in the pipeline are introduced and characterized with their new features, while perspective comments for future ADCs are proposed.

[Antibody-drug conjugates: design and clinical progress].

Antibody-drug conjugates, constructed with monoclonal antibodies, linker and cytotoxins, have distinctive advantages over chemotherapy drugs and antibody drugs in cancer therapy. In this review, the strategy of developing ADCs, and the important progress in past decade are well summarized. The representative ADCs in the pipeline are introduced and characterized with their new features. While, perspective for future directions of ADCs is proposed.

Human IP10-scFv and DC-induced CTL synergistically inhibit the growth of glioma in a xenograft model.

The epidermal growth factor receptor (EGFR) mutant of EGFRvIII is highly expressed in glioma cells, and the EGFRvIII-specific dendritic cell (DC)-induced tumor antigen-specific CD8(+) cytotoxic T lymphocytes (CTLs) may hold promise in cancer immunotherapy. Interferon (IFN)-γ-inducible protein (IP)-10 (IP-10) is a potent inhibitor of angiogenesis and can recruit CXCR3(+) T cells, including CD8(+) T cells, which are important for the control of tumor growth. In this study, we assessed if the combination of IP10-EGFRvIIIscFv with DC-induced CTLs would improve the therapeutic antitumor efficacy. IP10-scFv was generated by linking the human IP-10 gene with the DNA fragment for anti-EGFRvIIIscFv with a (Gly4Ser)3 flexible linker, purified by affinity chromatography, and characterized for its anti-EGFRvIII immunoreactivity and chemotactic activity. DCs were isolated from human peripheral blood monocyte cells and pulsed with EGFRvIII-peptide, then co-cultured with autologous CD8(+) T cells. BALB/c-nu mice were inoculated with human glioma U87-EGFRvIII cells in the brain and treated intracranially with IP10-scFv and/or intravenously with DC-induced CTLs for evaluating the therapeutic effect. Treatment with both IP10-scFv and EGFRvIII peptide-pulsed, DC-induced CTL synergistically inhibited the growth of glioma and prolonged the survival of tumor-bearing mice, which was accompanied by the inhibition of tumor angiogenesis and enhancement of cytotoxicity, thereby increasing the numbers of brain-infiltrating lymphocytes (BILs) and prolonging the residence time of CTLs in the tumor.

Bioactive tyrosine-derived cytochalasins from fungus Eutypella sp. D-1.

Three new cytochalasins Z24 , Z25 , Z26 (1-3, resp.) and one known compound, scoparasin B (4), were isolated from the fungus Eutypella sp. D-1 isolated from the soil of high latitude of the Arctic. The structures of 1-3 were elucidated from spectroscopic data (NMR, MS). These compounds were evaluated for cytotoxic activities against several human tumor cell lines. Among them, compound 1 exhibited moderate cytotoxicity toward human breast cancer MCF-7 cell line with IC50 of 9.33 µM.

Improving prediction of tacrolimus concentration using a combination of population pharmacokinetic modeling and machine learning in chinese renal transplant recipients.

Aims: The population pharmacokinetic (PPK) model-based machine learning (ML) approach offers a novel perspective on individual concentration prediction. This study aimed to establish a PPK-based ML model for predicting tacrolimus (TAC) concentrations in Chinese renal transplant recipients. Methods: Conventional TAC monitoring data from 127 Chinese renal transplant patients were divided into training (80%) and testing (20%) datasets. A PPK model was developed using the training group data. ML models were then established based on individual pharmacokinetic data derived from the PPK basic model. The prediction performances of the PPK-based ML model and Bayesian forecasting approach were compared using data from the test group. Results: The final PPK model, incorporating hematocrit and CYP3A5 genotypes as covariates, was successfully established. Individual predictions of TAC using the PPK basic model, postoperative date, CYP3A5 genotype, and hematocrit showed improved rankings in ML model construction. XGBoost, based on the TAC PPK, exhibited the best prediction performance. Conclusion: The PPK-based machine learning approach emerges as a superior option for predicting TAC concentrations in Chinese renal transplant recipients.

Eutypellaolides A-J, Sesquiterpene diversity expansion of the polar fungus Eutypella sp. D-1.

Eight new 12,8-eudesmanolide sesquiterpenes, eutypellaolides A-H (1-8), and two new eudesmane-type sesquiterpenes, eutypellaolides I-J (9-10), along with four known 12,8-eudesmanolide compounds 11-14, were isolated from the culture extract of the polar fungus Eutypella sp. D-1 by one strain many compounds (OSMAC) approach. The structures of these compounds were determined through comprehensive spectroscopic data and experimental and calculated ECD analysis. Antibacterial, immunosuppressive, and PTP1B inhibition activities of these compounds were evaluated. Compounds 1 and 11 exhibited strong inhibitory activities against Bacillus subtilis and Staphylococcus aureus, with each showing an MIC value of 2 µg/mL. Compound 9 displayed weak immunosuppressive activity against ConA-induced T-cell proliferation with an inhibitory rate of 61.7% at a concentration of 19.8 µM. Compounds 5, 11, and 14 exhibited weak PTP1B inhibition activities with IC50 values of 44.8, 43.2, and 49.5 µM, respectively.

A novel recombinant protein of IP10-EGFRvIIIscFv and CD8(+) cytotoxic T lymphocytes synergistically inhibits the growth of implanted glioma in mice.

The epidermal growth factor receptor (EGFR) mutant of EGFRvIII is highly expressed on glioma cells and has been thought to be an excellent target molecule for immunotherapy. IP-10 is a potent chemokine and can recruit CXCR3(+) T cells, including CD8(+) T cells that are important for the control of tumor growth. This study is aimed at investigating the therapeutic efficacy of a novel fusion protein of IP10-EGFRvIIIscFv (IP10-scFv) in combination with glioma lysate-pulsed DCs-activated CD8(+) cytotoxic T lymphocytes (CTLs) in a mouse model of glioma. A plasmid of pET-IP10-scFv was generated by linking mouse IP-10 gene with the DNA fragment for anti-EGFRvIIIscFv, a (Gly4Ser)3 flexible linker and a His-tag. The recombinant IP10-scFv in E. coli was purified by affinity chromatography and characterized for its anti-EGFRvIII immunoreactivity and chemotactic activity. C57BL/6 mice were inoculated with mouse glioma GL261 cells in the brain and treated intracranially with IP10-scFv and/or intravenously with CTL for evaluating the therapeutic effect. The glioma-specific immune responses were examined. The IP10-scFv retained anti-EGFRvIII immunoreactivity and IP-10-like chemotactic activity. Treatment with both IP10-scFv and CTL synergistically inhibited the growth of glioma and prolonged the survival of tumor-bearing mice, accompanied by increasing the numbers of brain-infiltrating lymphocytes (BILs) and the frequency of CXCR3(+)CD8(+) T cells, enhancing glioma-specific IFN-γ responses and cytotoxicity, and promoting glioma cell apoptosis in mice. Our novel data indicate that IP10-scFv and CTL have synergistic therapeutic effects on inhibiting the growth of mouse glioma in vivo.

A novel cold-adapted ß-galactosidase isolated from Halomonas sp. S62: gene cloning, purification and enzymatic characterization.

A novel 1,170 bp ß-galactosidase gene sequence from Halomonas sp. S62 (BGalH) was identified through whole genome sequencing and was submitted to GenBank (Accession No. JQ337961). The BGalH gene was heterologously expressed in Escherichia coli BL21(DE3) cells, and the enzymatic properties of recombinant BGalH were studied. According to the polyacrylamide gel electrophoresis results and the sequence alignment analysis, BGalH is a dimeric protein and cannot be classified into one of the known ß-galactosidase families (GH1, GH2, GH35, GH42). The optimal pH and temperature were determined to be 7.0 and 45 °C, respectively; the K m and K cat were 2.9 mM and 390.3 s(-1), respectively, for the reaction with the substrate ortho-nitrophenyl-ß-D-galactopyranoside. At 0-20 °C, BGalH exhibited 50-70 % activity relative to its activity under the optimal conditions. BGalH was stable over a wide range of pHs (6.0-8.5) after a 1 h incubation (>93 % relative activity) and was thermostable at 50 °C and below (>60 % relative activity). The enzyme hydrolyzes lactose completely in milk over 24 h at 7 °C. The characteristics of this novel ß-galactosidase suggest that BGalH may be a good candidate for medical researches and food industry applications.

Cloning, expression and functional characterization of a novel trehalose synthase from marine Pseudomonas sp. P8005.

Trehalose synthase (TreS) catalyzes the reversible interconversion of maltose and trehalose. A novel treS gene with a length of 3,369 bp, encoding a protein of 1,122 amino acid residues with a predicted molecular mass of 126 kDa, was cloned from a marine Pseudomonas sp. P8005 (CCTCC: M2010298) and expressed in Escherichia coli. The amino acid sequence identities between this novel TreS and other reported TreS is relatively low. The purified recombinant TreS showed an optimum pH and temperature of 7.2 and 37 °C, respectively. The enzyme displayed a high conversion rate (70 %) of maltose to trehalose during equilibrium and had a higher catalytic efficiency (k cat/K m) for maltose than for trehalose, suggesting its application in the production of trehalose. In addition to maltose and trehalose, this enzyme can also act on sucrose, although this activity is relatively low. Mutagenesis studies demonstrated that enzymatic activity was reduced dramatically by individually substitution with alanine for D78, Y81, H121, D219, E261, H331 or D332, which implied that these residues might be important in P8005-TreS. Experiments using isotope-labeled substrates showed that [(2)H2]trehalose combined with unlabeled trehalose was converted to [(2)H2]maltose and maltose, but without any production of [(2)H]maltose or [(2)H]trehalose and with no incorporation of exogenous [(2)H7]glucose into the disaccharides during the conversion catalyzed by this enzyme. This finding indicated the involvement of an intramolecular mechanism in P8005-TreS catalyzing the reversible interconversion of maltose and trehalose.

[Comparison of direct immune-fluorescent assay and real-time quantitative PCR in detecting the Hantavirus].

OBJECTIVE: To compare the differences between the direct immuno-fluorescent assay (DFA) and real-time quantitative PCR in detecting the Hantavirus (HV) in rat lungs. METHODS: From April to October in 2012, a total of 479 rats were caught by mouse-trap in residential or wild areas in Huxian, Jingyang, and Meixian of Shaanxi province, where haemorrhagic fever with renal syndrome (HFRS) was highly prevalent. The rats were dissected to take the two lungs, one was frozen and applied immuno-fluorescent assay to detect HV antigen while the other one was extracted its RNA and detected HV nucleic acid by real-time quantitative PCR. Then we compared the positive rate of the two methods. RESULTS: Out of the 479 rats, 105 were caught from residential areas and the other 374 were caught from wild areas. Among the 105 rats caught from residential areas, no HV were detected out neither by DFA nor by real-time quantitative PCR. Among the 374 wild rats, 13.1% (49/374) were detected HV positive by DFA and 14.7% (55/374) were detected HV positive by real-time quantitative PCR. The difference showed no statistical significance (χ(2) = 0.402, P = 0.526). When detecting each lung sample, the HV positive rate was 10.2% (49/479) under the detection by DFA while the HV positive rate was 11.5% (55/479) under the detection by real-time quantitative PCR. The difference had no statistical significance (χ(2) = 1.286, P = 0.257) and the consistency coefficient was 68.2% under the paired chi-square test analysis, which showed high consistency (u = 11.759, P < 0.05). The sensitivity of real-time quantitative PCR to detect HV was 77.6% (38/49) comparing with DFA as standard, and the specificity was 96.1% (413/430). Out of the 9 suspected HV positive sample detected by DFA, 6 were confirmed positive by real-time quantitative PCR and 3 were denied. CONCLUSION: Compared with the DFA, real-time quantitative PCR could also be used to detect the infection of HV in rats, and the result might be more stable.