Novel human topoisomerase I inhibitors, topopyrones A, B, C and D. I. Producing strain, fermentation, isolation, physico-chemical properties and biological activity

In the course of a screening program for specific inhibitors of human topoisomerase I using a recombinant yeast, we have discovered four new active compounds. All four compounds were isolated from the culture broth of a fungus, Phoma sp. BAUA2861, and two of them were isolated from the culture broth of a fungus, Penicillium sp. BAUA4206. We designated these compounds as topopyrones A, B, C and D. Topopyrones A, B, C and D selectively inhibited recombinant yeast growth dependent on expression of human topoisomerase I with IC50 values of 1.22, 0.15, 4.88 and 19.63 ng/ml, respectively. The activity and selectivity of topopyrone B were comparable to those of camptothecin. The relaxation of supercoiled pBR322 DNA by human DNA topoisomerase I was inhibited by these compounds, however they did not inhibit human DNA topoisomerase II. Topopyrones A, B, C and D were cytotoxic to all tumor cell lines when tested in vitro. Topopyrone B has potent inhibitory activity against herpesvirus, especially varicella zoster virus (VZV). It inhibited VZV growth with EC50 value of 0.038 microg/ml, which is 24-fold stronger than that of acyclovir (0.9 microg/ml). Topopyrones A, B, and C were inhibitory to Gram-positive bacteria.     

Bisabosquals, novel squalene synthase inhibitors. I. Taxonomy, fermentation, isolation and biological activities

In the course of screening for yeast squalene synthase inhibitors, bisabosqual A was isolated from the culture broth of Stachybotrys sp. RF-7260. The related compounds bisabosquals B, C and D were also isolated from Stachybotrys ruwenzoriensis RF-6853. Bisabosquals inhibited squalene synthases. IC50 values of bisabosqual A against the microsomal squalene synthases from Saccharomyces cerevisiae, Candida albicans, HepG2 cell and rat liver were 0.43, 0.25, 0.95 and 2.5 microg/ml, respectively. Bisabosqual C exhibited inhibitory activities similar to bisabosqual A. Bisabosqual A showed broad spectrum antifungal activity in vitro.     

Reveromycins, new inhibitors of eukaryotic cell growth. II. Biological activities.

Reveromycins A, B, C and D showed inhibitory activity against EGF-stimulated mitogen response in Balb/MK cells. Furthermore reveromycins A, C and D exhibited morphological reversion of srcts-NRK cells, antiproliferative activity against human tumor cell lines and antifungal activity. The effects of reveromycins A, C and D on eukaryotic cells were closely similar to each other, but those of reveromycin B were very weak. In vitro studies revealed that reveromycin A is a selective inhibitor of protein synthesis in eukaryotic cells.     

L-696,474, a novel cytochalasin as an inhibitor of HIV-1 protease. III. Biological activity.

L-696,474, an inhibitor of the HIV-1 protease, was discovered in extracts of the fungal culture Hypoxylon fragiforme (MF5511; ATCC 20995). L-696,474 is a novel cytochalasin with a molecular weight of 477 and an empirical formula of C30H39NO4. L-696,474 inhibited HIV-1 protease activity with an IC50 of 3 microM and the mode of inhibition was competitive with respect to substrate (apparent Ki = 1 microM). Furthermore, L-696,474 was not a slow-binding inhibitor. The inhibition due to L-696,474 was also independent of the HIV-1 protease concentration. L-696,474 was inactive against pepsin, another aspartyl protease; stromelysin, a zinc-metalloproteinase; papain, a cysteine-specific protease or human leucocyte elastase, a serine-specific protease. Two other novel cytochalasins (L-697,318 and L-696,475) isolated from the same culture were inactive against the HIV-1 protease. Commercially available cytochalasins B, C, D, E, F, H and J were inactive while cytochalasin A was as active as L-696,474 against the HIV-1 protease.     

Argyrins,immunosuppressive cyclic peptides from myxobacteria. I. Production,isolation, physico-chemical and biological properties

A group of cyclic peptides consisting of 8 amino acid residues, named argyrins A to H, were isolated from the culture broth of strains of the myxobacterium Archangium gephyra. Argyrin B was found to be a potent inhibitor of T cell independent antibody formation by murine B cells and strongly inhibited the two way murine mixed lymphocyte reaction. All argyrins had slight antibiotic activity, especially against Pseudomonas sp., and inhibited growth of mammalian cell cultures. The growth inhibition was often incomplete and varied highly with different cell lines.     

The myxalamids, new antibiotics from Myxococcus xanthus (Myxobacterales). I. Production, physico-chemical and biological properties, and mechanism of action

From the cell mass and culture supernatant of Myxococcus xanthus strain Mx X12 an antibiotic activity against yeasts, molds and some Gram-positive bacteria could be extracted. It consisted of 4 biologically active compounds which were named myxalamid A, B, C and D. The main component, myxalamid B, was shown to block in beef heart submitochondrial particles the respiratory chain at the site of complex I, i.e. NADH: ubiquinone oxidoreductase. The myxalamids are new antibiotics.     

Sperabillins, new antibacterial antibiotics with potent in vivo activity. Taxonomy, fermentation, isolation and biological activity.

A Gram-negative bacterium was found to produce new antibacterial antibiotics, sperabillins A, B, C and D, and the producing bacterium was characterized and identified as Pseudomonas fluorescens YK-437. Sperabillins were isolated by column chromatographies using cation-exchange resins, activated carbon and cation-exchange Sephadex, and preparative reverse-phase HPLC. Sperabillins showed antibacterial activity against Gram-negative and Gram-positive bacteria including antibiotic-resistant strains of Pseudomonas aeruginosa and Staphylococcus aureus. Sperabillin A inhibited DNA, RNA, protein, and cell wall biosynthesis in Escherichia coli. Sperabillins showed good protective effects in experimentally infected mice.     

Arborcandins A, B, C, D, E and F, novel 1,3-beta-glucan synthase inhibitors: production and biological activity

Arborcandins A, B, C, D, E and F, which possess potent 1,3-beta-glucan synthase inhibitory activity, were isolated from the culture broth of a filamentous fungus, strain SANK 17397. Arborcandins are novel cyclic peptides, that are structurally different from known glucan synthase inhibitors such as echinocandins. The 1,3-beta-glucan synthases of Candida albicans and Aspergillus fumigatus were inhibited by arborcandins with IC50 ranging from 0.012 to 3 microg/ml. The apparent Ki value of arborcandin C for C. albicans and A. fumigatus were 0.12 microM and 0.016 microM, respectively. The inhibition against these two 1,3-beta-glucan synthases by arborcandin C was noncompetitive. These compounds exhibited potent fungicidal activity against Candida spp. with MIC ranging from 0.25 to 8 microg/ml. The growth of A. fumigatus was suppressed by arborcandins with concentrations ranging from 0.063 to 4 microg/ml.     

Antibiotics GE23077, novel inhibitors of bacterial RNA polymerase. I. Taxonomy, isolation and characterization

GE 23077 factors A1, A2, B1 and B2 are novel antibiotics isolated from fermentation broths of an Actinomadura sp. strain. GE23077 antibiotics are cyclic peptides, which inhibit Escherichia coli RNA polymerase at nM concentrations. Both rifampicin-sensitive and rifampicin-resistant polymerases are inhibited, whereas E. coli DNA polymerase and wheat germ RNA polymerase are substantially not affected. In spite of the potent activity on the enzyme, the antibiotics generally show poor activity against whole cell bacteria. The spectrum of activity is restricted to Moraxella catarrhalis, including clinical isolates, with partial activity against Neisseria gonorrhoeae and Mycobacterium smegmatis.     

Pyloricidins, novel anti-helicobacterpylori antibiotics produced by Bacillus sp. I. Taxonomy, fermentation and biological activity

Novel anti-Helicobacter pylori antibiotics, pyloricidins A, A1, A2, B, C and D, were discovered in the culture broth of two bacilli strains. Pyloricidins selectively inhibited the growth of H. pylori. Pyloricidin B was efficacious in the treatment of gastric infection caused by H. pylori in Mongolian gerbils and may be promising for cure of H. pylori infection as a single agent.     

New analogues of rosaramicin isolated from a Micromonospora strain. I. Taxonomy, fermentation, isolation and physico-chemical and biological properties

Antibiotics 6108 A1, B, C and D, a new series of analogues of rosaramicin, were found together with rosaramicin, juvenimicin A4 and M-4365 A1 from the cultured broth of strain BA06108 which was assigned to be a new species of Micromonospora. 6108 A1 and C showed inhibitory activity against Gram-positive and some Gram-negative bacteria as potent as rosaramicin and exhibited low acute toxicity in mice. However, 6108 B showed less potent antimicrobial activity and 6108 D showed higher toxicity than those two antibiotics.     

Arisugacins C and D, novel acetylcholinesterase inhibitors and their related novel metabolites produced by Penicilium sp. FO-4259-11

The mutant of Penicillium sp. FO-4259, an arisugacins A and B producing strain, was found to produce a series of metabolites, designated arisugacins C, D, E, F, G and H, which were structurally related to arisugacins A and B. These compounds were isolated from the culture broth and the physico-chemical and biological properties were examined. The IC50 values of arisugacins C and D against acetylcholinesterase (AChE) were 2.5 microM and 3.5 microM, respectively. However arisugacins E, F, G and H did not inhibit AChE at 100 microM. Though they showed only weak or no activity against AChE compared with arisugacins A and B, they may be useful for the study of the structure-activity relationship.     

Formadicins, new monocyclic beta-lactam antibiotics of bacterial origin. I. Taxonomy, fermentation and biological activities

A Gram-negative bacterium produces new monocyclic beta-lactam antibiotics with a formylamino substituent, named formadicins A, B, C and D. The producing bacterium was taxonomically characterized and designated as Flexibacter alginoliquefaciens sp. nov. YK-49. Formadicins have narrow antibacterial spectra. They are highly active against some species of Pseudomonas, Proteus and Alcaligenes. Of the four, formadicin C shows the most potent antibacterial activity. Several amino acids such as glycine, D-alanine and D-leucine were antagonistic against formadicins. Formadicins, especially formadicins A and C having the formylamino substituent bound to the 3-position of a beta-lactam nucleus, were highly resistant to hydrolysis by various types of beta-lactamases. Formadicins A and C showed affinity for penicillin-binding proteins (PBPs) 1A and 1B in Pseudomonas aeruginosa IFO 3080, but formadicin B and nocardicin A showed affinity only for PBP 1B. Formadicins A and C did not lyse Escherichia coli LD-2 solely at their MICs, but when combined with mecillinam each induced a rapid lysis of this organism.     

A new antitumor substance BE-13793C, produced by a streptomycete. Taxonomy, fermentation, isolation, structure determination and biological activity.

A new antitumor substance, BE-13793C, which has topoisomerase inhibitory activity was isolated from the culture broth of a strain of actinomycetes. The producing strain, BA13793, isolated from a soil sample collected in Seto, Aichi Prefecture, Japan, has a resemblance to Streptoverticillium mobaraense. The active principle was extracted from the mycelium of strain BA13793 with methanol and purified by Sephadex LH-20 column chromatography. BE-13793C showed strong inhibitory activity against topoisomerases I and II and inhibited the growth of doxorubicin-resistant or vincristine-resistant P388 murine leukemia cell lines, as well as their parent P388 cell line.     

Aurafuron A and B, new bioactive polyketides from Stigmatella aurantiaca and Archangium gephyra (Myxobacteria). Fermentation, isolation, physico-chemical properties, structure and biological activity

New antibiotic polyketides, named aurafuron A (1) and B (2) were isolated from culture extracts of myxobacteria of the species Stigmatella aurantiaca and Archangium gephyra, strain Ar 10844. By multi-step chromatography 1 and 2 were separated from a variety of other non-related co-metabolites, and their structures elucidated by spectroscopic methods as new 5-alkenyl-3 3(2H)-furanones. Aurafurons inhibited the growth of some filamentous fungi and additionally, aurafuron B was weakly active against few Gram-positive bacteria. Both compounds also showed cytotoxic activity against the mouse fibroblast cell line L929.     

A80915, a new antibiotic complex produced by Streptomyces aculeolatus. Discovery, taxonomy, fermentation, isolation, characterization, and antibacterial evaluation.

New semi-naphthaquinone antibiotics A80915A, B, C, and D were isolated from the fermented broth of Streptomyces aculeolatus A80915 (NRRL 18422). Factors A and C, present in both the broth filtrate and mycelial methanol extract, and factors B and D, found predominantly in the broth filtrate, were recovered by extraction with ethyl acetate. Purification of the individual factors was accomplished by preparative reverse phase high performance liquid chromatograph on C18 bonded silica supports. Factors A through D show antimicrobial activity against Gram-positive aerobic and anaerobic organisms in vitro. Mechanism of action studies demonstrated nearly complete inhibition of macromolecular biosynthesis (protein, RNA, DNA, and cell wall) by A80915 factors A through D. A less highly cyclized semi-naphthaquinone, A80915 factor G, was isolated from the broth of the strain fermented in an alternate medium.     

Fibrostatins, new inhibitors of prolyl hydroxylase. I. Taxonomy, isolation and characterization

Fibrostatins, potent inhibitors of prolyl hydroxylase, were isolated as orange crystals from the culture broth of strain No. 23924, which was identified as Streptomyces catenulae subsp. griseospora. In vitro inhibitory activity (ID50 value) of fibrostatins A, B, C, D, E and F against prolyl hydroxylase of chick embryos was 23, 39, 29, 180, 10 and 14 microM, respectively.     

Cytotoxic steroids from Sarcococca saligna

Two new C??-steroidal esters, sarsaligates A(1) and B(2), and two new steroidal alkaloids, sarsaligenines A(3) and B(4), together with four known compounds (sarcovagine, sarcorucinine, dimethylamino-3 β-pregnane-20-one, and β-sitosterol 5- 8, respectively), were isolated from the leaves and stems of Sarcococca saligna. The structures of compounds 1-4 were elucidated by NMR and MS spectroscopic analysis. Of the compounds tested, 5 and 6 were the most cytotoxic against the cell lines K562, SK-BR-3, and PANC-1, with IC?? values in the range of 2.25-5.00?μM, while 3 and 4 selectively inhibited HL-60 cells with IC?? values of 2.87 and 3.61 μM, respectively. Compounds 3-6 therefore deserve further evaluation of their cytotoxic potentials.     

Stresgenin B, an inhibitor of heat-induced heat shock protein gene expression, produced by Streptomyces sp. AS-9

Stresgenin B was isolated as an inhibitor of heat-induced heat shock protein (HSP) gene expression from a culture broth of Streptomyces sp. AS-9 by silica gel chromatography and HPLC. The molecular formula of the novel compound was determined as C11H13NO5 by high resolution FAB-MS analysis, and the structure was determined by UV, 1H NMR, 13C NMR, HMQC, HMBC, and NOESY spectra. Stresgenin B inhibited heat-induced luciferase reporter-gene expression directed by the human hsp70B promoter in Chinese hamster ovary (CHO) cells at concentrations lower than the concentrations for inhibition of dexamethasone-induced luciferase reporter-gene expression directed by the mouse mammary tumor virus (MMTV)-LTR promoter. The inhibition of heat-induced reporter gene expression was evident even when cells were exposed to stresgenin B only during heat stress treatment. Moreover, the compound inhibited heat-induced syntheses of hsp72/73, hsp90, and hsp110 and thereby suppressed the induction of thermotolerance. Stresgenin B showed moderate cytotoxic activities against several neoplastic cell lines and also showed antibacterial activities against Micrococcus luteus, Bacillus subtilis and Staphylococcus aureus strains.