Anne W. Dombrowski

Discovery of Novel Antifungal (1,3)-β-d-Glucan Synthase Inhibitors

ABSTRACT The increasing incidence of life-threatening fungal infections has driven the search for new, broad-spectrum fungicidal agents that can be used for treatment and prophylaxis in immunocompromised patients. Natural-product inhibitors of cell wall (1,3)-β-d-glucan synthase such as lipopeptide pneumocandins and echinocandins as well as the glycolipid papulacandins have been evaluated as potential therapeutics for the last two decades. As a result, MK-0991 (caspofungin acetate; Cancidas), a semisynthetic analogue of pneumocandin Bo, is being developed as a broad-spectrum parenteral agent for the treatment of aspergillosis and candidiasis. This and other lipopeptide antifungal agents have limited oral bioavailability. Thus, we have sought new chemical structures with the mode of action of lipopeptide antifungal agents but with the potential for oral absorption. Results of natural-product screening by a series of newly developed methods has led to the identification of four acidic terpenoid (1,3)-β-d-glucan synthase inhibitors. Of the four compounds, the in vitro antifungal activity of one, enfumafungin, is comparable to that of L-733560, a close analogue of MK-0991. Like the lipopeptides, enfumafungin specifically inhibits glucan synthesis in whole cells and in (1,3)-β-d-glucan synthase assays, alters the morphologies of yeasts and molds, and produces a unique response in Saccharomyces cerevisiae strains with point mutations in FKS1, the gene which encodes the large subunit of glucan synthase.

Apicidins: Novel cyclic tetrapeptides as coccidiostats and antimalarial agents from Fusarium pallidoroseum

Abstract Apicidin is a cyclic tetrapeptide [cyclo-(N-O-Methyl-L-Trp-L-Ile-D-Pip-L-2-amino-8-oxo-decanoyl)] isolated from Fusarium pallidoroseum by bioassay guided separation. It is a potent inhibitor of apicomplexan histone deacetylase (IC50 1–2 nM), a broad spectrum antiparasitic agent in vitro against apicomplexan parasites and has shown in vivo efficacy against Plasmodium berghei malaria. Isolation, structure and stereochemistry are discussed.

Equisetin and a novel opposite stereochemical homolog phomasetin, two fungal metabolites as inhibitors of HIV-1 integrase

Abstract Integration is an essential step in HIV replication and is catalyzed by an enzyme called integrase. We have isolated equisetin ( 1a ), and a novel opposite stereochemical homolog, phomasetin ( 2a ), from Fusarium heterosporum and a Phoma sp. respectively. They inhibit the invitro recombinant integrase enzyme with IC 50 values of 7–20 μM. Unlike known inhibitors, these compounds also inhibit the integration reactions catalyzed by preintegration complexes isolated from HIV-1 infected cells.

Isolation and structure of nodulisporic acid A1 and A2, novel insecticides from a Nodulisporium sp.

The isolation and structure elucidation of two novel indole terpene insecticides nodulisporic acid A1(2) and A2 (3) from a Nodulisporium sp. are reported.

Structure and conformation of ophiobolin K and 6- epiophiobolin K from Aspergillus ustus as a nematocidal agent.

Abstract The structure, stereochemistry, solution conformation and nematocidal activity of Ophiobolin K (1a) and 6-epiophiobolin K (1b) isolated from Aspergillus ustus has been described. The molecular structure of these compounds were deduced from extensive use of 2D NMR spectroscopy. The stereochemistry and solution conformation was determined in CDCl3 by the application of extensive 1H-1H NOE difference spectroscopy. Significant difference in the conformation of A/B cis 1a and its A/B trans isomer 1b was observed including the folding of the side chain. Ophiobolin K showed a unique boat like conformation whereas the B ring of 6- epiophiobolin K must exist in at least two different conformations. Ophiobolin K exhibited nematocidal activity (ED50 10 μg/mL) against the free-living nematode Caenorhabditis elegans , but the A/B trans isomer 1b was found to be inactive.

Discovery of structurally diverse natural product antagonists of chemokine receptor CXCR3

The chemokines (CXCL9, CXCL10 and CXCL11) and associated CXCR3 receptor are expressed during the inflammatory process from multiple sclerosis, atherosclerosis or organ transplantation resulting in the recruitment of lymphocytes leading to tissue damage. It is hypothesized that blocking of the ligand/CXCR3 receptor interaction has potential to provide opportunity for development of agents that would block tissue rejection. In this paper, four classes of natural product inhibitors (IC50 ranging 0.1–41 μM) have been described that block the CXCR3 receptor interaction of IP-10 ligand. These include a cyclic thiopeptide (duramycin), polyketide glycosides (roselipins), steroidal glycosides (hypoglausin A and dioscin) and a novel alkyl pyridinium alkaloid that were isolated by bioassay-guided fractionation of the organic extracts derived from actinomycete, fungal, plant and marine sources and discovered using 125 I IP-10/CXCR3 binding assay. Duramycin was the most potent with an IC50 of 0.1 μM. Roselipins 2A, 2B and 1A showed IC50 values of 14.6, 23.5, and 41 μM, respectively. Diosgenin glycosides dioscin, hypoglaucin A and kallstroemin D exhibited IC50 values of 2.1, 0.47 and 3 μM, respectively. A novel cyclic 3-alkyl pyridinium salt isolated from a sponge displayed a binding IC50 of 0.67 μ M.

PREUSSOMERIN D FROM THE ENDOPHYTE HORMONEMA DEMATIOIDES

Preussomerin A, an aromatic bis-ketal, was first isolated from the coprophilous fungus Preussia isomera Cain during a study of interspecies competition among dung inhabiting fungi (Weber et al., 1990). Upon further investigation, a group of compounds closely related to preussomerin A was isolated from the same organism and identified as preussomerin B, C, D, E, and F, some of which exhibited significant antifungal and antibacterial properties (Weber and Gloer, 1991). We report the isolation and antibiotic activity of preussomerin D (FIG. 1) from the endophytic fungus Hormonema dematioides Lagerberg & Melin recovered from living plant tissue of a coniferous tree. During a comparative study of fungal endophytes from apparently healthy living leaves and twigs of Chamaecyparis thyoides (L.) B.S.P. (white Atlantic cedar) collected from the Pine Barrens in the intercoastal region of New Jersey, H. dematioides was isolated infrequently from four of five study sites (Bills and Polishook, 1992). One particular isolate JP 184 was recovered from disinfected living foliage collected near Dover Forge, Ocean Co., New Jersey during the spring of 1990. When submitted for natural products screening, this fungus produced in vitro a cytotoxic compound L-733,757 that later proved by silica gel chomatography, mass spectrometry, high pressure liquid chromatography (HPLC), and nuclear magnetic resonance (NMR) to be chemically identical to preussomerin D. Hormonema dematioides, a slow-growing black yeast usually associated with conifer wood, is characterized by basipetal conidiogenesis and dark, thick-walled hyphae with cells wider than long, often with longitudinal septa (FIG. 2) (Hermanides-Nijhof, 1977). It is morphologically similar to and often isolated from coniferous tissue along with Aureobasidium pullulans (de Bary)

Integracides: tetracyclic triterpenoid inhibitors of HIV-1 integrase produced by Fusarium sp.

HIV-1 integrase is a critical enzyme in the replication of HIV-1. It is absent in the host cells and therefore is a good target for treatment of HIV-1 infections. Integracides are members of the tetracyclic triterpenoids family that were isolated from the fermentation broth of a Fusarium sp. Integracide A, a sulfated ester, exhibited significant inhibitory activity against strand transfer reaction of HIV-1 integrase. The discovery, structure elucidation including single crystal X-ray structure and HIV-1 inhibitory activity of these compounds are described.

Biogeography and relatedness of Nodulisporium strains producing nodulisporic acid

Nodulisporic acid, a novel indole terpene with insecticidal properties, was first isolated from a fermentation broth of an endophytic Nodulisporium sp. Following an extensive culture screening effort, fermentations of 12 other strains of Nodulisporium also yielded nodulisporic acid. These strains came from a variety of environmental substrata collected from seven tropical regions in four continents. Cul- tural characteristics and microscopic features show that all the nodulisporic acid-producing Nodulispor- ium strains are morphologically very similar. AP-PCR and sequencing of the rDNA region consisting of the two internal transcribed spacers and the 5.8S gene revealed that the isolates were distributed into three groups, according to the length of the ITS1. The two groups with the longest sequences were not distin- guishable, based on nucleotide divergence data cal- culated from the common region of ITS1. The group of isolates with shorter sequences showed lower ho- mology with the other groups in the ITS1 region, but those strains could not be distinguished from the oth- er groups, according to ITS2 sequences. These data suggest that the nodulisporic acid producing isolates are very closely related and may constitute a single species, although divided into populations showing some degree of genetic differentiation. Comparison of the sequences obtained in this work with sequenc- es from other xylariaceous fungi with Nodulisporium- type anamorphs failed in determining the teleo- morph of the nodulisporic acid-producing Nodulis- porium species. However, it revealed that these iso- lates constitute a monophyletic group, clearly

Diterpenoid pyrones, novel blockers of the voltage-gated potassium channel Kv1.3 from fungal fermentations

Abstract The isolation, structure elucidation and chemical modification of nalanthalide, a novel diterpenoid pyrone blocker of the voltage-gated potassium channel Kv1.3 are reported. The structure–activity relationship of the derivatives with respect to various associated biological activities is also discussed.