Teppei Kawahara

JBIR-124: a novel antioxidative agent from a marine sponge-derived fungus Penicillium citrinum SpI080624G1f01

JBIR-124: a novel antioxidative agent from a marine sponge-derived fungus Penicillium citrinum SpI080624G1f01

JBIR-94 and JBIR-125, Antioxidative Phenolic Compounds from Streptomyces sp. R56-07

Two phenolic compounds, JBIR-94 (1) and JBIR-125 (2), were isolated from the fermentation broth of strain R56-07, which was identified by phylogenetic methods as a novel species of Streptomyces. The structures of 1 and 2 were assigned on the basis of 1D and 2D NMR spectroscopy and MS analyses. Compounds 1 and 2 exhibited 1,1-diphenyl-2-picrylhydrazyl radical scavenging activity with an IC(50) value of 11.4 and 35.1 μM, respectively. These compounds are the first examples of hydroxycinnamic acid amides containing putrescine or spermidine produced by actinomycetes.

Characterization of Giant Modular PKSs Provides Insight into Genetic Mechanism for Structural Diversification of Aminopolyol Polyketides

Polyketides form many clinically valuable compounds. However, manipulation of their biosynthesis remains highly challenging. An understanding of gene cluster evolution provides a rationale for reprogramming of the biosynthetic machinery. Herein, we report characterization of giant modular polyketide synthases (PKSs) responsible for the production of aminopolyol polyketides. Heterologous expression of over 150 kbp polyketide gene clusters successfully afforded their products, whose stereochemistry was established by taking advantage of bioinformatic analysis. Furthermore, phylogenetic analysis of highly homologous but functionally diverse domains from the giant PKSs demonstrated the evolutionary mechanism for structural diversification of polyketides. The gene clusters characterized herein, together with their evolutionary insights, are promising genetic building blocks for de novo production of unnatural polyketides.

Three new depsipeptides, JBIR-113, JBIR-114 and JBIR-115, isolated from a marine sponge-derived Penicillium sp. fS36

Three new depsipeptides, JBIR-113, JBIR-114 and JBIR-115, isolated from a marine sponge-derived Penicillium sp. fS36

Relative configuration of JBIR-129, a cytotoxic 34-membered glycosidic polyol macrolide from Streptomyces sp. RK74.

JBIR-129 was isolated as the potent cytotoxic compound, which consists of the 34-membered polyol macrolide skeleton with five sugar moieties. The relative configuration of the aglycone moiety (C7-C27 and C33-C39) was established by the J-based configuration analysis using vicinal (1)H-(1)H (from (1)H NMR and PS-DQF-COSY spectra) and long-range (1)H-(13)C coupling constants (from sge-HETLOC and several J-resolved HMBC spectra) with steric information obtained from ROESY.

JBIR-137 and JBIR-138, new secondary metabolites from Aspergillus sp. fA75.

Natural products are considered to be good sources for the screening of lead compounds of clinical drugs. We performed many drug screenings employing a variety of assay systems with crude extracts of microbial cultures as a traditional natural product library. In some assay systems, effective application of our crude extract library was difficult without making some improvements. From this viewpoint, we started to construct a purified natural compounds library from cultures of microorganisms. To achieve this, we established the highthroughput detection system for microbial secondary metabolites using UPLC-UV-evaporative light-scattering (ELS)-MS system, and more than 1000 compounds, which we have already isolated, were analyzed by the common analytic method and in our database. The registered compounds in microbial cultures are automatically identified with our system, which allows us easily to pick up unregistered compounds. The unregistered compounds are isolated from the cultures and store in our library. Because Aspergillus species are known to produce more than 950 documented bioactive compounds such as mevinolin, aflatoxin and citrinin,1 their secondary metabolites are an important source to obtain various bioactive compounds. Therefore, we attempted to obtain secondary metabolites from cultures of Aspergillus. During chemical screening based on our analytic system, we isolated a new janthitrem derivative named JBIR137 (1) and a novel metabolite JBIR-138 (2), together with the known compounds, a tremorgenic agent janthitrem B2 and 6-hydroxycyclopiamine B3 from the culture of Aspergillus sp. fA75 (Figure 1). This paper describes the fermentation, isolation, structural elucidation, and briefly, biological activity of 1 and 2. Aspergillus sp. fA75 was isolated from a soil sample collected in the forest at Noda, Chiba Prefecture, Japan. The strain was cultivated in 50-ml test tubes, each containing 15 ml potato dextrose medium (24 g l–1; BD Biosciences, San Jose, CA, USA). The test tubes were shaken reciprocally (320 r.p.m.) at 27 1C for 2 days. Aliquots (4 ml) of the culture were transferred to 500-ml Erlenmeyer flasks containing brown rice 15 g (Akitakomachi, Yamagata, Japan), bacto–yeast extract 30 mg (BD Biosciences), sodium tartarate 15 mg, K2HPO4 15 mg and water 45 ml. The flasks were incubated statically at 27 1C for 14 days. The culture (10 flasks) was extracted with 80% aq. Me2CO (200 ml per flask), and the extract was filtered. After concentration in vacuo, the aqueous residue was extracted with EtOAc (600 ml 3). The EtOAc layer was dried over anhydrous Na2SO4 and evaporated in vacuo, yielding a dark brown gum (815 mg). The extract was fractionated using normal-phase medium-pressure liquid chromatography (Purif-Pack SI-30, Shoko Scientific Co., Yokohama, Japan) with a gradient system of n-hexane–EtOAc (0–25% EtOAc) followed by the stepwise solvent system of CHCl3–MeOH (0, 2, 5, 10, 20, 30 and 100% MeOH) to obtain five fractions (5% fraction-1, 5% fraction-2, 5% fraction-3, 10 and 20–30%). The fractions were monitored by UPLC-UV-ELS-MS system. Compound 1 was isolated from the 5% MeOH fraction-1 (26.5 mg) by reversed-phase HPLC using a CAPCELL PAK C18 MG II column (5.0mm, 20 i.d. 150 mm; Shiseido, Tokyo, Japan) with 85% aq. MeOH containing 0.1% formic acid (flow rate 10 ml min–1, Retention time (Rt)1⁄4 11.6 min). From the 5% MeOH fraction-2 (89.1 mg), janthitrem B2 (5.0 mg) was isolated by HPLC preparation. The 5% MeOH fraction-3 (45.6 mg) was applied to gel filtration chromatography (Sephadex LH-20, GE Healthcare BioSciences AB, Uppsala, Sweden) eluting with CHCl3–MeOH (1:1) to yield crude 2 (45.6 mg). The obtained material was further purified by the HPLC (40% aq. MeOH containing 0.1% formic acid, Rt1⁄4 17.3 min) to yield pure 2 (9.9 mg). The 6-hydroxycyclopiamine B3 (3.7 mg) was purified from the 20–30% MeOH fraction (124.7 mg) using LH-20 column chromatography and HPLC. JBIR-137 (1) was obtained as a colorless amorphous solid: [a]22-66 (MeOH; c 0.13); UV lmax nm (e): 281 (30 300), 290 (32 200) and 374 (5800) in MeOH; IR (nmax): 3400 (hydroxy), 1618, 1455, 1371 (aromatic and pyrrole) cm 1. Its molecular formula was determined

JBIR-129 and -139, cytotoxic 34-membered polyol macrolides of microbial origin.

New 34-membered polyol macrolides JBIR-129 (1) and JBIR-139 (2) were isolated from the culture of the terrestrial Streptomyces RK74. The planar structures of 1 and 2 were established on the basis of 1D and 2D NMR, ESI-TOF-MS, IR, and UV spectra. The relative configurations of the sugar units were determined by analyzing vicinal ¹H-¹H coupling constants and steric information. Both 1 and 2 showed cytotoxic activity against human ovarian adenocarcinoma SKOV-3 cells with IC₅₀ values of 0.3 and 0.4 μM, respectively.

JBIR-120: a new growth inhibitor of hormone-refractory prostate cancer cells.

Prostate cancer is a common nondermatological cancer in older adult men. Androgen receptor (AR) signaling has a central role in prostate cancer cell growth and survival,1 and therefore, androgen ablation therapy is recognized as a standard regimen for the treatment of advanced and metastatic prostate cancers.2 However, most patients who undergo androgen ablation progress from being androgen-dependent to developing hormone-refractory prostate cancer within 2 years after initiating therapy. Although the recurrent tumors are often resistant to standard AR-targeting agents, which cause deprivation of androgens or block androgen–AR interaction, AR-mediated signaling still has a key role in the development and maintenance of hormone-refractory prostate cancers.3, 4, 5 Thus, identifying new therapeutic agents targeting the AR signaling pathway may possibly control the occurrence of hormone-refractory prostate cancers. Mashima et al.6 earlier reported that nigericin can block AR-mediated signaling in hormone-refractory prostate cancer cells. In the current study, we discovered a novel compound—JBIR-120 (1)—extracted from the culture broth of a new Streptomyces strain, RI104-LiC104. This paper describes the isolation, structure elucidation and biological activity of 1.

Foxo3a Inhibitors of Microbial Origin, JBIR-141 and JBIR-142.

JBIR-141 (1) and JBIR-142 (2) were discovered as potent Foxo3a inhibitors that consist of three quite unique substructures, a 1-((dimethylamino)ethyl)-5-methyl-4,5-dihydrooxazole-4-carboxylic acid that is originated from Ala-Thr amino acid residues, a 3-acetoxy-4-amino-7-(hydroxy(nitroso)amino)-2,2-dimethylheptanoic acid, and an α-acyl tetramic acid fused with a 2-methylpropan-1-ol moiety. Their structures involving absolute configurations were determined by spectroscopic data, chemical degradation, anisotropy methods, and LC-MS analyses of diastereomeric derivatives. Compounds 1 and 2 exhibited specific inhibition against Foxo3a transcriptional activity with IC50 values of 23.1 and 166.2 nM, respectively.

Establishment of the absolute configuration of the 34-membered polyol macrolide compound JBIR-129

Establishment of the absolute configuration of the 34-membered polyol macrolide compound JBIR-129