The Discovery of Weddellamycin, a Tricyclic Polyene Macrolactam Antibiotic from an Antarctic Deep-Sea-Derived Streptomyces sp. DSS69, by Heterologous Expression.

Polyene macrolactams are a special group of natural products with great diversity, unique structural features, and a wide range of biological activities. Herein, a cryptic gene cluster for the biosynthesis of putative macrolactams was disclosed from a sponge-associated bacterium, Streptomyces sp. DSS69, by genome mining. Cloning and heterologous expression of the whole biosynthetic gene cluster led to the discovery of weddellamycin, a polyene macrolactam bearing a 23/5/6 ring skeleton. A negative regulator, WdlO, and two positive regulators, WdlA and WdlB, involved in the regulation of weddellamycin production were unraveled. The fermentation titer of weddellamycin was significantly improved by overexpression of wdlA and wdlB and deletion of wdlO. Notably, weddellamycin showed remarkable antibacterial activity against various Gram-positive bacteria including MRSA, with MIC values of 0.10-0.83 µg/mL, and antifungal activity against Candida albicans, with an MIC value of 3.33 µg/mL. Weddellamycin also displayed cytotoxicity against several cancer cell lines, with IC50 values ranging from 2.07 to 11.50 µM.

Cyclopentenone-Containing Tetrahydroquinoline and Geldanamycin Alkaloids from Streptomyces malaysiensis as Potential Anti-Androgens against Prostate Cancer Cells.

Malaymycin (1), a new cyclopentenone-containing tetrahydroquinoline alkaloid, and mccrearamycin E (2), a geldanamycin analogue bearing a rare ring-contracted cyclopentenone moiety, and a C2-symmetric macrodiolide (7) were isolated from Streptomyces malaysiensis SCSIO41397. Their structures including absolute configurations were determined by detailed analyses of NMR and HRMS data and ECD calculations. The occurrence of mccrearamycin E (2) bearing a ring-contracted cyclopentenone is rare in the geldanamycin class. All isolated compounds were evaluated for their cytotoxicities against five cancer cell lines. As a result, compounds 1, 4, 5, and 7 showed cytotoxicity against some or all of the five cancer cell lines with IC50 values ranging from 0.067 to 7.2 µM. In particular, compound 1 inhibited the growth of C42B and H446 cell lines with IC50 values of 67 and 70 nM, respectively. Malaymycin (1) significantly induced cell cycle arrest at the G0/G1 phase in C42B cell lines and caused cell shrinkage and inhibited the expression of the androgen receptor (AR) at both the mRNA and protein levels in a dose-dependent manner. Further examination by qRT-PCR analysis showed that 1 strongly suppressed the expression of AR target genes KLK2 and KLK3 in the C42B and 22RV1 cell lines, which suggested that 1 might be a promising potential lead compound for the development of a treatment for the castration-resistant prostate cancer (CRPC).

Novel Macrolactams from a Deep-Sea-Derived Streptomyces Species.

Four polyene macrolactams including the previously reported niizalactam C (4), and three new ones, streptolactams A-C (1-3) with a 26-membered monocyclic, [4,6,20]-fused tricyclic and 11,23-oxygen bridged [14,16]-bicyclic skeletons, respectively, were isolated from the fermentation broth of the deep-sea sediment-derived Streptomyces sp. OUCMDZ-3159. Their structures were determined based on spectroscopic analysis, X-ray diffraction analysis, and chemical methods. The abiotic formation of compounds 2 and 4 from compound 1 were confirmed by a series of chemical reactions under heat and light conditions. Compounds 1 and 3 showed a selective antifungal activity against Candida albicans ATCC 10231.

Polycyclic Macrolactams Generated via Intramolecular Diels-Alder Reactions from an Antarctic Streptomyces Species.

Three new polycyclic macrolactams, cyclamenols B-D (1-3), together with a known macrolactam, cyclamenol A (4), were isolated from the Streptomyces sp. OUCMDZ-4348. Their structures including absolute configurations were determined on the basis of spectroscopic analysis, chemical methods, and ECD calculations. The biosynthetic pathways involving intramolecular Diels-Alder reactions were proposed. Compound 1 exhibited selective inhibition against the gastric carcinoma cell line N87 with an IC50 value of 10.8 µM.

New geldanamycin derivatives with anti Hsp properties by mutasynthesis.

Mutasynthetic supplementation of the AHBA blocked mutant strain of S. hygroscopicus, the geldanamycin producer, with 21 aromatic and heteroaromatic amino acids provided new nonquinoid geldanamycin derivatives. Large scale (5 L) fermentation provided four new derivatives in sufficient quantity for full structural characterisation. Among these, the first thiophene derivative of reblastatin showed strong antiproliferative activity towards several human cancer cell lines. Additionally, inhibitory effects on human heat shock protein Hsp90α and bacterial heat shock protein from H. pylori HpHtpG were observed, revealing strong displacement properties for labelled ATP and demonstrating that the ATP-binding site of Hsps is the target site for the new geldanamycin derivatives.

Divergolides T⁻W with Apoptosis-Inducing Activity from the Mangrove-Derived Actinomycete Streptomyces sp. KFD18.

Four new ansamycins, named divergolides T-W (1-4), along with two known analogs were isolated from the fermentation broth of the mangrove-derived actinomycete Streptomyces sp. KFD18. The structures of the compounds, including the absolute configurations of their stereogenic carbons, were determined by spectroscopic data and single-crystal X-ray diffraction analysis. Compounds 1-4 showed cytotoxic activity against the human gastric cancer cell line SGC-7901, the human leukemic cell line K562, the HeLa cell line, and the human lung carcinoma cell line A549, with 1 being the most active while compounds 5 and 6 were inactive against all the tested cell lines. Compounds 1 and 3 showed very potent and specific cytotoxic activities (IC50 2.8 and 4.7 µM, respectively) against the SGC-7901 cells. Further, the apoptosis-inducing effect of 1 and 3 against SGC-7901 cells was demonstrated by two kinds of staining methods for the first time.

DNA Binding and Molecular Dynamic Studies of Polycyclic Tetramate Macrolactams (PTM) with Potential Anticancer Activity Isolated from a Sponge-Associated Streptomyces zhaozhouensis subsp. mycale subsp. nov.

A sponge-associated actinomycete (strain MCCB267) was isolated from a marine sponge Mycale sp. collected in the Indian Ocean off the Southeast coast of India. Phylogenetic studies of this strain using 16S rRNA gene sequencing showed high sequence similarity to Streptomyces zhaozhouensis. However, strain MCCB267 showed distinct physiological and biochemical characteristic features and was thus designated as S. zhaozhouensis subsp. mycale. subsp. nov. A cytotoxicity-guided fractionation of the crude ethyl acetate extract of strain MCCB267 culture medium yielded four pure compounds belonging to the polycyclic tetramate macrolactam (PTM) family of natural products: ikarugamycin (IK) (1), clifednamide A (CF) (2), 30-oxo-28-N-methylikarugamycin (OI) (3), and 28-N-methylikarugamycin (MI) (4). The four compounds exhibited promising cytotoxic activity against NCI-H460 lung carcinoma cells in vitro, by inducing cell death via apoptosis. Flow cytometric analysis revealed that 1, 3, and 4 induced cell cycle arrest during G1 phase in the NCI-H460 cell line, whereas 2 induced cell arrest in the S phase. A concentration-dependent accumulation of cells in the sub-G1 phase was also detected upon treatment of the cancer cell line with compounds 1-4. The in vitro cytotoxicity studies were supported by molecular docking and molecular dynamic simulation analyses. An in silico study revealed that the PTMs can bind to the minor groove of DNA and subsequently induce the apoptotic stimuli leading to cell death. The characterization of the isolated actinomycete, the study of the mode of action of the four PTMs, 1-4, and the molecular docking and molecular dynamic simulations analyses are herein described.

Ansamycins with Antiproliferative and Antineuroinflammatory Activity from Moss-Soil-Derived Streptomyces cacaoi subsp. asoensis H2S5.

Three new 21-membered macrocyclic benzenoid ansamycins, trienomycins J-L (1-3), together with seven known analogues, trienomycins A-G (4-10), were isolated from liquid culture of the moss soil-derived actinomycete Streptomyces cacaoi subsp. asoensis H2S5. The structures of the new compounds were elucidated by extensive NMR spectroscopic analysis and HRESIMS data. The absolute configurations of trienomycins were established by Marfey's method. Antiproliferative assays showed that compound 1 had the greatest activity against HepG2 cells, with an IC50 value of 0.1 µM. The induction of apoptosis of HepG2 cells by 1 was investigated by flow cytometry and evaluation of nuclear morphology. In addition, all of the compounds inhibited nitric oxide production with IC50 values of 0.02 to 8.3 µM, and compounds 1, 4, and 7 were the most potent inhibitors. These findings will facilitate the development of new antineuroinflammatory agents.

Umezawamides, new bioactive polycyclic tetramate macrolactams isolated from a combined-culture of Umezawaea sp. and mycolic acid-containing bacterium.

New polycyclic tetramate macrolactams, Umezawamides A (1) and B (2) were isolated from a combined-culture of Umezawaea sp. RD066910 and mycolic-acid containing bacterium Tsukamurella pulmonis TP-B0596. Their planar structures and partial stereochemistries were determined based on the spectroscopic analysis, MMFF conformational search, and ECD calculations. Umezawamides are the first secondary metabolites isolated from the genus Umezawaea and they exhibited cytotoxicities to P388 murine leukemia cells. Furthermore, umezawamide A (1) showed growth inhibitory activity against Candida albicans.

JBIR-150, a novel 20-membered polyene macrolactam from marine-derived Streptomyces sp. OPMA00071.

During the course of constructing a natural product library for drug screening consisting of microbial culture extracts originated from marine samples, we evaluated natural product components profiles via UPLC TOF-MS and routine biological tests for cytotoxic and antibiotic activities for all of the culture extract samples. By combination of chemical screening and biological activities, we succeeded in discovering a 20-membered macrolactam antibiotic subsequently designated JBIR-150 (1) from a marine-derived actinomycete identified as Streptomyces sp. that was isolated from an Okinawan marine sediment. The chemical structure of 1 was determined by interpreting NMR spectroscopic and mass spectrometric data. Compound 1 exhibited moderate cytotoxicity against MESO-1 and Jurkat cells.

Gunnilactams A-C, Macrocyclic Tetralactams from the Mycelial Culture of the Entomogenous Fungus Paecilomyces gunnii.

Three novel macrocyclic tetralactams, gunnilactam A (1), gunnilactam B (2), and gunnilactam C (3), were isolated from the submerged fermentation broth of Paecilomyces gunnii, an entomogenous fungus identified as the anamorph of Cordyceps gunnii. Their structures were determined using NMR data, HREIMS, and single-crystal X-ray crystallography. Gunnilactam A exhibited selective cytotoxic activity against human prostate cancer C42B cells with an IC50 value of 5.4 µM.

Mccrearamycins A-D, Geldanamycin-Derived Cyclopentenone Macrolactams from an Eastern Kentucky Abandoned Coal Mine Microbe.

Four cyclopentenone-containing ansamycin polyketides (mccrearamycins A-D), and six new geldanamycins (Gdms B-G, including new linear and mycothiol conjugates), were characterized as metabolites of Streptomyces sp. AD-23-14 isolated from the Rock Creek underground coal mine acid drainage site. Biomimetic chemical conversion studies using both simple synthetic models and Gdm D confirmed that the mccrearamycin cyclopentenone derives from benzilic acid rearrangement of 19-hydroxy Gdm, and thereby provides a new synthetic derivatization strategy and implicates a potential unique biocatalyst in mccrearamycin cyclopentenone formation. In addition to standard Hsp90α binding and cell line cytotoxicity assays, this study also highlights the first assessment of Hsp90α modulators in a new axolotl embryo tail regeneration (ETR) assay as a potential new whole animal assay for Hsp90 modulator discovery.

New C-19-modified geldanamycin derivatives: synthesis, antitumor activities, and physical properties study.

Thiazinogeldanamycin (2) was identified from Streptomyces hygroscopicus 17997 at the late stage of the fermentation. The pH was firstly proposed as an important factor in the biosynthesis of it. It was verified that 2 was produced by direct chemical reactions between geldanamycin (1, GDM) and cysteine or aminoethanethiol hydrochloride at pH > 7 in vitro. The proposed synthesis pathway for compound 2 was also discussed. Eleven new C-19-modified GDM derivatives, including five stable hydroquinone form derivatives, were synthesized, most of which exhibited desirable properties such as lower cytotoxicity, increased water solubility, and potent antitumor activity. Especially, compounds 5 and 8 showed antitumor activities against HepG2 cell with IC50 values of 2.97-6.61 µM, lower cytotoxicity and at least 15-fold higher water solubility compared with 1 in pH 7.0 phosphate buffer.

A unique macrolactam derivative via a [4+6]-cycloaddition from Streptomyces niveus.

One new macrolactam derivative, nivelactam (1) and one new polyenoic acid derivative, niveamide (2), along with two other known 20-atom macrolactams (3 and 4) were isolated from the fermentation broth of Streptomyces niveus, which obtained from the forest soil in northeastern China. The structures of 1 and 2 were elucidated on the basis of HRESIMS, IR, and NMR spectroscopic data analyses. Compound 1 was proposed as an intramolecular [4+6]-cycloaddition product of 3 by S. niveus, and displayed moderate cytotoxic activity against a panel of human tumor cell lines in vitro, with IC50 values ranging from 3.76 ± 0.58 to 15.02 ± 2.81 µM.

Niizalactams A-C, Multicyclic Macrolactams Isolated from Combined Culture of Streptomyces with Mycolic Acid-Containing Bacterium.

A terrestrial bacterium, Streptomyces sp. NZ-6, produced niizalactams A-C (1-3), unprecedented di- and tricyclic macrolactams, by coculturing with the mycolic acid-containing bacterium Tsukamurella pulmonis TP-B0596. Their complete structures, including absolute configurations, were elucidated on the basis of spectroscopic data and chemical derivatization. Their unique skeletons are proposed to be biosynthesized from a common 26-membered macrolactam intermediate by SN2 cyclization or an intramolecular Diels-Alder reaction.

Bioactive 7-Oxabicyclic[6.3.0]lactam and 12-Membered Macrolides from a Gorgonian-Derived Cladosporium sp. Fungus.

One new bicyclic lactam, cladosporilactam A (1), and six known 12-membered macrolides (2-7) were isolated from a gorgonian-derived Cladosporium sp. fungus collected from the South China Sea. Their complete structural assignments were elucidated by comprehensive spectroscopic investigation. Quantum chemistry calculations were used in support of the structural determination of 1. The absolute configuration of 1 was determined by calculation of its optical rotation. Cladosporilactam A (1) was the first example of 7-oxabicyclic[6.3.0]lactam obtained from a natural source. Compound 1 exhibited promising cytotoxic activity against cervical cancer HeLa cell line with an IC50 value of 0.76 µM.

Bioactive Polycyclic Tetramate Macrolactams from Lysobacter enzymogenes and Their Absolute Configurations by Theoretical ECD Calculations.

Two new polycyclic tetramate macrolactams, lysobacteramides A (1) and B (2), together with HSAF (heat-stable antifungal factor, 3), 3-dehydroxy HSAF (4), and alteramide A (5) were isolated from a culture of Lysobacter enzymogenes C3 in nutrient yeast glycerol medium. Their structures were determined by MS and extensive NMR analysis. The absolute configurations of 1-5 were assigned by theoretical calculations of their ECD spectra. Although HSAF and analogues were reported from several microorganisms, their absolute configurations had not been established. The isolation and the absolute configurations of these compounds revealed new insights into the biosynthetic mechanism for formation of the polycycles. Compounds 1-4 exhibited cytotoxic activity against human carcinoma A549, HepG2, and MCF-7 cells with IC50 values ranging from 0.26 to 10.3 µM. Compounds 2 and 3 showed antifungal activity against Fusarium verticillioides with IC50 value of 47.9 and 6.90 µg/mL, respectively.

Heronamides D-F, polyketide macrolactams from the deep-sea-derived Streptomyces sp. SCSIO 03032.

Three new macrolactams, heronamides D-F (1-3), were isolated from the deep-sea-derived Streptomyces sp. SCSIO 03032 upon changing cultivation conditions. The planar structures of heronamides D-F (1-3) were elucidated by extensive MS and NMR spectroscopic analyses and comparisons with the closely related heronamides A-C. The relative configurations of 1-3 were deduced by detailed analysis of (3)JHH values and NOESY data. The absolute configurations of 1 and 2 were determined by chemical modifications and application of the modified Mosher's method. None of the compounds exhibited obvious antimicrobial or cytotoxic activities.

19-[(1'S,4'R)-4'-Hydroxy-1'-methoxy-2'-oxopentyl]geldanamycin, a natural geldanamycin analogue from Streptomyces hygroscopicus 17997.

A novel natural geldanamycin analogue was discovered in Streptomyces hygroscopicus 17997. Its 4,5-dihydro form was also identified in the gdmP gene disruption mutant of Streptomyces hygroscopicus 17997. The structures of the two compounds were determined to be 19-[(1'S,4'R)-4'-hydroxy-1'-methoxy-2'-oxopentyl]geldanamycin (1) and 19-[(1'S,4'R)-4'-hydroxy-1'-methoxy-2'-oxopentyl]-4,5-dihydrogeldanamycin (2), respectively, by extensive spectroscopic data analysis, including 2D NMR, modified Mosher's method, and electronic circular dichroism. Compared to geldanamycin, 1 and 2 showed increased water solubility and decreased cytotoxicity against HepG2 cells.