Summary of Research Project Results Under the JSPS FY2000
"Research for the future Program"



1.Research Institution Kitasato University
 
2.Research Area Integrated Fields
 
3.Research Field Production of Novel Useful Substances through Integration of Life Sciences and Chemistry
 
4.Term of Project FY1996〜FY2000
 
5.Project Number 96I00304
 
6.Title of Project Discovery and production of lead compounds for medicines from microorganisms

7.Projetct Leader
Name Institution,Department Title of Position
Satoshi Omura Kitasato Univ., Grad. Sch. Pharm. Sciences Professor

8.Core Members

Names Institution,Department Title of Position
Haruo Tanaka Kitasao Univ., School of Pharmaceutical Sciences Professor
Haruo Ikeda Kitasao Univ., School of Pharmaceutical Sciences Assoc. Professor
Hiroshi Tomoda The Kitasato Institute, Res. Ctr. Biological Function Assoc. Director

9.Cooperating Researchers

Names Institution,Department Title of Position
Masahiko Hayashi The Kitasato Institute, Ctr. Basic Research Res. Head
Mun Chual Rho JSPS Researcher
Ichiji Namatame JSPS Researcher

10.Summary of Research Results

This research project focused on two main themes as follows; 1. Discovery and production of new lead compounds for diseases of human immunodeficiency virus (HIV), cancer, atherosclerosis and inflammation from microorganisms. --- In order to discover lead compounds from microorganisms, original screening systems were conducted by utilizing intact cells with special functions. 1) From screening for anti-HIV substances in a syncytium formation assay system, actinohivin (AH) was isolated from the culture broth of an actinomycete strain K97-0003. AH consists of a 114-amino acid chain that exhibits internal sequence triplication. AH inhibited both T-cell and macrophage tropic syncytium formations. AH was found to bind gp120 by selectively recognizing high-mannose polysaccharide chain of glycoproteins. 2) From screening for anti-atherosclerotic agents in a foam cell formation assay sytem, beauveriolides, phenochalasins and K97-0239 were isolated from fungal and actinomycete culture broths. These inhibitors reduced the number and the size of lipid droplets formed in macrophages. Beauveriolides inhibited the synthesis of cholesteryl ester by blocking acyl-CoA:cholesterol acyltransferase activity specifically. Importantly, beauveriolide proved to be orally active in both LRL receptor knockout mice and apo E knockout mice, reducing atheroma lesion in the aota and heart. 3) From screening for tumor metastasis inhibitors in a cell-cell adhision assay system, macrosphelides (MS) were isolated from the fungal culture broth. MS-A inhibited the binding of sialyl LewisX to E-selectin. Intraperitoneal injection of MS-A also inhibited the pulmonary metastasis of B16/BL6 melanoma in mice. On the other hand, MS-B was found to show immunosuppressive activity in mouse models. The mechanism of immunosuppression of MS-B may differ from those of FK506 and cyclosporine A.
2. Analysis of the biosyhthetic gene of anthelmintic avermectin in Streptomyces avermitilis. --- The entire avermectin biosynthetic gene cluster was cloned and characterized. The gene consists of 18 ORFs in a distance of 82 kb containing polyketide synthases organized as two sets of six modular repeats and post-polyketide modification enzymes. Based on the findings, unnatural avermectin analogs like ivermectin were biosynthesized by manipulating the gene.

11.Key Words

(1)Microbial metabolites、(2)Lead compounds for medicines、(3)Anti-HIV actinohivin
(4)Anti-atherosclerotic beauveriolide、(5)Anti-metastatic macrosphelide、(6)Anthelmintic
(7)Avermectin、(8)Polyketide biosynthetic gene、(9)Gene manupulation

12.References

[Reference Articles]
Author Title of Article
S. Takamatsu et al. A new inhibitor of melanogenesis, albocycline K3, produced by Streptomyces sp. OH-3984.
Journal Volume Year Pages Concerned
J. Antibiot 49 1996 485-486

Author Title of Article
M. Hayashi, et al Chlovalicin, a new cytocidal antibiotic produced by Sphorothrix sp. FO-4649. I. Taxonomy, fermentation, isolation and biological activities.
Journal Volume Year Pages Concerned
J. Antibiot. 49 1996 631-634

Author Title of Article
S. Takamatsu, et al Chlovalicin, a new cytocidal antibiotic produced by Sphorothrix sp. FO-4649. II. Physicochemical properties and structural elucidation.
Journal Volume Year Pages Concerned
J. Antibiot. 49 1996 635-638

Author Title of Article
Y. Takahashi, et al Physiological regulation of sporulation of Kitasatosporia setae in submerged culture.
Journal Volume Year Pages Concerned
Actinomycetol. 10 1996 43-49

Author Title of Article
H. Tomoda, et al Amidepsine E, an inhibitor of diacylglycerol acyltransferase produced by Humicola sp. FO-5969.
Journal Volume Year Pages Concerned
J. Antibiot. 49 1996 929-931

Author Title of Article
H. Miura, et al Lactacystin increase LDL receptor level on Hep G2 cells.
Journal Volume Year Pages Concerned
Biochem. Biophys. Res. Commun. 227 1996 684-687

Author Title of Article
Y. Takahashi, et al Rare actinomycetes isolated from desert soils.
Journal Volume Year Pages Concerned
Actinomycetol. 10 1996 91-97

Author Title of Article
H. Gouda, et al. Stereostructure of (-)-chloropeptin I, a novel inhibitor of gp120-CD4 binding, via high-temperature molecular dynamics, Monte Carlo conformational searching and NMR spectroscopy.
Journal Volume Year Pages Concerned
J. Am. Chem. Soc. 118 1996 13087-13088

Author Title of Article
H. Tanaka, et al. Chloropeptins, new anti-HIV antibiotics inhibiting gp120-CD4 binding from Streptomyces sp. I. Taxonomy, fermentation, isolation and physico-chemical properties and biological activities.
Journal Volume Year Pages Concerned
J. Antibiot 50 1997 58-65

Author Title of Article
K. Matsuzaki, et al. Chloropeptins, new anti-HIV antibiotics inhibiting gp120-CD4 binding from Streptomyces sp. II. Structure elucidation of chloropeptin I.
Journal Volume Year Pages Concerned
J. Antibiot. 50 1997 66-69

Author Title of Article
H. Tomoda, et al. Synthesis of four chiral isomers of β-lactone DU-6622 and inhibition of HMG-CoA synthase by the specific (2R, 3R)-isomer.
Journal Volume Year Pages Concerned
J. Org. Chem. 62 1997 2161-2165

Author Title of Article
S. Takamatsu, et al. Macrosphelides C and D, novel inhibitors of cell adhesion.
Journal Volume Year Pages Concerned
J. Antibiot. 50 1997 878-880

Author Title of Article
Y. -P. Kim, et al. Pyridindolols K1 and K2, new alkaloids from Streptomyces sp. K93-0711.
Journal Volume Year Pages Concerned
J. Antibiot. 50 1997 189-193

Author Title of Article
N. Tabata, et al. Xanthohumols, diacylglycerol acyltransferase inhibitors, from Humulus lupulus.
Journal Volume Year Pages Concerned
Phytochemistry 46 1997 683-687

Author Title of Article
T. Sunazuka, et al. Relative and absolute stereochemistries and total synthesis of(+)-macrosphelides A and B, potent, orally bioavailable inhibitor of cell-cell adhesion.
Journal Volume Year Pages Concerned
J. Am. Chem. Soc. 119 1997 10247-10248

Author Title of Article
H. Ikeda, et al. Genetic analysis of avermectin B O-methyltransferase in avermectin-producing Streptomyces avermitilis.
Journal Volume Year Pages Concerned
Gene 206 1998 175-180

Author Title of Article
M. -C. Rho, et al. Enhancement of drug accumulation by andrastin A produced by Penicillium sp. FO-3929 in vincristine-resistant KB cells.
Journal Volume Year Pages Concerned
J. Antibiot 51 1998 68-72

Author Title of Article
K. Matsuzaki, et al. New brominated and halogen-less derivatives and structure-activity relationship of azaphilones inhibiting gp120-CD4 binding.
Journal Volume Year Pages Concerned
J. Antibiot. 51 1998 1004-1011

Author Title of Article
I. Namatame, et al. Beauveriolides, specific inhibitors of lipid droplet formation in mouse macrophages, produced by Beauveria sp. FO-6979.
Journal Volume Year Pages Concerned
J. Antibiot. 52 1999 1-6

Author Title of Article
I. Namatame, et al. Structure elucidation of fungal beauveriolide III, a novel inhibitor of lipid droplet formation in mouse macrophages.
Journal Volume Year Pages Concerned
J. Antibiot. 52: 1999 7-12

Author Title of Article
H. Zhang, et al Zelkovamycin, a new cyclic peptide antibiotic from Streptomyces sp. K96-0670. I. Production, isolation and biological properties.
Journal Volume Year Pages Concerned
J. Antibiot. 52 1999 29-33

Author Title of Article
N. Tabata, et al. Zelkovamycin, a new cyclic peptide antibiotic from Streptomyces sp. K96-0670. II. Structure elucidation.
Journal Volume Year Pages Concerned
J. Antibiot. 52 1999 34-39

Author Title of Article
S. Omura, et al. Roselipins, novel fungal metabolites having a highly methylated fatty acid modified with a mannose and an arabinitol
Journal Volume Year Pages Concerned
J. Antibiot. 52 1999 586-589

Author Title of Article
H. Tomoda, et al. Roselipins, inhibitor of diacylglycerol acyltransferase, produced by Gliocladium roseum KF-1040.
Journal Volume Year Pages Concerned
J. Antibiot. 52 1999 689-694

Author Title of Article
N. Tabata, et al. Structure elucidation of roselipins, inhibitors of diacylglycerol acyltransferase produced by Gliocladium roseum KF-1040.
Journal Volume Year Pages Concerned
J. Antibiot 52 1999 815-826

Author Title of Article
H. Tomoda, et al. Phenochalasins, inhibitors of lipid droplet formation in mouse macrophages, produced by Phomopsis sp. FT-0211.
Journal Volume Year Pages Concerned
J. Antibiot. 52 1999 851-856

Author Title of Article
H. Tomoda, et al. Structure elucidation of fungal phenochalasins, novel inhibitors of lipid droplet formation in mouse macrophages.
Journal Volume Year Pages Concerned
J. Antibiot. 52 1999 857-861

Author Title of Article
H. Tomoda, et al. Differential inhibition of HMG-CoA synthase and pancreatic lipase by the specific chiral isomers of β-lactone DU-6622.
Journal Volume Year Pages Concerned
Biochem. Biophys. Res. Commun. 265 1999 536-540

Author Title of Article
I. Namatame, et al. Complete inhibition of mouse macrophage-derived foam cell formation by triacsin C.
Journal Volume Year Pages Concerned
J. Biochem. 125 1999 319-327

Author Title of Article
H. Ikeda. Genetic analysis of biosynthesis of polyketide anthelmintic macrolide avermectin in Streptomyces avermitilis.
Journal Volume Year Pages Concerned
Actinomycetologica 13 1999 94-112

Author Title of Article
H. Ikeda, et al. Organization of the biosynthetic gene cluster for the polyketide anthelmintic macrolide avermectin in Streptomyces avermitilis.
Journal Volume Year Pages Concerned
Proc. Natl. Acad. Sci. USA 96 1999 9509-9514

Author Title of Article
I. Namatame, et al. Effect of fungal metabolites cytochalasans on lipid droplet formation in mouse macrophages.
Journal Volume Year Pages Concerned
J. Antibiot. 53 2000 19-25

Author Title of Article
R. Obata, et al. Structure-activity relationships study of pyripyropenes: reversal of cancer cell multidrug resistance.
Journal Volume Year Pages Concerned
J. Antibiot 53 2000 422-425

Author Title of Article
M. -C. Rho, et al. Reversal of multidrug resistance by 7-O-benzoylpyripyropene A in multidrug-resistant tumor cells.
Journal Volume Year Pages Concerned
J. Antibiot. 53 2000 1201-1206

Author Title of Article
A. Fukami, et al. A new antimicrobial antibiotic from Actinoplanes capillaceus sp. K95-5561T.
Journal Volume Year Pages Concerned
J. Antibiot. 53 2000 1212-1214

Author Title of Article
A. Matsumoto, et al. Actinoplanes capillaceus sp. nov., a new species of the genus Actinoplanes.
Journal Volume Year Pages Concerned
Antonie van Leeuwenhoek 78 2000 107-115

Author Title of Article
S. Funayama, et al. Cytotoxic alkaloids of Pachsandra terminalis.
Journal Volume Year Pages Concerned
Biol. Pharm. Bull. 23 2000 262-264

Author Title of Article
H. Chiba, et al. Actinohivin, a novel anti-HIV protein that inhibits syncytium formation: isolation, characterization, and biological activities.
Journal Volume Year Pages Concerned
Biochem. Biophys. Res. Commun. 282 2000 595-601

Author Title of Article
J. Inokoshi, et al. Molecular cloning of actinohivin, a novel anti-HIV protein from an actinomycete, and its expression in Escherichia coli.
Journal Volume Year Pages Concerned
Biochem. Biophys. Res. Commun. 281 2000 1261-1265

Author Title of Article
S. Omura & K. Shiomi Production of medicines and microbial secondary metabolites
Journal Volume Year Pages Concerned
Bioscience and Industry 54 1996 633-635

Author Title of Article
S. Omura Antibiotics in the 21st century.
Journal Volume Year Pages Concerned
Jpn J. Antibiot 49 1996 755-765

Author Title of Article
H. Ikeda and S. Omura Production of useful unnatural products by manupulating biosynthtic genes of microbial secondary metabolites.
Journal Volume Year Pages Concerned
Kagakutoseibutsu 34 1996 761-771

Author Title of Article
H. Ikeda and S. Omura Avermectin biosynthesis.
Journal Volume Year Pages Concerned
Chem. Rev. 97 1997 2591-2609

Author Title of Article
H. Tanaka, et al. gp120-CD4 binbing inhibitors exhibiting anti-HIV activity.
Journal Volume Year Pages Concerned
Nippon Nougeikagaku Zassi 71 1997 530-534

Author Title of Article
H. Ikeda and S. Omura Combinatorial biosynthesis of polyketides.
Journal Volume Year Pages Concerned
Tanpakusitsu Kakusan Kouso 43 1998 1265-1277

Author Title of Article
K. Shiomi, et al. Meroterpenoids with various biological activities produced by fungi.
Journal Volume Year Pages Concerned
Pure Appl. Chem. 71 1999 1059-1064

Author Title of Article
S. Omura Anti-infective agents into the 21st century.
Journal Volume Year Pages Concerned
Nippon Saikingaku Zassi 54 1999 795-813

Author Title of Article
I. Namatame, et al. Discovery of anti-atherosclerotic agents from microorganisms.
Journal Volume Year Pages Concerned
Bioscience and Industry 58 2000 193-194

[Reference Books]
Author Title of Book
H. Tanaka and S. Omura Biotechnology / (Screening of novel receptor-activity compounds of microbial origin)
Publisher Year Pages
VCH Verlargsgesellschaft mbH 1997 107-132

Author Title of Book
H. Tomoda and S. Omura Enzyme Technologies for Pharmaceutical and Biotechnological Applications/(Screening for inhibitors of lipid metabolism)
Publisher Year Pages
Marcel Dekker Inc., New York 2001 343-378


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