title: Exploitation of heparanase inhibitors from microbial metabolites using an efficient visual screening system.
abstract: In this paper we describe the establishment of an efficient visual method for screening heparanase inhibitors, and we present the results of screening 10,000 microbial culture broths. Heparanase-overexpressing stable clones of the human hepatocellular carcinoma HepG2 cells were established and used as an enzyme source. Digestion of heparan sulfate (HS) was detected using novel HS-containing tablets or SDS-polyacrylamide gel electrophoresis. This method was able to find suramin, a known heparanase inhibitor, from a library of typical enzyme inhibitors. By screening 10,000 culture broths of microorganisms (actinomycetes, fungi, and bacteria) an actinomycete strain, RK99-A234, was found to have heparanase inhibitory activity. RK-682 was identified in the fermentation broth as a heparanase inhibitor, IC50 = 17 microM.
authors.affiliation: Antibiotics Laboratory, RIKEN Discovery Research Institute, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan.
authors.forename: Keisuke
authors.initials: K
authors.lastname: Ishida
authors.forename: Takayuki
authors.initials: T
authors.lastname: Teruya
authors.forename: Siro
authors.initials: S
authors.lastname: Simizu
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Osada
mesh_terms: D039903:Actinobacteria / Q000378:metabolism; D000818:Animals; D015153:Blotting, Western; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy / Q000473:pathology; D045744:Cell Line, Tumor; D002999:Clone Cells; D003470:Culture Media; D018076:DNA, Complementary / Q000096:biosynthesis / Q000235:genetics; D004353:Drug Evaluation, Preclinical; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology*; D005966:Glucuronidase / Q000037:antagonists & inhibitors* / Q000096:biosynthesis; D006497:Heparan Sulfate / Q000378:metabolism; D006801:Humans; D008113:Liver Neoplasms / Q000188:drug therapy / Q000473:pathology; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology*; D014162:Transfection; D000099175:Heparanase
chemical_list: D003470:Culture Media; D018076:DNA, Complementary; D004791:Enzyme Inhibitors; D005966:Glucuronidase; D006497:Heparan Sulfate; D010749:Phosphoprotein Phosphatases; D000099175:Heparanase; C094097:RK 682