Jun Katada

Cyclo-oxygenase-2 enhances basic fibroblast growth factor-induced angiogenesis through induction of vascular endothelial growth factor in rat sponge implants

Angiogenesis is reportedly enhanced by prostaglandins (PGs). In the present study, we investigated whether or not cyclo‐oxygenase (COX)‐2 mediated angiogenesis in chronic and proliferate granuloma. In rat sponge implants, angiogenesis was gradually developed over a 14‐day experimental period as granuloma formed. This angiogenesis was enhanced by topical injections of human recombinant basic fibroblast growth factor (bFGF). In sponge granuloma, mRNA of COX‐1 was constitutively expressed, whereas that of COX‐2 was increased with neovascularization in parallel with that of vascular endothelial growth factor (VEGF). Topical injections of bFGF increased the expression of COX‐2 mRNA. bFGF‐stimulated angiogenesis was inhibited by indomethacin or selective COX‐2 inhibitors, NS‐398, nimesulide, and JTE‐522. The levels of PGE2 and 6‐keto‐PGF1α in the sponge granuloma were increased with bFGF 13 fold and 9 fold, respectively, and these levels were markedly reduced by NS‐398. The expression of VEGF mRNA in the granuloma was also enhanced by bFGF, and was reduced by NS‐398. Topical injections of PGE2 and beraprost sodium, a PGI2 analogue, increased the expression of VEGF mRNA, with angiogenesis enhancement. The enhanced angiogenesis by bFGF was significantly inhibited by topical injections of VEGF anti‐sense oligonucleotide. These results suggested that COX‐2 may enhance bFGF‐induced neovascularization in sponge granuloma by PG‐mediated expression of VEGF, and that a COX‐2 inhibitor would facilitate the management of conditions involving angiogenesis.

(−)-Phenylahistin: A new mammalian cell cycle inhibitor produced by aspergillus ustus

Abstract (−)-Phenylahistin is a fungal diketopiperazine metabolite consisting of L -phenylalanine and isoprenylated dehydrohistidine, and it showed an inhibitory activity on the cell cycle progression of P388 cells in the G2/M phase.

AT2 receptor‐dependent vasodilation is mediated by activation of vascular kinin generation under flow conditions

Physiological roles of angiotensin II type 2 receptor (AT2) are not well defined. This study was designed to investigate the mechanisms of AT2‐dependent vascular relaxation by studying vasodilation in pressurized and perfused rat mesenteric arterial segments. Perfusion of angiotensin II in the presence of AT1 antagonist elicited vascular relaxation, which was completely dependent on AT2 receptors on endothelium. FR173657 (>1 μM), a bradykinin (BK) B2‐specific antagonist, significantly suppressed AT2‐dependent vasodilation (maximum inhibition: 68.5% at 10 μM). Kininogen‐deficient Brown Norway Katholiek rats showed a significant reduction in AT2‐mediated vasodilatory response compared with normal wild‐type Brown Norway rats. Indomethacin (>1 μM), aprotinin (10 μM) and soybean trypsin inhibitor (10 μM) also reduced AT2‐dependent vasodilation. Our results demonstrated that stimulation of AT2 receptors caused a significant vasodilation through local production of BK in resistant arteries of rat mesentery in a flow‐dependent manner. Such vasodilation counterbalances AT1‐dependent vasoconstriction to regulate the vascular tone.

Chymase mediates mast cell-induced angiogenesis in hamster sponge granulomas

We investigated the contribution of mast cell chymase in mast cell-dependent angiogenesis using the hamster sponge-implant model, where angiogenesis in the granulation tissue surrounding the subcutaneously implanted sponge was evaluated by measuring the hemoglobin content. Daily local injection of compound 48/80 (3-100 microg/site/day), a potent mast cell activator, induced formation of granulomas and angiogenesis in time- and dose-dependent manners. This angiogenic response was inhibited by chymase inhibitors including chymostatin (> or = 1 nmol/site/day), soybean trypsin inhibitor (SBTI; > or = 1.4 nmol/site/day) and lima bean trypsin inhibitor (LBTI; > or = 3.3 nmol/site/day), but not by a tryptase inhibitor like leupeptin (> or = 700 nmol/site/day). Although pyrilamine (> or = 2,580 nmol/site/day), a histamine H1 receptor antagonist, and protamine (300 microg/site/day) also inhibited angiogenesis, these effects were much less pronounced than those by chymase inhibitors. Furthermore, antigen-induced angiogenesis in hamsters pre-sensitized with ovalbumin was also inhibited by the chymase inhibitors by 60-70%. Our results suggest that chymase is a major mediator in mast cell-mediated angiogenesis.

Synthesis and biological activities of phenylahistin derivatives

X-ray crystallographic analysis was performed and several phenylahistin derivatives were synthesized to elucidate the structural components necessary for the anti-microtubule activity of phenylahistin. We primarily focused on the unique isoprenylated dehydrohistidine structure. Our results showed that a uniplanar pseudo-three-ring structure formed by the hydrogen bonding of diketopiperazine and imidazole rings is important for the anti-microtubule activity of phenylahistin.

Cytotoxic effects of NSL-1406, a new thienopyrimidine derivative, on leukocytes and osteoclasts.

We synthesized a series of thienopyrimidine derivatives and examined their cytotoxic effects on several cell lines. One of the derivatives, NSL-1406, was shown to exert potent cytotoxic effects on leukemia cell line including P388 cells and J774 cells. It was also inhibitory on mouse osteoclasts and suppressed the in vitro bone resorption by osteoclasts at nanomolar concentrations.

Development of the new potent non-peptide gpIIb/IIIa antagonist NSL-95301 by utilizing combinatorial technique

Abstract The synthetic study of new non-peptide gpIIb/IIIa antagonist, NSL-95301, is presented. The combinatorial technique was engaged in the lead compound discovery process and optimization process to find (+)-NSL-95301. The IC50 value of collagen induced platelet aggregation inhibitory activity is 92 nM.

Symmetrical anhydride-type serine protease inhibitors: Structure-activity relationship studies of human chymase inhibitors

We prepared a potent and relatively selective human chymase inhibitor 9 (-), based on the study of SAR of a symmetrical anhydride-type serine protease inhibitor 1. Kinetic studies suggested that 9 (-) reacts with the Ser residue at the active site of the enzyme, forming a stable acyl enzyme complex. We also showed the importance of the tri-substituted beta-amino acid structure for the potent anti-enzymatic activity.

Structure-activity relationship studies of chloromethyl ketone derivatives for selective human chymase inhibitors.

Based on the SAR study of a classical chloromethyl ketone derivative, Z-PheCH2Cl 1, a series of compounds were synthesized. Among all the derivatives, compound 21 was found to be a potent human chymase inhibitor with no inhibitory activity against human leukocyte cathepsin G.

Discovery and structure--activity relationship studies of a novel and specific peptide motif, Pro-X-X-X-Asp-X, as a platelet fibrinogen receptor antagonist.

A novel hexapeptide, H-Pro-Ser-Nva-Gly-Asp-Trp-OH 6, a specific antagonist of platelet fibrinogen receptor (GpIIb/IIIa), was discovered in a structure-activity relationship (SAR) study where the role of the N-terminal Pro moiety of an RGD-containing peptide, H-Pro-Ser-Arg-Gly-Asp-Trp-OH 1, which is a potent but not specific antagonist toward GpIIb/IIIa integrin, was investigated. This novel peptide 6 exhibits very high activity as a human platelet aggregation inhibitor (IC50 0.59 microM, human PRP/collagen) as well as marked specificity for GpIIb/IIIa. A series of substitutions at the third position (Nva residue) in this hexapeptide, focused on the conformational rigidity, led to compounds which are superior to the original novel peptide 6 with regard to anti-platelet activity. The peptides, H-Pro-Ser-Hyp-Gly-Asp-Trp-OH 17 and H-Pro-Ser-delta Pro-Gly-Asp-Trp-OH 18 with the 5-membered ring structure, which restricted the conformation of the peptide backbone at the third position, inhibited the aggregation of human platelets at submicromolar concentrations (IC50 0.39 and 0.30 microM, respectively). Further structure-activity relationship studies at each position of the peptide sequence suggest a novel motif sequence, Pro-X1-X2-X3-Asp-X4, for specific GpIIb/IIIa integrin recognition, in which the N-terminal free Pro residue and the Asp residue at the fifth position are essential to the activity. This motif sequence is summarized as follows: (1) a small amino acid such as Ser, Ala or Gly is preferable at X1 position; (2) X2 may be any amino acid, preferably a bulky amino acid such as Tle or a cyclic amino acid such as Pro; (3) X3 must be a small amino acid such as Gly; and (4) X4 is preferably an amino acid with an aromatic side chain.