Mamoru Koketsu

Occurrence of a sialylglycopeptide and free sialylglycans in hen's egg yolk

Free sialylglycans (FSGs) and a sialylglycopeptide (SGP) as components of hens egg yolk were found and their chemical structures were determined. SGP and FSGs were isolated from fresh egg yolk by treatment with phenol, gel filtration and successive chromatographies on columns of anion- and cation-exchangers. They were localized in the yolk plasma. The glycan moiety of SGP, which was liberated by PNGase digestion, was studied for the chemical structure by HPLC mapping with p-aminobenzoic ethylester-derivatization, sugar composition analysis, 1H nuclear magnetic resonance and matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry and the glycomoiety was found to be an N-linked disialyl-biantennary glycan. The amino acid sequence of the peptide moiety of SGP was determined to consist of Lys-Val-Ala-Asn-Lys-Thr, the Asn of which is modified with the disialylglycan moiety. FSGs were determined to be two free disialyl-biantennary glycans whose reducing end was either Man beta1-4GlcNAc (FSG-I) or Man beta1-4GlcNAc beta1-4GlcNAc (FSG-II). Since the molar value of SGP present in one egg yolk (2.8 micromol) is comparable to those of well-known major yolk proteins, low density lipoprotein, lipovitellins and phosvitin, it can be considered that SGP is one of the major components in hens egg yolk.

Synthesis of 1,3-thiazine derivatives and their evaluation as potential antimycobacterial agents.

A series of eight 5,6-dihydro-4H-1,3-thiazine derivatives was synthesized by the BF3 x Et2O-catalyzed reaction of selected alpha,beta-unsaturated ketones with thiobenzamide at room temperature. The antimycobacterial activities of these compounds were determined against Mycobacterium tuberculosis H37Rv (ATCC 27294) using the Alamar blue susceptibility assay. Three compounds, 5-hydroxy-3-phenyl-4-aza-2-thiabicyclo[3.3.1]none-3-ene 3a, 4-hydroxy-4-methyl-6-pentyl-2-phenyl-5,6-dihydro-4H-1,3-thiazine 3b, and 4-ethyl-4-hydroxy-2-phenyl-5,6-dihydro-4H-1,3-thiazine 3c exhibited inhibitory activities of 97, 77 and 76%, respectively, at a concentration of 6.25 microg/ml. The actual MIC99 for the most active of these compounds, 3a, was also determined to be >6.25 microg/ml. These results, and especially those for 3a, suggest that 1,3-thiazines are potential lead compounds in the search for new antitubercular agents.

Isoselenocyanates: A Powerful Tool for the Synthesis of Selenium-Containing Heterocycles

Selenium-containing heterocyclic compounds have been well recognized, not only because of their remarkable reactivities and chemical properties, but also because of their diverse pharmaceutical applications. In this context, isoselenocyanates have been emerged as a powerful tool for the synthesis of selenium-containing heterocycles, since they are easy to prepare and store and are safe to handle. In this review the recent advances in the development of synthesis methods for selenium-containing heterocycles from isoselenocyanates are presented and discussed.

Synthesis of 3-selena-1-dethiacephems and selenazepines via iodocyclization.

A convenient approach to synthesize novel selenium-beta-lactams, 3-selena-1-dethiacephems and selenazepines, was accomplished via the regioselective iodocyclization reaction. The substituent of allenyl moieties dramatically influenced the regiochemical outcome in the iodocyclization of allene-selenourea derivatives.

Novel compounds, '1,3-selenazine derivatives' as specific inhibitors of eukaryotic elongation factor-2 kinase.

The inhibitory activities of 5,6-dihydro-4H-1,3-selenazine derivatives on protein kinases were investigated. In a multiple protein kinase assay using a postnuclear fraction of v-src-transformed NIH3T3 cells, 4-ethyl-4-hydroxy-2-p-tolyl-5, 6-dihydro-4H-1,3-selenazine (TS-2) and 4-hydroxy-6-isopropyl-4-methyl-2-p-tolyl-5,6-dihydro-4H-1, 3-selenazine (TS-4) exhibited selective inhibitory activity against eukaryotic elongation factor-2 kinase (eEF-2K) over protein kinase A (PKA), protein kinase C (PKC) and protein tyrosine kinase (PTK). In further experiments using purified kinases, TS-2 (IC(50)=0.36 microM) and TS-4 (IC(50)=0.31 microM) inhibited eEF-2K about 25-fold more effectively than calmodulin-dependent protein kinase-I (CaMK-I), and about 6-fold (TS-2) or 33-fold (TS-4) more effectively than calmodulin-dependent protein kinase-II (CaMK-II), respectively. TS-2 and TS-4 showed much weaker inhibitory activity toward PKA and PKC, while TS-4, but not TS-2, moderately inhibited immunoprecipitated v-src kinase. TS-2 (10.7-fold) and TS-4 (12.5-fold) demonstrated more potent and more specific eEF-2K inhibitory activity than rottlerin, a previously identified eEF-2K inhibitor. TS-2 inhibited ATP or eEF-2 binding to eEF-2K in a competitive or non-competitive manner, respectively. In cultured v-src-transformed NIH3T3 cells, TS-2 also decreased phospho-eEF-2 protein level (IC(50)=4.7 microM) without changing the total eEF-2 protein level. Taken together, these results suggest that TS-2 and TS-4 are the first identified selective eEF-2K inhibitors and should be useful tools for studying the function of eEF-2K.

5-Chloroacetyl-2-amino-1,3-selenazoles attenuate microglial inflammatory responses through NF-κB inhibition

Microglia are the prime effector cells involved in immune and inflammatory responses in the central nervous system (CNS). In pathological conditions, microglia are activated to restore CNS homeostasis. However, chronic microglial activation endangers neuronal survival through the release of various toxic proinflammatory molecules. Thus, negative regulators of microglial activation have been identified as potential therapeutic candidates in many neurological diseases. A number of selenium-containing compounds show antioxidant activity. In this study, we investigated 2-amino-1,3-selenazole derivatives with regard to anti-inflammatory activity or inhibitory effects on microglial activation. Among 26 derivatives of 2-amino-1,3-selenazole and bis-(2-amino-5-selenazoyl) ketones, we observed that 5-chloroacetyl-2-piperidino-1,3-selenazole (CS1) and 5-chloroacetyl-2-morpholino-1,3-selenazole (CS2) strongly inhibited lipopolysaccharide (LPS)-induced nitric oxide (NO) release from BV2 microglial cells. In rat primary cultured microglia, CS1 and CS2 significantly reduced LPS-induced production of NO, tumor necrosis factor (TNF)-alpha, and prostaglandin E(2). Real-time reverse transcription-polymerase chain reaction analysis revealed that the pretreatment of primary microglial cells with CS1 and CS2 attenuated LPS-induced mRNA expression for inducible NO synthase, TNF-alpha, interleukin-1beta, and cyclooxygenase-2. In addition, CS1 and CS2 suppressed LPS-induced activation of nuclear factor-kappaB and Akt. These results suggest that CS1 and CS2 may provide neuroprotection by suppressing the proinflammatory pathway in activated microglia.

Dietary Tricin Suppresses Inflammation-Related Colon Carcinogenesis in Male Crj: CD-1 Mice

The flavone 4′,5,7-trihydroxy-3′,5′-dimethoxyflavone (tricin) present in rice, oats, barley, and wheat exhibits antigrowth activity in several human cancer cell lines and anti-inflammatory potential. However, the chemopreventive activity has not yet been elucidated in preclinical animal models of colorectal cancer. This study was designed to determine whether dietary tricin exerts inflammation-associated colon carcinogenesis induced by azoxymethane and dextran sulfate sodium in mice. Male Crj: CD-1 mice were initiated with a single i.p. injection of azoxymethane (10 mg/kg body weight) and followed by a 1-week exposure to dextran sulfate sodium (1.5%, w/v) in drinking water to induce colonic neoplasms. They were then given the experimental diet containing 50 or 250 ppm tricin. The experiment was terminated at week 18 to determine the chemopreventive efficacy of tricin. In addition, the effects of dietary tricin on the expression of several inflammatory cytokines, including tumor necrosis factor (TNF)-α, were assayed. The development of colonic adenomas and adenocarcinomas was significantly reduced by feeding with 50 and 250 ppm tricin, respectively. Dietary tricin also significantly reduced the proliferation of adenocarcinoma cells as well as the numbers of mitoses/anaphase bridging in adenocarcinoma cells. The dietary administration with tricin significantly inhibited the expression of TNF-α in the nonlesional cypts. Our findings that dietary tricin inhibits inflammation-related mouse colon carcinogenesis by suppressing the expression of TNF-α in the nonlesional cyrpts and the proliferation of adenocarcinomas suggest a potential use of tricin for clinical trials of colorectal cancer chemoprevention.

1,3-Selenazine derivatives induce cytotoxicity and DNA fragmentation in human HT-1080 fibrosarcoma cells

The inhibitory effects of a series of 5,6-dihydro-4H-1,3-selenazine derivatives, 1,3-selenazole, and 5,6-dihydro-4H-1,3-thiazine derivatives on the proliferation of human HT-1080 fibrosarcoma cells were investigated. The compounds 4-ethyl-4-hydroxy-2-p-tolyl-5, 6-dihydro-4H-1,3-selenazine (TS-2) and 4-hydroxy-4-methyl-6-propyl-2-p-tolyl-5,6-dihydro-4H-1,3-selenazine++ + (TS-6) exhibited the strongest inhibitory effect among 1, 3-selenazine derivatives, and the EC(50) of TS-2 and TS-6 was 7.76 and 8.40 microM, respectively. On the other hand, 1,3-selenazole and 5,6-dihydro-4H-1,3-thiazines had no inhibitory effects. TS-2 and TS-6 inhibited the proliferation of HT-1080 cells time- and dose-dependently. They induced dose-dependent DNA fragmentation in HT-1080 cells, revealing a typical apoptosis characteristics. The present study demonstrated that TS-2 and TS-6 inhibited HT-1080 proliferation through the induction of DNA fragmentation.

Preparation of N,N-unsubstituted selenoureas and thioureas from cyanamides

Abstract Reaction of cyanamides with LiAlHSeH and LiAlHSH in the presence of HCl in diethyl ether provided the corresponding N , N -unsubstituted selenoureas and thioureas in moderate to high yields, respectively.

Selenium-Containing Heterocycles Using Selenoamides, Selenoureas, Selenazadienes, and Isoselenocyanates

In recent years, considerable attention has been devoted to the synthesis of selenium-containing heterocycles because of their interesting reactivities and potential pharmaceutical applications. The significant starting materials for the selenium-containing heterocycles synthesis are selenoamides, selenoureas, selenazadienes, and isoselenocyanates. This review article introduces the developed synthetic methods mainly on our findings.