Yumi Kidachi

Anti-Tumor Activity of an Enzymatically Synthesized α-1,6 Branched α-1,4-Glucan, Glycogen

Oral administration of an enzymatically synthesized α-1,4:1,6-glycogen (ESG) at a dose of 50 μg/ml significantly prolonged the survival time of Meth A tumor-bearing mice. ESG also significantly stimulated macrophage-like cells (J774.1), leading to augmented production of nitric oxide (NO) and tumor necrosis factor-α (TNF-α). The weight-average degree of polymerization (DPw) and the ratio of branch linkage (BL) of ESG were 149,000 and 8.1% respectively. β-Amylase-treated ESG, however, lost J774.1-activating activity although inhibited subcutaneous growth of Meth A tumor cells admixed with it. Its DPw and BL changed to 126,000 and 20% respectively. Partially degraded amylopectin [(AP), DPw: 110,000, BL; 5.1] was also effective at stimulating J774.1, but its activity was lower than that of ESG. Other α-glucans [cycloamylose (CA), enzymatically synthesized amylose (ESA), highly branched cyclic dextrin (HBCD), and β-amylase-treated HBCD], of which DPw was lower than that of ESG, showed no J774.1-activating activity and weaker anti-tumor activity.

Novel effects of glycyrrhetinic acid on the central nervous system tumorigenic progenitor cells: induction of actin disruption and tumor cell-selective toxicity.

Licorice extracts are used worldwide in foods and medicines, and glycyrrhetinic acid (GA) is a licorice component that has been reported to induce various important biological activities. In the present study, we show that GA induces actin disruption and has tumor cell-selective toxic properties, and that its selectivity is superior to those of all the clinically available antitumor agents tested. The cytotoxic activity of GA and the tested antitumor agents showed better correlation with the partition coefficient (log P) values rather than the polar surface area (PSA) values. For selective toxicity against tumor cells, GA was most effective at 10 microM that was the same concentration as the previously reported maximum plasma GA level reached in humans ingesting licorice. These results suggest that GA could be utilized as a promising chemopreventive and therapeutic antitumor agent. The underlying mechanisms involved in the selective toxicity to tumor cells by GA are also preliminarily discussed.

Anti-nitric oxide production activity of isothiocyanates correlates with their polar surface area rather than their lipophilicity

There is increasing demand for novel anti-inflammatory drugs with different mechanisms of action. We synthesized a series of isothiocyanates 2b-h based on 6-(methylsulfinyl)hexyl isothiocyanate (6-MITC) found in the pungent spice Wasabia japonica. Inhibitory activities against in-vitro growth of tumor cells and production of nitric oxide (NO) using the mouse macrophage-like cell line J774.1 were noted. All isothiocyanates were optimized by Hartree-Fock/3-21G model, and the logP values and the polar surface area (PSA) values were calculated. Substitution of the methylsulfinyl group (CH(3)S(O)-R) in 6-MITC with a formyl (CHO-R), a methylsulfanyl (CH(2)S-R) or a methyl (CH(3)-R) group reduced the activities of the parent isothiocyanate. Substitution with a formyl group resulted in lower lipophilicity (logP value) whereas substitution with a methylsulfanyl or methyl group resulted in a lower PSA value. The inhibitory activity of isothiocyanates showed better correlation with their PSA values rather than their partition coefficient (logP) values. Isothiocyanates with higher PSA values and some degree of logP value may have potent biological activity.

Larval mesenchyme cell specification in the primitive echinoid occurs independently of the double-negative gate

Echinoids (sea urchins) are divided into two major groups – cidaroids (a ‘primitive’ group) and euechinoids (a ‘derived’ group). The cidaroids are a promising model species for understanding the ancestral developmental mechanisms in echinoids, but little is known about the molecular mechanisms of cidaroid development. In euechinoids, skeletogenic mesenchyme cell specification is regulated by the double-negative gate (DNG), in which hesC represses the transcription of the downstream mesenchyme specification genes (alx1, tbr and ets1), thereby defining the prospective mesenchyme region. To estimate the ancestral mechanism of larval mesenchyme cell specification in echinoids, the expression patterns and roles of mesenchyme specification genes in the cidaroid Prionocidaris baculosa were examined. The present study reveals that the expression pattern and function of hesC in P. baculosa were inconsistent with the DNG model, suggesting that the euechinoid-type DNG is not utilized during cidaroid mesenchyme specification. In contrast with hesC, the expression patterns and functions of alx1, tbr and ets1 were similar between P. baculosa and euechinoids. Based on these results, we propose that the roles of alx1, tbr and ets1 in mesenchyme specification were established in the common ancestor of echinoids, and that the DNG system was acquired in the euechinoid lineage after divergence from the cidaroid ancestor. The evolutionary timing of the establishment of the DNG suggests that the DNG was originally related to micromere and/or primary mesenchyme cell formation but not to skeletogenic cell differentiation.

Selective cytotoxicity of glycyrrhetinic acid against tumorigenic r/m HM-SFME-1 cells: potential involvement of H-Ras downregulation.

With the intensive need for the development of more effective and safer agents for chemoprevention and therapy of human cancer, natural products from plants have been expected to play significant roles in creating new and better chemopreventive and therapeutic agents. Selectivity is also an important issue in cancer prevention and therapy. In the present study, normal serum-free mouse embryo (SFME) and tumorigenic human c-Ha-ras and mouse c-myc cotransfected highly metastatic serum-free mouse embryo-1 (r/m HM-SFME-1) cells were treated with various concentrations of clinically available antitumor agents or glycyrrhetinic acid (GA), and the antiproliferative effects of these compounds were determined by the MTT assay. Western blotting analysis, RT-PCR, fluorescence staining and confocal laser scanning microscopic observation were adopted to analyze H-Ras regulation. GA exhibited the tumor cell-selective toxicity through H-Ras downregulation, and its selectivity was superior to those of all the clinically available antitumor agents examined. For the selective toxicity of tumor cells, GA was most effective at 10 microM. Interestingly, this concentration was the same as the previously reported maximum plasma GA level reached in humans ingesting licorice. These results in the present study suggest that GA with its cytotoxic effects could be utilized as a promising chemopreventive and therapeutic antitumor agent.

Toluene at environmentally relevant low levels disrupts differentiation of astrocyte precursor cells

Abstract Recent findings that describe endocrine disruption caused by exposure to low levels of certain chemicals in the environment have led to a paradigm shift in the way toxicology studies are designed. Toluene at high levels damages the human central nervous system; however, the effects of toluene at low levels have not been studied. The authors used serum-free mouse embryo cells—a precursor of astrocytes—to predict the effect of chemicals on developing brain cells. When serum-free mouse embryo cells were exposed to low levels of toluene, induction of glial fibrillary acidic protein was inhibited. This study demonstrated that environmentally relevant low levels of toluene could disrupt normal prenatal brain development.

Homology modeling and structural analysis of human P-glycoprotein.

Homology modeling and structural analysis of human P-glycoprotein (hP-gp) were performed with a software package the Molecular Operating Environment (MOE). A mouse P-gp (mP-gp; PDB code: 3G5U) was selected as a template for the 3D structure modeling of hP-gp. The modeled hP-gp showed significant 3D similarities at the drug biding site (DBS) to the mP-gp structure. The contact energy profiles of the hP-gp model were in good agreement with those of the mP-gp structure. Ramachandran plots revealed that only 3.5% of the amino acid residues were in the disfavored region for hP-gp. Further, docking simulations between 6-(methylsulfinyl)hexyl isothiocyanate (6-MITC) and the P-gp models revealed the similarity of the ligand-receptor bound location between the hP-gp and mP-gp models. These results indicate that the hP-gp model was successfully modeled and analyzed. To the best of our knowledge, this is the first report of a hP-gp model with a naturally occurring isothiocyanate, and our data verify that the model can be utilized for application to target hP-gp for the development of antitumor drugs. Abbreviations ABC - ATP-binding cassette, ASE-Dock - alpha sphere and excluded volume-based ligand-protein docking, DBS - drug biding site, MDR - multidrug resistance, MOE - Molecular Operating Environment, ITC - isothiocyanate, P-gp - P-glycoprotein.

Micromere formation and expression of endomesoderm regulatory genes during embryogenesis of the primitive echinoid Prionocidaris baculosa.

Blastomere composition and expression profiles of wnt8 and hox11/13b orthologues were examined in the primitive indirect‐developing echinoid Prionocidaris baculosa. We found that blastomere composition in the 16‐cell‐stage Prionocidaris embryos was different from that of the indirect‐developing echinoids belonging to Euechinoidea, a derived group of the echinoids. The sizes of the blastomeres in the 16‐cell‐stage embryo varied, and no embryos formed a “micromere quartet,” a group of four equal‐sized micromeres. The smallest blastomere was usually located around the vegetal pole. We also found significant differences in early expression profiles of wnt8 orthologues of the Prionocidaris and euechinoids. Unlike euechinoids, the expression of wnt8 orthologue of Prionocidaris was not detected at the 16‐cell stage; it began at the 32‐cell stage in the broad area containing the vegetal pole. However, in later stages, the expression profiles of hox11/13b and wnt8 orthologues of Prionocidaris were similar to that of euechinoid orthologues. The present study suggests that there are considerable differences between Prionocidaris and euechinoids in early developmental mechanisms in the vicinity of the vegetal pole.

Structural insight into the ligand-receptor interaction between glycyrrhetinic acid (GA) and the high-mobility group protein B1 (HMGB1)-DNA complex

Structural analysis of the high-mobility group protein B1 (HMGB1)-DNA complex and a docking simulation between glycyrrhetinic acid (GA) and the HMGB1-DNA complex were performed with a software package the Molecular Operating Environment (MOE). An HMGB1-DNA (PDB code: 2GZK) was selected for the 3D structure modeling of the HMGB1-DNA complex. The Site Finder module of the MOE identified 16 possible ligand-binding sites in the modeled HMGB1-DNA complex. The docking simulation revealed that GA possibly inhibits functions of HMGB1 interfering with Lys90, Arg91, Ser101, Tyr149, C230 and C231 in the HMGB1-DNA complex. To the best of our knowledge, this is the first report of an HMGB1-DNA complex with GA, and our data verify that the GA-HMGB1-DNA model can be utilized for application to target HMGB1 for the development of antitumor drugs. Abbreviations ASE-Dock - alpha sphere and excluded volume-based ligand-protein docking, CNS - central nervous system, GA - glycyrrhetinic acid, GL - glycyrrhizin, HMGB1 - high-mobility group protein B1, LBS - ligand-biding site, MOE - Molecular Operating Environment, SRY - sex-determining region on the Y chromosome.

New Cytotoxic Phenolic Derivatives from Matured Fruits of Magnolia denudata

Magnolia species are widely cultivated in Japan as garden plants, and have been found to contain various compounds, including alkaloids, terpenoids, lignans, and neolignans. The constituents of the mature fruits of M. denudata were investigated, and two new phenolic derivatives, named denudalide and denudaquinol, were isolated and characterized, together with a known neolignan compound (denudatin A). Denudalide and denudaquinol showed cytotoxicity against the SFME and r/mHM-SFME-1 cell lines.