Tatsuhiko Kodama

The University of Tokyo, Tōkyō, Japan

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Publications (452)2535.85 Total impact

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    ABSTRACT: ROBO1, fibronectin type-III domain (Fn)-containing protein, is a novel immunotherapeutic target for hepatocellular carcinoma in humans. The crystal structure of the antigen-binding fragment (Fab) of B2212A, the monoclonal antibody against the third Fn domain (Fn3) of ROBO1, was determined in pursuit of antibody drug for hepatocellular carcinoma. This effort was conducted in the presence or absence of the antigen, with the chemical features being investigated by determining the affinity of the antibody using molecular dynamics and thermodynamics. The structural comparison of B2212A Fab between the complex and the free form revealed that the interfacial Tyr(L) 50 (superscripts L, H, and F stand for the residues in the light chain, heavy chain, and Fn3, respectively) played important roles in Fn3 recognition. That is, the aromatic ring of Tyr(L) 50 pivoted toward Phe(F) 68, forming a CH/π interaction and a new hydrogen bond with the carbonyl O atom of Phe(F) 68. Molecular dynamics simulations predicted that the Tyr(L) 50-Phe(F) 68 interaction almost entirely dominated Fab-Fn3 binding, and Ala-substitution of Tyr(L) 50 led to a reduced binding of the resultant complex. On the contrary, isothermal titration calorimetry experiments underscored that Ala-substitution of Tyr(L) 50 caused an increase of the binding enthalpy between B2212A and Fn3, but importantly, it induced an increase of the binding entropy, resulting in a suppression of loss in the Gibbs free energy in total. These results suggest that mutation analysis considering the binding entropy as well as the binding enthalpy will aid in the development of novel antibody drugs for hepatocellular carcinoma. This article is protected by copyright. All rights reserved. Copyright © 2014 The Protein Society.
    Protein Science 03/2015; 24(3). DOI:10.1002/pro.2619 · 2.86 Impact Factor
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    ABSTRACT: We synthesized two new biotin analogues, biotin carbonate 5 and biotin carbamate 6. These molecules were designed to reversibly bind with streptavidin by replacing the hydrogen bond donor NH group(s) of biotin's cyclic urea moiety to oxygen. Biotin carbonate 5 was synthesized from L-arabinose (7), which furnishes the desired stereochemistry at the 3,4-cis-dihydroxy groups, in 11% overall yield (10 steps). Synthesis of biotin carbamate 6 was accomplished from L-cysteine-derived chiral aldehyde 33 in 11% overall yield (7 steps). Surface plasmon resonance analysis of water-soluble biotin carbonate analogue 46 and biotin carbamate analogue 47 revealed that KD values of these compounds for binding to streptavidin were 6.7x10-6 M (46) and 1.7x10-10 M (47), respectively. These values were remarkably greater than that of biotin (KD = 10-15 M), and thus indicate the importance of the nitrogen atoms of biotin's cyclic urea motif for the strong binding between biotin and streptavidin. Copyright © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
    Chemistry - An Asian Journal 02/2015; DOI:10.1002/asia.201500120 · 4.57 Impact Factor
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    ABSTRACT: For a multistep pre-targeting method using antibodies, a streptavidin mutant with low immunogenicity, termed LISA-314, was produced previously as a drug delivery tool. However, endogenous biotins with high affinity (Kd < 10(-10) M) for the binding pocket of LISA-314 prevents access of exogenous biotin-labeled anticancer drugs. In the present study, we improve the binding pocket of LISA-314 to abolish its affinity for endogenous biotin species, therefore ensuring that the newly designed LISA-314 binds only artificial biotin analogue. The replacement of three amino acid residues was performed in two steps to develop a mutant termed V212, which selectively binds to 6-(5-((3aS,4S,6aR)-2-iminohexahydro-1H-thieno[3,4-d]imidazol-4-yl)pentanamido)hexanoic acid (IMNtail). Surface plasmon resonance results showed that V212 has a Kd value of 5.9 × 10(-7) M towards IMNtail, but no binding affinity for endogenous biotin species. This V212/IMNtail system will be useful as a novel delivery tool for anticancer therapy. © The Authors 2015. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.
    Journal of Biochemistry 02/2015; DOI:10.1093/jb/mvv004 · 3.07 Impact Factor
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    ABSTRACT: The streptavidin/biotin interaction has been widely used as a useful tool in research fields. For application to a pre-targeting system, we previously developed a streptavidin mutant that binds to an iminobiotin analog while abolishing affinity for natural biocytin. Here, we design a bivalent iminobiotin analog that shows 1000-fold higher affinity than before, and determine its crystal structure complexed with the mutant protein.
    Bioscience Biotechnology and Biochemistry 01/2015; DOI:10.1080/09168451.2014.991692 · 1.27 Impact Factor
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    ABSTRACT: In this study, we propose a supercomputer-assisted drug design approach involving all-atom molecular dynamics (MD)-based binding free energy prediction after the traditional design/selection step. Because this prediction is more accurate than the empirical binding affinity scoring of the traditional approach, the compounds selected by the MD-based prediction should be better drug candidates. In this study, we discuss the applicability of the new approach using two examples. Although the MD-based binding free energy prediction has a huge computational cost, it is feasible with the latest 10 petaflop-scale computer. The supercomputer-assisted drug design approach also involves two important feedback procedures: The first feedback is generated from the MD-based binding free energy prediction step to the drug design step. While the experimental feedback usually provides binding affinities of tens of compounds at one time, the supercomputer allows us to simultaneously obtain the binding free energies of hundreds of compounds. Because the number of calculated binding free energies is sufficiently large, the compounds can be classified into different categories whose properties will aid in the design of the next generation of drug candidates. The second feedback, which occurs from the experiments to the MD simulations, is important to validate the simulation parameters. To demonstrate this, we compare the binding free energies calculated with various force fields to the experimental ones. The results indicate that the prediction will not be very successful, if we use an inaccurate force field. By improving/validating such simulation parameters, the next prediction can be made more accurate.
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    ABSTRACT: Polycomb repressive complex 1 (PRC1) plays an essential role in the epigenetic repression of gene expression during development and cellular differentiation via multiple effector mechanisms including ubiquitination of H2A and chromatin compaction. However, whether it regulates the step-wise progression of adipogenesis is unknown. Here we show that FBXL10/KDM2B is an anti-adipogenic factor that is up-regulated during the early phase of 3T3-L1 preadipocyte differentiation and in adipose tissue in a diet induced model of obesity. Interestingly, inhibition of adipogenesis does not require the JmjC demethylase domain of FBXL10 but it does require the F-box and leucine rich repeat (LRRs) domains which we show recruit a non-canonical polycomb repressive complex 1 (PRC1) containing RING1B, SKP1, PCGF1 and BCOR. Knockdown of either RING1B or SKP1 prevented FBXL10 mediated repression of 3T3-L1 preadipocyte differentiation indicating that PRC1 formation mediates the inhibitory effect of FBXL10 on adipogenesis. Using ChIP-seq we show that FBXL10 recruits RING1B to key specific genomic loci surrounding the key cell-cycle and adipogenic genes Cdk1, Uhrf1, Pparg1 and Pparg2 to repress adipogenesis. These results suggest that FBXL10 represses adipogenesis through targeting a non-canonical PRC1 complex to repress key genes (e.g. Pparg) that control conversion of pluripotent cells into the adipogenic lineage. Copyright © 2014, The American Society for Biochemistry and Molecular Biology.
    Journal of Biological Chemistry 12/2014; DOI:10.1074/jbc.M114.626929 · 4.60 Impact Factor
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    ABSTRACT: The Tokyo Medical University Hospital in Japan and the Lund University hospital in Sweden have recently initiated a research program with the objective to impact on patient treatment by clinical disease stage characterization (phenotyping), utilizing proteomics sequencing platforms. By sharing clinical experiences, patient treatment principles, and biobank strategies, our respective clinical teams in Japan and Sweden will aid in the development of predictive and drug related protein biomarkers. Data from joint lung cancer studies are presented where protein expression from Neuro- Endocrine lung cancer (LCNEC) phenotype patients can be separated from Small cell- (SCLC) and Large Cell lung cancer (LCC) patients by deep sequencing and spectral counting analysis. LCNEC, a subtype of large cell carcinoma (LCC), is characterized by neuroendocrine differentiation that small cell lung carcinoma (SCLC) shares. Pre-therapeutic histological distinction between LCNEC and SCLC has so far been problematic, leading to adverse clinical outcome. An establishment of protein targets characteristic of LCNEC is quite helpful for decision of optimal therapeutic strategy by diagnosing individual patients. Proteoform annotation and clinical biobanking is part of the HUPO initiative (http://www.hupo.org) within chromosome 10 and chromosome 19 consortia.
    12/2014; 3(1):61. DOI:10.1186/s40169-014-0038-x
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    ABSTRACT: We previously created a low-immunogenic core streptavidin mutant No. 314 (LISA-314) by replacing six amino-acid residues for use as a delivery tool for an antibody multistep pre-targeting process (Yumura et al., Protein science, 22, 213-221, 2013). Here, we performed high-resolution X-ray structural analyses of LISA-314 and wild-type streptavidin to investigate the effect of substitutions on the protein function and the three-dimensional structure. LISA-314 forms a tetramer in the same manner as wild-type streptavidin. The binding mode of d-biotin in LISA-314 is also completely identical to that in wild-type streptavidin, and conformational changes were observed mostly at the side chains of substituted sites. Any large conformational changes corresponding to the reduction of B factors around the substituted sites were not observed. These results demonstrated the LISA-314 acquired low immunogenicity without losing structural properties of original wild-type streptavidin. Copyright © 2014. Published by Elsevier B.V.
    Journal of Bioscience and Bioengineering 11/2014; DOI:10.1016/j.jbiosc.2014.10.025 · 1.74 Impact Factor
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    ABSTRACT: Pentraxins belong to the superfamily of conserved proteins that are characterized by a cyclic multimeric structure. Pentraxin 3 (PTX3) is a long pentraxin which can be produced by different cell types upon exposure to various inflammatory signals. Inside the neutrophil PTX3 is stored in form of granules localized in the cytoplasm. Neutrophilic granules are divided into three types: azurophilic (primary) granules, specific (secondary) granules and gelatinase (tertiary) granules. PTX3 has been considered to be localized in specific (secondary) granules. Immunofluorescent analyses using confocal laser microscopic examination were performed to clarify the localization of all three groups of granules within the cytoplasm of the mature neutrophils and neutrophils stimulated with IL-8. Furthermore, PTX3 was localized in primary granules of promyelocyte cell line HL-60. As a result, we suggest that PTX3 is localized not only in specific granules, but is also partly expressed in primary and tertiary granules. After the stimulation with IL-8, irregular reticular structures called neutrophil extracellular traps (NETs) were formed, three types of granules were trapped by NETs and PTX3 showed partial colocalization with these granular components. PTX3 localized in all three types of granules in neutrophils may play important roles in host defense. Copyright © 2014. Published by Elsevier Inc.
    Experimental and Molecular Pathology 11/2014; 98(1). DOI:10.1016/j.yexmp.2014.11.009 · 2.88 Impact Factor
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    ABSTRACT: VEGF is a key regulator of endothelial cell migration, proliferation and inflammation, which leads to activation of several signaling cascades including the calcineurin-NFAT pathway. NFAT is important for not only immune responses but also cardiovascular development and pathogenesis of Down syndrome. We recently showed that the VEGF-calcineurin-NFAT signaling axis regulates tumor angiogenesis and tumor metastasis by using Down syndrome model mice and clinical patient samples. However, the connection between genome-wide views of the NFAT mediated gene regulation, and the downstream gene function in endothelium has not been extensively studied. Here, we performed comprehensive mapping of genome-wide NFATc1 binding in VEGF-stimulated primary cultured endothelial cells, and elucidate functional consequences of the VEGF-NFATc1 mediated phenotypic changes. Comparison of NFATc1 ChIP-sequence profile and epigenetic histone marks revealed that predominant NFATc1-occupied peaks overlapped with promoter-associated histone marks. Moreover, we identified two novel NFATc1 regulated genes, CXCR7 and RND1. CXCR7 knockdown abrogated SDF-1- and VEGF-mediated cell migration and tube formation. SiRNA treatment of RND1 impaired vascular barrier function, caused RhoA hyper-activation, and further stimulated VEGF-mediated vascular outgrowth from aortic rings. Taken together, these findings suggest that dynamic NFATc1-binding to target genes is critical for VEGF-mediated endothelial cell activation. CXCR7 and RND1 are NFATc1 target-genes with multiple functions including regulation of cell migration, tube formation, and barrier formation in endothelial cells.
    Journal of Biological Chemistry 08/2014; 289(42). DOI:10.1074/jbc.M114.555235 · 4.60 Impact Factor
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    ABSTRACT: Macrophages are important for maintaining intestinal immune homeostasis. Here, we show that PPARβ/δ (peroxisome proliferator-activated receptor β/δ) directly regulates CD300a in macrophages that express the immunoreceptor tyrosine based-inhibitory motif (ITIM)-containing receptor. In mice lacking CD300a, high-fat diet (HFD) causes chronic intestinal inflammation with low numbers of intestinal lymph capillaries and dramatically expanded mesenteric lymph nodes. As a result, these mice exhibit triglyceride malabsorption and reduced body weight gain on HFD. Peritoneal macrophages from Cd300a-/- mice on HFD are classically M1 activated. Activation of toll-like receptor 4 (TLR4)/MyD88 signaling by lipopolysaccharide (LPS) results in prolonged IL-6 secretion in Cd300a-/- macrophages. Bone marrow transplantation confirmed that the phenotype originates from CD300a deficiency in leucocytes. These results identify CD300a-mediated inhibitory signaling in macrophages as a critical regulator of intestinal immune homeostasis.
    Scientific Reports 06/2014; 4:5412. DOI:10.1038/srep05412 · 5.08 Impact Factor
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    ABSTRACT: BackgroundROBO1 is a membrane protein that functions in axon guidance. ROBO1 contributes to tumour metastasis and angiogenesis and may have potential as a target protein of immunotherapy because ROBO1 is specifically expressed at high levels in hepatocellular carcinoma. In this study, we examined biodistribution and radioimmunotherapy (RIT) using a radioisotope-labelled anti-ROBO1 monoclonal antibody (MAb) against hepatocellular carcinoma models.MethodsROBO1-positive HepG2 human hepatocellular carcinoma xenograft nude mice were used in this study. We conjugated anti-ROBO1 MAb with 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA), and the conjugates were labelled with 111In and 90Y. To study biodistribution, the 111In-DOTA-anti-ROBO1 MAb was injected into HepG2 xenograft mice via the tail vein. To evaluate any antitumour effect, a RIT study was performed, and the 90Y-DOTA-anti-ROBO1 MAb was injected via the tail vein. Tumour volume, mouse weight, and blood cell count were periodically measured throughout the experiments. The tumours and organs of mice were collected, and a histopathological analysis was carried out.ResultsThe tumour uptake of 111In-anti-ROBO1 MAb in HepG2 xenograft mice was 15.0% ± 0.69% injected dose per gram at 48 h after injection.Immunotherapy with cold-anti-ROBO1 MAb (70 μg) did not cause a significant antitumour effect. RIT with 6.7 MBq of 90Y-anti-ROBO1 MAb caused significant tumour growth suppression. Transient body weight loss and bone-marrow suppression were observed. Histopathological analyses of tumours revealed the fatal degeneration of tumour cells, significant reduction of the Ki-67 index, and an increase of the apoptosis index. Normal organs showed no significant injury, but a transient reduction of hematopoietic cells was observed in the spleen and in the sternal bone marrow.ConclusionsThese results suggest that RIT with 90Y-anti-ROBO1 MAb is a promising treatment for ROBO1-positive hepatocellular carcinoma.
    06/2014; 4:29. DOI:10.1186/s13550-014-0029-3
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    ABSTRACT: Statins exert atheroprotective effects through the induction of specific transcriptional factors in multiple organs. In endothelial cells, statin-dependent atheroprotective gene up-regulation is mediated by Kruppel-like factor (KLF) family transcription factors. To dissect the mechanism of gene regulation, we sought to determine molecular targets by performing microarray analyses of human umbilical vein endothelial cells (HUVECs) treated with pitavastatin, and KLF4 was determined to be the most highly induced gene. In addition, it was revealed that the atheroprotective genes induced with pitavastatin, such as nitric oxide synthase 3 (NOS3) and thrombomodulin (THBD), were suppressed by KLF4 knockdown. Myocyte enhancer factor-2 (MEF2) family activation is reported to be involved in pitavastatin-dependent KLF4 induction. We focused on MEF2C among the MEF2 family members and identified a novel functional MEF2C binding site 148 kb upstream of the KLF4 gene by chromatin immunoprecipitation along with deep sequencing (ChIP-seq) followed by luciferase assay. By applying whole genome and quantitative chromatin conformation analysis {chromatin interaction analysis with paired end tag sequencing (ChIA-PET), and real time chromosome conformation capture (3C) assay}, we observed that the MEF2C-bound enhancer and transcription start site (TSS) of KLF4 came into closer spatial proximity by pitavastatin treatment. 3D-Fluorescence in situ hybridization (FISH) imaging supported the conformational change in individual cells. Taken together, dynamic chromatin conformation change was shown to mediate pitavastatin-responsive gene induction in endothelial cells.
    PLoS ONE 05/2014; 9(5):e96005. DOI:10.1371/journal.pone.0096005 · 3.53 Impact Factor
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    ABSTRACT: Synergistic transcriptional activation by different stimuli has been reported along with a diverse array of mechanisms, but the full scope of these mechanisms has yet to be elucidated. We present a detailed investigation of hypoxia-inducible factor (HIF) 1 dependent gene expression in endothelial cells which suggests the importance of crosstalk between the peroxisome proliferator-activated receptor (PPAR) beta/delta and HIF signaling axes. A migration assay shows a synergistic interaction between these two stimuli, and we identify angiopoietin-like 4 (ANGPTL4) as a common target gene by using a combination of microarray and ChIP-seq analysis. We profile changes of histone marks at enhancers under hypoxia, PPARbeta/delta agonist and dual stimulations and these suggest that the spatial proximity of two response elements is the principal cause of the synergistic transcription induction. A newly developed quantitative chromosome conformation capture assay shows the quantitative change of the frequency of proximity of the two response elements. To the best of our knowledge, this is the first report that two different transcription factors cooperate in transcriptional regulation in a synergistic fashion through conformational change of their common target genes.
    Genome biology 04/2014; 15(4):R63. DOI:10.1186/gb-2014-15-4-r63 · 10.47 Impact Factor
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    ABSTRACT: Cell adhesion mediated by cadherins depends critically on the homophilic trans-dimerization of cadherin monomers from apposing cells, generating the so-called strand-swap dimer (ss-dimer). Recent evidence indicates that the ss-dimer is preceded by an intermediate species known as the X-dimer. Until now, the stabilized form of the X-dimer had only been observed in E-cadherin among the classical type I cadherins. Herein we report the isolation and characterization of the analogous X-dimer of human P-cadherin. Small-angle X-ray scattering (SAXS) and site directed mutagenesis data indicates that the overall architecture of the X-dimer of human P-cadherin is similar to that of E-cadherin. The X-dimerization is triggered by Ca2+ and governed by specific protein-protein interactions. The attachment of three molecules of Ca2+ with high affinity (KD = 9 μM) stabilizes the monomeric conformation of P-cadherin (ΔTM = 17 °C). The Ca2+-stabilized monomer subsequently dimerizes in the X-configuration by establishing protein-protein interactions requiring the first two extracellular domains of the cadherin. The homophilic X-dimerization is very specific, since the presence of the highly homologous E-cadherin does not interfere with the self-recognition of P-cadherin. These data suggest that the X-dimer could play a key role in the specific cell-cell adhesion mediated by human P-cadherin.
    Biochemistry 02/2014; 53(11). DOI:10.1021/bi401341g · 3.38 Impact Factor
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    ABSTRACT: Although thousands of long noncoding RNAs (lncRNAs) are localized in the nucleus, only a few dozen have been functionally characterized. Here we show that nuclear enriched abundant transcript 1 (NEAT1), an essential lncRNA for the formation of nuclear body paraspeckles, is induced by influenza virus and herpes simplex virus infection as well as by Toll-like receptor3-p38 pathway-triggered poly I:C stimulation, resulting in excess formation of paraspeckles. We found that NEAT1 facilitates the expression of antiviral genes including cytokines such as interleukin-8 (IL8). We found that splicing factor proline/glutamine-rich (SFPQ), a NEAT1-binding paraspeckle protein, is a repressor of IL8 transcription, and that NEAT1 induction relocates SFPQ from the IL8 promoter to the paraspeckles, leading to transcriptional activation of IL8. Together, our data show that NEAT1 plays an important role in the innate immune response through the transcriptional regulation of antiviral genes by the stimulus-responsive cooperative action of NEAT1 and SFPQ.
    Molecular cell 02/2014; 53(3):393-406. DOI:10.1016/j.molcel.2014.01.009 · 14.46 Impact Factor
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    ABSTRACT: Hepatitis C virus (HCV) nonstructural protein 5B (NS5B) has RNA-dependent RNA polymerase (RdRp) activity. Because NS5B recognizes various RNA motifs besides the HCV genome, NS5B has the potential of interacting with host RNA molecules. In this study, an RNA pool enriched with the 3′-UTR sequences was generated and mRNA molecules with high affinity binding to NS5B were selected by iterative selection. Among the high binding mRNA 3′-UTR segments, we analyzed the housekeeping ribosomal protein S4, X-linked [RPS4X] mRNA 3′-UTR and the 3′-UTR of galectin-1 (GAL-1) mRNA, which is known to be one of the genes upregulated in HCV-infected liver cells and to have a wide spectrum of biological properties. By means of IP-RT-PCR, it was demonstrated that both of the mRNA molecules bind to NS5B in the cytoplasm. Interestingly, GAL-1 and RPS4X mRNA can serve as templates for NS5B RdRp, suggesting these RNA molecules are regulated in vivo by NS5B.
    Virology 02/2014; s 450–451:13–23. DOI:10.1016/j.virol.2013.11.036 · 3.28 Impact Factor
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    ABSTRACT: Cardiovascular disease poses a major challenge for the 21st century, exacerbated by the pandemics of obesity, metabolic syndrome and type 2 diabetes. While best standards of care, including high-dose statins, can ameliorate the risk of vascular complications, patients remain at high risk of cardiovascular events. The Residual Risk Reduction Initiative (R3i) has previously highlighted atherogenic dyslipidaemia, defined as the imbalance between proatherogenic triglyceride-rich apolipoprotein B-containing-lipoproteins and antiatherogenic apolipoprotein A-I-lipoproteins (as in high-density lipoprotein, HDL), as an important modifiable contributor to lipid-related residual cardiovascular risk, especially in insulin-resistant conditions. As part of its mission to improve awareness and clinical management of atherogenic dyslipidaemia, the R3i has identified three key priorities for action: i) to improve recognition of atherogenic dyslipidaemia in patients at high cardiometabolic risk with or without diabetes; ii) to improve implementation and adherence to guideline-based therapies; and iii) to improve therapeutic strategies for managing atherogenic dyslipidaemia. The R3i believes that monitoring of non-HDL cholesterol provides a simple, practical tool for treatment decisions regarding the management of lipid-related residual cardiovascular risk. Addition of a fibrate, niacin (North and South America), omega-3 fatty acids or ezetimibe are all options for combination with a statin to further reduce non-HDL cholesterol, although lacking in hard evidence for cardiovascular outcome benefits. Several emerging treatments may offer promise. These include the next generation peroxisome proliferator-activated receptoralpha agonists, cholesteryl ester transfer protein inhibitors and monoclonal antibody therapy targeting proprotein convertase subtilisin/kexin type 9. However, long-term outcomes and safety data are clearly needed. In conclusion, the R3i believes that ongoing trials with these novel treatments may help to define the optimal management of atherogenic dyslipidaemia to reduce the clinical and socioeconomic burden of residual cardiovascular risk.
    Cardiovascular Diabetology 01/2014; 13(1):26. DOI:10.1186/1475-2840-13-26 · 3.71 Impact Factor
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    ABSTRACT: The computational structure-based drug design (SBDD) mainly aims at generating or discovering new chemical compounds with sufficiently large binding free energy. In any de novo drug design methods and virtual screening methods, drug candidates are selected by approximately evaluating the binding free energy (or the binding affinity). This approximate binding free energy, usually called "empirical score," is critical to the success of the SBDD. The purpose of this work is to yield physical insight into the approximate evaluation method in comparison with an exact molecular dynamics (MD) simulation-based method (named MP-CAFEE), which can predict binding free energies accurately. We calculate the binding free energies for 58 selected drug candidates with MP-CAFEE. Here, the compounds are generated by OPMF, a novel fragment-based de novo drug design method, and the ligand-protein interaction energy is used as an empirical score. The results show that the correlation between the binding free energy and the interaction energy is not strong enough to clearly distinguish compounds with nM-affinity from those with µM-affinity. This implies that it is necessary to take into account the natural protein motion with explicitly surrounded by water molecules to improve the efficiency of the drug candidate selection procedure.
    CHEMICAL & PHARMACEUTICAL BULLETIN 01/2014; 62(7):661-7. DOI:10.1248/cpb.c14-00132 · 1.38 Impact Factor
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    ABSTRACT: Autoimmune diseases often result from an imbalance between regulatory T (Treg) cells and interleukin-17 (IL-17)-producing T helper (TH17) cells; the origin of the latter cells remains largely unknown. Foxp3 is indispensable for the suppressive function of Treg cells, but the stability of Foxp3 has been under debate. Here we show that TH17 cells originating from Foxp3(+) T cells have a key role in the pathogenesis of autoimmune arthritis. Under arthritic conditions, CD25(lo)Foxp3(+)CD4(+) T cells lose Foxp3 expression (herein called exFoxp3 cells) and undergo transdifferentiation into TH17 cells. Fate mapping analysis showed that IL-17-expressing exFoxp3 T (exFoxp3 TH17) cells accumulated in inflamed joints. The conversion of Foxp3(+)CD4(+) T cells to TH17 cells was mediated by synovial fibroblast-derived IL-6. These exFoxp3 TH17 cells were more potent osteoclastogenic T cells than were naive CD4(+) T cell-derived TH17 cells. Notably, exFoxp3 TH17 cells were characterized by the expression of Sox4, chemokine (C-C motif) receptor 6 (CCR6), chemokine (C-C motif) ligand 20 (CCL20), IL-23 receptor (IL-23R) and receptor activator of NF-κB ligand (RANKL, also called TNFSF11). Adoptive transfer of autoreactive, antigen-experienced CD25(lo)Foxp3(+)CD4(+) T cells into mice followed by secondary immunization with collagen accelerated the onset and increased the severity of arthritis and was associated with the loss of Foxp3 expression in the majority of transferred T cells. We observed IL-17(+)Foxp3(+) T cells in the synovium of subjects with active rheumatoid arthritis (RA), which suggests that plastic Foxp3(+) T cells contribute to the pathogenesis of RA. These findings establish the pathological importance of Foxp3 instability in the generation of pathogenic TH17 cells in autoimmunity.
    Nature medicine 12/2013; DOI:10.1038/nm.3432 · 28.05 Impact Factor

Publication Stats

20k Citations
2,535.85 Total Impact Points


  • 1982–2015
    • The University of Tokyo
      • • Research Center for Advanced Science and Technology
      • • Division of Internal Medicine
      Tōkyō, Japan
  • 2005–2011
    • Beth Israel Deaconess Medical Center
      • • Division of Molecular and Vascular Medicine
      • • Center for Vascular Biology Research
      Boston, Massachusetts, United States
  • 2005–2010
    • Tokyo Medical and Dental University
      • Department of Cell Signaling
      Edo, Tōkyō, Japan
  • 2009
    • National Institute of Advanced Industrial Science and Technology
      Tsukuba, Ibaraki, Japan
    • Showa General Hospital
      Edo, Tōkyō, Japan
  • 2003–2007
    • Harvard University
      Cambridge, Massachusetts, United States
  • 2001–2007
    • Niigata University
      • Division of Cellular and Molecular Pathology
      Niahi-niigata, Niigata, Japan
  • 1992–2007
    • Kumamoto University
      • • Department of Cell Pathology
      • • Department of Medical Biochemistry
      • • Department of Metabolic Medicine
      Kumamoto, Kumamoto, Japan
  • 2006
    • Yokohama City University
      • Department of Gastroenterology
      Yokohama, Kanagawa, Japan
  • 2003–2005
    • University of Tsukuba
      • Institute of Clinical Medicine
      Tsukuba, Ibaraki-ken, Japan
  • 2004
    • Saitama University
      Saitama, Saitama, Japan
    • Kyorin University
      Edo, Tōkyō, Japan
  • 1996–2002
    • University of Oxford
      • Sir William Dunn School of Pathology
      Oxford, ENG, United Kingdom
  • 1999
    • The University of Tokushima
      Tokusima, Tokushima, Japan
  • 1998
    • Dartmouth College
      • Department of Biochemistry
      Hanover, New Hampshire, United States
    • Leiden University
      • Leiden Amsterdam Center for Drug Research
      Leiden, South Holland, Netherlands
  • 1995–1996
    • Jichi Medical University
      • Division of Anatomy
      Totigi, Tochigi, Japan
  • 1994
    • National Institute of Health and Nutrition
      Edo, Tōkyō, Japan