Welcome Address

Dean Prof. Yoshinori Moriyama
Faculty of Pharmaceutical Sciences,
Okayama University

 First of all, I would like to express my sincere thanks to the members of Sungkyunkwan University in South Korea who visited us at Okayama University.  When we visited School of Pharmacy, Sungkyunkwan University, in this September, I found an agreement form which was exchanged more than 10 years ago on display in a safe place in the dean’s room.  I was very pleased to learn that the faculty members of Sungkyunkwan University consider us as an important partner.  Now, we have opportunity to show off our research to each other at Okayama University.   I hope that all of participants enjoy this symposium, providing important progress for mutual understanding, collaboration and the development of both faculties

1 (OU-1)

Molecular imaging technology on the research of pharmaceutical sciences
Shuichi Enomoto1,2,3

1 Professor, Department of Pharmaceutical Analytical Chemistry, Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, Okayama University, Okayama, Japan,
2 Professor, Okayama Medical Innovation Center of Molecular Imaging, Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, Okayama University, Okayama, Japan,
3 Head, Next-generation Imaging Team, Center for Life Science Technologies, RIKEN Kobe Institute, Japan

Rapidly occurring discoveries and innovations in molecular biology, imaging and computing technology and driving a new scientific specialty referred to as "molecular imaging". Although mmolecular imaging is broadly defined as the characterization and measurement of biological processes in living animals, model systems and humans at the cellular and molecular level, using remote imaging detection methods, such as PET, SPECT, MRI and optical imaging and is helping us to better understand many of the basic molecular mechanisms of disease, the ability to use imaging to drive "personalized medicine" in patients is still a distant goal. Translational imaging research will facilitate the goal. Our mission is to: a) Translate advances in cellular and molecular biology, chemistry, physics, computer sciences, engineering, instrumentation and animal models into improvements in care for patients, b) Provide further advances in understanding of the molecular basis of disease and, c) Train student on undergraduate or graduate school and investigators in the use of modern molecular imaging techniques. The focus of our laboratory is the dynamics of bio-molecules on living organisms, as measured using various molecular imaging modalities, PET, SPECT, CT, MRI, Optical imaging technologies and our newly developed modality “Gamma-ray Emission Imaging (GREI)” which
development of RIKEN’s original semiconductor Compton camera GREI, and the world’s first demonstration of multiple molecular imaging. Recently we have successfully developed several new anti-body- and peptide-based probes for diagnostic imaging of caner, inflammation, Alzheimer disease and diabetes. And we also have successfully developed aminobenzopyrano-xanthene (ABPX) dyes that exhibits fluorescence emission in both dilute solution and the aggregates.

Our recent topics
1.         T. Miyamoto, S. Kamino, A. Odani, M. Hiromura, S. Enomoto, Basicity of N-terminal amine in TCUN peptide regulates stability constant of albumin-like Cu2+ complex. Chem. Lett., 42, 1099-1101 (2013).
2.         Y. Shirasaki, S. Kamino, M. Tanioka, K. Watanabe, Y. Takeuchi, S. Komeda, and S. Enomoto, New aminobenzopyranoxanthene-based colorimetric sensor for Cu2+ with dual-color signal detection system., Chem. Asian J., 8, 2609-2613 (2013).
3.         S. Motomura, Y. Kanayama, M. Hiromura, T. Fukuchi, T. Ida, H. Haba, Y. Watanabe, and S. Enomoto, Improved imaging performance of semiconductor Compton camera GREI makes for a new methodology to integrate bio-metal analysis and molecular imaging technology in living organisms., J. Anal. At. Spectrom. , 28, 934-939(2013).
4.         S. Kadowaki, M. Munekane, Y. Kitamura, M. Hiromura, S. Kamino, Y. Yoshikawa, H. Saji, S. Enomoto., Development of new zinc dithiosemicarbazone complex for use as oral antidiabetic agent , Biol. Trace Elem. Res., 154, 111-119 (2013).
5.         M. Taniguchi, A. Fukunaka, M. Hagihara, K. Watanabe, S. Kamino, T. Kambe, S. Enomoto, M. Hiromura, Essential Role of the Zinc Transporter ZIP9/SLC39A9 in Regulating the Activations of Akt and Erk in B-Cell Receptor Signaling Pathway in DT40 Cells, PLoSOne, vol. 8; 3 e58022. (2013).
6.         S. Kamino, A. Muranaka, M. Murakami, A. Tatsumi, N. Nagaoka, Y. Shirasaki, K. Watanabe, K. Yoshida, J. Horigome, S. Komeda, M. Uchiyama, S. Enomoto, A Red-Emissive aminobenzopyrano-xanthene Dye: Elucidation of Fluorescence Emission Mechanism in Solution and Aggregate State, Phys. Chem. Chem. Phys., 15, 2131-2140 (2013).
7.         S. Takeda, H. Odaka, S. Ishikawa, S. Watanabe, H. Aono, T. Takahashi, Y. Kanayama, M. Hiromura and S. Enomoto, Demonstration of in-vivo multi-probe tracker based on a Si/CdTe semiconductor Compton camera, IEEE Trans. Nucl. Sci., 59, 70-76 (2012).
8.         K. Higashikawa, N. Akada, K. Yagi, K. Watanabe, S. Kamino, Y. Kanayama, M. Hiromura and S. Enomoto, Exploration of target molecules for molecular imaging of inflammatory bowel disease, Biochem. Biophys. Res. Commun., 410, 416-421 (2011).
9.         H. Fujishiro, M. Doi, S. Enomoto and S. Himeno, High sensitivity of RBL-2H3 cells to cadmium and manganese: an implication of the role of ZIP8, Metallomics, 3, 710-718 (2011).
10.      A. Fukunaka, Y. Kurokawa, F. Teranishi, I. Sekler, K. Oda, M. L. Ackland, V. Faundez, M. Hiromura, S. Masuda, M. Nagao, S. Enomoto and T. Kambe, Tissue nonspecific alkaline phosphatase is activated via a two-step mechanism by zinc transport complexes in the early secretory pathway, J. Biol. Chem., 286, 16363-16373 (2011).
11.      T. Fukuchi, Y. Arai, F. Watanabe, S. Motomura, S. Takeda, Y. Kanayama, H. Haba, Y. Watanabe and S. Enomoto, A digital signal processing module for Ge semiconductor detectors, IEEE Trans. Nucl. Sci., 58, 461-467 (2011).
12.      M. Kidera, Y. Seto, K. Takahashi, S. Enomoto, S. Kishi, M. Makita, T. Nagamatsu, T. Tanaka and M. Toda, New method for comprehensive detection of chemical warfare agents using an electron-cyclotron-resonance ion-source mass spectrometer, Spectroc. Acta Pt. A-Molec. Biomolec. Spectr., 78, 1215-1219 (2011).
13.      H. Fujishiro, K. Kubota, D. Inoue, A. Inoue, T. Yanagiya, S. Enomoto and S. Himeno, Cross-resistance of cadmium-resistant cells to manganese is associated with reduced accumulation of both cadmium and manganese, Toxicology, 280, 118-125 (2011).

, Ph.D. Shuichi Enomoto

 (Email: senomoto@ pharm.okayama-u.ac.jp)

Research Interests

Non-natural amino acids, Epigenetic drug discovery, Molecular design, Fragment-based drug discovery, In-silico screening for drug candidates, ADMET (absorption, distribution, metabolism, excretion and toxicity, Single cell analysis, Intracellular network, Discrimination between normal cells and abnormal cells, Data analysis, Data base integration, Nanofabrication and nanodevice, Labeling chemistry, Analysis of metabolite dynamics, Clinical test/clinical trials, Comparative study of the normal and disease-model animals, Comparative study of the human and nonhuman-primates, Normal stem cells vs cancer stem cells, Central Nervous System Disorder, Regenerative medicine, Microdosing and exploratory clinical study, Pharmacokinetics, Efficacy evaluation, Biomarkers of pathophysiology, Molecular imaging, Multiple molecular imaging, Development of quantitative diagnosis of presbyopia, PET, SPECT, MRI, Optical imaging, Metallomics.

2 (SKKU-1)

Recent Progress in CH Activation and CC Bond Formation
In Su Kim, Associate Professor
School of Pharmacy, Sungkyunkwan University, Suwon 440-746, Republic of Korea

CH bond functionalization has been a longstanding goal in organic synthesis since it obviates the prefunctionalization of substrates. The combination of transition metal and directing group is a useful strategy to facilitate CH bond cleavage, which affords valuable transformations of CH bond to CC, CX, CO and CN bonds.
Since the pioneering discovery of CH bond activation by Murai, recent progress has been focused on dehydrogenative cross-coupling between sp2 or sp3 CH bonds and sp2 CH bonds of arenes or alkenes. In this area of research, various directing groups such as ketones, carboxylic acids, amides, pyrrole/pyridine, anilides, carbamate, urea and azine-N-oxide can provide an anchor for catalytic ortho-metalation of aromatic rings. Although the reactions using arenes or alkenes as coupling partners have been well documented, the reactions between the aromatic CH bonds and aldehydes or α-keto acids remain relatively unexplored. From the synthetic point of view, the direct and catalytic introduction of carbonyl functional group into aromatic compounds via CH bond cleavage is among the greatest challenges in synthetic chemistry. This new method is complementary to Friedel-Crafts acylation and directed lithiation/acylation process.
Recently, our groups reported the rhodium or palladium-catalyzed oxidative acylation of benzamides, N-benzyltriflamides, oximes, phenoxypyrdines and hydrazones with aldehydes or alcohols to afford the corresponding aryl ketones or 3-hydroxyisoindolin-1-ones, which are important structural units and synthetic intermediates in pharmaceuticals, natural products, and functional materials. Also, we found the palladium-catalyzed decarboxylative acylation of phenylacetamides, O-phenylcarbamtes and oximes using α-keto acids in the presence of ammonium persulfate as a convenient oxidant affording aryl ketones in moderate to good yields.
In this presentation, we describe our recent achievements about the development of transition metal-catalyzed new C-C bond formation reactions including oxidative acylation, decarboxylative acylation, tandem indole synthesis, C-2 allylation of indoles, and etc.

In Su Kim
, Ph.D.
Email: insukim@skku.edu)
Associate Professor  (School of Pharmacy)

Research Interests 

C−H Activation, C−C Bond Formation, oxidative acylation,
decarboxylative acylation, catalytic ortho-metalation

3 (OU-2)

Toward molecular understanding of sensory systems by structural biology Atsuko Yamashita, Professor
Structural Biology

Sense, such as vision, taste sensation, olfaction, audition, and touch sensation, is an important function for living organisms to receive, transduce, integrate, recognize and process the environmental information. For the first step of a molecular-level approach to the question of how we recognize environments, our lab are addressing the structure biology of sensory receptor proteins at the front line toward the environment, which recognize chemical or mechanical stimuli and respond to them. We are currently performing the structural and functional analyses of taste receptors, as representatives of chemoreceptors, and TRP channels, as representatives of mechanoreceptors.
The pivot of our study is structural analyses of the proteins by X-ray crystallography. However, our target proteins are eukaryotic membrane proteins, which are ones of the most difficult targets in the field of structural biology. In order to achieve structural analyses of these targets, we simultaneously carry out research and development of methods to facilitate sample preparation, a bottleneck for membrane protein crystallography. Our main strategies for this is utilizing the GFP-fusion technologies, recently applied in the field of structural biology for sample evaluation and screening.
[Results and Discussion]
Sample preparation of our target proteins, taste receptors and TRP channels, has so far been difficult even for the partial regions of the receptors, because of the poor expression level and protein unstability. By making use of the GFP-fusion strategies, we have succeeded the protein preparation of functional regions of these receptors, and structural and functional analyses of them are currently in progress. At this joint symposium, I will present our recent achievement about the analyses of the regulatory region of a fungus TRP channel (Ihara et al. J. Biol. Chem., 288, 15303, 2013), providing clues to understand the mechanism of multimodal responses of TRP channels

Atsuko Yamashita, Ph.D. (Email: a_yama@ pharm.okayama-u.ac.jp)
Professor (Structural Biology)

Research Interests

Membrane Proteins, Sensory Receptors, Transporters, Ion Channels, Taste, Mechanosensation

4 (SKKU-2)

Structure-based Virtual Screening of Chemical Database to Discover Novel Ligands for Various Biomacromolecules
Hyun-Ju Park, Professor
School of Pharmacy, Sungkyunkwan University

Virtual screening is generally recognized as a valuable tool to reduce the size of a chemical library containing a huge number of compounds to a target-focused compound library. To overcome the major bottlenecks in this area is to find suitable method for database filtering and scoring methods for docking. Using various available computational programs, we setup a reliable strategy for virtual screening and applied it to the identification of small molecule hit ligands for the targets of interest.
Promising results obtained from our study will be introduced. We identified ligands for
histone deacetylase, one of epigenetic regulators; RNA secondary structures involved in -1 ribosomal frameshifting of many viruses; and G-quadruplex DNA structure in the c-Myc oncogenic promoter.

Hyun-Ju Park, Ph.D. (Email: hyunju85@ skku.edu)
School of Pharmacy, Sungkyunkwan University

Research Interests 

structure-based virtual screening, high-content fluorescence-based screening, QSAR/QSPR modeling

5 (OU-3)

Network dysfunction by Amyloid β oligomers
Tsuyoshi Inoue, Associate Professor
Biophysical Chemistry

Alzheimer disease is a common form of dementia, characterized by memory impairment. Amyloid β (Aβ), a peptide of 42 amino acids, is the cause of Alzheimer disease. Although Aβ peptides are highly aggregated in the brain of Alzheimer disease, recent studies have shown that a low aggregated form of Aβ, called Aβ oligomers, is toxic to the brain. Actually, direct injection of Aβ oligomers into the mouse brain induces memory impairment (Cleary et al, Nat Neurosci, 2005), and impairs long-term potentiation in the hippocampus, a synaptic process of memory (Walsh et al, Nature, 2002). However, it is still unknown how neural activities are impaired by Aβ oligomers.

To address this question, we used electrophysiological techniques in awake mice. Under anesthesia, a recording electrode was implanted into the hippocampus, and an injection cannula was implanted into the lateral ventricle. Several days later, neural activities in the hippocampus were recorded in an awake condition, Aβ oligomers were injected into the lateral ventricle, and changes in the hippocampal activities by Aβ oligomers were observed. To examine learning ability in mice, we used a novel object recognition test as a behavioral test.
[Results and Discussion]
Theta oscillations (5-8 Hz) are characteristic electrical activities in the hippocampus, which are important for memory acquisition. We found that the theta oscillations were reduced by direct injection of Aβ oligomers. We further found a peptide, which could recover the reduced theta oscillations by Aβ oligomers. Interestingly, learning ability in mice was impaired by Aβ oligomers, and the memory impairment was also recovered by the peptide. Our results indicate that Aβ oligomers reduce theta oscillations in the hippocampus, and also that the network dysfunction and the memory impairment by Aβ oligomers can be both recovered by the peptide we found.

Tsuyoshi Inoue, Ph.D. (Email: tinoue@pharm.okayama-u.ac.jp)
Associate Professor (Biophysical Chemistry)

Research Interests

Electrophysiology, Neuroscience, Alzheimer disease

6 (SKKU-3)

Host-pathogen interaction during host invasion of pneumococcus
Dong-Kwon Rhee, Professor
Molecular Microbiology Lab

Streptococcus pneumoniae (pneumococcus) is the major cause of community-associated pneumonia, otitis media, septicemia, and meningitis. Pneumonia has one of the highest morbidity and mortality rates from infections (≥ 2 million deaths every year), and is the sixth-leading cause of death in the United States. It is also responsible for a large number of deaths following influenza epidemics. Pneumococci produce a thick polysaccharide capsule that shields pneumococci from host phagocytes, however this capsule is removed during invasion into the host cells. Moreover, the host cells respond rapidly to the invading pathogens to avoid or neutralize invasion of the pneumococcus. Thus understanding of pathogen-host interaction could p
rovide information on chemotherapeutic measures useful for prevention and treatment of pneumococcal diseases.
Our lab found that 1)
Pep27, an autolysis-inducing factor of S. pneumoniae, was thus expected to effect cytotoxicity. The loss of Pep27 had a much larger than expected decrease in toxicity and has made the pep27 mutant strain sufficiently non-toxic to be used as a live vaccine. 2) ClpL, a major heat shock protein (HSP) in pneumococci, generates antibiotic resistance; Penicillin resistance in S. pneumoniae was induced by ClpL. A mutant lacking ClpL was more susceptible to penicillin and had a thinner cell wall than the parental type, whereas a ClpL overexpressing strain shows a higher resistance to penicillin and a thicker cell wall in the wild type. Heat shock induced a ClpL-dependent increase in the mRNA levels and protein synthesized by the major cell wall synthesis gene pbp2x. Fractionation and electron micrograph data revealed that ClpL induced by heat shock is localized at the cell wall, and the DclpL showed significantly reduced net translocation of PBP2x into the cell wall. 3) ClpL inhibits pneumococcal adherence to A549 cells through activation of the small Rho-GTPases via Rap1 up-regulation. ClpL-induced Rap1 activates Rac1, which in turn phosphorylates cofilin and inactivates actin cytoskeleton rearrangement. These findings may shed light on how the HSP100 family member ClpL can modulate virulence at the early stages of infection. Taken together, genes induced by pneumococcal invasion or by the host cells could be valuable biomarkers for diagnosis, preventive, and therapeutic measures to overcome pneumococcal diseases.

Dong-Kwon Rhee, Ph.D. (Email: dkrhee@skku.edu)
Professor (Molecular Microbiology Lab, School of Pharmacy)

Research Interests 

Pneumococcal pathogenesis, Host-cell invasion, Signaling, Host-pathogen interaction, Stress response

7 (OU-4)

A novel antimicrobial agent from Nuphar japonicum against Methicillin resistant Staphylococcus aureus
Teruo KURODA, Associate Professor
Department of Environmental and Applied Microbiology

The emergence of drug resistant bacteria is a growing clinical problem that can cause therapeutic failures throughout the world. In particular, methicillin-resistant Staphylococcus aureus (MRSA) has recently become one of the most important pathogenic bacteria. To develop anti-MRSA drugs, we have been screening active compounds which have anti-MRSA activity, and investigate their action mechanisms.
The active compound named “compound K” was isolated from methanol extract of Nuphar rhizome (the rhizome of Nuphar japonicum) with liquid-liquid separation and silica gel column chromatography. Minimum inhibitory concentration of compound K was determined microdilution method according to the recommendations of the Japanese Society of Chemotherapy.
[Results & Discussion]
Compound K showed potent growth inhibition of S. aureus. MICs of compound K against several MRSA and methicillin-sensitive S. aureus (MSSA) were 1-4 mg/mL. In addition, compound K showed antimicrobial activity for vancomycin intermediate-resistant S. aureus (VISA). It indicated that compound K was effective against S. aureus irrespective of sensitivity to oxacillin and vancomycin. When compound K was added at one-fourth concentration of MIC, MICs of gentamicin and/or arbekacin were significantly decreased. This synergy effect was observed for several clinical isolated MRSA. Compound K also showed inhibition for topoisomerase IV of S. aureus which have essential role in DNA replication. However, compound K did not exhibit cross-resistance to norfloxacin-resistant S. aureus, which indicated that the action site in topoisomerase IV was different from that of quinolone resistance-determining regions (QRDR) and that it inhibited other targets besides topoisomerase IV.

Teruo KURODA, Ph.D. (Email: tkuroda@cc.okayama-u.ac.jp)
Associate Professor (Department of Environmental and Applied Microbiology)

Research Interests 

MRSA, antimicrobial agents, antiseptic agents, multidrug efflux pump, Pseudomonas aeruginosa

8 (SKKU-4)

Organic-inorganic hybrid nanoparticles for siRNA delivery
Ji Hoon Jeong, Associate Professor
School of Pharmacy, Sungkyunkwan University, Republic of Korea

Gene therapy based on small interference RNA (siRNA) holds enormous potential for therapeutic intervention of a broad range of genetic diseases, including infectious diseases, gene-related disorders, and cancer. A number of cationic carriers forming nanoparticulates mostly via electrostatic interactions with oppositely charged nucleic acid have been developed for siRNA delivery. However, due to its rigid double-stranded structure, siRNA has often failed to generate nanoparticulates as tight as those formed with plasmid DNA with conventional cationic carriers. Therefore, improving transfection efficiency siRNA is prerequisite for its successful use in clinical settings. Inorganic materials such as hydroxyapatite and gold have a wide range of applications in biomedical systems. In this study, we demonstrated bio-organic templated inorganic nanoparticles for siRNA delivery. The new modalities of the organic-inorganic hybrid nanoparticles would give a chance to cope with the siRNA encapsulation problem and enable concurrent cellular siRNA delivery and imaging. The possibilities of using the nanoparticles for therapeutic and imaging purposes were assessed.

Ji Hoon Jeong, Ph.D. (Email: jhjeong@skku.edu)
Associate Professor (School of Pharmacy)

Research Interests 

gene and drug delivery, drug dissolution and stabilization, microneedles, stem cell engineering

9 (OU-5)

Mast Cell: A critical regulator of inflammation
Satoshi Tanaka
Professor, Immunobiology

  Mast cells are found in nearly all the vascularized tissues, although the physiological roles of mast cells remain to be fully clarified. Accumulating evidence indicates that mast cells play critical roles in inflammatory responses, such as immediate allergy, whereas mast cells have recently been found to be involved in suppression of immune responses, such as immune tolerance. Mast cells originate in the hematopoietic stem cells in the bone marrow and undergo terminal differentiation in the tissues, in which they are ultimately resident. The characteristics of tissue mast cells are, therefore, profoundly affected by their microenvironmental factors, including the neighboring cells, extracellular matrix, and local cytokine/growth factor levels. Although recent studies demonstrated using the genetically-mast cell-deficient models that mast cells should be required for a variety of physiological and pathological responses, it remains unknown how tissue mast cells regulate these responses.
[Results and Discussion]
  We established a culture model of murine cutaneous mast cells, in which IL-3-dependent bone marrow-derived cultured mast cells (BMMCs) are co-cultured with Swiss 3T3 fibroblasts in the presence of stem cell factor (Takano et al., FEBS Lett., 2008). This model reflects the characteristics of cutaneous mast cells, such as sensitivity to substance P and increased granule storage of histamine, heparin and neutral proteases. We extracted genes, of which expression levels were drastically changed during the co-culture period. We focused on the up-regulation of CD44, one of the primary receptor for hyaluronan, because hyaluronan is abundant in the cutaneous tissues and might be associated with cutaneous mast cells. We demonstrated that CD44 regulates the process of proliferation of mast cells in the cutaneous tissues (Takano et al., Lab. Invest., 2009). We previously found the impaired granule maturation in the mature peritoneal mast cells obtained from the gene-targeted mice that lacks the ability of histamine synthesis. By using our cutaneous mast cell model, we revealed that histamine synthesis should be required for granule maturation of mast cells, of which process is largely independent of specific histamine receptors (Nakazawa et al., Eur. J. Immunol., 2013). Our findings will contribute to development of novel therapeutic compounds that prevent proliferation and maturation of tissue mast cells for chronic inflammatory diseases.

Satoshi Tanaka, Ph.D. (Email: tanaka@ pharm.okayama-u.ac.jp)
Professor, Department of Immunobiology

Research Interests

Mast Cell, Inflammation, Allergy, Histamine, Microenvironment