Takatsugu Kasahara, Tsung-Che Chang, Hiromasa Yoshioka, Sayaka Urano, Yasuko Egawa, Michiko Inoue, Tsuyoshi Tahara, Koji Morimoto, R Ambara Pradipta and Katsunori Tanaka : Anticancer approach by targeted activation of a global inhibitor of sialyltransferases with acrolein., Chemical Science, Vol.15, No.25, 9566-9573, 2024.
(要約)
-Neu5Ac (PFN) in cancer cells using the reaction between phenyl azide and endogenous acrolein, which is overproduced in most cancer cells. The prodrug significantly suppressed tumor growth in mice as effectively as PFN without causing kidney dysfunction, which is associated with PFN. The use of sialylated glycans as immune checkpoints is gaining increasing attention, and the proposed method for precisely targeting aberrant sialylation provides a novel avenue for expanding current cancer treatments.
Takashi Tamura, Tsuyoshi Tahara, Michiko Inoue, Ryota Nanjo, Hirotaka Onoe, Takako Yamamoto and Shin Kawamata : Study on the Extrapolability of Current Tumorgenicity Test With Mice by Comparing the Syngeneic or Allogeneic Mouse Transplantation Model., Stem Cells Translational Medicine, 2024.
(要約)
The extrapolability of the current tumorigenicity test performed by transplanting human cell product into immunodeficient (NOG) mice was investigated. For this purpose, the susceptibility to form teratomas of NOG mice was assessed by transplanting undifferentiated human-induced pluripotent stem cells (hiPSCs) as positive control cells via the liver, striatum, or tail vein and evaluating the TPD50 value (dose required to form teratomas in half of the transplanted mice). This was then compared to the TPD50 of syngeneic or allogeneic mouse models. The TPD50 of C57/BL/6(B6)-iPSC or 129/Ola(129)-embryonic stem cell (ESC) transplanted into the liver of syngeneic mice was 4.08×105 and 4.64×104 cells, respectively, while the TPD50 of hiPSC administered into the liver of NOG mice was 4.64×104 cells. The TPD50 of B6-miPSC-synergic, 129-mESC-synergic, or 129-cell/B6 allogeneic transplantation into the striatum was 5.09×102, 1.0×104, and 3.73×104 cells, respectively, while that of hiPSC/NOG mice was 1.0×103 cells. The TPD50 for B6-miPSC or 129-mESC syngeneic tail vein infusion was 3.16×106 or 5.62×106 cells, respectively, while no incidence was observed from 1×107 B6-miPSCs in 129 mice or hiPSCs in NOG mice infusion study. Although the number of data sets was limited, these data indicate that the teratoma formation from transplanted undifferentiated hiPSCs via the liver or striatum in NOG mice is comparable to that in syngeneic or allogeneic mouse transplantation model, suggesting that the result of the current tumorigenicity test in NOG mice would provide useful information to infer the incidence of teratoma from residual undifferentiated hPSCs in hPSC-derived products after transplantation.
Danxi Li, Di Hu, Yuta Ochi, Wakako Arakaki, Aya Mawatari, Mika Shigeta, Yuping Wu, Emi Hayashinaka, Hiroyuki Neyama, Tsuyoshi Tahara, Yasuhiro Wada, Feng Li, Hisashi Doi, Yasuyoshi Watanabe and Yilong Cui : Regional neuroinflammation induced by peripheral infection contributes to fatigue-like symptoms: a [18F]DPA-714 positron emission tomography study in rats, Frontiers in Immunology, Vol.14, 1261256, 2023.
(要約)
A series of symptoms, including fever, widespread pain, fatigue, and even ageusia, have frequently been reported in the context of various infections, such as COVID-19. Although the pathogenic mechanisms underlying an infection causing fever and pain have been well established, the mechanisms of fatigue induced by infection in specific brain regions remain unclear. To elucidate whether and how the peripheral infection cause fatigue via regional neuroinflammation, we performed a brain-wide investigation of neuroinflammation in a peripheral pseudoinfection rat model using [F]DPA-714 positron emission tomography (PET) imaging analysis, in which the polyriboinosinic: polyribocytidylic acid (poly I:C) was intraperitoneally injected. Transient fever lasting for several hours and subsequent suppression of spontaneous activity lasting a few days were induced by poly I:C treatment. Significant increase in plasma interleukin (IL)-1β, IL-6 and tumour necrosis factor (TNF)-α were observed at 2 and 4 h following poly I:C treatment. PET imaging analysis revealed that the brain uptake of [F]DPA-714 was significantly increased in several brain regions one day after poly I:C treatment, such as the dorsal raphe (DR), parvicellular part of red nucleus (RPC), A5 and A7 noradrenergic nucleus, compared with the control group. The accumulation of [F]DPA-714 in the DR, RPC and A5 was positively correlated with subsequent fatigue-like behavior, and that in the A7 tended to positively correlate with fever. These findings suggest that peripheral infection may trigger regional neuroinflammation, which may cause specific symptoms such as fatigue. A similar mechanism might be involved in COVID-19.
Takashi Niwa, Tsuyoshi Tahara, CE Chase, RG Fang, Takayoshi Nakaoka, Satsuki Irie, Emi Hayashinaka, Yasuhiro Wada, Hidefumi Mukai, Kenkichi Masutomi, Yasuyoshi Watanabe, Yilong Cui and Takamitsu Hosoya : Synthesis of 11C-Radiolabeled Eribulin as a Companion Diagnostics PET Tracer for Brain Glioblastoma, Bulletin of the Chemical Society of Japan, Vol.96, No.3, 283-290, 2023.
(要約)
<jats:title>Abstract</jats:title> <jats:p>The successful 11C-radiolabeling of eribulin, an analog of the marine natural product halichondrin B, and an approved anticancer drug for the treatment of breast cancer and liposarcoma, is reported. A rapid sequence involving a nitroaldol reaction with [11C]nitromethane and subsequent reduction of the nitro group enabled the introduction of a carbon-11 atom at the C35-position of eribulin. Optimization of the reaction and purification conditions led to a reproducible synthetic method for [35-11C]eribulin with 248 ± 104 MBq of radioactivity, 88.2 ± 5.8% radiochemical purity, and 132 ± 32 MBq/nmol molar activity. The total synthetic time was 38.0 ± 1.3 min (n = 12). PET imaging using mice bearing brain tumors revealed a specific accumulation of [35-11C]eribulin in tumors without any significant metabolic changes. These results indicate the applicability of [35-11C]eribulin for the quantitative measurement of eribulin migration into tumor tissue, which would be beneficial for exploring the application of eribulin for glioblastoma treatment and estimating the appropriate dosage for each patient.</jats:p>
Tsuyoshi Tahara, Shuhei Takatani, Mieko Tsuji, Nina Shibata, Nami Hosaka, Michiko Inoue, Masahiro Ohno, Daiki Ozaki, Aya Mawatari, Yasuyoshi Watanabe, Hisashi Doi and Hirotaka Onoe : Characteristic Evaluation of a 11C-Labeled Leucine Analog, l-α-[5-11C]methylleucine, as a Tracer for Brain Tumor Imaging by Positron Emission Tomography, Molecular Pharmaceutics, Vol.20, No.3, 1842-1849, 2023.
(要約)
Amino acid transporters are upregulated in many cancer cells, and system L amino acid transporters (LAT1-4), in particular, LAT1, which preferentially transports large, neutral, and branched side-chain amino acids, are considered a primary target for cancer positron emission tomography (PET) tracer development. Recently, we developed a C-labeled leucine analog, l-α-[5-C]methylleucine ([5-C]MeLeu), via a continuous two-step reaction of Pd-mediated C-methylation and microfluidic hydrogenation. In this study, we evaluated the characteristics of [5-C]MeLeu and also compared the sensitivity to brain tumors and inflammation with l-[C]methionine ([C]Met) to determine its potential for brain tumor imaging. Competitive inhibition experiments, protein incorporation, and cytotoxicity experiments of [5-C]MeLeu were performed in vitro. Further, metabolic analyses of [5-C]MeLeu were performed using a thin-layer chromatogram. The accumulation of [5-C]MeLeu in tumor and inflamed regions of the brain was compared with [C]Met and C-labeled ()-ketoprofen methyl ester by PET imaging, respectively. Transporter assay with various inhibitors revealed that [5-C]MeLeu is mainly transported via system L amino acid transporters, especially LAT1, into A431 cells. The protein incorporation assay and metabolic assay in vivo demonstrated that [5-C]MeLeu was neither used for protein synthesis nor metabolized. These results indicate that MeLeu is very stable in vivo. Furthermore, the treatment of A431 cells with various concentrations of MeLeu did not change their viability, even at high concentrations (∼10 mM). In brain tumors, the tumor-to-normal ratio of [5-C]MeLeu was more elevated than that of [C]Met. However, the accumulation levels of [5-C]MeLeu were lower than those of [C]Met (the standardized uptake value (SUV) of [5-C]MeLeu and [C]Met was 0.48 ± 0.08 and 0.63 ± 0.06, respectively). In brain inflammation, no significant accumulation of [5-C]MeLeu was observed at the inflamed brain area. These data suggested that [5-C]MeLeu was identified as a stable and safe agent for PET tracers and could help detect brain tumors, which overexpress the LAT1 transporter.
Miho Shukuri, Aya Mawatari, Shuhei Takatani, Tsuyoshi Tahara, Michiko Inoue, Wakako Arakaki, Masahiro Ohno, Hisashi Doi and Hirotaka Onoe : Synthesis and preclinical evaluation of 18F-FKTP methyl ester as a radiotracer for COX-1 imaging in neuroinflammation, Journal of Nuclear Medicine, Vol.63, No.11, 1761-1767, 2022.
(要約)
Cyclooxygenase (COX) is a rate-limiting enzyme in the synthesis of proinflammatory prostanoids from arachidonic acid. In vivo imaging of COX by PET is a potentially powerful tool for assessing the inflammatory response to injury, infection, and disease. We previously reported on a promising PET probe for COX imaging, C-labeled ketoprofen methyl ester, which can detect COX-1 activation in models of neuroinflammation and neurodegenerative disorders. In the current study, we aimed to design a fluorine-substituted benzoyl group of ketoprofen (FKTP) and to evaluate its racemate and enantiomers (F-labeled ketoprofen methyl ester, [F]FKTP-Me) as PET proradiotracers, potential radiopharmaceuticals for in vivo PET study of COX-1. We performed nucleophilic aromatic F-fluorination to obtain the desired racemic radiolabeled probe, ()-[F]FKTP-Me, at a radiochemical yield of 11%-13%. Subsequent high-performance liquid chromatography separation with a chiral column yielded the desired enantiomerically pure ()- and ()-[F]FKTP-Me. We examined the in vivo properties of ()-, ()-, and ()-[F]FKTP-Me in PET studies using rats in which hemispheric inflammation was induced by intrastriatally injecting a lipopolysaccharide. Racemic ()-[F]FKTP-Me and enantiomeric ()- or ()-[F]FKTP-Me were synthesized with radiochemical and chemical purities of more than 99%. The metabolite analysis revealed that the racemic ()-[F]FKTP-Me crossed the blood-brain barrier and entered the brain, where it was subsequently hydrolyzed to its pharmacologically active acid form. PET images revealed a high accumulation of ()-, ()-, and ()-[F]FKTP in the inflamed regions in rat brain. Moreover, the accumulated radioactivity of ()-[F]FKTP-Me was higher than that of ()-[F]FKTP-Me and ()-[F]FKTP-Me, which was correlated with the stereospecific inhibitory activity of FKTP against COX-1. From the results of this study, we conclude that racemic ()-[F]FKTP-Me and its enantiomers could act as proradiotracers of neuroinflammation in rat brain by the association of their hydrolyzed acid forms with COX-1 in inflamed regions. In particular, ()-[F]FKTP-Me demonstrated suitable properties as a COX-1-specific probe in PET imaging of neuroinflammation.
Shuhei Takatani, Tsuyoshi Tahara, Mieko Tsuji, Daiki Ozaki, Nina Shibata, Yoshinobu Hashizume, Masaaki Suzuki, Hirotaka Onoe, Yasuyoshi Watanabe and Hisashi Doi : C-Methylation and Microfluidic Hydrogenation: Potentiality of Leucine PET Probes for Tumor Imaging., ChemMedChem, 2021.
(要約)
I, radiochemical purity of 95-99 %, and chemical purity of 95-99 %. The Pd impurities in the injectable solution measured using inductively coupled plasma mass spectrometry met the international criteria for human use. Positron emission tomography scanning after an intravenous injection of L-[5-
Kenward Vong, Tsuyoshi Tahara, Sayaka Urano, Igor Nasibullin, Kazuki Tsubokura, Yoichi Nakao, Almira Kurbangalieva, Hirotaka Onoe, Yasuyoshi Watanabe and Katsunori Tanaka : Disrupting tumor onset and growth via selective cell tagging (SeCT) therapy., Science Advances, Vol.7, No.17, 2021.
(要約)
This study presents the early framework of selective cell tagging (SeCT) therapy, which is the concept of preferentially labeling specific cells in vivo with chemical moieties that can elicit a therapeutic response. Using glycosylated artificial metalloenzyme (GArM)-based protein labeling, this study reports two separate functional strategies. In one approach, early tumor onset can be suppressed by tagging cancer cells in living mice with an integrin-blocking cyclic-Arg-Gly-Asp (cRGD) moiety, thereby disrupting cell adhesion onto the extracellular matrix. In another approach, tumor growth in mice can be reduced by tagging with a cytotoxic doxorubicin moiety. Subsequent cell death occurs following internalization and drug release. Overall, experiments have shown that mouse populations receiving the mixture of SeCT labeling reagents exhibited a significant delay/reduction in tumor onset and growth compared with controls. Highlighting its adaptability, this work represents a foundational step for further development of SeCT therapy and its potential therapeutic applications.
Ivan Smirnov, Regina Sibgatullina, Sayaka Urano, Tsuyoshi Tahara, Peni Ahmadi, Yasuyoshi Watanabe, R Ambara Pradipta, Almira Kurbangalieva and Katsunori Tanaka : A Strategy for Tumor Targeting by Higher-Order Glycan Pattern Recognition: Synthesis and In Vitro and In Vivo Properties of Glycoalbumins Conjugated with Four Different N-Glycan Molecules., Small, Vol.16, No.46, 2020.
(要約)
Natural glycoconjugates that form glycocalyx play important roles in various biological processes based on cell surface recognition through pattern recognition mechanisms. This work represents a new synthesis-based screening strategy to efficiently target the cancer cells by higher-order glycan pattern recognition in both cells and intact animals (mice). The use of the very fast, selective, and effective RIKEN click reaction (6π-azaelectrocyclization of unsaturated imines) allows to synthesize and screen various structurally well-defined glycoalbumins containing two and eventually four different N-glycan structures in a very short time. The importance of glycan pattern recognition is exemplified in both cell- and mouse-based experiments. The use of pattern recognition mechanisms for cell targeting represents a novel and promising strategy for the development of diagnostic, prophylactic, and therapeutic agents for various diseases including cancers.
Akihiro Ogura, Sayaka Urano, Tsuyoshi Tahara, Satoshi Nozaki, Regina Sibgatullina, Kenward Vong, Takehiro Suzuki, Naoshi Dohmae, Almira Kurbangalieva, Yasuyoshi Watanabe and Katsunori Tanaka : A viable strategy for screening the effects of glycan heterogeneity on target organ adhesion and biodistribution in live mice., Chemical Communications, Vol.54, No.63, 8693-8696, 2018.
(要約)
This work represents the first broad study of testing diverse heterogenous glycoconjugates (7 different glycoalbumins) for their differential in vivo binding (11 different cancer cell types) in both cell- and animal-based studies. As a result, various changes in biodistribution, excretion, and even tumor adhesion were observed.
(キーワード)
Animals / Cell Adhesion Molecules / Cell Line, Tumor / Glycoconjugates / Humans / Mice / Organ Specificity / Serum Albumin / Tissue Distribution
Kazuki Tsubokura, H Kenward K Vong, R Ambara Pradipta, Akihiro Ogura, Sayaka Urano, Tsuyoshi Tahara, Satoshi Nozaki, Hirotaka Onoe, Yoichi Nakao, Regina Sibgatullina, Almira Kurbangalieva, Yasuyoshi Watanabe and Katsunori Tanaka : In Vivo Gold Complex Catalysis within Live Mice., Angewandte Chemie International Edition, Vol.56, No.13, 3579-3584, 2017.
(要約)
complex was designed and developed that enables organ-specific, localized propargyl ester amidation with nearby proteins within live mice. The targeted reactivity can be imaged through the use of Cy7.5- and TAMRA-linked propargyl ester based fluorescent probes. This targeting system could enable the exploitation of other metal catalysis strategies for biomedical and clinical applications.
(キーワード)
Animals / Catalysis / Coordination Complexes / Fluorescent Dyes / Gold / Mice / Mice, Inbred BALB C / Mice, Nude / Optical Imaging / Serum Albumin / Tissue Distribution
Liliya Latypova, Regina Sibgatullina, Akihiro Ogura, Katsumasa Fujiki, Alsu Khabibrakhmanova, Tsuyoshi Tahara, Satoshi Nozaki, Sayaka Urano, Kazuki Tsubokura, Hirotaka Onoe, Yasuyoshi Watanabe, Almira Kurbangalieva and Katsunori Tanaka : Sequential Double "Clicks" toward Structurally Well-Defined Heterogeneous N-Glycoclusters: The Importance of Cluster Heterogeneity on Pattern Recognition In Vivo., Advanced Science, Vol.4, No.2, 2016.
(要約)
Structurally well-defined heterogeneous N-glycoclusters are prepared on albumin via a double click procedure. The number of glycan molecules present, in addition to the spatial arrangement of glycans in the heterogeneous glycoclusters, plays an important role in the in vivo kinetics and organ-selective accumulation through glycan pattern recognition mechanisms.
Ayumi Tsutsui, Akihiro Ogura, Tsuyoshi Tahara, Satoshi Nozaki, Sayaka Urano, Mitsuko Hara, Soichi Kojima, Almira Kurbangalieva, Hirotaka Onoe, Yasuyoshi Watanabe, Naoyuki Taniguchi and Katsunori Tanaka : In vivo imaging of advanced glycation end products (AGEs) of albumin: first observations of significantly reduced clearance and liver deposition properties in mice., Organic & Biomolecular Chemistry, Vol.14, No.24, 5755-5760, 2016.
(要約)
Advanced glycation end products (AGEs) are associated with various diseases, especially during aging and the development of diabetes and uremia. To better understand these biological processes, investigation of the in vivo kinetics of AGEs, i.e., analysis of trafficking and clearance properties, was carried out by molecular imaging. Following the preparation of Cy7.5-labeled AGE-albumin and intravenous injection in BALB/cA-nu/nu mice, noninvasive fluorescence kinetics analysis was performed. In vivo imaging and fluorescence microscopy analysis revealed that non-enzymatic AGEs were smoothly captured by scavenger cells in the liver, i.e., Kupffer and other sinusoidal cells, but were unable to be properly cleared from the body. Overall, these results highlight an important link between AGEs and various disorders associated with them, which may serve as a platform for future research to better understand the processes and mechanisms of these disorders.
Kaneyasu Nishimura, Daisuke Doi, Bumpei Samata, Shigeo Murayama, Tsuyoshi Tahara, Hirotaka Onoe and Jun Takahashi : Estradiol Facilitates Functional Integration of iPSC-Derived Dopaminergic Neurons into Striatal Neuronal Circuits via Activation of Integrin α5β1., Stem Cell Reports, Vol.6, No.4, 511-524, 2016.
(要約)
For cell transplantation therapy for Parkinson's disease (PD) to be realized, the grafted neurons should be integrated into the host neuronal circuit to restore the lost neuronal function. Here, using wheat-germ agglutinin-based transsynaptic tracing, we show that integrin α5 is selectively expressed in striatal neurons that are innervated by midbrain dopaminergic (DA) neurons. In addition, we found that integrin α5β1 was activated by the administration of estradiol-2-benzoate (E2B) in striatal neurons of adult female rats. Importantly, we observed that the systemic administration of E2B into hemi-parkinsonian rat models facilitates the functional integration of grafted DA neurons derived from human induced pluripotent stem cells into the host striatal neuronal circuit via the activation of integrin α5β1. Finally, methamphetamine-induced abnormal rotation was recovered earlier in E2B-administered rats than in rats that received other regimens. Our results suggest that the simultaneous administration of E2B with stem cell-derived DA progenitors can enhance the efficacy of cell transplantation therapy for PD.
Multivalent interactions play an essential role in molecular recognition in living systems. These effects were employed to target tumor cells using albumin clusters bearing 10 molecules of asparagine-linked glycans (N-glycans). Noninvasive near-infrared fluorescence imaging clearly revealed A431 tumors implanted in BALB/cA-nu/nu mice after 1h in an N-glycan structure-dependent manner, thereby demonstrating the efficient use of glycan multivalency effects for tumor targeting in vivo.
Akihiro Ogura, Tsuyoshi Tahara, Satoshi Nozaki, Koji Morimoto, Yasuhiko Kizuka, Shinobu Kitazume, Mitsuko Hara, Soichi Kojima, Hirotaka Onoe, Almira Kurbangalieva, Naoyuki Taniguchi, Yasuyoshi Watanabe and Katsunori Tanaka : Visualizing Trimming Dependence of Biodistribution and Kinetics with Homo- and Heterogeneous N-Glycoclusters on Fluorescent Albumin., Scientific Reports, Vol.6, 2016.
(要約)
A series of N-glycans, each sequentially trimmed from biantennary sialoglycans, were homo- or heterogeneously clustered efficiently on fluorescent albumin using a method that combined strain-promoted alkyne-azide cyclization and 6π-azaelectrocyclization. Noninvasive in vivo kinetics and dissection analysis revealed, for the first time, a glycan-dependent shift from urinary to gall bladder excretion mediated by sequential trimming of non-reducing end sialic acids. N-glycoalbumins that were trimmed further, in particular, GlcNAc- and hybrid biantennary-terminated congeners, were selectively taken up by sinusoidal endothelial and stellate cells in the liver, which are critical for diagnosis and treatment of liver fibrillation. Our glycocluster strategy can not only reveal the previously unexplored extracellular functions of N-glycan trimming, but will be classified as the newly emerging glycoprobes for diagnostic and therapeutic applications.
Tsuyoshi Tahara, Zhouen Zhang, Masahiro Ohno, Yukako Hirao, Nami Hosaka, Hisashi Doi, Masaaki Suzuki and Hirotaka Onoe : A novel (11)C-labeled thymidine analog, [(11)C]AZT, for tumor imaging by positron emission tomography., EJNMMI Research, Vol.5, No.1, 2015.
(要約)
In PET studies, C6 and HeLa tumors in mice were clearly visualized using [(11)C]AZT. Time-activity curves using [(11)C]AZT showed that the accumulation of radioactivity in tumors plateaued at 10 min after injection and persisted for 60 min, while most of the radioactivity in other tissues was rapidly excreted into the urine. In various tissues of the body, tumor tissue showed the highest radioactivity at 80 min after injection (five to six times higher uptake than that of blood). Compared with tumor tissue, uptake was lower in other proliferative tissues such as the spleen, intestine, and bone marrow, resulting in a high tumor-to-bone marrow ratio. Cellular uptake of [(11)C]AZT in C6 cells was completely blocked by the application of thymidine, strongly indicating the specific involvement of nucleoside transporters. In contrast, the time-activity curve of [(11)C]d4T in the tumor showed transient and rapid excretion with almost no obvious tumor tissue accumulation.
Vascular smooth muscle cells (VSMCs) exhibit shrinkage-induced activation of Na(+)/H(+) exchanger isoform 1 (NHE-1) and Na(+), K(+), 2Cl(-) cotransporter (NKCC) under hyperosmotic conditions. To investigate the roles of these ion transporters in vascular smooth muscle force induced by hyperosmotic stress, we tested the effects of 5-(N, N-dimethyl)-amiloride (DMA; NHE inhibitor), cariporide (a selective NHE-1 inhibitor), and bumetanide (NKCC inhibitor) on the contractile response of rat aortic rings to hyperosmolar solutions. NHE inhibitors significantly augmented the maximum force response and contractile sensitivity to hyperosmolar sucrose, NaCl, and glucose in endothelium-denuded rings. Bumetanide elicited a comparatively modest increase in sensitivity. NHE inhibitors blocked the increase in intracellular pH and enhanced the cell volume decrease of cultured VSMCs after exposure to hyperosmolar sucrose. However, DMA had no effect on the increase in cytosolic free Ca(2+) concentration ([Ca(2+)]i) in rat VSMCs and on the increases in phosphorylation of myosin phosphatase target subunit 1 and myosin light chain (MLC) in aortic rings in response to hyperosmolar sucrose. Hyperosmolar sucrose-induced force was significantly attenuated by cytochalasin B in the presence or absence of DMA. Exposure to hyperosmolar sucrose increased the ratio of F- to G-actin; the ratio was further elevated by DMA. These results suggest that the potentiation of hyperosmotic shrinkage by NHE inhibition promotes actin polymerization in VSMCs and augments force production independent of changes in [Ca(2+)]i and MLC phosphorylation.
Takayuki Iwata, Katsunori Tanaka, Tsuyoshi Tahara, Satoshi Nozaki, Hirotaka Onoe, Yasuyoshi Watanabe and Koichi Fukase : A conformationally fixed analog of the peptide mimic Grb2-SH2 domain: synthesis and evaluation against the A431 cancer cell., Molecular BioSystems, Vol.9, No.5, 1019-1025, 2013.
(要約)
A small peptide mimic of the Grb2-SH2 domain, which was previously prepared through the template-assisted click approach and exhibited selective A431 tumor growth inhibition both in vitro and in vivo, was further elaborated on to enhance the interaction with target phosphorylated proteins. A conformationally fixed analog was efficiently synthesized by solid-supported ring-closing metathesis and Cu(i)/His-mediated self-activating Huisgen [3+2] cycloadditon as the key steps, and exhibited a 10-fold enhanced affinity to a phosphorylated peptide, a truncated peptide analog of the Grb2-SH2-interacting phosphoproteins. A stronger interaction with the target phosphorylated proteins gave this cyclic analog cytotoxic activity in A431 cells.
Shigeyuki Yamamoto, Yasuomi Ouchi, Daisaku Nakatsuka, Tsuyoshi Tahara, Kei Mizuno, Seiki Tajima, Hirotaka Onoe, Etsuji Yoshikawa, Hideo Tsukada, Masao Iwase, Kouzi Yamaguti, Hirohiko Kuratsune and Yasuyoshi Watanabe : Reduction of [11C](+)3-MPB binding in brain of chronic fatigue syndrome with serum autoantibody against muscarinic cholinergic receptor., PLoS ONE, Vol.7, No.12, 2012.
(要約)
The present results demonstrate that serum autoantibody against the mAChR can affect the brain mAChR without altering acetylcholinesterase activity and cognitive functions in CFS patients.
Katsunori Tanaka, Sanae Shirotsuki, Takayuki Iwata, Chika Kageyama, Tsuyoshi Tahara, Satoshi Nozaki, O Eric R Siwu, Satoru Tamura, Shunsuke Douke, Nobutoshi Murakami, Hirotaka Onoe, Yasuyoshi Watanabe and Koichi Fukase : Template-assisted and self-activating clicked peptide as a synthetic mimic of the SH2 domain., ACS Chemical Biology, Vol.7, No.4, 637-645, 2012.
(要約)
A new synthetic strategy for obtaining artificial receptors that selectively regulate and/or control specific protein/protein interactions was developed based on the template-assisted and the self-activating click reaction applied to a combinatorial library. Synthetic mimics of the Grb2-SH2 domain, examined as a model case, selectively bound to a target signaling protein to induce cytotoxicity and inhibit tumor growth in vivo.
Tsuyoshi Tahara, Masayoshi Yamamoto, Reiko Akagi, Hideo Harigae and Shigeru Taketani : The low expression allele (IVS3-48C) of the ferrochelatase gene leads to low enzyme activity associated with erythropoietic protoporphyria., International Journal of Hematology, Vol.92, No.5, 769-771, 2010.
(キーワード)
Alleles / Asian Continental Ancestry Group / Ferrochelatase / Gene Expression Regulation, Enzymologic / Genetic Association Studies / Humans / Mutation / Protoporphyria, Erythropoietic
Katsunori Tanaka, Kaori Minami, Tsuyoshi Tahara, Eric OC Siwu, Koichi Koyama, satoshi Nozaki, Hirotaka Onoe, Yasuyoshi Watanabe and Koichi Fukase : A Combined 6π-Azaelectrocyclization/Staudinger Approach to Protein and Cell Engineering: Noninvasive Tumor Targeting by N-Glycan-Engineered Lymphocytes, Journal of carbohydrate chemistry, Vol.29, No.3, 118-132, 2010.
26.
Satoshi Nozaki, Hiroshi Mizuma, Masaaki Tanaka, Guanghua Jin, Tsuyoshi Tahara, Kei Mizuno, Masanori Yamato, Kaori Okuyama, Asami Eguchi, Kouji Akimoto, Takahito Kitayoshi, Noriko Mochizuki-Oda, Yosky Kataoka and Yasuyoshi Watanabe : Thiamine tetrahydrofurfuryl disulfide improves energy metabolism and physical performance during physical-fatigue loading in rats., Nutrition Research, Vol.29, No.12, 867-872, 2009.
(要約)
Impaired energy metabolism is considered a possible cause of fatigue. The thiamine derivative, thiamine tetrahydrofurfuryl disulfide (TTFD), is prescribed and is also an over-the-counter drug for the attenuation of fatigue. It is readily absorbed from the intestinal tract and converted into thiamine pyrophosphate (TPP), which plays an important role as a cofactor for enzymes of metabolic pathways involved in the production of adenosine triphosphate (ATP). We postulated that TTFD has an anti-fatigue effect by improving energy metabolism during physical-fatigue loading. Here, we initially used the forced swimming test to determine whether daily TTFD or thiamine for 5 days has anti-fatigue effects on weight-loaded rats. The swimming duration of TTFD-, but not of thiamine-treated rats, was significantly longer than that of control rats (P < .05). Based on these findings, we examined changes in the levels of thiamine and its phosphate esters in various organs and the effect of TTFD on ATP levels in skeletal muscle after forced swimming, to determine the cellular mechanisms of the anti-fatigue effect of TTFD. Daily TTFD resulted in a characteristic distribution of thiamine and its phosphate esters in rat skeletal muscle, liver, kidney, heart, brain, and plasma. Furthermore, daily TTFD attenuated the decrease in ATP content in the skeletal muscle caused by forced swimming with a weight load for a defined period (150 s). These results indicate that TTFD exerts anti-fatigue effects by improving energy metabolism during physical fatigue.
(キーワード)
Adenosine Triphosphate / Animals / Energy Metabolism / Fatigue / Fursultiamin / Male / Muscle, Skeletal / Organ Specificity / Phosphorylation / Physical Endurance / Physical Exertion / Rats / Rats, Sprague-Dawley / Swimming / Thiamine / Vitamin B Complex
This study compared the effects of placebo with a carotenoid compound, crocetin, as well as an antioxidant, ascorbic acid, on physical fatigue in humans. In this double-blind, placebo-controlled, 3-way crossover study, 14 Japanese healthy volunteers (7 men and 7 women) were randomized to oral administration of crocetin (15 mg), ascorbic acid (3,000 mg), or placebo for 8 days. Subjects performed workload tests on a bicycle ergometer at fixed workloads for 120 minutes at 2 times (a total of 240 minutes) as a fatigue-inducing physical task. During the physical task, subjects performed nonworkload tests at maximum velocity (MV) of 10 seconds at 30 minutes (30-minute test) after the start of the physical task and at 30 minutes before the end of the task (210-minute test). The change in MV from the 30- to the 210-minute test was significantly higher in men who received crocetin compared with men who received placebo (P < .05). This effect of crocetin was specific to males. Administration of ascorbic acid did not change in MV from the 30-minute to the 210-minute test on males or females. These results suggest that daily administration of crocetin may attenuate physical fatigue in men.
Some mental or physical fatigue-related biochemical changes were determined. Various biochemical alterations reflecting homeostatic perturbation and its responses might be shown. We believe that our results contribute to clarifying the mechanism of fatigue, developing evaluation methods, and establishing a basis for treatment.
Suzuka Ataka, Masaaki Tanaka, Satoshi Nozaki, Hiroshi Mizuma, Kei Mizuno, Tsuyoshi Tahara, Tomohiro Sugino, Tomoko Shirai, Yoshitaka Kajimoto, Hirohiko Kuratsune, Osami Kajimoto and Yasuyoshi Watanabe : Effects of oral administration of caffeine and D-ribose on mental fatigue., Nutrition, Vol.24, No.3, 233-238, 2008.
(要約)
Because plasma branched-chain amino acid levels are decreased by mental fatigue, these results suggest that administration of caffeine improved task performance through the enhancement of central nervous system activity without increasing the sensation of fatigue. However, further decreases in branched-chain amino acid levels indicate that caffeine might promote deeper fatigue than placebo. Unfortunately, research subsequent to our study design has shown that D-ribose dosing higher than we used is needed to see a clinical effect and therefore no conclusions can be made from this study as to the efficacy of D-ribose.
(キーワード)
Administration, Oral / Adrenocorticotropic Hormone / Adult / Amino Acids, Branched-Chain / Caffeine / Central Nervous System Stimulants / Cross-Over Studies / Double-Blind Method / Fatigue / Female / Humans / Hydrocortisone / Male / Mental Processes / Reaction Time / Ribose / Task Performance and Analysis / Treatment Outcome
Oral administration of coenzyme Q10 improved subjective fatigue sensation and physical performance during fatigue-inducing workload trials and might prevent unfavorable conditions as a result of physical fatigue.
Katsunori Tanaka, Tatsuro Masuyama, Koki Hasegawa, Tsuyoshi Tahara, Hiroshi Mizuma, Yasuhiro Wada, Yasuyoshi Watanabe and Koichi Fukase : A submicrogram-scale protocol for biomolecule-based PET imaging by rapid 6pi-azaelectrocyclization: visualization of sialic acid dependent circulatory residence of glycoproteins., Angewandte Chemie International Edition, Vol.47, No.1, 102-105, 2008.
Gerhard Kelter, Daniel Steinbach, Badireenath Venkata Konkimalla, Tsuyoshi Tahara, Shigeru Taketani, Heinz-Herbert Fiebig and Thomas Efferth : Role of transferrin receptor and the ABC transporters ABCB6 and ABCB7 for resistance and differentiation of tumor cells towards artesunate., PLoS ONE, Vol.2, No.8, 2007.
(要約)
The anti-malarial artesunate also exerts profound anti-cancer activity. The susceptibility of tumor cells to artesunate can be enhanced by ferrous iron. The transferrin receptor (TfR) is involved in iron uptake by internalization of transferrin and is over-expressed in rapidly growing tumors. The ATP-binding cassette (ABC) transporters ABCB6 and ABCB7 are also involved in iron homeostasis. To investigate whether these proteins play a role for sensitivity towards artesunate, Oncotest's 36 cell line panel was treated with artesunate or artesunate plus iron(II) glycine sulfate (Ferrosanol). The majority of cell lines showed increased inhibition rates, for the combination of artesunate plus iron(II) glycine sulfate compared to artesunate alone. However, in 11 out of the 36 cell lines the combination treatment was not superior. Cell lines with high TfR expression significantly correlated with high degrees of modulation indicating that high TfR expressing tumor cells would be more efficiently inhibited by this combination treatment than low TfR expressing ones. Furthermore, we found a significant relationship between cellular response to artesunate and TfR expression in 55 cell lines of the National Cancer Institute (NCI), USA. A significant correlation was also found for ABCB6, but not for ABCB7 in the NCI panel. Artesunate treatment of human CCRF-CEM leukemia and MCF7 breast cancer cells induced ABCB6 expression but repressed ABCB7 expression. Finally, artesunate inhibited proliferation and differentiation of mouse erythroleukemia (MEL) cells. Down-regulation of ABCB6 by antisense oligonucleotides inhibited differentiation of MEL cells indicating that artesunate and ABCB6 may cooperate. In conclusion, our results indicate that ferrous iron improves the activity of artesunate in some but not all tumor cell lines. Several factors involved in iron homeostasis such as TfR and ABCB6 may contribute to this effect.
Tsuyoshi Tahara, Masaaki Tanaka, Satoshi Nozaki, Guanghua Jin, Hirotaka Onoe and Yasuyoshi Watanabe : Decrease of hepatic delta-aminolevulinate dehydratase activity in an animal model of fatigue., Biochemical and Biophysical Research Communications, Vol.353, No.4, 1068-1073, 2006.
(要約)
Fatigue can be defined physiologically as inability to maintain the expected power output. At present, no standard of fatigue are yet available. In order to find biomarkers of fatigue, we investigated the level of delta-aminolevulinic acid (ALA), the first intermediate metabolite in the heme biosynthetic pathway, in the plasma and urine of an animal model of fatigue. To prepare fatigued animals, we kept rats for 5 days in a cage filled with water to a height of 1.5 cm. As a result, the plasma and urinary ALA levels were increased in the fatigued animals as compared with those in the control animals. One day after the rats had been returned to their normal cages, these increased levels were restored to the control ones. We also examined the activity of the enzyme ALA dehydratase (ALAD), which is the second enzyme in the heme biosynthetic pathway, and ALAD gene expression during the fatigue and its recovery sessions. The ALAD activity, as well as its gene expression, in the liver of the fatigued animals was decreased as compared with those of the control animals. Both activity and gene expression of ALAD were recovered to their respective control levels after the rats had been allowed to rest in their normal cages for 1 day. Furthermore, the activity of ALA synthase (ALAS), the rate-limiting enzyme in the heme biosynthesis, in the liver was increased after the fatigue session for 5 days. Although this level of increase in the plasma concentration of ALA may not induce fatigue, increase in plasma and urinary ALA levels can be biomarkers of fatigue.
Reiko Akagi, Rikako Inoue, Shikibu Muranaka, Tsuyoshi Tahara, Shigeru Taketani, E Karl Anderson, D John Phillips and Shigeru Sassa : Dual gene defects involving delta-aminolaevulinate dehydratase and coproporphyrinogen oxidase in a porphyria patient., British Journal of Haematology, Vol.132, No.2, 237-243, 2006.
(要約)
Summary A Caucasian male had symptoms of acute porphyria, with increases in urinary delta-aminolaevulinic acid (ALA), porphobilinogen (PBG) and coproporphyrin that were consistent with hereditary coproporphyria (HCP). However, a greater than expected increase in ALA, compared with PBG, and a substantial increase in erythrocyte zinc protoporphyrin, suggested additional ALA dehydratase (ALAD) deficiency. Nucleotide sequence analysis of coproporphyrinogen oxidase (CPO) cDNA of the patient, but not of the parents, revealed a novel nucleotide transition G835-->C, resulting in an amino acid change, G279R. The mutant CPO protein expressed in Escherichia coli was unstable, and produced about 5% of activity compared with the wild-type CPO. Erythrocyte ALAD activity was 32% of normal in the proband. Nucleotide sequence analysis of cloned ALAD cDNAs from the patient revealed a C36-->G base transition (F12L amino acid change). The F12L ALAD mutation, which was found in the mother and a brother, was previously described, and is known to lack any enzyme activity. This patient thus represents the first case of porphyria where both CPO and ALAD deficiencies were demonstrated at the molecular level.
Tsuyoshi Tahara, Jiying Sun, Kazuhiko Igarashi and Shigeru Taketani : Heme-dependent up-regulation of the alpha-globin gene expression by transcriptional repressor Bach1 in erythroid cells., Biochemical and Biophysical Research Communications, Vol.324, No.1, 77-85, 2004.
(要約)
The transcriptional factor Bach1 forms a heterodimer with small Maf family, and functions as a repressor of the Maf recognition element (MARE) in vivo. To investigate the involvement of Bach1 in the heme-dependent regulation of the expression of the alpha-globin gene, human erythroleukemia K562 cells were cultured with succinylacetone (SA), a heme biosynthetic inhibitor, and the level of alpha-globin mRNA was examined. A decrease of alpha-globin mRNA was observed in SA-treated cells, which was restored by the addition of hemin. The heme-dependent expression of alpha-globin occurred at the transcriptional level since the expression of human alpha-globin gene promoter-reporter gene containing hypersensitive site-40 (HS-40) was decreased when K562 cells were cultured with SA. Hemin treatment restored the decrease of the promoter activity by SA. The regulation of the HS-40 activity by heme was dependent on the NF-E2/AP-1 (NA) site, which is similar to MARE. The NA site-binding activity of Bach1 in K562 increased upon SA-treatment, and the increase was diminished by the addition of hemin. The transient expression of Bach1 and mutated Bach1 lacking CP motifs suppressed the HS-40 activity, and cancellation of the repressor activity by hemin was observed when wild-type Bach1 was expressed. The expression of NF-E2 strengthened the restoration of the Bach1-effect by hemin. Interestingly, nuclear localization of Bach1 increased when cells were treated with SA, while hemin induced the nuclear export of Bach1. These results indicated that heme plays an important role in the induction of alpha-globin gene expression through disrupting the interaction of Bach1 and the NA site in HS-40 enhancer in erythroid cells.
Tsuyoshi Tahara, Jiying Sun, Katsuyuki Nakanishi, Masafumi Yamamoto, Hajime Mori, Takeshi Saito, Hiroyoshi Fujita, Kazuhiko Igarashi and Shigeru Taketani : Heme positively regulates the expression of beta-globin at the locus control region via the transcriptional factor Bach1 in erythroid cells., The Journal of Biological Chemistry, Vol.279, No.7, 5480-5487, 2003.
(要約)
The transcription factor Bach1 heterodimerizes with small Maf proteins to repress Maf recognition element (MARE)-dependent gene expression. The repressor activity of Bach1 is inhibited by the direct binding of heme. To investigate the involvement of Bach1 in the heme-dependent regulation of the expression of the beta-globin gene, mouse erythroleukemia (MEL) cells were cultured with succinylacetone (SA), a specific inhibitor of heme biosynthesis, and the level of beta-globin mRNA was examined. A marked decrease of beta-globin mRNA in SA-treated cells was observed, and this decrease was reversed by the addition of hemin. An iron chelator, desferrioxamine, also lowered the level of beta-globin mRNA. The heme-dependent expression of beta-globin is a transcriptional event since the expression of the human beta-globin gene promoter-reporter gene containing the microlocus control region (microLCR) was inhibited when human erythroleukemia K562 cells and MEL cells were cultured with SA. Hemin treatment restored the decrease in promoter activity caused by SA. The control of the microLCR-beta-globin promoter reporter gene by heme was dependent on DNase I-hypersensitive site 2 (HS2), which contains MARE. The MARE binding activity of Bach1 in K562 and MEL cells increased upon SA treatment, and the increase was diminished by the treatment with hemin. Transient expression of Bach1 suppressed the microLCR activity, and this repressor activity was cancelled by treatment with hemin. The expression of a mutated Bach1 lacking heme-binding sites led to a loss in the heme responsiveness of the microLCR. Furthermore, chromatin immunoprecipitation experiments revealed that Bach1 bound to the MARE of HS2 increased by the treatment of MEL cells with SA, and this was cancelled by hemin. We propose that heme positively regulates the beta-globin gene expression by blocking the interaction of Bach1 with the MARE in the LCR.
Yumiko Yasui, Shikibu Muranaka, Tsuyoshi Tahara, Ryo Shimizu, Sonoko Watanabe, Yutaka Horie, Eiji Nanba, Hiroshi Uezato, Atsushi Takamiyagi, Shigeru Taketani and Reiko Akagi : A new ferrochelatase mutation combined with low expression alleles in a Japanese patient with erythropoietic protoporphyria., Clinical Science, Vol.102, No.5, 501-506, 2002.
(要約)
We investigated the molecular defect of the ferrochelatase gene in a Japanese patient with erythropoietic protoporphyria (EPP), and identified a novel 16 base pair (574-589) deletion within exon 5. This deletion resulted in a frame-shift mutation and created a premature stop codon at amino acid position 198. The same molecular defect was also identified in his mother and a brother who had symptomatic EPP, but not in his father who was asymptomatic. The subjects with EPP were homozygous for the low expression haplotype, while his father was heterozygous for this haplotype. These results indicate that the combination of a 16 base pair deletion and low expression of the wild-type allelic variant is responsible for EPP in this pedigree.
(キーワード)
Adult / Base Sequence / Cells, Cultured / DNA Mutational Analysis / DNA, Complementary / Ferrochelatase / Humans / Male / Molecular Sequence Data / Mutation / Pedigree / Polymorphism, Genetic / Porphyria, Hepatoerythropoietic
(文献検索サイトへのリンク)
● PubMed @ National Institutes of Health, US National Library of Medicine (PMID): 11980567
Chiho Shinozaki, Yutaka Kohmura, Tetsuro Yoshimaru, Tsuyoshi Tahara, Masaya Denda, Hidefumi Mukai, Kohta Mohri, Yi Long Chen, Toyomasa Katagiri and Akira Otaka : Study on a lipidated anti-cancer peptide allowing long-lasting duration in mice model, AIMECS 2023, Seoul, Jun. 2023.
国内講演発表:
1.
田原 強 : 小動物を用いたがんPETイメージング, SMART2023, 2023年8月.
2.
Takashi Niwa, 田原 強, Chase E. Charles, Fang G. Francis, Nakaoka Takayoshi, Irie Satsuki, Hayashinaka Emi, Wada Yasuhiro, Mukai Hidefumi, Masutomi Kenkichi, Watanabe Yasuyoshi, Cui Yi-Long ), Hosoya Takamitsu : Synthesis of 11C-Radiolabeled Eribulin via Henry Reaction, 日本分子イメージング学会誌, 2022年5月.