Shin-ichi Tsunoda

National Institute of Biomedical Innovation, Ibaragi, Ōsaka, Japan

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Publications (162)486.28 Total impact

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    ABSTRACT: Ephrin receptor A10 (EphA10) is a relatively uncharacterized protein which is expressed in many breast cancers but not expressed in normal breast tissues. Here, we examined the potential of EphA10 as a drug target in breast cancer. Immunohistochemical staining of clinical tissue sections revealed that EphA10 was expressed in various breast cancer subtypes, including triple negative breast cancers (TNBCs), with no expression observed in normal tissues apart from testis. Ligand-dependent proliferation was observed in EphA10-transfected MDA-MB-435 cells (MDA-MB-435(EphA10)) and native TNBC cells (MDA-MB-436). However, this phenomenon was not observed in parental MDA-MB-435 cells which express a low level of EphA10. Finally, tumor growth was significantly suppressed by administration of an anti-EphA10 monoclonal antibody in a xenograft mouse model. These results suggest that inhibition of EphA10 signaling may be a novel therapeutic option for management of breast cancer, including TNBCs which are currently not treated with molecularly targeted agents.
    Journal of controlled release : official journal of the Controlled Release Society. 06/2014;
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    ABSTRACT: We recently identified Eph receptor A10 (EphA10) as a novel breast cancer-specific protein. Moreover, we also showed that an in-house developed anti-EphA10 monoclonal antibody (mAb) significantly inhibited proliferation of breast cancer cells, suggesting EphA10 as a promising target for breast cancer therapy. However, the only other known report for EphA10 was its expression in the testis at the mRNA level. Therefore, the potency of EphA10 as a drug target against cancers other than the breast is not known. The expression of EphA10 in a wide variety of cancer cells was studied and the potential of EphA10 as a drug target was evaluated. Screening of EphA10 mRNA expression showed that EphA10 was overexpressed in breast cancer cell lines as well as in prostate and colon cancer cell lines. Thus, we focused on prostate cancers in which EphA10 expression was equivalent to that in breast cancers. As a result, EphA10 expression was clearly shown in clinical prostate tumor tissues as well as in cell lines at the mRNA and protein levels. In order to evaluate the potential of EphA10 as a drug target, we analyzed complement-dependent cytotoxicity effects of anti-EphA10 mAb and found that significant cytotoxicity was mediated by the expression of EphA10. Therefore, the idea was conceived that the overexpression of EphA10 in prostate cancers might have a potential as a target for prostate cancer therapy, and formed the basis for the studies reported here.
    Biochemical and Biophysical Research Communications 06/2014; · 2.28 Impact Factor
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    ABSTRACT: Asian dust is a springtime meteorological phenomenon that originates in the deserts of China and Mongolia. The dust is carried by prevailing winds across East Asia where it causes serious health problems. Most of the information available on the impact of Asian dust on human health is based on epidemiological investigations, so from a biological standpoint little is known of its effects. To clarify the effects of Asian dust on human health, it is essential to assess inflammatory responses to the dust and to evaluate the involvement of these responses in the pathogenesis or aggravation of disease. Here, we investigated the induction of inflammatory responses by Asian dust particles in macrophages. Treatment with Asian dust particles induced greater production of inflammatory cytokines interleukin-6 and tumor necrosis factor- α (TNF- α ) compared with treatment with soil dust. Furthermore, a soil dust sample containing only particles ≤10 μ m in diameter provoked a greater inflammatory response than soil dust samples containing particles >10 μ m. In addition, Asian dust particles-induced TNF- α production was dependent on endocytosis, the production of reactive oxygen species, and the activation of nuclear factor- κ B and mitogen-activated protein kinases. Together, these results suggest that Asian dust particles induce inflammatory disease through the activation of macrophages.
    Research Journal of Immunology 01/2014; 2014:856154.
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    ABSTRACT: Although amorphous silica nanoparticles are widely used in the production of food products (e.g., as anticaking agents), there is little information available about their absorption and biological effects after oral exposure. Here, we examined the in vitro intestinal absorption and in vivo biological effects in mice of orally administered amorphous silica particles with diameters of 70, 300, and 1,000 nm (nSP70, mSP300, and mSP1000, respectively) and of nSP70 that had been surface-modified with carboxyl or amine groups (nSP70-C and nSP70-N, respectively). Analysis of intestinal absorption by means of the everted gut sac method combined with an inductively coupled plasma optical emission spectrometer showed that the intestinal absorption of nSP70-C was significantly greater than that of nSP70. The absorption of nSP70-N tended to be greater than that of nSP70; however, the results were not statistically significant. Our results indicate that silica nanoparticles can be absorbed through the intestine and that particle diameter and surface properties are major determinants of the degree of absorption. We also examined the biological effects of the silica particles after 28-day oral exposure in mice. Hematological, histopathological, and biochemical analyses showed no significant differences between control mice and mice treated with the silica particles, suggesting that the silica nanoparticles evaluated in this study are safe for use in food production.
    Nanoscale Research Letters 01/2014; 9(1):532. · 2.52 Impact Factor
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    ABSTRACT: Eph receptor A10 (EphA10) is a valuable breast cancer marker that is highly expressed in breast cancer tissues by comparison with normal breast tissues, as we previously reported. However, the role of EphA10 expression in breast cancer is not well understood. Here, we have analyzed the expression of EphA10 at the mRNA- and protein-level in clinical breast cancer tissues and then evaluated the relationship with clinicopathological parameters for each sample. EphA10 mRNA expression was quantified by real-time polymerase chain reaction using complimentary DNA (cDNA) samples derived from breast cancer patients. Lymph node (LN) metastasis and stage progression were significantly correlated with EphA10 expression at the mRNA level (P = 0.0091 and P = 0.034, respectively). Furthermore, immunohistochemistry (IHC) staining of breast cancer tissue microarrays (TMAs) revealed that EphA10 expression at the protein level was also associated with LN metastasis and stage progression (P = 0.016 and P = 0.011, respectively). These results indicate that EphA10 expression might play a role in tumor progression and metastasis. Our findings will help elucidate the role of EphA10 in clinical breast cancer progression.
    Cancer Medicine 12/2013; 2(6):972-977.
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    ABSTRACT: Detection of drug-target proteins and biomarkers that are expressed in cancer tissue has significant potential for both diagnosis and treatment of cancer. However, current immuno-histochemical and cytogenetic analyses of biopsy specimens for pre-operational diagnosis are highly invasive and often difficult to apply to lung cancer patients. The purpose of this study was to evaluate the possible utility of determining epidermal growth factor receptor (EGFR) expression on exosomal membranes using a targeted ELISA with an anti-CD81 antibody as a capture antibody for lung cancer diagnosis. While soluble EGFR (sEGFR) levels in plasma were not remarkably different between lung cancer patients and normal controls, significantly higher exosomal EGFR expression levels were observed in 5/9 cancer cases compared to normal controls. These results suggest that measurement of exosomal protein levels could be useful for in vitro diagnosis, and that exosomal EGFR is a possible biomarker for characterization of lung cancer.
    Pharmazie 12/2013; 68(12):969-73. · 0.96 Impact Factor
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    ABSTRACT: Although nanomaterials are being used in various fields, their safety is not yet sufficiently understood. We have been attempting to establish a nanomaterials safety-assessment system by using biomarkers to predict nanomaterial-induced adverse biological effects. Here, we focused on microRNAs (miRNAs) because of their tissue-specific expression and high degree of stability in the blood. We previously showed that high intravenous doses of silica nanoparticles of 70 nm diameter (nSP70) induced liver damage in mice. In this study, we compared the effectiveness of serum levels of liver-specific or -enriched miRNAs (miR-122, miR-192, and miR-194) with that of conventional hepatic biomarkers (alanine aminotransferase (ALT) and aspartate aminotransferase (AST)) as biomarkers for nSP70. After mice had been treated with nSP70, their serum miRNAs levels were measured by using quantitative RT-PCR. Serum levels of miR-122 in nSP70-treated mice were the highest among the three miRNAs. The sensitivity of miR-122 for liver damage was at least as good as those of ALT and AST. Like ALT and AST, miR-122 may be a useful biomarker of nSP70. We believe that these findings will help in the establishment of a nanomaterials safety-assessment system.
    Nanotechnology 09/2013; 24(40):405102. · 3.84 Impact Factor
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    ABSTRACT: Nanomaterials with particle sizes <100 nm have been already applied in various applications such as cosmetics, medicines, and foods. Therefore, ensuring the safety of nanomaterials is becoming increasingly important. Here we examined the localization and biological responses of intranasally administered amorphous nanosilica particles in mice, focusing on the coagulation system. We used nanosilica particles with diameters of 30, 70, or 100 nm (nSP30, nSP70, or nSP100 respectively), and conventional microscale silica particles with diameters of 300 or 1000 nm (mSP300 or mSP1000, respectively). BALB/c mice were intranasally exposed to nSP30, nSP70, nSP100, mSP300, or mSP1000 at concentrations of 500 mug/mouse for 7 days. After 24 hours of last administration, we performed the in vivo transmission electron microscopy analysis, hematological examination and coagulation tests. In vivo transmission electron microscopy analysis showed that nanosilica particles with a diameter <100 nm were absorbed through the nasal cavity and were distributed into liver and brain. Hematological examination and coagulation tests showed that platelet counts decreased and that the activated partial thromboplastin time was prolonged in nSP30 or nSP70-treated groups of mice, indicating that nanosilica particles might have activated a coagulation cascade. In addition, in in vitro activation tests of human plasma, nanosilica particles had greater potential than did conventional microscale silica particles to activate coagulation factor XII. In nanosilica-particle-treated groups, the levels of soluble CD40 ligand, and von Willebrand factor which are involved in stimulating platelets tended to slightly increase with decreasing particle size. These results suggest that intranasally administered nanosilica particles with diameters of 30 and 70 nm could induce abnormal activation of the coagulation system through the activation of an intrinsic coagulation cascade. This study provides information to advance the development of safe and effective nanosilica particles.
    Particle and Fibre Toxicology 08/2013; 10(1):41. · 9.18 Impact Factor
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    ABSTRACT: Monoclonal antibodies (mAbs) that are internalized into cells are a current focus in the development of antibody-drug conjugates (ADCs). We describe a phage display-based high-throughput screening system to rapidly isolate cell-internalizing mAbs. We simultaneously examined the cell-internalizing activities of several hundred independent mAbs, and successfully isolated cell-internalizing mAbs against the tumor endothelial markers Roundabout homolog 4 (Robo4) and vascular endothelial growth factor receptor 2 (VEGFR2). Tumor accumulation of mAbs with high cell-internalizing activity was significantly higher than that of mAbs with low cell-internalizing activity. Furthermore, the anti-tumor effects of ADCs of mAbs with high cell-internalizing activity were significantly stronger than those of mAbs with low cell-internalizing activity. While anti-VEGFR2 therapy caused a significant loss of body weight, anti-Robo4 therapy did not. These findings indicate that cell-internalizing activity has an important role in the biodistribution and therapeutic effects of ADCs. Further, Robo4 can be an effective marker for tumor vascular targeting.
    Blood 01/2013; · 9.78 Impact Factor
  • Shin-Ichi Tsunoda
    YAKUGAKU ZASSHI 01/2013; 133(9):923-4. · 0.37 Impact Factor
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    ABSTRACT: We previously identified the E693Δ mutation in amyloid precursor protein (APP) in patients with Alzheimer's disease (AD) and then generated APP-transgenic mice expressing this mutation. As these mice possessed abundant Aß oligomers from 8 months of age but no amyloid plaques even at 24 months of age, they are a good model to study pathological effects of amyloid ß (Aß) oligomers. The two-dimensional fluorescence difference in gel electrophoresis (2D-DIGE) technology, using a mixed-sample internal standard, is now recognized as an accurate method to determine and quantify proteins. In this study, we examined the proteins for which levels were altered in the hippocampus of 12-month-old APP(E693Δ)-transgenic mice using 2D-DIGE and liquid chromatography-tandem mass spectrometry (LC-MS/MS). Fourteen proteins were significantly changed in the hippocampus of APP(E693Δ)-transgenic mice. Actin cytoplasmic 1 (ß-actin), heat shock cognate 71kDa, γ-enolase, ATP synthase subunit ß, tubulin ß-2A chain, clathrin light chain B (clathrin) and dynamin-1 were increased. Heat shock-related 70kDa protein 2, neurofilament light polypeptide (NFL), stress-induced-phosphoprotein 2, 60kDa heat shock protein (HSP60), α-internexin, protein kinase C and casein kinase substrate in neurons protein 1 (Pacsin 1), α-enolase and ß-actin were decreased. Western Blotting also validated the changed levels of HSP60, NFL, clathrin and Pacsin 1 in APP(E693Δ)-transgenic mice. The identified proteins could be classified as cytoskeleton, chaperons, neurotransmission, energy supply and signal transduction. Thus, proteomics by 2D-DIGE and LC-MS/MS has provided knowledge of the levels of proteins in the early stages of AD brain.
    Neuroscience Letters 12/2012; · 2.03 Impact Factor
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    ABSTRACT: The cytokine lymphotoxin-α (LTα) is a promising candidate for use in cancer therapy. However, the instability of LTαin vivo and the insufficient levels of tumor necrosis factor receptor 1 (TNFR1)-mediated bioactivity of LTα limit its therapeutic potential. Here, we created LTα mutants with increased TNFR1-mediated bioactivity by using a phage display technique. We constructed a phage library displaying lysine-deficient structural variants of LTα with randomized amino acid residues. After affinity panning, we screened three clones of lysine-deficient LTα mutant, and identified a LTα mutant with TNFR1-mediated bioactivity that was 32times that of the wild-type LTα (wtLTα). When compared with wtLTα, the selected clone showed augmented affinity to TNFR1 due to slow dissociation rather than rapid association. In contrast, the mutant showed only 4times the TNFR2-mediated activity of wtLTα. In addition, the LTα mutant strongly and rapidly activated caspases that induce TNFR1-mediated cell death, whereas the mutant and wtLTα activated nuclear factor-kappa B to a similar extent. Our data suggest that the kinetics of LTα binding to TNFR1 play an important role in signal transduction patterns, and a TNFR1-selective LTα mutant with augmented bioactivity would be a superior candidate for cancer therapy.
    Cytokine 12/2012; · 2.52 Impact Factor
  • Pharmazie 11/2012; 67(11):958-9. · 0.96 Impact Factor
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    ABSTRACT: Practical uses of nanomaterials are rapidly spreading to a wide variety of fields. However, potential harmful effects of nanomaterials are raising concerns about their safety. Therefore, it is important that a risk assessment system is developed so that the safety of nanomaterials can be evaluated or predicted. Here, we attempted to identify novel biomarkers of nanomaterial-induced health effects by a comprehensive screen of plasma proteins using two-dimensional differential in gel electrophoresis (2D-DIGE) analysis. Initially, we used 2D-DIGE to analyze changes in the level of plasma proteins in mice after intravenous injection via tail veins of 0.8 mg/mouse silica nanoparticles with diameters of 70 nm (nSP70) or saline as controls. By quantitative image analysis, protein spots representing >2.0-fold alteration in expression were found and identified by mass spectrometry. Among these proteins, we focused on hemopexin as a potential biomarker. The levels of hemopexin in the plasma increased as the silica particle size decreased. In addition, the production of hemopexin depended on the characteristics of the nanomaterials. These results suggested that hemopexin could be an additional biomarker for analyzing the biological responses associated with exposure to silica nanoparticles. We believe that this study will contribute to the development of biomarkers to ensure the safety of silica nanoparticles.
    Nanoscale Research Letters 10/2012; 7(1):555. · 2.52 Impact Factor
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    ABSTRACT: Recently, nanomaterials have been utilized in various fields. In particular, amorphous nanosilica particles are increasingly being used in a range of applications, including cosmetics, food technology, and medical diagnostics. However, there is concern that the unique characteristics of nanomaterials might induce undesirable effects. The roles played by the physical characteristics of nanomaterials in cellular responses have not yet been elucidated precisely. Here, by using nanosilica particles (nSPs) with a diameter of 70nm whose surface was either unmodified (nSP70) or modified with amine (nSP70-N) or carboxyl groups (nSP70-C), we examined the relationship between the surface properties of nSPs and cellular responses such as cytotoxicity, reactive oxygen species (ROS) generation, and DNA damage. To compare the cytotoxicity of nSP70, nSP70-N, or nSP70-C, we examined in vitro cell viability after nSP treatment. Although the susceptibility of each cell line to the nSPs was different, nSP70-C and nSP70-N showed lower cytotoxicity than nSP70 in all cell lines. Furthermore, the generation of ROS and induction of DNA damage in nSP70-C- and nSP70-N-treated cells were lower than those in nSP70-treated cells. These results suggest that the surface properties of nSP70 play an important role in determining its safety, and surface modification of nSP70 with amine or carboxyl groups may be useful for the development of safer nSPs. We hope that our results will contribute to the development of safer nanomaterials.
    Biochemical and Biophysical Research Communications 10/2012; · 2.28 Impact Factor
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    ABSTRACT: Nanomaterials (NMs) exhibit unique physicochemical properties and innovative functions, and they are increasingly being used in a wide variety of fields. Ensuring the safety of NMs is now an urgent task. Recently, we reported that amorphous silica nanoparticles (nSPs), one of the most widely used NMs, enhance antigen-specific cellular immune responses and may therefore aggravate immune diseases. Thus, to ensure the design of safer nSPs, investigations into the effect of nSPs on antigen presentation in dendritic cells, which are central orchestrators of the adaptive immune response, are now needed. Here, we show that nSPs with diameters of 70 and 100nm enhanced exogenous antigen entry into the cytosol from endosomes and induced cross-presentation, whereas submicron-sized silica particles (>100nm) did not. Furthermore, we show that surface modification of nSPs suppressed cross-presentation. Although further studies are required to investigate whether surface-modified nSPs suppress immune-modulating effects in vivo, the current results indicate that appropriate regulation of the characteristics of nSPs, such as size and surface properties, will be critical for the design of safer nSPs.
    Biochemical and Biophysical Research Communications 09/2012; 427(3):553-6. · 2.28 Impact Factor
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    ABSTRACT: Mesothelioma is a highly malignant tumor with a poor prognosis and limited treatment options. Although cisplatin (CDDP) is an effective anticancer drug, its response rate is only 20%. Therefore, discovery of biomarkers is desirable to distinguish the CDDP-susceptible versus resistant cases. To this end, differential proteome analysis was performed to distinguish between mesothelioma cells of different CDDP susceptibilities, and this revealed that expression of annexin A4 (ANXA4) protein was higher in CDDP-resistant cells than in CDDP-susceptible cells. Furthermore, ANXA4 expression levels were higher in human clinical malignant mesothelioma tissues than in benign mesothelioma and normal mesothelial tissues. Finally, increased susceptibility was observed following gene knockdown of ANXA4 in mesothelioma cells, whereas the opposite effect was observed following transfection of an ANXA4 plasmid. These results suggest that ANXA4 has a regulatory function related to the cisplatin susceptibility of mesothelioma cells and that it could be a biomarker for CDDP susceptibility in pathological diagnoses.
    Biochemical and Biophysical Research Communications 04/2012; 421(1):140-4. · 2.28 Impact Factor
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    ABSTRACT: It has gradually become evident that nanomaterials, which are widely used in cosmetics, foods, and medicinal products, could induce substantial inflammation. However, the roles played by the physical characteristics of nanomaterials in inflammatory responses have not been elucidated. Here, we examined how particle size and surface modification influenced the inflammatory effects of nanosilica particles, and we investigated the mechanisms by which the particles induced inflammation. We compared the inflammatory effects of silica particles with diameters of 30-1,000 nm in vitro and in vivo. In macrophages in vitro, 30- and 70-nm nanosilica particles (nSP30 and nSP70) induced higher production of tumor necrosis factor-α (TNFα) than did larger particles. In addition, intraperitoneal injection of nSP30 and nSP70 induced stronger inflammatory responses involving cytokine production than did larger particles in mice. nSP70-induced TNFα production in macrophage depended on the production of reactive oxygen species and the activation of mitogen-activated protein kinases (MAPKs). Furthermore, nSP70-induced inflammatory responses were dramatically suppressed by surface modification of the particles with carboxyl groups in vitro and in vivo; the mechanism of the suppression involved reduction in MAPK activation. These results provide basic information that will be useful for the development of safe nanomaterials.
    Archives of Toxicology 03/2012; 86(8):1297-307. · 5.22 Impact Factor
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    ABSTRACT: Amorphous nanosilica particles (nSP) are being utilized in an increasing number of applications such as medicine, cosmetics, and foods. The reduction of the particle size to the nanoscale not only provides benefits to diverse scientific fields but also poses potential risks. Several reports have described the in vivo and in vitro toxicity of nSP, but few studies have examined their effects on the male reproductive system. The aim of this study was to evaluate the testicular distribution and histologic effects of systemically administered nSP. Mice were injected intravenously with nSP with diameters of 70 nm (nSP70) or conventional microsilica particles with diameters of 300 nm (nSP300) on two consecutive days. The intratesticular distribution of these particles 24h after the second injection was analyzed by transmission electron microscopy. nSP70 were detected within sertoli cells and spermatocytes, including in the nuclei of spermatocytes. No nSP300 were observed in the testis. Next, mice were injected intravenously with 0.4 or 0.8 mg nSP70 every other day for a total of four administrations. Testes were harvested 48 h and 1 week after the last injection and stained with hematoxylin-eosin for histologic analysis. Histologic findings in the testes of nSP70-treated mice did not differ from those of control mice. Taken together, our results suggest that nSP70 can penetrate the blood-testis barrier and the nuclear membranes of spermatocytes without producing apparent testicular injury.
    Biochemical and Biophysical Research Communications 03/2012; 420(2):297-301. · 2.28 Impact Factor
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    ABSTRACT: Since metastasis is one of the most important prognostic factors in colorectal cancer, development of new methods to diagnose and prevent metastasis is highly desirable. However, the molecular mechanisms leading to the metastatic phenotype have not been well elucidated. In this study, a proteomics-based search was carried out for metastasis-related proteins in colorectal cancer by analyzing the differential expression of proteins in primary versus metastasis focus-derived colorectal tumor cells. Protein expression profiles were determined using a tissue microarray (TMA), and the results identified Rho GDP-dissociation inhibitor alpha (Rho GDI) as a metastasis-related protein in colon and prostate cancer patients. Consequently, Rho GDI may be useful as a diagnostic biomarker and/or a therapeutic to prevent colon and prostate cancer metastasis.
    Pharmazie 03/2012; 67(3):253-5. · 0.96 Impact Factor

Publication Stats

1k Citations
486.28 Total Impact Points

Institutions

  • 2006–2014
    • National Institute of Biomedical Innovation
      • Laboratory of Biopharmaceutical Research
      Ibaragi, Ōsaka, Japan
  • 1995–2013
    • Osaka University
      • • School of Pharmaceutical Sciences
      • • Division of Molecular Pharmaceutical Science
      Ibaraki, Osaka-fu, Japan
    • Kobe Gakuin University
      • Faculty of Pharmaceutical Sciences
      Suita, Osaka-fu, Japan
  • 1995–2001
    • Osaka University of Pharmaceutical Sciences
      • Faculty of Pharmaceutical Sciences
      Ōsaka, Ōsaka, Japan
  • 1997–1999
    • Chugai pharmceutical
      Japan