Faculty Profiles - HANAFUSA Tadashi

Papers - HANAFUSA Tadashi

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The current status and novel advances of boron neutron capture therapy clinical trials. International journal

Tianyun Zhou, Kazuyo Igawa, Tomonari Kasai, Takuya Sadahira, Wei Wang, Tomofumi Watanabe, Kensuke Bekku, Satoshi Katayama, Takehiro Iwata, Tadashi Hanafusa, Abai Xu, Motoo Araki, Hiroyuki Michiue, Peng Huang

American journal of cancer research 14 ( 2 ) 429 - 447 2024

Language：English Publishing type：Research paper (scientific journal)

Boron neutron capture therapy (BNCT) is a treatment method that focuses on improving the cure rate of patients with cancer who are difficult to treat using traditional clinical methods. By utilizing the high neutron absorption cross-section of boron, material rich in boron inside tumor cells can absorb neutrons and release high-energy ions, thereby destroying tumor cells. Owing to the short range of alpha particles, this method can precisely target tumor cells while minimizing the inflicted damage to the surrounding normal tissues, making it a potentially advantageous method for treating tumors. Globally, institutions have progressed in registered clinical trials of BNCT for multiple body parts. This review summarized the current achievements in registered clinical trials, Investigator-initiated clinical trials, aimed to integrate the latest clinical research literature on BNCT and to shed light on future study directions.

Development example of internal exposure calculation system in medium-sized radiation facilities Reviewed

IMADA Yui, ISOBE Midori, NAGAMATSU Tomohiro, TERATO Hiroaki, HANAFUSA Tadashi

Japanese Journal of Radiation Safety Management 23 ( 2 ) 78 - 82 2024

Authorship：Corresponding author Language：Japanese Publisher：Japanese Society of Radiation Safety Management

When calculating internal exposure dose, a method of extracting and evaluating representatives is recommended. We have developed an evaluation system that calculates internal doses using records of radioisotopes use and radiation workers’ entry into controlled areas. By using this system, it is possible to evaluate internal doses based on correct working hours and to evaluate internal doses of non-representatives, thereby improving the accuracy of radiation safety management.

DOI： 10.11269/jjrsm.23.78

Particle and Heavy Ion Transport Code System‐Based Microdosimetry for the Development of Boron Agents for Boron Neutron Capture Therapy Reviewed

Takafumi Shigehira, Tadashi Hanafusa, Kazuyo Igawa, Tomonari Kasai, Shuichi Furuya, Hisakazu Nishimori, Yoshinobu Maeda, Hiroyuki Michiue, Atsushi Fujimura

Advanced Theory and Simulations 2023.4

Practical Peport of the Group-based Management That Restricts Simultaneous Use of Radioisotopes Reviewed

Yui Imada, Midori Isobe, Tomohiro Nagamatsu, Hiroaki Terato, Tadashi Hanafusa

Japanese Journal of Radiation Safety Management 21 ( 2 ) 64 - 68 2022.11

In Vitro Studies to Define the Cell-Surface and Intracellular Targets of Polyarginine-Conjugated Sodium Borocaptate as a Potential Delivery Agent for Boron Neutron Capture Therapy Reviewed

4. Atsushi Fujimura, Seiji Yasui, Kazuyo Igawa, Ai Ueda, Kaori Watanabe, Tadashi Hanafusa, Yasuaki Ichikawa, Sachiko Yoshihashi, Kazuki Tsuchida, Atsunori Kamiya and Shuichi Furuya

Cells 9 ( 10 ) 2149 - 2149 2020.9

Language：English Publishing type：Research paper (scientific journal) Publisher：MDPI AG

Boron neutron capture therapy (BNCT) requires pharmaceutical innovations and molecular-based evidence of effectiveness to become a standard cancer therapeutic in the future. Recently, in Japan, 4-borono-L-phenylalanine (BPA) was approved as a boron agent for BNCT against head and neck (H&N) cancers. H&N cancer appears to be a suitable target for BPA-BNCT, because the expression levels of L-type amino acid transporter 1 (LAT1), one of the amino acid transporters responsible for BPA uptake, are elevated in most cases of H&N cancer. However, in other types of cancer including malignant brain tumors, LAT1 is not always highly expressed. To expand the possibility of BNCT for these cases, we previously developed poly-arginine peptide (polyR)-conjugated mercaptoundecahydrododecaborate (BSH). PolyR confers the cell membrane permeability and tumor selectivity of BSH. However, the molecular determinants for the properties are not fully understood. In this present study, we have identified the cluster of differentiation 44 (CD44) protein and translational machinery proteins as a major cell surface target and intracellular targets of BSH-polyR, respectively. CD44, also known as a stem cell-associated maker in various types of cancer, is required for the cellular uptake of polyR-conjugated molecules. We showed that BSH-polyR was predominantly delivered to a CD44High cell population of cancer cells. Once delivered, BSH-polyR interacted with the translational machinery components, including the initiation factors, termination factors, and poly(A)-biding protein (PABP). As a proof of principle, we performed BSH-polyR-based BNCT against glioma stem-like cells and revealed that BSH-polyR successfully induced BNCT-dependent cell death specifically in CD44High cells. Bioinformatics analysis indicated that BSH-polyR would be suitable for certain types of malignant tumors. Our results shed light on the biochemical properties of BSH-polyR, which may further contribute to the therapeutic optimization of BSH-BNCT in the future.

DOI： 10.3390/cells9102149

The accelerator-based boron neutron capture reaction evaluation system for head and neck cancer Reviewed

Shou Wang, Kazuyo Igawa, Ryo Ogawara, Mitsuru Suda, Tsuyoshi Hamano, Soichiro Ibaragi, Tadashi Hanafusa, Yasuaki Ichikawa, Akira Sasakia.

Applied Radiation and Isotopes 165 2020.7

Radiocesium concentration in stems, leaves, and panicles of rice grown in a sandy soil replacement paddy field treated with different rates of cattle manure compost in Kawamata, Fukushima Reviewed

3. Nittaya WAKAI, Morihiro MAEDA, Toshiro ONO, Tadashi HANAFUSA, Jun YAMASHITA and Kuniyuki SAITOH

Journal of Environmental Science for Sustainable Society (JESSS) 2019 ( 9 ) 1 - 10 2019.7

Factors affecting 137Cs concentration in wild plants and soils of different land use in Iitate village after the Fukushima nuclear power plant accident. Reviewed

4. Nittaya WAKAI, Jun YAMASHITA, Takashi ENOMOTO, Tadashi HANAFUSA, Toshiro ONO, Morihiro MAEDA

Radiation Safety Management 2018.9

In vitro analysis of radioprotective effect of monoterpenes Reviewed

Ken-ichi Kudo, Tadashi Hanafusa, Toshiro Ono

JOURNAL OF RADIOANALYTICAL AND NUCLEAR CHEMISTRY 313 ( 1 ) 169 - 174 2017.7

Language：English Publishing type：Research paper (scientific journal) Publisher：SPRINGER

Monoterpenes are naturally occurring hydrocarbons composed of two units of isoprenes. They exhibit antioxidant activity to scavenge reactive oxygen species, such as hydroxyl radicals. We investigated the potential of monoterpenes such as thymol, linalool, and menthol to act as radioprotectants. The proliferation of EL4 cells, a mouse lymphoma cell line, treated with linalool at a concentration of 500 mu M or more was not affected by X-ray irradiation. Plasmid-nicking assay performed using formamidopyrimidine-DNA glycosylase showed that linalool prevented single strand breaks and oxidized purines on pUC19 plasmid DNA. These findings indicate that linalool has the ability to scavenge reactive oxygen species and is a potential radioprotector.

DOI： 10.1007/s10967-017-5268-0

Oncogenic function of the homeobox A13-long noncoding RNA HOTTIP-insulin growth factor-binding protein 3 axis in human gastric cancer Reviewed

Sophie S. W. Wang, Kenly Wuputra, Chung-Jung Liu, Yin-Chu Lin, Yi-Ting Chen, Chee-Yin Chai, Chen-Lung Steve Lin, Kung-Kai Kuo, Ming-Ho Tsai, Shin-Wei Wang, Ker-Kong Chen, Hiroyuki Miyoshi, Yukio Nakamura, Shigeo Saito, Tadashi Hanafusa, Deng-Chyang Wu, Chang-Shen Lin, Kazunari K. Yokoyama

ONCOTARGET 7 ( 24 ) 36049 - 36064 2016.6

Language：English Publishing type：Research paper (scientific journal) Publisher：IMPACT JOURNALS LLC

To study the mechanisms of gastric tumorigenesis, we have established CSN cell line from human normal gastric mucosa, and CS12, a tumorigenic and invasive gastric cancer cell line from CSN passages. Many stem cell markers were expressed in both CSN and CS12 cells, but LGR5 and NANOG were expressed only in CS12 cells. Increased expression of homeobox A13 (HoxA13) and its downstream cascades was significant for the tumorigenic activity of CS12 cells, and was associated with recruitment of E2F-1 to HoxA13 promoter accompanied with increased trimethylation of histone H3 lysine 4 (H3K4me3) at the hypomethylated E2F motifs. Knockdown of HoxA13 caused the downregulation of long non-coding RNA HOTTIP and insulin growth factor-binding protein 3 (IGFBP-3) genes, indicating that both were targets of HoxA13. Concurrent regulation of HoxA13-HOTTIP was mediated by the mixed lineage leukemia-WD repeat domain 5 complex, which caused the trimethylation of H3K4 and then stimulated cell proliferation. HoxA13 transactivated the IGFBP-3 promoter through the HOX-binding site. Activation of IGFBP-3 stimulated the oncogenic potential and invasion activity. Increased expression of HoxA13 (63.2%) and IGFBP-3 (28.6%) was detected in human gastric cancer tissues and was found in the gastric cancer data of The Cancer Genome Atlas. Taken together, the HoxA13 HOTTIP IGFBP-3 cascade is critical for the carcinogenic characteristics of CS12 cells.

DOI： 10.18632/oncotarget.9102

Adsorption and removal of strontium in aqueous solution by synthetic hydroxyapatite Reviewed

Yuichi Nishiyama, Tadashi Hanafusa, Jun Yamashita, Yoko Yamamoto, Toshiro Ono

JOURNAL OF RADIOANALYTICAL AND NUCLEAR CHEMISTRY 307 ( 2 ) 1279 - 1285 2016.2

Language：English Publishing type：Research paper (scientific journal) Publisher：SPRINGER

Hydroxyapatite (HAP) is a main mineral constituent of bone and tooth and has an outstanding bio-compatibility. HAP is a possible sorbent for heavy metals in wastewater due to its high adsorption capacity and low water solubility. We developed a removal system of Sr-90 from aqueous solution by HAP column procedure. More than 90 % of Sr-90 was adsorbed and removed from the Sr-90 containing solution. Divalent cations, Ca2+, had little effect on the removal of Sr-90 up to a concentration of 1 mmol L-1. This clearly indicates that the HAP column technique is advantageous with respect to the capacity to adsorb Sr-90 from water present in the environment.

DOI： 10.1007/s10967-015-4228-9

An Evaluation of Radiation Exposure and Contamination in PET Facility Reviewed

Hidekazu SAKUNO, Keiko YAMATO, Tadashi HANAFUSA, Toshiro ONO

Japanese Journal of Radiation Safety Management 14 ( 2 ) 91 - 95 2015

A simple method for sampling and measurement of radiocarbon by a passive diffusion sampling technique Radiation Reviewed

Tomohiro NAGAMATSU, Tadashi HANAFUSA, Toshiro ONO

Radiation Safety Management 14 ( 1 ) 9 - 14 2015

Language：English Publishing type：Research paper (scientific journal) Publisher：Japanese Society of Radiation Safety Management

Passive diffusion sampling has been widely used to monitor organic and inorganic pollutants in the environment. We herein developed a new and simple passive diffusion sampling method to monitor ¹⁴C in the indoor working environment of radioisotope facilities. The badge-type Shikata sampler is an open-end type with no diffusion cap that fits a collection filter with a diameter of 25 mm. This sampler could efficiently collect indoor ¹⁴C with good reliability and a collection efficiency that was 10-fold higher than that of the conventional active pumped method. The Shikata sampler is small, simple, and easy to handle. It may be subsidiary used to monitor indoor ¹⁴C concentrations in radioisotope working rooms, in which radioactivity must be measured once a month as recommended by the Industrial Safety and Health Law.<br>

DOI： 10.12950/rsm.14.9

Other Link： http://search.jamas.or.jp/link/ui/2016061476

Estimation of soil-to-plant transfer factors of radiocesium in 99 wild plant species grown in arable lands 1 year after the Fukushima 1 Nuclear Power Plant accident Reviewed

Jun Yamashita, Takashi Enomoto, Masao Yamada, Toshiro Ono, Tadashi Hanafusa, Tomohiro Nagamatsu, Shoji Sonoda, Yoko Yamamoto

JOURNAL OF PLANT RESEARCH 127 ( 1 ) 11 - 22 2014.1

Language：English Publishing type：Research paper (scientific journal) Publisher：SPRINGER JAPAN KK

One year after the deposition of radionuclides from the Fukushima 1 Nuclear Power Plant (A formal name is Fukushima Daiichi Nuclear Power Station) in March 2011, radiocesium (Cs-134, Cs-137) concentrations ([Cs]) were comprehensively investigated in the wild plants of 99 species most of which were annual or summer green perennial herbs and started to grow from April 2012 at the heavily contaminated fields of paddy (three study sites) and upland (one study site) in Fukushima Prefecture. The survey was conducted three times (April, July and October) in the year. In each site, soils (soil cores of 5-cm depth) and plants (aerial shoots) were collected for determination of [Cs] on a dry weight basis, and then the transfer factor (TF) of radiocesium from soil to plant ([Cs](plant)/[Cs](soil)) was estimated in each species. The [Cs] values of both soils and plants largely varied. However, some species exhibited relatively high TF values (more than 0.4) (e.g., Athyrium yokoscense, Dryopteris tokyoensis, and Cyperus brevifolius), while others exhibited almost negligible values (less than 0.01) (e.g., Salix miyabeana, Humulus scandens, and Elymus tsukushiensis). In addition, judging from the 11 species grown in both paddy and upland fields, TF values were generally higher in the paddy fields. The estimation of phytoextraction efficiency of soil radiocesium by weed communities in the paddy fields suggests that the weed community is not a practical candidate for phytoremediation technique.

DOI： 10.1007/s10265-013-0605-z

Practice of the safe handling of radioisotopes using naturally occurring radioactive materials Reviewed

HANAFUSA Tadashi, NAGAMATSU Tomohiro, ONO Toshiro

Japanese Journal of Radiation Safety Management 13 ( 2 ) 166 - 172 2014

Authorship：Lead author Language：Japanese Publisher：Japanese Society of Radiation Safety Management

We proposed a practical protocol for the safe handling of radiation and radioisotopes using the naturally occurring radioactive materials (NORMs) ⁸⁷Rb and ⁴⁰K. The protocol can be utilized for the education and training course for radiation handling workers. Rubidium chloride solutions were used for the practice of dilution of unsealed radioisotopes instead of ³²P. Rubidium chloride and potassium chloride solid radiation sources were used for the practice of monitoring of the surface contamination using the GM survey meter instead of ³²P and ¹⁴C. Rubidium chloride solutions were also used for the practice of the contamination check by the smear method. Substitution of the radioisotopes under the Laws Concerning the Prevention from Radiation Hazards due to Radioisotope and others to NORMs can be successfully achieved without notable demerits. We supposed that most of the practice for the safe handling of radioisotopes could be carried out outside a controlled area.<br>

DOI： 10.11269/jjrsm.13.166

Other Link： https://jlc.jst.go.jp/DN/JALC/10041555346?from=CiNii

A purification system for 64Cu produced by a biomedical cyclotron for antibody PET imaging Reviewed

Teruaki Toyota, Tadashi Hanafusa, Takashi Oda, Iwane Koumura, Takanori Sasaki, Eiji Matsuura, Hiromi Kumon, Tsuneo Yano, Toshiro Ono

Journal of Radioanalytical and Nuclear Chemistry 298 ( 1 ) 295 - 300 2013

Language：English Publishing type：Research paper (scientific journal)

Ion exchange is a simple and efficient method for separating no-carrier-added 64Cu from an irradiated Ni target. We developed a semi-automated two-round 64Cu separation system equipped with a strong-base anion exchange resin column. We first verified the efficiency of the system using a non-radioactive substitute consisting of 25 mg of Ni and 127 ng of Cu, and confirmed that Cu was completely eluted at the second round of the separation step. After the bombardment, separation of 64Cu from the Ni target was achieved with high radiochemical purity. 64Cu produced and separated in this study had an extremely low level of Ni impurity. It could be used for labeling monoclonal antibodies for antibody positron emission tomography imaging and synthesizing radiopharmaceuticals. © 2012 The Author(s).

DOI： 10.1007/s10967-012-2340-7

Serological identification of Tektin5 as a cancer/testis antigen and its immunogenicity Reviewed

Tadashi Hanafusa, Ali Eldib Ali Mohamed, Shohei Domae, Eiichi Nakayama, Toshiro Ono

BMC CANCER 12 ( 1 ) 520 2012.11

Authorship：Lead author Language：English Publishing type：Research paper (scientific journal) Publisher：BIOMED CENTRAL LTD

Background: Identification of new cancer antigens is necessary for the efficient diagnosis and immunotherapy. A variety of tumor antigens have been identified by several methodologies. Among those antigens, cancer/testis (CT) antigens have became promising targets.
Methods: The serological identification of antigens by the recombinant expression cloning (SEREX) methodology has been successfully used for the identification of cancer/testis (CT) antigens. We performed the SEREX analysis of colon cancer.
Results: We isolated a total of 60 positive cDNA clones comprising 38 different genes. They included 2 genes with testis-specific expression profiles in the UniGene database, such as TEKT5 and a CT-like gene, A kinase anchoring protein 3 (AKAP3). Quantitative real-time RT-PCR analysis showed that the expression of TEKT5 was restricted to the testis in normal adult tissues. In malignant tissues, TEKT5 was aberrantly expressed in a variety of cancers, including colon cancer. A serological survey of 101 cancer patients with different cancers by ELISA revealed antibodies to TEKT5 in 13 patients, including colon cancer. None of the 16 healthy donor serum samples were reactive in the same test.
Conclusion: We identified candidate new CT antigen of colon cancer, TEKT5. The findings indicate that TEKT5 is immunogenic in humans, and suggest its potential use as diagnostic as well as an immunotherapeutic reagent for cancer patients.

DOI： 10.1186/1471-2407-12-520

Isolation and characterization of human lung cancer antigens by serological screening with autologous antibodies Reviewed

Tadashi Hanafusa, Ali Eldib Ali Mohamed, Kenta Kitaoka, Yoshihiro Ohue, Eiichi Nakayama, Toshiro Ono

CANCER LETTERS 301 ( 1 ) 57 - 62 2011.2

Authorship：Lead author Language：English Publishing type：Research paper (scientific journal) Publisher：ELSEVIER IRELAND LTD

Serological analysis of a recombinant cDNA expression library (SEREX) derived from two lung adenocarcinoma cancer cell lines using autologous sera led to the isolation of 41 positive cDNA clones comprising 28 different antigens. They coded for a variety of nuclear and cytoplasmic proteins. Among the antigens, nucleoporin 107 (NUP107) was isolated most frequently (5 of 41 clones). The second most frequently isolated antigen was coded for by C21orf58 (4 of 41 clones). During serological analysis of selected antigens based on their reactivity to sera from normal individuals and lung cancer patients, none of the antigens showed a cancer-restricted recognition pattern. However, five genes including NUP107 showed higher expression when we examined the changes in gene expression in five different adenocarcinoma cell lines, including those used in SEREX, compared with their levels in normal lung tissues by cDNA microarray analysis. On the other hand, the expression levels of five genes including C21orf58 were down regulated in all adenocarcinoma cell lines. This SEREX study combining comprehensive gene expression assays has added to the growing list of lung cancer antigens, which may aid the development of diagnostic and immunotherapeutic reagents for patients with lung cancer. (C) 2010 Elsevier Ireland Ltd. All rights reserved.

DOI： 10.1016/j.canlet.2010.10.024

Proposal of a Utilization of a Luminous Bacterium in the Teaching and Learning of Radiation Safety Reviewed

Tadashi Hanafusa, Akihiro Sakoda, Tomohiro Nagamatsu, Ikuo Kinno, Toshiro Ono

Radiation Safety Management 10 ( 1 ) 8 - 13 2011

An Assessment of Radioactivity Level in 51Cr-contaminated Dry Solid Waste Generated from a Research Facility for Verification of Clearance Levels Reviewed

Tomohiro Nagamatsu, Tadashi Hanafusa, Toshiro Ono, Kiyonori Yamaoka.

Radiation Safety Management 9 ( 1 ) 7 - 12 2010

Language：English Publishing type：Research paper (scientific journal) Publisher：Japanese Society of Radiation Safety Management

Radioactive waste generated from research laboratories and other facilities is regulated by the Law Concerning Prevention from Radiation Hazards due to Radioisotopes etc. (Prevention Law). However, the Prevention Law does not provide the level of clearance or the procedures to follow for compliance monitoring. To assess radioactivity amounts for making decisions about clearance levels, the radioactivity levels in dry solid semi-combustible wastes generated from biomedical research, such as ⁵¹Cr-release assays, were measured and evaluated. Radioactivity of semi-combustible waste was 1.42-6.32% of the initial level. In comparison, records for the past 8 years in the Shikata Laboratory, Department of Radiation Research, Okayama University Advanced Science Research Center, indicated 7% to 90% of the initial radioactivity remained in the waste and was differed widely among researchers. This study determined an accurate radioactivity level in dry solid waste, which could lead to savings in disposal costs.<br>

DOI： 10.12950/rsm.9.7

Selective delivery of beta cell antigen to dendritic cells in vivo leads to deletion and tolerance of autoreactive CD8(+) T cells in NOD mice Reviewed

Arunika Mukhopadhaya, Tadashi Hanafusa, Irene Jarchum, Yi-Guang Chen, Yoshiko Iwai, David V. Serreze, Ralph M. Steinman, Kristin V. Tarbell, Teresa P. DiLorenzo

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA 105 ( 17 ) 6374 - 6379 2008.4

Authorship：Lead author Language：English Publishing type：Research paper (scientific journal) Publisher：NATL ACAD SCIENCES

Type 1 diabetes (T1D) is an autoimmune disease resulting from defects in central and peripheral tolerance and characterized by T cell-mediated destruction of islet beta cells. Cytotoxic CD8(+) T cells, reactive to beta cell antigens, are required for T1D development in the NOD mouse model of the disease, and CD8(+) T cells specific for beta cell antigens can be detected in the peripheral blood of T1D patients. It has been evident that in nonautoimmune-prone mice, dendritic cells (DCs) present model antigens in a tolerogenic manner in the steady state, e.g., in the absence of infection, and cause T cells to proliferate initially but then to be deleted or rendered unresponsive. However, this fundamental concept has not been evaluated in the setting of a spontaneous autoimmune disease. To do so, we delivered a mimotope peptide, recognized by the diabetogenic CD8(+) T cell clone A14 to DCs in NOD mice via the endocytic receptor DEC-205. Proliferation of transferred antigen-specific T cells was initially observed, but this was followed by deletion. Tolerance was achieved because rechallenge of mice with the mimotope peptide in adjuvant did not induce an immune response. Thus, targeting of DCs with P cell antigens leads to deletion of autoreactive CD8+ T cells even in the context of ongoing autoimmunity in NOD mice with known tolerance defects. Our results provide support for the development of DC targeting of self antigens for treatment of chronic T cell-mediated autoimmune diseases.

DOI： 10.1073/pnas.0802644105

Methodological Approach for Assessment of the Radioactivity Level in Dry Solid Waste Reviewed

Tomohiro Nagamatsu, Akihiro Sakoda, Katsumi Hanamoto, Ikuo Kinno, Tadashi Hanafusa, Kiyonori Yamaoka, Toshiro Ono

Radiation Safety Management 7 ( 1 ) 6 - 10 2008

Aberrant promoter methylation of insulin-like growth factor binding protein-3 gene in human cancers Reviewed

Kunitoshi Tomii, Kazunori Tsukuda, Shinichi Toyooka, Hideaki Dote, Tadashi Hanafusa, Hiroaki Asano, Minoru Naitou, Hiroyoshi Doihara, Takumi Kisimoto, Hideki Katayama, Harvery I. Pass, Hiroshi Date, Nobuyoshi Shimizu

INTERNATIONAL JOURNAL OF CANCER 120 ( 3 ) 566 - 573 2007.2

Language：English Publishing type：Research paper (scientific journal) Publisher：WILEY-LISS

Insulin-like growth factor binding protein-3 (IGFBP-3) is postulated to be a mediator of growth suppression signals. Here, we examined the methylation status of IGFBP-3 to correlate to clinicopathological factors in human cancers. The methylation status of IGFBP-3 was determined by bisulfite DNA sequencing and was correlated with expression semi-quantified by real-time RT-PCR to develop a methylation-specific PCR (MSP) assay for IGFBP-3. Using the MSP assay, we examined the methylation status of IGFBP-3 in gastric cancer (GC), colorectal cancer (CRC), breast cancer (BC) and malignant mesothelioma (MM). IGFBP-3 methylation was detected in 6 of 13 (46%) and 16 of 24 (67%) GC cell lines and tumors, respectively; 4 of 8 (50%) and 15 of 26 (58%) CRC cell lines and tumors, respectively; 3 of 11 (27%) and 7 of 39 (18%) BC cell lines and tumors, respectively and 1 of 5 (20%) and 18 of 56 (32%) MM cell lines and tumors, respectively. Interestingly, the methylation status of MM specimens from Japanese patients (75%, 12 out of 16 patients) was significantly higher than those from the USA (15%, 6 out of 40 patients) (p < 0.0001), suggesting the presence of ethnic differences in the IGFBP-3 methylation status. We also found that IGFBP-3 methylation was preferentially present in GCs arising in the lower-third of the stomach (p = 0.079). In summary, our results showed that IGFBP-3 methylation played an important role in the silencing of its expression, suggesting that IGFBP-3 may act as a tumor suppressor gene in several human cancers examined. (c) 2006 Wiley-Liss, Inc.

DOI： 10.1002/ijc.22341

IGFBP-3 expression in hepatocellular carcinoma involves abnormalities in TGF-beta and/or Rb signaling pathways Reviewed

E Yumoto, H Nakatsukasa, T Hanafusa, Y Yumoto, K Nouso, E Matsumoto, T Onishi, Y Takuma, H Tanaka, T Fujikawa, M Suzuki, M Uemura, Y Shiratori

INTERNATIONAL JOURNAL OF ONCOLOGY 27 ( 5 ) 1223 - 1230 2005.11

Language：English Publishing type：Research paper (scientific journal) Publisher：PROFESSOR D A SPANDIDOS

Insulin-like growth factor binding protein-3 (IGFBP-3) is a mediator of growth suppression signals and a putative tumor suppressor gene. The growth suppression mechanisms of IGFBP-3 have not been well clarified. We examined the expression of IGFBP-3 transcripts in human hepatocellular carcinoma (HCC) and the relationship between IGFBP-3 expression and the transforming growth factor-beta (TGF-beta) and/or retinoblastoma (Rb) signaling pathways. In situ hybridization revealed IGFBP-3 transcripts in cancer cells in 6 of 57 (10%) HCCs, including moderately and poorly differentiated HCCs with intrahepatic metastasis. In contrast, all lung metastatic nodules of 4 HCCs showed IGFBP-3 transcripts in cancer cells. The cDNA microarray showed that genes for the TGF-beta pathway and Rb were up-regulated in IGFBP-3-expressing HCCs. In 6 HCCs presenting IGFBP-3, immunohistochemical analyses showed abnormalities in the TGF-beta and/or Rb pathways; the loss of phosphorylated-Smad2 was observed in 2, and overexpression of phosphorylated-Rb was observed in the remaining 4 HCCs. The present study suggests that IGFBP-3 mediates growth suppression signals via the TGF-beta and/or Rb pathways in HCC.

Altered expression of vascular endothelial growth factor, fibroblast growth factor-2 and endostatin in patients with hepatocellular carcinoma Reviewed

S Uematsu, T Higashi, K Nouso, K Kariyama, SI Nakamura, M Suzuki, H Nakatsukasa, Y Kobayashi, T Hanafusa, T Tsuji, Y Shiratori

JOURNAL OF GASTROENTEROLOGY AND HEPATOLOGY 20 ( 4 ) 583 - 588 2005.4

Language：English Publishing type：Research paper (scientific journal) Publisher：BLACKWELL PUBLISHING ASIA

Background: Advanced hepatocellular carcinoma (HCC) in humans is characterized by hypervascularity. In the present study, the expressions of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (FGF-2) and endostatin were analyzed in patients with chronic liver disease to clarify the effect of these major angiogenic factors.
Methods: Serum concentrations of VEGF, FGF-2 and endostatin in 24 patients with HCC, 16 patients with liver cirrhosis (LC) and 13 healthy volunteers were measured by enzyme-linked immunosorbent assay. The expression of VEGF in 21 surgically resected HCC samples was analyzed by immunohistochemistry, and that of VEGF isoforms in 15 HCC samples was assessed by reverse transcriptase-polymerase chain reaction (RT-PCR).
Results: Serum VEGF, FGF-2 and endostatin concentrations were significantly elevated in patients with HCC compared with healthy volunteers; but there was no significant difference between patients with HCC and those with non-HCC liver disease. Immunohistochemical analysis showed that VEGF protein was strongly expressed in both well-differentiated HCC cells and non-cancerous hepatocytes, whereas in moderately and poorly differentiated HCC the expression was stronger in the endothelial cells (EC) lining intratumor vessels than in the cancer cells. On RT-PCR for VEGF isoforms it was found that VEGF-121, VEGF-165 and VEGF-189 were expressed in all but one of the HCC samples and in all corresponding non-HCC samples.
Conclusions: The results suggest that VEGF, FGF-2, and endostatin concentrations are elevated prior to the emergence of HCC and that the distribution of VEGF changes dynamically during the development of HCC. (C) 2005 Blackwell Publishing Asia Pty Ltd.

DOI： 10.1111/j.1400-1746.2005.03726.x

Functional promoter upstream p53 regulatory sequence of IGFBP3 that is silenced by tumor specific methylation Reviewed

T Hanafusa, T Shinji, H Shiraha, K Nouso, Y Iwasaki, E Yumoto, T Ono, N Koide

BMC CANCER 5 2005.1

Authorship：Lead author Language：English Publishing type：Research paper (scientific journal) Publisher：BIOMED CENTRAL LTD

Background: Insulin-like growth factor binding protein (IGFBP)-3 functions as a carrier of insulin-like growth factors (IGFs) in circulation and a mediator of the growth suppression signal in cells. There are two reported p53 regulatory regions in the IGFBP3 gene; one upstream of the promoter and one intronic. We previously reported a hot spot of promoter hypermethylation of IGFBP-3 in human hepatocellular carcinomas and derivative cell lines. As the hot spot locates at the putative upstream p53 consensus sequences, these p53 consensus sequences are really functional is a question to be answered.
Methods: In this study, we examined the p53 consensus sequences upstream of the IGFBP-3 promoter for the p53 induced expression of IGFBP-3. Deletion, mutagenesis, and methylation constructs of IGFBP-3 promoter were assessed in the human hepatoblastoma cell line HepG2 for promoter activity.
Results: Deletions and mutations of these sequences completely abolished the expression of IGFBP-3 in the presence of p53 overexpression. In vitro methylation of these p53 consensus sequences also suppressed IGFBP-3 expression. In contrast, the expression of IGFBP-3 was not affected in the absence of p53 overexpression. Further, we observed by electrophoresis mobility shift assay that p53 binding to the promoter region was diminished when methylated.
Conclusion: From these observations, we conclude that four out of eleven p53 consensus sequences upstream of the IGFBP-3 promoter are essential for the p53 induced expression of IGFBP-3, and hypermethylation of these sequences selectively suppresses p53 induced IGFBP-3 expression in HepG2 cells.

DOI： 10.1186/1471-2407-5-9

Telomerase reverse transcriptase gene amplification in hepatocellular carcinoma Reviewed

Y Takuma, K Nouso, Y Kobayashi, S Nakamura, H Tanaka, E Matsumoto, T Fujikawa, M Suzuki, T Hanafusa, Y Shiratori

JOURNAL OF GASTROENTEROLOGY AND HEPATOLOGY 19 ( 11 ) 1300 - 1304 2004.11

Language：English Publishing type：Research paper (scientific journal) Publisher：BLACKWELL PUBLISHING ASIA

Background and Aim: Telomerase activation is essential for the immortality of cancer cells. The expression of telomerase reverse transcriptase (hTERT), the catalytic component of the telomerase complex, regulates telomerase activity in human cancers. Amplification of the hTERT gene, located at chromosome 5p, is thought to be a potential genetic event contributing to telomerase activation in sporadic tumors.
Methods: The amplification of the hTERT gene was examined in 46 surgically resected hepatocellular carcinomas (HCC) by real-time polymerase chain reaction and the status was compared with the expression of hTERT mRNA and clinicopathological parameters.
Results: Amplified hTERT genes were found in 21.7% (10/46) of HCC. The incidence of amplified hTERT genes in poorly differentiated HCC (6/12, 50%) was significantly higher than that in highly to moderately differentiated HCC (4/34, 11.8%; P = 0.012). Tumor size in those cases with hTERT gene amplification was larger compared to those cases with no amplification (P = 0.047). Amplification of the hTERT gene was not observed in non-cancerous tissues. The hTERT mRNA level did not correlate with the number of hTERT genes.
Conclusions: Based on these results, it is thought that hTERT gene amplification is a cancer-specific event, and may furthermore contribute to the dedifferentiation and development of HCC. However, hTERT gene overexpression was rarely due to an increased hTERT gene copy number in HCC. (C) 2004 Blackwell Publishing Asia Pry Ltd.

DOI： 10.1111/j.1400-1746.2004.03447.x

Decreased expression of B7 costimulatory molecules and major histocompatibility complex class-I in human hepatocellular carcinoma Reviewed

K Fujiwara, T Higashi, K Nouso, H Nakatsukasa, Y Kobayashi, M Uemura, SI Nakamura, S Sato, T Hanafusa, Y Yumoto, Naito, I, Y Shiratori

JOURNAL OF GASTROENTEROLOGY AND HEPATOLOGY 19 ( 10 ) 1121 - 1127 2004.10

Language：English Publishing type：Research paper (scientific journal) Publisher：BLACKWELL PUBLISHING ASIA

Background and Aim: We analyzed the expression of antigen-processing and antigen-presenting molecules in surgically resected fresh samples of human hepatocellular carcinoma (HCC) tissue to elucidate a mechanism of immune escape. We also examined the expression of interleukin (IL)-10 protein, which might act to downregulate expression of antigen-processing and antigen-presenting molecules.
Methods: Twenty-eight HCC samples obtained by surgical resection were analyzed for the expression of beta2-microglobulin, heat-shock protein (HSP)-70, human leukocyte antigen (HLA) class-I, CD80 (B7-1), CD86 (B7-2) and IL-10 by immunostaining.
Results beta2-Microglobulin and HSP-70 were preserved in all samples. In contrast, the expression of HLA class-I molecules was significantly reduced according to lowering in the histological grading of tumor differentiation (P = 0.024). Furthermore, B7-1 and B7-2 expression was reduced in tumor cells compared with corresponding areas of liver tissue without malignant involvement irrespective of the histological grading of tumors (21% and 36%, respectively). Although IL-10 protein was expressed in 54% of HCC, no relationship between the expression of IL-10 and downregulation of B7-1, B7-2, and HLA class-I was evident.
Conclusion: These findings suggest the potential role of B7 co-stimulatory molecules and HLA class-I molecules in facilitating HCC escape from immune surveillance without the involvement of IL-10. (C) 2004 Blackwell Publishing Asia Pty Ltd.

DOI： 10.1111/j.1400-1746.2004.03467.x

Cellular distribution of telomerase reverse transcriptase in human hepatocellular carcinoma Reviewed

T Onishi, K Nouso, T Higashi, N Toshikuni, H Nakatsukasa, Y Kobayashi, M Uemura, E Yumoto, K Fujiwara, S Sato, S Nakamura, J Yokoyama, T Hanafusa, Y Shiratori

JOURNAL OF GASTROENTEROLOGY AND HEPATOLOGY 18 ( 10 ) 1168 - 1174 2003.10

Language：English Publishing type：Research paper (scientific journal) Publisher：BLACKWELL PUBLISHING ASIA

Background and Aim: Telomerase is the enzyme that synthesizes telomeric DNA, and the activation of telomerase is closely related to cellular immortality. Telomerase activity has been reported in many human cancer tissues and is regulated by the expression of human telomerase reverse transcriptase ( hTERT). The aim of the present study was to identify hTERT-expressing cells in human liver tissues and evaluate the feasibility of the hTERT promoter for gene therapy of hepatocellular carcinoma (HCC).
Methods: The authors examined the cellular distribution of hTERT transcripts in surgically resected HCC by in situ hybridization.
Results: Among 20 samples, hTERT expression was observed in 15 HCC. Transcripts of hTERT were homogenously distributed in the cytoplasm of HCC cells in nine of 15 cases; six of 15 cases displayed a heterogeneous staining pattern. All poorly differentiated HCC that expressed hTERT showed a homogenous pattern of staining. None of the non-cancerous hepatocytes were positive for the transcripts, but infiltrating lymphocytes were faintly stained. The homogenous expression of hTERT was also observed in the vascular invasion of HCC.
Conclusions: The results indicate that most HCC cells express hTERT RNA and that the promoter is a good candidate as a target for gene therapy. However, careful consideration must be taken concerning the potential effects on lymphocytes. (C) 2003 Blackwell Publishing Asia Pty Ltd.

DOI： 10.1046/j.1440-1746.2003.03137.x

Identification of the antigens predominantly reacted with serum from patients with hepatocellular carcinoma Reviewed

M Uemura, K Nouso, Y Kobayashi, H Tanaka, S Nakamura, T Higashi, T Ono, E Nakayama, T Hanafusa, Y Shiratori

CANCER 97 ( 10 ) 2474 - 2479 2003.5

Language：English Publishing type：Research paper (scientific journal) Publisher：JOHN WILEY & SONS INC

BACKGROUND. To identify antigens specifically recognized by the immune surveillance system in patients with hepatocellular carcinoma (HCC), the authors examined two complementary DNA (cDNA) libraries of moderately differentiated HCC by serologic analysis of recombinant cDNA expression libraries (SEREX).
METHODS. The libraries were screened with autologous patients' sera, and sequences of the reacted clones were determined. To study the immunoreactivity of the antigens, sera from 20 patients with HCC, from 20 healthy volunteers, and from 16 patients with chronic viral hepatitis were examined.
RESULTS. Twenty-seven antigens were identified. They included SART1, p57Kip2, ROCK-1, gamma-catenin, and heat shock proteins, which are classified as tumor-associated genes. Three of 27 antigens-Tat-binding protein-1 (TBP-1), beta4 integrin-binding protein (p27[BBP]), and ribosomal protein L30 (rpL30)-were reacted predominantly with sera from patients with HCC (55% of patients, 45% of patients, and 20% of patients, respectively). Patients in the control group had no antibodies against these three antigens. Seventy percent of patients with HCC had the antibody against at least one of these antigens.
CONCLUSIONS. Disease specific humoral immune response against TBP-1, p27(BBP), and rpL30 was induced in patients with HCC, and the antibodies against these antigens also may be used as tumor markers. (C) 2003 American Cancer Society.

DOI： 10.1002/cncr.11374

Quantitative analysis of p40/p46 and p69/p71 forms of 2 ',5 '-oligoadenylate synthetase mRNA by competitive PCR and its clinical application Reviewed

A Takahashi, Y Iwasaki, J Miyaike, H Taniguchi, H Shimomura, T Hanafusa, Y Yumoto, A Moriya, N Koide, T Tsuji

CLINICAL CHEMISTRY 48 ( 9 ) 1551 - 1559 2002.9

Language：English Publishing type：Research paper (scientific journal) Publisher：AMER ASSOC CLINICAL CHEMISTRY

Background: 2',5.-Oligoadenylate synthetases (2-5AS) are type I interferon (IFN)-induced proteins with antiviral capacity. Three major forms of 2-5AS with distinct enzymatic activities have been described in IFN-treated human cells. We. measured, distinct forms of 2-5AS MRNA to analze the relationship with its enzymatic activity and response to IFN therapy in chronic hepatitis C.
Methods: We established a method to quantify p40/p46 and p69/p71 forms of 2-5AS mRNA by use of reverse transcription followed by competitive PCR. The 2-5AS mRNA concentrations, were measured in peripheral blood mononuclear cells from 40 patients with chronic hepatitis C and 28 control individuals.
Results: Reconstitution experiments and comparison with Northern blot analyses revealed that our method accurately. and linearly quantified 2-5AS mRNA. 2-5AS mRNA concentrations and 2-5AS enzymatic activity were correlated (P <0.03). Our data demonstrated a correlation in 2-5AS mRNA between p40/p46 and p69/p71 (P <0.02), indicating a. similar regulation of the expression of these genes. Our data also demonstrated that pretreatment concentrations of 2-5AS mRNA correlated with responses to IFN therapy in chronic hepatitis C.
Conclusions: Our method for measuring 2-5AS mRNA concentrations could provide art important marker for selecting patients for IFN therapy and may be useful for the development of more effective therapeutic strategies for chronic hepatitis C. (C) 2002 American Association for Clinical Chemistry.

Genome aberrations observed by restriction landmark genome scanning analysis of chromosomal DNA in various types of primary hepatocellular carcinoma Reviewed

T Hanafusa, Y Yumoto, T Mimura, T Shinji, H Hada, N Koide, T Tsuji

HEPATO-GASTROENTEROLOGY 49 ( 44 ) 482 - 486 2002.3

Authorship：Lead author Language：English Publishing type：Research paper (scientific journal) Publisher：H G E UPDATE MEDICAL PUBLISHING S A

Background/Aims: The genes responsible for hepatocellular carcinoma have not been identified. To identify the relevant genes of hepatocellular carcinoma, detailed and comprehensive information of genomic aberrations must be obtained. To reveal the chromosomal aberrations in hepatocellular carcinoma, we carried out a restriction landmark genome scanning analysis of various types of hepatocellular carcinoma.
Methodology: Samples of various types of hepatocellular carcinoma, including two with multinodular-hepatocellular carcinomas, one hepatocellular carcinoma showing nodules in a nodule pattern, one hepatocellular carcinoma metastasized to different tissues, three small (<2.0 cm) hepatocellular carcinomas and four large (>5.0 cm) hepatocellular carcinomas were examined by the restriction landmark genome scanning method with corresponding nonhepatocellular carcinoma tissues. Restriction enzyme (N) under bar(o) under bar(t) under bar I was used as a landmark enzyme, (E) under bar(c) under bar(o) under bar RV was used as a fragmentation enzyme, and (H) under bar(i) under bar(n) under bar fI was used as a digestion enzyme in the gel for two-dimensional electorophoresis.
Results: We observed spot aberrations with different origins. Frequently observed spot aberrations originated from the change in the methylation status of repetitive sequences. No clear correlation between the pathological grade and the number or type of spot aberrations was observed.
Conclusions: We demonstrated that major aberrations of restriction landmark genome scanning spots originated from the change of methylation status in hepatocellular carcinoma.

Serum gamma-interferon-inducing factor (IL-18) and IL-10 levels in patients with acute hepatitis and fulminant hepatic failure Reviewed

E Yumoto, T Higashi, K Nouso, H Nakatsukasa, K Fujiwara, T Hanafusa, Y Yumoto, T Tanimoto, M Kurimoto, N Tanaka, T Tsuji

JOURNAL OF GASTROENTEROLOGY AND HEPATOLOGY 17 ( 3 ) 285 - 294 2002.3

Language：English Publishing type：Research paper (scientific journal) Publisher：BLACKWELL PUBLISHING ASIA

Background and Aims: The aim was to determine the role of T-helper (Th)1/Th2 cytokine responses in the clinical outcome of patients with acute liver injury.
Methods: The serum levels of the cytokines, interleukin (IL)-18, gamma-interferon (IFN-gamma), IL-10 and IL-4 were measured in 20 fulminant hepatic failure (FHF), 18 acute hepatitis (AH), 30 chronic viral hepatitis and 20 liver cirrhosis (LC) patients. Thirteen cases were from the intensive care unit (ICU) and there were 21 healthy volunteers. Immunohistochemical staining of liver biopsies for IL-18 expression was also performed.
Results: Serum IL-18 levels in patients with FHF were significantly more elevated than in patients with other liver diseases, ICU cases and healthy volunteers. Furthermore, serum IFN-gamma levels in patients with FHF were also significantly higher than in patients with chronic viral hepatitis, LC and healthy volunteers. We found a positive correlation between the levels of IL-18 and IFN-gamma. However, no relationship was observed between these and clinical outcome. In immunohistochemical staining, CD68+ macrophage cells and IL-18-positive cells were observed in portal zones. Elevated serum IL-10 levels were restricted to patients presenting with FHF, and were significantly higher in surviving cases (P < 0.01). Furthermore, serum IL-10 levels, but not IL-4 levels, were inversely correlated with serum total bilirubin concentrations (P = 0.045) and the death rate (p) outlined in Japan (P = 0.030).
Conclusion: These results suggest that IL-18 and IFN-gamma are involved in the pathogenesis of acute hepatic injury in humans, and that, in particular, elevated serum levels of IL-10 may be predictive of improved outcomes for these patients. (C) 2002 Blackwell Science Asia Pty Ltd.

DOI： 10.1046/j.1440-1746.2002.02690.x

Reduced expression of insulin-like growth factor binding protein-3 and its promoter hypermethylation in human hepatocellular carcinoma Reviewed

T Hanafusa, Y Yumoto, K Nouso, H Nakatsukasa, T Onishi, T Fujikawa, M Taniyama, S Nakamura, M Uemura, Y Takuma, E Yumoto, T Higashi, T Tsuji

CANCER LETTERS 176 ( 2 ) 149 - 158 2002.2

Authorship：Lead author Language：English Publishing type：Research paper (scientific journal) Publisher：ELSEVIER SCI IRELAND LTD

Insulin-like growth factor binding protein-3 (IGFBP-3) is postulated to be a mediator of growth suppression signals. Reduced expression Of the IGFBP-3 was observed in nine out of 12 human hepatocellular carcinomas (HCC) (75%). Promoter hypermethylation of the IGFBP-3 was detected in four out of 12 HCCs (33%) although mutations were not identified. The expression of IGFBP-3 was restored by the demethylating agent 5-aza-2'-deoxycytidine in HCC cell line with promoter hypermethylation (HepG2). As IGFBP-3 functions like a tumor suppressor gene, it may be used as a therapeutic target for HCC. (C) 2002 Elsevier Science Ireland Ltd. All rights reserved.

DOI： 10.1016/S0304-3835(01)00736-4

Replication of hepatitis B virus which carries foreign DNA in vitro Reviewed

Tadashi Hanafusa, Yasuhiro Yumoto, Hajime Hada, Toshiyuki Shinji, Norio Koide, Takao Tsuji

Biochemical and Biophysical Research Communications 262 ( 2 ) 530 - 533 1999.8

Authorship：Lead author Language：English Publishing type：Research paper (scientific journal) Publisher：Academic Press Inc.

Targeting a specific DNA sequence to the desired tissues is an important step in gene therapy. The hepatitis B virus (HBV) is the only DNA virus that has hepatocyte specificity. We attempted to construct an HBV-based vector for targeting the liver. We observed the replication and secretion of virus particles in an HBV construct that lacks X gene and carries an extra 63 bp DNA fragment in vitro. Replication was observed in the cell line HuH-7 but not HepG2. From this construct, we designed an HBV-based vector that could carry foreign DNA. HBV based vectors provide for the possibilities of generating therapeutic agents for individual patients. Our host vector system may be used to clear out the HBV from the HBV carrier or chronic hepatitis B patients by introducing a genetically engineered HBV into these patients.

DOI： 10.1006/bbrc.1999.1224

Expression of the midkine gene in human hepatocellular carinomas. (jointly worked) Reviewed

Koide N, Hada H, Shinji T, Ujike K, Hirasaki S, Yumoto Y, Hanafusa T, Kadomatsu K, Muramatsu H, Muramatsu T, Tsuji T

Hepato-Gastroenterology 46 ( 30 ) 3189 - 3196 1999.4

Volume reduction by the incineration of the combustible radioactive solid samples from radioisotope usase at the utilization facility : Estimation of the distribution of low energy β emitter using the imaging plate. (jointly worked) Reviewed

Yumoto Y, Hanafusa T, Nagamatsu T, Okada S

Radioisotopes 48 ( 5 ) 320 - 326 1999.4

Language：English Publisher：Japan Radioisotope Association

We want to establish a system of volume reduction by the incineration of the combustible radioactive solid wastes from radioisotope usage at the utilization facility. We have been performing experiments using an experimental incineration system to examine the distribution of radionuclides during incineration and to collect basic data. To reproduce the realistic conditions of incineration of low-level radioactive wastes in an experimental system, we adopted new incineration methods in this study. Low level radioactive samples (LLRS) were set up in a mesh container of stainless steel and incinerated at high temperature (over 800°C) generated by two sets of high calorie gas burners. Low energy β-emitters ³⁵S, ⁴⁵Ca, ³³P, and a high energy β-emitter ³²P were used for the experiment. Their translocation percentages in exhaust air and dust were estimated using the Imaging Plate. Distribution of radionuclides during the incineration was similar to that estimated by conventional methods by our study or to that reported in incineration of liquid scintillation cocktail waste. We concluded that the use of the Imaging Plates is a simple and reliable method for estimation of the distribution of low energy β-emitters in incineration gas and ash.

DOI： 10.3769/radioisotopes.48.320

Other Link： https://jlc.jst.go.jp/DN/JALC/00060829443?from=CiNii

Point mutations in the S and pre-S2 senes observed in two hepatitis B virus carriers positive for antibody to hepatitis B surface antigen. (jointly worked) Reviewed

Shinji T, Koide N, Hanafusa T, Hada H, Oka T, Takayama N, Shiraha H, Nakamura M, Ujike K,Yumoto Y, Tsuji T

Hepato Gastroenterology 45 ( 20 ) 500 - 502 1998.4

Reciprocal Gene Expression of Rat Fibro glycan and β-actin during the coruse of Regeneration after-D-Galactosamine Liver Injury (jointly worked) Reviewed

Shinji T, Koide H, Hada H, Sasaki S, Oka T, Takayama N, Shiraha H, Ujike K, Nakamura M, Hanafusa T, Yumoto Y, Tsuji T

Hepato・Gastroenterdogy 44 239 1997.4

Setting up Experimental Incineration System for Low-level Radioactive Samples and Combustion Experiments. Reviewed

YUMOTO Yasuhiro, HANAFUSA Tadashi, NAGAMATSU Tomohiro, OKADA Shigeru

RADIOISOTOPES 46 ( 7 ) 443 - 449 1997

Language：Japanese Publisher：Japan Radioisotope Association

An incineration system was constructed which were composed of a combustion furnace (AP-150R), a cyclone dust collector, radioisotope trapping and measurement apparatus and a radioisotope storage room built in the first basement of the Radioisotope Center.<BR>Low level radioactive samples (LLRS) used for the combustion experiment were composed of combustible material or semi-combustible material containing 500 kBq of ^99mTcO₄ or 23.25 kBq of ¹³¹INa. The distribution of radioisotopes both in the inside and outside of combustion furnace were estimated. We measured radioactivity of a smoke duct gas in terminal exit of the exhaust port. In case of combustion of LLRS containing ^99mTcO₄ or ¹³¹INa, concentration of radioisotopes at the exhaust port showed less than legal concentration limit of these radioisotopes. In cases of combustion of LLRS containing ^99mTcO₄ or ¹³¹INa, decontamination factors of the incineration system were 120 and 1.1, respectively.

DOI： 10.3769/radioisotopes.46.443

Promoter-independent loss of mRNA and protein of the Rb gene in a human hepatocellular carcinoma Reviewed

H Hada, N Koide, T Morita, H Shiraha, T Shinji, M Nakamura, K Ujike, N Takayama, T Oka, T Hanafusa, Y Yumoto, K Hamazaki, T Tsuji

HEPATO-GASTROENTEROLOGY 43 ( 11 ) 1185 - 1189 1996.9

Language：English Publishing type：Research paper (scientific journal) Publisher：H G E UPDATE MEDICAL PUBL LTD.

Background: Inactivation of the retinoblastoma (Rb) gene is considered to play a fundamental role in the genesis and progression of several human cancers. In retinoblastoma, the inactivation of Rb promoter by mutations or hypermethylation has been reported. Although genetic changes of Rb gene have been described in hepatocellular carcinoma (HCC) an, epigenetic change such as hypermethylation of the Rb promoter as reported in, retinoblastoma has not been described.
Materials and Methods: We examined the hypermethylation in the promoter region of Rb gene by restriction fragment length polymorphism in 19 HCCs, as well as the expression of Rb mRNA and protein by RT-PCR and by immunoblotting, respectively.
Results: We found no evidence of hypermethylation. in the promoter region, of the Rb gene in all HCCs analyzed. However, the expression of Rb mRNA and protein was lost in one HCC, and no mutation was detected in the Rb promoter region of this patient. The inactivation of Rb promoter by hypermethylation or by inhibition of binding of transcription factors due to point mutations did not contribute to the loss of mRNA and protein in. the patient.
Conclusions: Hypermethylation in the Rb promoter region appeared to have little causal effect on HCC.

Loss of heterozygosity and analysis od mutation of p53 in hepatocellular carcinoma (jointly worked) Reviewed

Yumoto Y, Hanafusa T, Hada H, Morita T, Ooguch S, Shinji T, Mitani T, Hamaya K, Koide N, Tsuji T

Journal of Gastroenterology and Hepatology 10 179 - 185 1995.4

Classification of hepatitis C viras into subgroups on the basis of Sequence variations in the envelope protein (jointly worked) Reviewed

Hada H, Koide N, Hanafusa T, Takabatake H, Sakaguchi K, Shinji T, Sasaki S, Oka T, Takayama N, Yumoto Y, and Tsuji T

Journal of Gastroenterology and Hepatology 8,70-74 70 - 74 1993.4

Isolation and characterization of DBEscherichia(/)-DB DBcoli(/)-DB DBhag(/)-DB operator mutants whose hag-48 expression has become repressible by a DBSalmonella(/)-DB DBHl(/)-DB repressor (jointly worked) Reviewed

Hanafusa T, Sakai A, Tominaga A, Enomoto M

Mol. Gen. Genet. 216 1993.4

Nucleotide sequence and regulated expression of the Salmonella fljA gene encoding a repressor of the phase 1 flagellin gene Reviewed

Tadashi Hanafusa, Kenji Saito, Akira Tominaga, Masatoshi Enomoto

MGG Molecular & General Genetics 236 ( 2-3 ) 260 - 266 1993.1

Authorship：Lead author Language：English Publishing type：Research paper (scientific journal) Publisher：Springer-Verlag

The nucleotide sequence of Salmonella abortus-equi fljA, which together with the phase 2 flagellin gene constitutes the fljBA operon and encodes the repressor for the phase 1 flagellin gene fliC, was determined. The repressor was predicted to be a basic protein consisting of 179 amino acid residues (Mr = 20419 Da) encoded by ORFII. This was confirmed by the fact that host fliC is repressed by plasmid-encoded ORFII, which indeed expresses a 20 kDa product as determined by urea SDS-polyacrylamide gel electrophoresis. An amino acid sequence capable of forming a helix-turn-helix type of structure was predicted in the C-terminal region of FljA. A rho-independent intercistronic terminator was detected between fljB and ftjA. Chloramphenicol acetyltransferase (CAT) assays of fusions indicated that the terminator is capable of reducing expression of fljA to the level of a few percent, relative to fljB in broth cultures and to 1 % in M9 glycerol cultures. © 1993 Springer-Verlag.

DOI： 10.1007/BF00277121

Integration of SCPl, a giant linear plasmid, into the DBStreptomyces(/)-DB DBcoelicolor(/)-DB chromosome (jointly worked) Reviewed

Kinashi H, Shimaji-Murayama M and Hanafusa T

Gene 115 35 - 41 1992.4

The structure of an integrated copy of the giant linear plasmid SCPl in the chromosome of DBStreptomyces(/)-DB DBcoelicolor(/)-DB 2612 (jointly worked) Reviewed

HANAFUSA T.

Mol. Gen. Genet. 231 363 - 368 1992.4

Detection by western blotting of an antibody to the hepatitis C virus E1 envelope protein in sera of patients with chronic liver disease.

H. Hada, N. Koide, T. Hanafusa, K. Sakaguchi, T. Shinji, S. Sasaki, T. Oka, N. Takayama, Y. Yumoto, T. Tsuji

Acta medica Okayama 46 ( 5 ) 365 - 370 1992

Language：English Publishing type：Research paper (scientific journal)

We detected an antibody to HCV envelope protein (E1) in sera of patients with HCV-related chronic liver diseases (20 patients with chronic hepatitis and 5 patients with liver cirrhosis) by Western blotting using the fusion protein of E1 envelope protein and beta-galactosidase as an antigen. The antibody to HCV E1 (anti-HCV E1) was detected in 8 (42%) of 19 patients positive for HCV-RNA (16 were positive and 3 were negative for antibody to C100-3) and in 1 (17%) of 6 patients negative for HCV-RNA but positive for antibody to C100-3. HCV-RNA was detected in 8 (89%) of 9 anti-HCV E1 positive sera. The value of alanine aminotransferase was significantly higher in patients positive for anti-HCV E1 than in patients negative for the antibody. Although an antibody to the envelope protein of HCV is suspected to be one of the candidates of virus-neutralizing antibodies, our results suggest this hypothesis appears to be unlikely.

Sequence Variations in the Envelope Protein of the Hepatitis C Virus : Comparison with Partial cDNA Sequence of a New Variant Virus Obtained by the Polymerase Chain Reaction (jointly worked) Reviewed

Hada H, Koide N, Takabatake H, Hanafusa T and Tsuji T

Acta Med Okayama 45 ( 5 ) 347 - 355 1991.4

Nucleotide sequence analysis of the unusually long terminal inverted repeats of a giant linear plasmid, SCPl (jointly worked) Reviewed

Kinashi H, Shimaji-Murayama M and Hanafusa T

Plasmid 26 123 - 130 1991.4

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