2000-2001

Aichi Cancer Center
Research Institute
Scientific Report
2000-2001
Chikusa-ku, Nagoya 464-8681
Japan

(The Cover)
An aerial photograph of Aichi Cancer Center
campus and buildings, standing in the setting
of the luxuriant verdure of the Kano-ko
Garden Park on the shore of Lake
Neko-ga-hora.
Published by
Dr. Toshitada Takahashi
Director
Aichi Cancer Center Research Institute
1-1 Kanokoden, Chikusa-ku, Nagoya 464-8681, Japan
Telephone: 052-762-6111
Facsimile: 052-763-5233
Editorial Committee
Dr. Reiji Kannagi, Chief (Division of Molecular Pathology)
Dr. Tetsuo Kuroishi (Division of Epidemiology and Prevention)
Dr. Hirotaka Osada (Division of Molecular Medicine)
Dr. Masatoshi Fujita (Division of Virology)
Dr. Hiroshi Kumimoto (Central Laboratory & Radiation Biology)
Mr. Morio Terashima, Photographer (Central Service Unit)
Dr. Malcolm A. Moore, English Editor
Printed by
Nagoya University COOP
1 Furoucho, Chikusa-ku, Nagoya 464-0814, Japan

Contents
______________________________________________________________________
Preface
Takahashi Toshitada 1
Organization of the Aichi Cancer Center Research Institute 2
SCIENTIFIC REPORTS
Division of Epidemiology and Prevention
General summary 5
1. Descriptive epidemiologic studies on cancer incidence and mortality
Inoue, M., Kuroishi, T., Hirose, K. Tominaga, S. and Tajima, K. 5
2. HERPACC studies on risk and protective factors for main sites of cancer
Hirose K., Saito, T., Inoue, M., Takezaki, T., Hamajima, N., Kuroishi, T.,
Tajima, K., Miura, S., Kuzuya, K., Sugiura, T., Mitsudomi, T., Okuma, K.,
Ogawa, H., Nishiwaki, K., Sakai, S., Yanagi, T., Ariyoshi, Y., Stellman, S.,
Wynder, E. and Aoki, K. 7
3. Development of cancer prevention programs
Hamajima, N., Hirose, K., Tajima, K. and Miura S. 10
4. Evaluation of secondary prevention of cancers
Kuroishi, T., Hirose,K., Suzuki, T., Tominaga, S., Sagawa, M., Fujimura,
S. and The Research Group for Lung Cancer Screening 11
5. Ethnoepidemiological studies on cancer
Takezaki, T., Hamajima, N., Inoue, M., Hirose, K., Tajima, K., Gao, C-M.,
Wu, J-Z., Wang, Y-M., TMo, B-Q., Wang, X-R., Yoo, K-Y., Ahn, Y-O.,
Kim, J-S., Zhou, Z-Y., Cao, J., Li C., Gao, F-C., Tokudome, Y., Sonoda,
S., Yashiki, S., Fujiyosh, T., Li, H-C., Zhao, S-H., Horai, S., Chiba, H.,
Senoh, H. and Tretli, S. 12
Division of Oncological Pathology
General summary 17
1. Reversibility of Heterotopic Proliferative Glands in Glandular Stomach
of Helicobacter pylori-infected Mongolian Gerbils on Eradication
Tatematsu, M., Nozaki, K., Shimizu, N., Tsukamoto, T., Inada, K., Cao, X.,
Ikehara, Y., Kaminishi, M. and Sugiyama, A. 18
2. Real-time observation of micrometastasis formation in the living mouse
liver using a GFP gene-tagged rat tongue carcinoma cell line
Nakanishi, H., Itoh, H., Ikehara, Y., Kato, T., Nakao, A. and Tatematsu, M. 18
3. Intestine specific homeobox genes, Cdx1 and Cdx2, are key molecules
in intestinalization of gastric carcinoma cells
Inada, K., Mizoshita, T., Tsukamoto, T., Nakanishi, H. and Tatematsu, M. 19
4. Different susceptibilities of p53 knockout (-/-), (+/-) and (+/+) mice to
induction of stomach adenocarcinomas by N-methyl-N-nitrosourea and
esophageal squamous cell carcinomas by methyl-n-amylnitrosamine
Tsukamoto, T., Yamamoto, M., Shirai, N., Sakai, H., Iidaka, T.,
i

ii
Donehower, L.A. and Tatematsu, M. 19
5. Polymorphisms of two fucosyltransferase genes (Lewis and Secretor genes)
involving type I Lewis antigens are associated with the presence of anti-
Helicobacter pylori IgG antibodies
Ikehara,Y., Nishihara, S., Yasutomi, H., Kitamura, T., Matsuo, K., Shimizu,
N., Inada, K., Kodera, Y., Yamamura, Y., Narimatsu, H., Hamajima, N.
and Tatematsu, M. 20
Division of Molecular Oncology
General summary 23
1. Multi-faceted analyses of a highly metastatic human lung cancer cell line
NCI-H460-LNM35 suggest mimicry of inflammatory cells in metastasis
Kozaki, K., Koshikawa, K., Osada, H., Konishi, H., Tatematsu, Y., Miyaishi,
O., Saito, H., Hida, T., Mitsudomi, T. and Takahashi, Ta. 23
2. Gene silencing by aberrant DNA methylation and abnormalities in chromatin
configuration in human lung cancer cells
Osada, H., Tatematsu, Y., Yatabe, Y., Masuda, A., Konishi, H., Harano, T.,
Nakagawa, T., Saito, T., Sugiyama, M., Yanagisawa, K., Takada, M. and
Takahashi, Ta. 24
3. Persistent increase in chromosome instability in lung cancers
Haruki, N., Masuda, A., Harano, T., Kiyono, T., Takahashi, Takao Tatematsu,
Y., Shimizu, S., Mitsudomi, T., Konishi, H., Osada, H., Fujii, Y. and
Takahashi, Ta. 25
4. Identification of frequent G2 checkpoint impairment and a homozygous
deletion of 14-3-3ε at 17p13.3 in small cell lung cancers
Konishi, H., Nakagawa, T., Harano, T., Mizuno, K., Saito, H., Masuda, A.,
Osada, H. and Takahashi, Ta. 25
5. In vitro molecular analysis of carcinogenesis of human lung adenocarcinomas
with the aim of clinical applications
Masuda, A., Konishi, H., Yatabe, Y., Hida, H., Saito, T. and Takahashi, Ta. 26
Division of Molecular Medicine
General summary 29
1. Search for MALT1-associated proteins using yeast two-hybrid strategy
Hosokawa, Y., Suzuki, H. and Seto, M. 29
2. Detection of API2-MALT1 chimeric transcripts involved in mucosa-
associated lymphoid tissue lymphomas by a single touchdown multiplex
polymerase chain reaction
Suguro, M., Suzuki, R., Nakamura, T., Suzuki, H., Hosokawa, Y.,
Nakamura, S. and Seto, M. 30
3. Detection of cyclin D1 overexpression by real-time reverse-transcriptase
mediated quantitative polymerase chain reaction for the diagnosis of mantle
cell lymphoma
Suzuki, R., Takemura, K., Tsutsumi, M., Nakamura, S., Hamajima, N. and
Seto, M. 30
4. Molecular Cytogenetic Analysis of the Breakpoint Region at 6q21-22 in
T-Cell Lymphoma/Leukemia Cell Lines
Tagawa, H., Miura, I., Suzuki, R., Hosokawa, Y. and Seto, M. 31

Division of Immunology
General summary 33
1. Production of a single chain variable fragment (scFv) antibody against type
III mutant EGFR
Yoshikawa, K., Nakayashiki, N., 2, Takasu, S., 2, Okamoto, K., Nakamura,
K., Hanai, N., Okamoto, S., Mizuno, M., Wakabayashi, T., Saga, S.,
Yoshida, J. and Takahashi, To. 33
2. Immunogenic gene products in cancer patients
Obata, Y., Takahashi, To., Tamaki, H., Tajima, K., Yoshida, M., Miura,
S., Iwase, T., Iwata, H, Mitsudomi, T., Takahashi, M., Sakamoto, J.,
Chen, Y.-T., Stockert, E. and Old, L.J. 34
3. Targeted cloning of cytotoxic T cells specific for minor histocompatibility
an-tigens restricted by HLA class I molecules of interest
Akatsuka, Y., Kondo, E., Nishida, T., Taji, H., Morishima, Y., Obata, Y.,
Kodera Y. and Takahashi, To. 34
4. Binding of thymus leukemia (TL) anti-gen tetramers to normal intestinal
intra-epithelial lymphocytes and thymocytes
Tsujimura, K., Obata, Y., Matsudaira, Y., Ozeki, S., Yoshikawa, K.,
Saga, S. and Takahashi, To. 35
Division of Virology
General summary 37
1. The Epstein-Barr Virus Pol catalytic subunit physically interacts with the
BBLF4/BSLF1/BBLF2/3 Complex.
Fujii, K., Yokoyama, N., Kiyono, T., Kuzushima, K., Fujita, M. and
Tsurumi, T. 37
2. Purification of the product of the Epstein-Barr virus BZLF1 gene
Nakasu, S. and Tsurumi, T. 38
3. Mechanisms by which Cdc2 kinase inhibits re-replication during late S-G2-M
phase in mammalian cells
Fujita, M. and Tsurumi, T. 38
4. Immortalization of Human Cells by HPV
Kiyono, T. and Tsurumi, T. 39
5. Longitudinal dynamics of Epstein-Barr Virus-specific cytotoxic T lymphocytes
in the posttransplant lymphoproliferative disorder
Kuzushima, K., Kimura, H., Hoshino, Y., Yoshimi, A., Tsuge, I., Horibe, K.,
Morishima, T., Kojima, S. and Tsurumi, T. 39
6. Identification of HLA A*2402-restricted cytomegalovirus-specific CD8 + T
cell epitopes by a computer algorithm and an enzyme-linked immunospot
assay
Kuzushima, K., Hayashi, N., Kimura, H. and Tsurumi, T. 40
Division of Molecular Pathology
General summary 43
1. Study of ligand specificity of three selectin family cell adhesion molecules, E-,
P- and L-selectins, using genetically engineered cells
Kanamori, A., Ohmori, K, Goto, Y., Uchimura, K., Muramatsu, T., Kiso, M.,
Tamatani, T. and Kannagi, R. 43
2. Expression of sialyl 6-sulfo Lewis X, a new ligand for cell adhesion molecules
iii

iv
of the selectin family, in human colon and cultured colon cancer cells
Izawa, M., Kumamoto, K., Kanamori, A., Kanda, K., Goto, Y., Ishida, H.,
Nakamura, S. and Kannagi, R. 44
3. Regulatory mechanisms for expression of functional carbohydrate
determinants on malignant and non-malignant cells:
3-1. Roles of sugar nucleotide transporters in the enhanced expression of
carbohydrate ligands for selectins, sialyl Lewis X and sialyl Lewis A,
on solid tumors
Kumamoto, K., Goto, Y., Ishida, N., Kawakita, M. and Kannagi, R. 45
3. Regulatory mechanisms for expression of functional carbohydrate
determinants on malignant and non-malignant cells:
3-2. A T-box transcriptional factor that synergizes with HTLV-1 Tax in
transactivating the selectin-ligand synthesizing enzyme, fucosyltrans-
ferase VII
Hiraiwa, N., and Kannagi, R. 46
4. A murine model of tumor suppression by vaccination with MUC1 DNA and
dendritic cells
Kontani, K. and Taguchi, O. 46
Division of Biochemistry
General summary 49
1. Aurora B and Rho-kinase regulate cleavage furrow-specific vimentin
phosphorylation in the cytokinetic process
Yasui, Y., Goto, H., Kawajiri, A., Nigg, E.A., Terada, Y., Tatsuka, M.,
Matsui, S., Manser, E., Lim, L., Nagata, K. and Inagaki, M. 49
2. Aurora B phosphorylation of histone H3 at serine28 prior to the mitotic
chromosome condensation
Goto, H., Yasui, Y., Nigg, E.A. and Inagaki, M. 50
3. Keratin attenuates tumor necrosis factor-induced cytotoxicity through
association with TRADD
Inada, H., Izawa, I., Nishizawa, M., Fujita, E., Kiyono, T., Takahashi, T.,
Momoi, T. and Inagaki, M. 50
4. ERBIN associates with p0071, an armadillo protein, at cell-cell junctions
of epithelial cells
Izawa, I., Nishizawa, M., Tomono, Y., Ohtakara, K., Takahashi, T. and
Inagaki, M. 51
5. Characterization of a mammalian septin MSF-A
Nagata, K., Kawajiri, A., Saito, N., Togashi,H., Takagishi, M., Matsui, S.,
Hotani, H. and Inagaki, M. 51
6. Functional analysis of DREF using transgenic flies
Hirose, F., Ohshima, N., Kwon, E-J., Yoshida, H., Inoue, Y.H., Matsukage,
A. and Yamaguchi, M. 52
7. Functional analysis of BEAF32A using transgenic flies
Yamaguchi, M., Yoshida, H., Hirose, F., Inoue, Y.H., Hayashi, Y.,
Yamagishi, M., Nishi Y., Tamai, K., Sakaguchi, K. and Matsukage, A. 52
Division of Central Laboratory & Radiation Biology
General summary 55
1. Analysis of a candidate tumor suppresor gene, LATS2, on 13q11-12 in

esophageal squamous cell carcinoma
Ishizaki, K., Fujimoto, J., Kumimoto, H., Nishimoto, Y., Shimada, Y.,
Shinoda, M. and Yamamoto, T. 55
2. Different susceptibility of each L-myc genotype to risk factors for
esophageal cancer or lung cancer
Kumimoto, H., Nishimoto , Y., Hamajima, N., Matsuo, K. and Ishizaki, K. 56
3. Establishment of immortal normal and ataxia telangiectasia fibroblast cell
lines by introduction of the hTERT gene
Nakamura, H., Fukami, H., Kiyono, T. and Ishizaki, K. 57
Central Service Unit
General summary
Tanabe, K., Nakamura, H., Terashima, M., Minoura, Y., Yamamoto, M. and
Hagino, M. 59
Librarians
Adachi, K. Mori, S., Teratani, M. and Yasuda, T. 60
Researches Supported by Special Project Programme
1. Identification of tumor-associated antigens recognized by T cells infiltrating
Epstein-Barr virus-positive gastric carcinomas
Kuzushima, K., Nakamura, S., Nakamura, T., Yamamura, Y., Hayashi, N.
and Tsurumi, T. 61
Publications
1 Journals 63
2 Reviews and books 78
3. Abstract for international conferences 81
Records of seminars 85
Records of symposia 87
Author index for research reports and publications 95
v

From left to right
The first row; Dr. S. Tominaga (Director, until March 2001, President, as of April 2001) and Dr. To.
Takahashi (Director, as of April 2001)
The second row; Mrs. M. Hosokawa (Adachi) and Ms. H. Tamaki.

Preface
_______________________________________________________________________________________
First of all, let me introduce myself to you. My name is Toshitada Takahashi, promoted to be the Director
of the Institute on April, 2001, as the former director, Dr. Suketami Tominaga, M.D., M.P. H., became
President of the Aichi Cancer Center.
It is my pleasure to share with you the 17 th Scientific Report (2000-2001) of the Aichi Cancer Center
Research Institute. Since the establishment of the Research Institute in 1965, the Scientific Reports have
been published biennially to document major research activities and highlight progress in and contributions
to cancer research at the Institute.
As illustrated on the following page, the organization of the Research Institute was remodeled to be 9
Divisions, consisting of three study groups, i.e. cancer prevention/epidemiology, preclinical/ experimental
therapy, and carcinogenesis/ molecular biology. A total of 59 full-time staff members, 35 researchers and 24
research assistants, as well as 5 research residents, are now conducting a wide range of cancer studies,
together with 2 of graduate school students affiliated with Nagoya University School of Medicine, Nagoya,
and approximately 30 visiting research fellows. The major areas being pursued are as follows:
- descriptive and analytical epidemiology of cancers
- primary and secondary prevention of cancer
- molecular pathogenesis of gastrointestinal cancers
- molecular oncology of lung cancer
- molecular biology of translocation-junction genes of hematopoietic tumors
- basic studies for cancer immunotherapy
- oncogenicity and molecular biology of DNA tumor viruses
- glycobiology of cancer cells in relation to metastasis
- molecular mechanisms of cell proliferation and movement
- involvement of repair mechanisms in carcinogenesis
More detailed descriptions of the research topics of each Division appear in the contents of the report. It is
our sincere hope that the activities of the Institute will make a major contribution to elucidation of the
mechanisms of carcinogenesis and to development of clinical applications in cancer diagnosis, treatment and
prevention.
Finally, I would like to express my deep appreciation to the Aichi Prefectural Government, not only for
the continuous support received, but also for the new research building (approximately 7100m 2 ), for which
construction was completed in January, 2002. Granting support from the Ministry of Education, Science,
Sports, Culture and Technology, the Ministry of Health, Labor, and Welfare, and other related organizations
is also gratefully acknowledged.
April, 2002
Toshitada Takahashi, M.D., D.M.Sci.
Director
1

SCIENTIFIC REPORTS

Division of Epidemiology and Prevention
________________________________________________________________________________
Kazuo Tajima, M.D. Ph.D. M.P.H. Chief
Tetsuo Kuroishi, M.S. Ph.D. Section Head (until March 2002)
Nobuyuki Hamajima, M.D. Ph.D. M.P.H. Section Head
Toshiro Takezaki, M.D. D.M.Sc. Section Head (as of April 2001)
Manami Inoue, M.D. Ph.D. S.M. Senior Researcher
Kaoru Hirose, B.P., Ph.D. Senior Research Assistant
Toshiko Saito, Research Assistant
Visiting Trainees
Keitaro Matsuo, M.D. Nagoya University Graduate School of Medicine (until July, 2002)
Xinen Huang, M.D. Nagoya City University Medical School
Hidemichi Yuasa, D.D.S. Central Hospital of Tokai Medical Institute
Kosuke Amano, M.Ed. Showa University School of Medicine
Hidemi Ito, M.D. Nagoya City University Medical School (as of April 2001)
Asahi Hishida, M.D. Nagoya University Graduate Medical of School (as of October 2001)
Chika Nozaki, M.D., Nagoya University Graduate Medical of School (as of April 2001)
Lucy Sayuri Ito, M.D. Santa Cruse Hospital, Sao Pauro, Brazil (July 2000-June 2001)
Chuanxia Yang, M.D. West China University of Medical School, Chengdu, China (April 2001-March 2002)
General Summary
The current research activities of the Laboratory of Epidemiology and Prevention cover the following five
subjects: 1) descriptive epidemiological studies of cancer incidence and mortality, with special reference to
improvement of Aichi Prefectural Cancer Registry; 2) development of the hospital-based epidemiologic research
program at Aichi Cancer Center (HERPACC) to determine risk and protective factors for main sites
of cancer, with special reference to molecular epidemiologic studies on environmental and host-specific factors;
3) development of a cancer prevention program for the general populace according to "Healthy People
in Japan 21st, Aichi"; 4) evaluation studies of mass screening programs for main sites of cancer in Japan; 5)
ethnoepidemiologic studies in the Asian-Pacific area.
Descriptive epidemiologic studies include data from a nation-wide vital statistics, “Vital Statistics Japan”.
At the end of 2000 a large-scale HERPACC study was completed and a more advanced version, HER-
PACC-II for molecular epidemiology, was started to clarify gene-environment interactions for modification
of carcinogenicity. For prevention measures against cancers, primary prevention trials were conducted for
control of the smoking habit and obesity. Secondary prevention programs were evaluated by orthodox epidemiological
methods.
To promote cancer control programs in the three Northeast Asian countries, a unique international collaborative
study, KOJACH (Korea, Japan and China), was planned in 2000 and a common nutritional and
molecular epidemiologic study of gastro-intestinal cancers is now running in Nagoya, Seoul, Nanjing,
Chongqin and Benxi. Furthermore, ethnoepidemiological studies on tumor viruses (HTLV and HBV) among
Mongoloids were continued for Tibetans in China and Sahme in Northland. Cancer prevention is the final
goal of epidemiological studies. Recently we are concentrating attention on molecular epidemiology to clarify
interaction between host-specific characters and lifestyle exposure to risk factors, with regard to actual
function of metabolic and detoxifying enzymes associated with genetic polymorphisms.
1. Descriptive epidemiologic studies on
cancer incidence and mortality
Inoue, M., Kuroishi, T., Hirose, K. Tominaga, S. *1 and
Tajima, K.
Probability of developing cancer over the
whole life span of a Japanese: To obtain a relevant
index of the impact of cancer on the Japanese
population, considering curable cases as well as
mortalities, the probability of developing cancer in
the entire life span of a Japanese was estimated.
Data sources were the estimates of cancer incidence
in Japan for 1994 by the research group for Population-based
Cancer Registration in Japan, and death
5

statistics of all sites and major sites of cancer for
1994 and 1996 obtained from the vital statistics of
Japan published by the Ministry of Health and
Welfare. Two methods were employed for estimation,
one based on incidence / death (I/D) ratios of
cancer and the other on the cumulative risk. The I/D
ratio method gave a lifetime probability of developing
cancer in any site of 58 % for males and 51 %
for females in 1996. With the cumulative risk, the
values up to 85+ years of age were 52 % in males
and 31 % in females, and for the average life
expectancy of Japanese, 77 years old for males and
84 years old for females, 32 % and 26 %
respectively. The estimated probabilities provide
reasonable and practical indices of the impact of
cancer today, allowing for the limitations of both
methods. This approach can be also applied to local
estimation if population-based cancer registry data
are available.
Estimation of the cancer incidence in Aichi
prefecture: Use of a model area with good quality
registry data: In Japan, population-based cancer
registries are organized by local government
and the quality of the registration remains modest,
mainly due to the voluntary-based operations without
legal restrictions and the insufficient budget.
The population of Aichi prefecture is estimated to
be seven million, and therefore the registry of the
6
prefecture covers a relatively large population.
However, its quality has not reached the level required
internationally, for the above-stated reasons,
and the derived incidences tend to be underestimates.
On the other hand, there is a geographically
continuous area, “Central Aichi”, with a good quality
of registry data, covering a sufficient population,
including both urban and rural areas. The present
study was aimed at trying to estimate the total cancer
incidence of Aichi Prefecture, using this good
quality area as a model.
The materials were data on cancer incidence and
deaths in 1990-1998 in this central area of Aichi
prefecture, with a population of approximately one
million, under the jurisdiction of four public health
centers covering nine municipalities. The DCN (%)
for all sites was around 14%, which was lower than
that of other representative registries in Japan, even
with the same population size. The Incidence /
Death ratio was around 1.9 in Central Aichi. Estimated
age-standardized incidences were found to be
around 270 (per 100,000) for males and 180 for females,
these values being high compared with those
estimated using data for the whole area of the prefecture,
but quite close to incidences for other areas
with the highest quality of data (Figure 1). Our
model area has typical demographic features of Aichi
prefecture. Therefore, it is suggested that the
Figure 1. Age-standardized cancer incidence rates per 100,000 population in Central Aichi, Aichi and all-Japan (standard
population: world population).

Figure 2. Trends in the age-standardized death rates per 100,000 population from cancer for selected sites in Japan,
1950-1999 (standard population: Japanese population in 1985).
cancer incidence in the prefecture is indeed being
underestimated and that the actual figures may be
closer to the present estimates.
Trends in cancer mortality in Japan: Cancer
mortality statistics in Japan (1950-1999) were calculated
from the 'Vital Statistics, Japan' series. The
age-standardized death rate for all sites of cancer
has gradually been increasing in males in recent
years, and gradually decreasing in females (Figure
2). The mortality rate from stomach cancer has been
decreasing in males and females, while that from
lung cancer has been increasing. In 1999 the number
of deaths from lung cancer was 37,934 for men,
accounting for 21.6% of all male cancer deaths, and
14,243 for women, accounting for 12.4 % of all female
cancer deaths, and the total number of these
deaths in both sexes ranked top among all sites of
cancer. The average age at death from all sites of
cancer in males rose 11.1 years over the last 45
years (70.7 years old in 1995 vs. 59.6 years in
1950), and 13.7 years in females (71.4 vs. 57.7).
*1 Director
2. HERPACC studies on risk and protective
factors for main sites of cancer
Hirose, K., Saito, T., Inoue, M., Takezaki, T., Hamajima,
N., Kuroishi, T., Tajima, K., Miura, S. *1 , Kuzuya, K. *2 ,
Sugiura, T. *3 , Mitsudomi, T. *4 , Okuma, K. *5 , Ogawa,
H. *6 , Nishiwaki, K. *7 , Sakai, S. *8 , Yanagi, T. *9 , Ariyoshi,
Y. *10 , Stellman, S. *11 , Wynder, E. *12 and Aoki, K. *13
Chronic atrophic gastritis and subsequent
gastric cancer: Gastric cancer is still one of the
most common cancers in Japan. Chronic atrophic
gastritis is regarded as an important factor, associated
with Helicobacter pylori infection. The purpose
of the present study was to elucidate chronological
change in cumulative risk of gastric cancer
occurrence with various degrees of chronic atrophic
gastritis by long-term follow-up of a large-scale
cohort. A total of 5,373 subjects without cancer
who underwent gastroscopic examination and completed
a life-style questionnaire at Aichi Cancer
Center between 1985-1989 were prospectively followed
until December 1999, by mail survey, hospital
record, and hospital- and population-based cancer
registry. Relative risks of gastric cancer associated
with baseline endoscopic findings were esti-
7

Figure 3. Hazard ratio for gastric cancer by baseline
endoscopic findings (n=5,373)
mated using hazard ratios and their 95% confidence
intervals with the Cox proportional hazard model,
adjusting for gender, age and gastric cancer family
history. After an average of 10 years of follow-up,
117 gastric cancer cases were identified. The risk
was greatest among the subjects with moderate atrophy
at baseline (hazard ratio=2.2) (Figure 3), especially
after 4-6 years of follow-up (hazard ratio=4.6-5.0).
After this time point, risk attenuation
with the length of follow-up period was observed.
Our study suggests the possibility that incomplete
chronic atrophic gastritis and the processes occurring
in atrophy are associated with development of
gastric cancer.
Risk factors for gastric cancer among Japanese
postmenopausal women: analysis by subsite
and histological subtype: To clarify whether reproductive
factors have an impact on gastric cancer
in Japanese females, a case-control study was conducted
using data from the HERPACC. The study
subjects comprised 365 postmenopausal women
with gastric cancer and 1,825 age-class frequency-
matched non-cancer outpatients presenting at Aichi
Cancer Center in 1988-1998. Cases were further divided
with regard to the anatomic subsites (upper
third, middle third, lower third) and histological
subtypes (differentiated, non-differentiated), and
associations were evaluated using odds ratios (ORs)
estimated by the logistic regression model, adjusting
for potential confounding factors. A high body
weight and corresponding body mass index at age
20 moderately increased the risk of gastric cancer,
especially for middle third and non-differentiated
cancers. Risk fluctuation with early or late age at
menarche and menopause and total duration of fertility
was not consistent. Individuals with a high age
at first parity tended to show a decreased risk of
8
cancer, irrespective of the subsite or histological
subtype. ORs were decreased with a short average
period of breastfeeding, especially for upper third
and non-differentiated cancers. From these results,
however, it appears that height, weight, menstrual
and reproductive factors have less impact on gastric
cancer than environmental factors such as smoking
and dietary habits or a family history of gastric
cancer.
Protective and risk factors for hormone related
cancer in women: To confirm the protective
effects of regular exercise on female hormone related
cancers, we undertook a case-referent comparative
study using HERPACC data. The case
group consisted of 2,367, 222 and 149 women who
had first been diagnosed as having breast, endometrial
and ovarian cancers. The referents were
24,620 female first-visit outpatients who had not
previously been diagnosed with any type of cancer.
The odds ratios (ORs) and their 95% confidence
intervals (95%CI) were estimated using an unconditional
logistic regression model.
From the cross analysis between frequency of
exercise and other selected factors among noncancer
referents, more frequent exercise was associated
with eating more fruits and vegetables and with less
cigarette smoking. This result indicated that persons
continuing regular exercise for health are much
concerned about their health maintenance and they
usually contrive to improve their life style. To clarify
the independent effects of regular exercise for
health on the risk reduction, we adjusted for the influence
of other related factors. Regular exercise
showed a negative association with breast cancer.
When women without regular exercise were referenced,
the adjusted ORs were 0.89 (95%CI:
0.75-1.05) for women exercising 3-4 times per
Figure 4. Adjusted ORs for hormone related cancers
according to level of exercise

month and 0.71 (95%CI: 0.62-0.81) for twice a
week or more. The downward trend in the risk of
breast cancer with regular exercise was statistically
significant (P10 times
higher than the OR of 3.5 for current smokers in
Japanese relative to hospital controls and six times
higher than in Japanese relative to community controls
(OR = 6.3). There were no substantial differ-
Figure 5. Consumption of cooked/raw fish and the odds
ratio (OR) for lung cancer by sex, with reference to
the histological type: adenocarcinomas (AD), and
squamous cell and small cell carcinomas (SQ &
SCC).
ences in the mean number of years of smoking or
average daily number of cigarettes smoked between
United States and Japanese cases or between United
States and Japanese controls, but American cases
began smoking on average 2.5 years earlier than
their Japanese counterparts.
To investigate risk modification for lung cancer
with diet in Japanese, we also conducted a
case-control study and evaluated variation in influence
with the histological type at Aichi Cancer
Center Hospital. We recruited 367 male and 240
female cases with adenocarcinomas, 381 male and
57 female cases with squamous cell and small cell
carcinomas, and 2964 male and 1189 female cancer-free
outpatients as controls. We found decreased
ORs for adenocarcinomas in both males
(OR=0.51) and females (OR=0.48) who consumed
cooked/raw fish (Figure 5), but not dried/salted fish
at the highest quartile frequency, compared with the
lowest. Decreased ORs for squamous cell and small
cell carcinomas were observed in males with frequent
consumption of raw and green vegetables,
fruit and milk. This study suggests cooked/raw fish
consumption lowers the risk of adenocarcinoma of
the lung in Japanese.
These findings partially support our hypothesis
that smoking and dietary habits contribute to differences
in lung cancer mortalities between the two
countries.
Gene-environmental interactions for cancers:
Genetic polymorphisms may modify the effects of
environmental risk factors on cancer occurrence.
We launched a comprehensive epidemiologic project,
HERPACC-II, including both lifestyle and
polymorphism data, following HERPACC-I which
solely concentrated on lifestyle. As of December
2001, about 5,300 samples of DNA were stored to
conduct case-control studies. Genotyping of about
60 polymorphisms was conducted at the Division
of Epidemiology and Prevention. Significant findings
were found for: 1) gene-environment interaction
for esophageal cancer between heavy drinking
and aldehyde dehydrogenase 2 (ALDH2); 2) interaction
for cancers of the esophagus and lung between
smoking and polymorphisms of L-myc and
NQO1; 3) altered malignant lymphoma risk with
methylenetetrahydrofalate reductase (MTHFR) and
methionine synthase (MS); 4) altered cancer risk
for breast and colorectum with B2- and
B3-adrenoceptors; 5) interactions between smoking
and two polymorphisms of interleukin 1B (IL-1B)
and myeloperoxidase (MPO) for Helicobacter pylori
infection; and 6) smoking habits with dopa-
9

mine receptor D2 (DRD2) and IL-1B. Further studies
on the interactions with polymorphisms are
continuing to be conducted, using larger sample
sizes.
The rapid progress in these polymorphism studies
has facilitated by a newly developed PCR
method, PCR-CTPP (polymerase chain reaction
with confronting two-pair primers) in our laboratory.
As shown in Figure 6, two-pair primers produce allele
specific bands for single nucleotide polymorphisms,
which allows electrophoresis directly after
PCR. This technique requires half of the cost and
time for genotyping by PCR-RFLP.
*1
Department of Breast Surgery, Aichi Cancer Center
Hospital
*2
Department of Gynecology, Aichi Cancer Center
Hospital
*3
Department of Respiratory Diseases
*4
Department of Thoracic Surgery
*5
Department of Hospital Laboratory
*6
Division of Human Science, Aichi Mizuho University
*7
Department of Respiratory Diseases, Nagoya National
Hospital
*8
Department of Respiratory Diseases, Red Cross Nagoya
First Hospital
*9
Vice Director, Red Cross Nagoya Second Hospital
*10
Director, Aichi Prefectural Hospital
*11
Division of Epidemiology, American Health Founda-
10
Figure 6. Logic of the polymerase chain reaction with confronting two-pair primers. At the 3’ ends of the inner
primers R1 and F2, the base specific to each allele is included. The difference between a-bp and b-bp
should be large enough to be distinguishable by electrophoresis.
tion, New York, US
*12
Previous President, American Health Foundation
(deceased), New York, US
*13
Emeritus President
3. Development of cancer prevention programs
Hamajima, N., Hirose, K., Tajima, K. and Miura, S. *1
Smoking cessation for hospital patients: In
order to measure smoking cessation rate among
those who visit medical facilities in Japan, a
large-scale follow-up study was conducted. Subjects
were self-reported smokers who visited a cancer
hospital, a general hospital, or one of four health
checkup facilities in 1997-98. Their smoking habits
were followed by two postal surveys. The first was
two months after the visit to hospital or attendance
at a health checkup screening, and the second was
one year thereafter. In total, 3,552 smokers participated
in the present study; 1,131 first visit outpatients
at a cancer hospital, 214 first visit outpatients
at a general hospital, and 2,207 examinees at four
health checkup facilities. The response rate for the
first follow-up varied from 57.3% to 80.2% of the
eligible participants in the six facilities, and that for
the second from 50.0 to 67.1%. When
non-respondents were classified as non-quitters, the
cessation rate two months after their participation
was 11.7% (95% confidence interval, 7.4-16.0%)

for the general hospital and 2.7% (2.1-3.5%) for the
four health checkup facilities, and that for one year
after was 9.8% (6.2-14.6%) and 6.0% (5.1-7.1%),
respectively. In the cancer hospital, the rate for
self-reported cancer patients was 74.6% (68.5-
80.0%) after two months and 51.3% (44.7-57.9%)
after one year. The smoking cessation rate was
found to be smaller in the health checkup examinees
than in the patients. Outpatients seemed to be
more amenable to smoking cessation, being a more
appropriate target for cessation programs. Based on
these baseline cessation rates, intervention studies
are now on going.
Obesity control trial for hospital patients: In
Japan, the mortality rate from female breast cancer
began to increase in 1965 and the increasing trend
has become more remarkable in recent years. Future
estimations of cancer mortality and incidence predict
that breast cancer will become the leading cancer
in Japanese women in the 21st century. A number
of risk factors for breast cancer related to reproductive
events have been established, e.g, early
menarche, nulliparity, late age at first birth and late
natural menopause. A family history of breast cancer
is associated with an overall increment of risk
around twofold and a high Body Mass Index (BMI)
increases the risk of breast cancer after menopause.
A case-referent study using HERPACC data provided
clear evidence that the risk of breast cancer in
post-menopausal Japanese women is markedly increased
by obesity.
With the change of nutrient intake after the
World War II, obesity is becoming one of the most
serious health problems in Japan, therefore, we
planned an intervention trial for obese women. After
obtaining informed consent, we recruited patients
over 30 years old with a BMI of 24 or more, a
total of 40 being randomly assigned into study
groups A (28) and B (12). Group A started the prevention
program at the entry and group B started
three months thereafter, according to the protocol.
At baseline, three, six and twelve months, participants
were checked for body size, dietary intake
and serum chemistry. It was stressed that they
should record their daily food intake and physical
activity for 3 months. Every weekend they returned
their diaries by mail and we gave them appropriate
comments by telephone or by mail after reviewing
them. This trial was designed to evaluate effectiveness
of the intervention trial in the group A during
the first 3 month by comparing with group B. After
follow up for 3 months, we observed significant
improvement in BMI and waist size. For group A
the average energy intake per day was about 2,000
Kcal at baseline and, then, 1,600 Kcal after three
months. Changes in key biomarkers (serum triglyceride,
total cholesterol and HDL cholesterol) related
to the reduction of BMI in three months were not
remarkable in the present interim analysis (Figure
7).
*1
Department of Breast Surgery, Aichi Cancer Center
Hospital
Figure 7. Comparison of the change between group A and B during first 3 months.
11

4. Evaluation of Secondary Prevention of
Cancers
Kuroishi, T., Hirose,K., Suzuki, T *1 ., Tominaga, S *2 ,
Sagawa, M. *3 , Fujimura, S.* 4 and The Research Group of
Lung Cancer Screening
In Japan the mortality rate from lung cancer has
been increasing in recent years. In 1999 lung cancer
was the commonest cause of cancer death in both
sexes.
Lung cancer screening has been conducted in
Japan mainly by chest X-ray examination plus sputum
cytology, the standard method for lung cancer
screening according to the Law of Health Services
for the Elderly. The purpose of this study was to
evaluate the effectiveness of mass screening for
lung cancer in Japan. We calculated the average
coverage-rates for lung cancer screening per year
from 1986 to 1995 for persons aged 40-69 years for
all of the 3,255 municipalities in Japan, selecting
"high coverage-rate" municipalities with average
coverage-rates of 50%, 60%, 70%, 80% or more.
Two municipalities were selected as "controls" for
each high coverage-rate municipality, and were
matched for population, national health insurance
Figure 8. Percentage change in the age-adjusted death
rate of cancer of the lung from 1986-90 to
1991-95 for 127 high coverage-rate municipalities
with an average coverage-rate of 70%
and over and for 254 comparable control municipalities.
12
rate, and the age-adjusted death rate from cancer of
the lung in the period 1986-90. We compared the
change in the age-adjusted death rate from 1986-90
to 1991-95 of the high coverage-rate municipalities
and the comparable controls. The percent reduction
in the average age-adjusted death rate from
cancer of the lung for those aged 40-69 years, target
age-groups of lung cancer screening, in the high
coverage-rate municipalities with an average coverage-rate
of 70%, 80% or more was contrasted
with the increase in control municipalities. In the
case of the high coverage-rate municipalities with
average coverage-rate of 70 % and over, the reduction
was statistically significantly greater than those
in the controls (Figure 8). These results suggest that
mass screening for lung cancer, mainly by chest
X-ray examination plus sputum cytology, can contribute
to a reduction in mortality from lung cancer.
*1
Department of Cancer Control and Statistics, Osaka
Medical Center for Cancer and Cardiovascular Diseases
*2
Director
*3
Department of Thoracic Surgery, Kanazawa Medical
College, Kanazawa, Japan,
*4
Tohoku Kosei Nenkin Hospital, Sendai, Japan
5. Ethnoepidemiological studies on cancer
Takezaki, T., Hamajima, N., Inoue, M., Hirose, K.,
Tajima, K., Gao, C-M. *1 ,Wu, J-Z. *1 , Wang, Y-M. *2 , Mo,
B-Q. *2 , Wang, X-R. *2 , Yoo, K-Y. *3 , Ahn, Y-O. *3 , Kim,
J-S. *4 , Zhou, Z-Y. *5 , Cao, J. *5 , Li, C. *6 , Gao, F-C. *6 ,
Tokudome, Y. *7 , Sonoda, S. *8 , Yashiki, S. *8 , Fujiyoshi,
T. *8 , Li, H-C. *8 , Zhao, S-H. *9 , Horai, S. *10 , Chiba, H. *11 ,
Senoh, H. *12 and Tretli, S. *13
Comparative epidemiological study on
GI-tract cancers focusing on Korea, Japan and
China (KOJACH study): China is one of the
highest risk areas for esophageal and gastric cancer
in the world. To clarify the environmental and host
factors associated with risk of esophageal and
stomach cancers, we have been conducting a comparative
epidemiological study in Jiangsu Province,
China, since 1996, focusing high and low risk-areas
for these cancers. We found frequent consumption
of garlic, in addition to other anticancer foods, to
lower the risk of these cancers in a case-control
study of a low-risk area, Pizhou, concordant with
our previous ecological findings for the general
population in high- and low-risk areas, and a
case-control study of a high-risk area. Frequent

Figure 9. Drinking habit and the odds ratio (OR)
for stomach cancer, with reference to the
hOGG1 Ser326Cys polymorphism.
consumption of these foods thus appears to be a
factor in low mortality from esophageal and stomach
cancer.
To investigate the gene and environmental interaction
for risk of esophageal and stomach cancers,
we conducted a case-control study in a
high-risk area, Huaian, recruiting 191 cases and 196
population-based controls. We selected gene polymorphisms
for cytochrome P-450 2E1 (CYP2E1),
hOGG1, GSTM1 and GSTT1, because CYP2E1 is
involved in metabolic activation of environmental
chemical carcinogens; hOGG1 encodes an enzyme
which repairs 8-hydroxyguanine adducts produced
by oxidative stress; and the GSTs are primarily responsible
for detoxication of xenobiotics. We found
a significant positive interaction between heterozy-
gous and homozygous RsaI rare alleles for CYP2E1
and ever-smoking in the odds ratio (OR) for stomach
cancer. A frequent drinking habit and pickled
vegetable consumption elevated the OR for stomach
cancers in individuals with the Cys/Cys genotype of
hOGG1, as compared to Ser/Ser and Ser/Cys carriers
(Figure 9). The GSTM1 null genotype was associated
with an increased OR for esophageal cancer,
but not for stomach cancer. A combined effect was
also observed between smoking and the GSTM1
null genotype with regard to esophageal risk. These
findings suggest that the polymorphisms are involved
in determining susceptibility to esophageal
and stomach cancer development.
To establish cancer prevention measurement in
northeastern countries bearing a historically common
cultural background, we started a case-referent
study on colorectal cancer, based on a standardized
epidemiological approach, in Korea (Seoul), Japan
(Nagoya) and China (Nanjing, Chongqing and
Benxi), the so called KOJACH Study, in 2000. First
we developed a semi-quantitative food frequency
questionnaire (SQFFQ) for the five cities according
to the Tokudome’s method (Tokudome et al, Jpn J
Clin Oncol 28: 679, 1998), and evaluated their validity
and reproducibility for further study. We
started collecting lifestyle data by standardized
questionnaire and blood samples for plasma and
DNA after obtaining informed consent from colorectal
cancer cases, hospital referents and population-based
referents. The intent is to analyze risk
and protective factors for colorectal cancer using
Figure 10. Worldwide distribution of HBV genotypes among various ethnic groups. Seventeen ethnic groups
(Usuda S, et al., 1999; Bowyer SM and Sim JGM. 2000; Nakamo T, et al., 2001; Bowyer SM, et al.,
1997; Arauz-Ruiz P, et al., 1997; Blitz L, et al., 1998; Norder H, et al., 1993) were used in the comparison,
including the Tibetans.
13

data for a total of 1,000 cases and 2,000 referents.
Immunogenetic study on Mongoloid populations:
Human T-cell leukemia virus type 1
(HTLV-1), the main cause of adult T-cell leukemia/lymphoma,
spread throughout the world but
microgeographical clusters of hyperendemicity.
Epidemiologic studies among Mongoloids showed
that HTLV-1 is highly endemic in South Japan (one
million carriers) and in the Andes district of South
America. On the other hand HTLV-II (also a risk
factor for adult T-cell leukemia/lymphoma) is
broadly distributed in the whole of South America,
except the Andes line. We now know that there are
no other HTLV-I/II clusters in the Asian Pacific
except among Aborigines in north Australia and
Melanesians in Papua New Guinea.
Hepatitis B virus (HBV), distributed throughout
the world, is classified into seven geographically
separated genotypes designated A to G. Since the
prevalence of HBV infection in isolated ethnic Tibetan
populations in China, and the HBV genotypes
involved have hitherto remained unclear, we collected
262 blood samples from four isolated villages
in east and west regions of Tibet (Photo). The
prevalence of HBV infection was estimated by EIA
for HBV Ag and HBV Ab, and the HBV genotypes
were determined by a PCR-microwell plate hybridization
method using plasma DNA. The prevalence
of HBV Ag and HBV Ab positives was
19.1% and 29.0%, respectively (Figure 10). We detected
only the C genotype, known to be a pre-
14
dominant among Mongoloid populations in Asia.
The evidence including ethnoepidemiologic findings
for HTLV-I opens doors to a new paradigm for
virus anthropology.
*1
Division of Epidemiology, Cancer Institute of Jiangsu
Province, Nanjing, China
*2
Nanjing Medical University, Nanjing, China
*3
Department of Preventive Medicine, College of
Medicine, Seoul National University, Seoul, Korea
*4
Department of Food and Nutrition in Oriental Medicine,
Semyung University, Semyung, Korea
*5
Laboratory of Molecular Toxicology, Third Military
Medical University, Chongqing, China
*6
Bengan General Hospital, Benxi, China
*7
Department of Life Science, Nagoya Bunri College
*8
Department of Virology, Faculty of Medicine, Kagoshima
University, Kagoshima
*9
Department of Blood Transfusion, Southwest Hospital,
Chongqing, P.R.China
*10
Department of Biosystems Sciences, The Graduate
University for Advanced Studies, Kanagawa
*11
Department of Laboratory Medicine, Hokkaido University
School of Medicine, Sapporo
*12
Department of Anatomy, Akita University School of
Medicine, Akita, Japan,
*13
Institute of Population-based Cancer Research, Oslo,
Norway
Photograph: Dr. Liang is collecting blood samples from local people in Eastern Tibet.

Dr. Parry J. Guilford, from University of Otago, New Zealand, giving us the lecture entitled "E-cadherin
Germline Mutations in Familial Gastric Cancer" in the 8th Aichi Cancer Center International
Symposium held on February 16, 2002 (see p. 88).
15

From left to right
First row; Mr. H. Tanaka , Dr. K. Inada, Dr. M. Tatematsu, Dr. H. Nakanishi, Dr. T. Tsukamoto and Dr. A. Tanaka.
Second row; Dr. M. Goto, Dr. X. Cao, Dr. T. Mizoshita, Dr. N. Ogasawara, Ms. C. Tomita, Ms. N. Yamada and Ms. M.
Yamamoto.
Third row; Dr Y. Tsukamoto, Dr. K. Matsumoto, Ms. S. Tokumasu, Ms. H. Ban and Ms. H. Maejima.
Inset; Dr. K. Nozaki, Dr. Y. Ikehara, Dr. N. Shirai, Dr. T. Iidaka, Ms. R. Haruta and Dr. A. Hirata.
16

Division of Oncological Pathology
________________________________________________________________________________
Tatematsu Masae, M.D. Chief
Hayao Nakanishi, M.D. Section Head
Ken-ichi Inada, M.D. Senior Researcher
Tetsuya Tsukamoto, M.D. Senior Researcher
Yuzuru Ikehara, M.D. Researcher
Sachiko Tokumasu, B.D., Research Assistant
Masami Yamamoto, D.V.M., Research Assistant
Harunari Tanaka, B.P., Research Assistant
Michiyo Tominaga, Semi-regular Employee
Nami Yamada, Semi-regular Employee
Hisayo Ban, Semi-regular Employee
Visiting Scientists
Malcolm A. Moore, Ph.D. Asian Pacific Organization for Cancer Prevention
Kato Kazuo, M.D., Fujita Health University School of Medicine
Visiting Trainees
Koji Nozaki, M.D., Research Resident
Hirofumi Yuasa, D.V.M., Tanabe Seiyaku Co., Ltd.
Takasuke Yamachika, M.D., Kita Hospital
Kiyoshi Kobayashi, D.V.M., Mitsubishi-Tokyo Pharmaceuticals, Inc.
Hiroki Sakai, D.V.M., Gifu University
Norimitsu Shirai, D.V.M., Pfizer Pharmaceuticals Inc.
Azusa Tanaka, D.D.S., School of Dentistry, Aichi-Gakuin University
Tsutomu Mizoshita, M.D., School of Medicine, Nagoya City University
Seiji Ito, M.D., Department Surgery II, Nagoya University School of Medicine
Yoshinari Mochizuki, M.D., Department Surgery II, Nagoya University School of Medicine
Takeshi Iidaka, D.V.M., Pfizer Pharmaceuticals Inc.
Yoshitaka Tsukamoto, D.D.S., School of Dentistry, Aichi-Gakuin University
Xueyuan Cao, M.D., Faculty of Medicine, University of Tokyo
General Summary
The responsibilities of the Division of Oncological Pathology include both autopsy and research activities.
From its establishment in 1965 up through the end of 2001, our laboratory performed a total of 2,477 autopsies.
Postmortem examinations are a source of valuable information on the behavior of neoplasms and their
response to therapy.
Research activities in our laboratory are divided into two main areas. The first deals with the molecular
basis of chemical carcinogenesis in the gastrointestinal tract of man, the rat, mouse and Mongolian gerbils,
along with the mechanisms regulating differentiation of stomach epithelium, especially intestinal metaplasia.
During 2000-2001, the research focused on various issues: a) heterotopic proliferative glands (HPGs) related
to Helicobacter pylori (Hp) infection, which frequently develop in the glandular stomach of infected gerbils
with slightly dysplastic change - an eradication experiment revealed the reversibility of this lesion, implying
that it is an entity distinct from a true neoplasm; b) Hp attachment to the gastric mucosa through adhesin,
which binds to Lewis b (Le(b)) or H type I carbohydrate structures - polymorphisms of the secretor (Se) and
Lewis (Le) genes, both involved in type I Le antigen synthesis, were found to alter the risk of Hp infection;
c) susceptibility of p53 nullizygote (–/–), heterozygote (+/–), and wild type (+/+) mice to N-methyl-
N-nitrosourea (MNU) gastric carcinogenesis and to methyl-n-amylnitrosamine (MNAN) esophageal carcinogenesis
- p53 may not be a direct target of MNU but rather play an important role as a gatekeeper in
mouse stomach carcinogenesis, whereas p53 mutations may be involved in the development of esophageal
SCCs induced by MNAN; d) intestine specific homeobox genes, Cdx1 and Cdx2, candidate genes for directing
intestinal development and differentiation of intestinal phenotypes in the gastrointestinal tract epithelium
- human gastric cancer tissues were examined and a positive correlation was demonstrated between
17

may reside in their capacity to attach stably to the
vessel wall rather than their potential for initial cell
arrest or subsequent growth. The system used in the
present study provides a powerful tool for analyzing
targets of various anti-metastatic agents in the sequential
process of metastasis.
* 1 Department of Gastroenterological Surgery, Aichi
Cancer Center Hospital
* 2 Department of Surgery II, Nagoya University of
Medicine
3. Intestine specific homeobox genes,
Cdx1 and Cdx2, are key molecules in
intestinalization of gastric carcinoma
cells
Inada, K., Mizoshita, T.* 1 , Tsukamoto, T., Nakanishi, H.
and Tatematsu, M.
Caudal-type homeobox genes Cdx1 and Cdx2
are candidates for directing intestinal development,
differentiation, and maintenance of the intestinal
phenotype in the human and mouse gastrointestinal
tract. The aim of this study was to assess relationships
among expression of Cdx1 and Cdx2 mRNAs,
histological classification and phenotypic expression
of carcinoma cells. Fresh human gastric cancer
tissues were collected from surgically resected
specimens from 70 patients after obtaining informed
consent. Northern hybridization was performed
against Cdx1 and Cdx2 mRNAs after extracting
RNAs. In addition, the carcinoma tissues
were evaluated both histologically and phenotypically
using mucin histochemical and immunohistochemical
methods. Neither Cdx1 nor Cdx2 expression
had any relation with the Lauren’s histological
classification (“intestinal” types: n=32, “diffuse”
types: n=38). On the other hand, Cdx1 was apparently
associated with intestinal phenotypic differentiation
both in the “intestinal” and “diffuse” type
{gastric phenotype: n=15, gastric and intestinal-mixed
phenotype: n=18, intestinal phenotype:
n=17, unclassified (null) type: n=20, p< 0.05}.
Cdx2 was also related with the phenotypically intestinal
gastric cancers only in intestinal type of
Lauren’s histological classification. The results
suggest that both Cdx1 and Cdx2 might play important
roles in expression of the intestinal phenotype,
not only in the normal intestine but also in
gastric neoplasms.
* 1 Department of Internal Medicine, School of Medicine,
Nagoya City University
4. Different susceptibilities of p53 knockout
(–/–), (+/–) and (+/+) mice to induction
of stomach adenocarcinomas by
N-methyl-N-nitrosourea and esophageal
squamous cell carcinomas by
methyl-n-amylnitrosamine
Tsukamoto, T., Yamamoto, M., Shirai, N.* 2 , Sakai, H.* 1 ,
Iidaka, T.* 2 , Donehower, L.A.* 3 and Tatematsu, M.
Mutations of the p53 tumor suppressor gene
constitute one of the most frequent molecular
changes in a wide variety of human cancers. Mice
deficient in p53 have recently attracted attention for
their potential to identify chemical genotoxins. In
this study we investigated the susceptibility of p53
nullizygote (–/–) , heterozygote (+/–), and wild type
(+/+) mice to N-methyl-N-nitrosourea (MNU) gastric
carcinogenesis and to methyl-n-amylnitrosa-
mine (MNAN), which specifically induces esophageal
tumors in mice. In the first experiment,
p53 knockout mice were treated with 30 ppm MNU
in drinking water one week on and one week off
and killed after 5 weeks. The numbers of pepsinogen
altered pyloric glands (PAPG), putative preneoplastic
lesions, were 1.8, 1.7 and 22.6 in p53
(+/+), (+/–), and (–/–) mice, respectively. In a
15-week experiment, adenomas were found in 0 of
19 (+/+) (0%), 2 of 21 (+/–) (9.5%), and 6 of 10
(–/–) (60.0%) animals. Also one well differentiated
adenocarcinoma was observed in a p53 (–/–)
mouse. After forty weeks treatment with 120 or
30 ppm MNU, there was no significant difference
in the incidence of gastric tumors between p53
(+/+) and (+/–) mice, but mortality from carcinogen-induced
lymphomas, leukemias and sarcomas
was very much greater in the latter group. Homozygous
KO animals could not be maintained
long-term. PCR-single strand conformation
polymorphism analysis of exons 5-8 of the p53
gene demonstrated no mutations, in DNA extracts
from 68 gastric tumors from 16 and 20 p53 (+/+)
and (+/–) mice receiving 30 ppm, respectively, and
14 and 18 given 120 ppm. For esophageal carcinogenesis,
the p53 (+/–), and (+/+) mice were
treated with 5 or 15 p.p.m. MNAN in their drinking
water for 8 weeks then maintained without further
treatment for an additional 7 or 17 weeks, being
killed at experimental weeks 15 or 25. An additional
group of p53 (–/–) mice were given 5p.p.m.
MNAN for 8 weeks and killed at week 15. In the
5p.p.m. groups, squamous cell carcinomas (SCCs)
were observed in 10/12 (83.3%) p53 (–/–) and 1/15
(6.7%) p53 (+/–) mice, but in none of the p53 (+/+)
mice. With 15 p.p.m., 2/14 (14.3%) p53 (+/–) and
19

Cdx1 expression and intestinal phenotypic differentiation.
Our second research area involves the molecular basis of cancer metastasis and applications in diagnosis
and treatment, especially for micrometastases. Several micrometastasis models featuring tagged with green
fluorescence protein (GFP) gene were established, including a rat tongue cancer cell line, human gastric and
colonic cancer cell lines with different metastatic potentials. Sequential steps in hematogenous, peritoneal
and lymphogenous metastasis in living mouse were then documented by an intravital videomicroscopy technique.
In addition, a rapid quantitative method for detection of micrometastases in peritoneal washes of patients
with gastric carcinoma was developed using real-time RT-PCR and prognostic potential demonstrated.
1. Reversibility of heterotopic proliferative
glands in glandular stomach of Helicobacter
pylori-infected mongolian gerbils
on eradication
Tatematsu, M., Nozaki, K., Shimizu, N., Tsukamoto, T.,
Inada, K., Cao, X., Ikehara, Y., Kaminishi, M. *1 and
Sugiyama, A. *2
Helicobacter pylori (Hp) infection is an important
factor in human gastric disorders. Mongolian
gerbils can be easily infected with Hp and provide
an excellent experimental model for clarifying the
role of the bacterium in chronic active gastritis,
peptic ulceration, intestinal metaplasia, and gastric
carcinoma development. We have proved the enhancing
effects of Hp infection on all histological
types of gastric cancers in Mongolian gerbils exposed
to chemical carcinogens. Heterotopic proliferative
glands (HPGs) also frequently develop
with Hp infection in the glandular stomach of infected
gerbils, with a slightly dysplastic change of
constituent cells. Distinguishing reversible inflammatory
lesions from true neoplasms upon eradication
is necessary for further biological or histochemical
investigations using this model. For this
purpose, we employed an experimental model of
long-term Hp infection and eradication in gerbils.
HPGs finally developed with a phenotypic shift of
intestinalization, including generation of Paneth
cells. After eradication, HPGs were obviously reduced,
and gastric lesions in mucosa also improved
with few remnants of the former injury. This shows
that reversible HPGs are frequently induced solely
by Hp infection in this animal species, and are related
to severe gastritis, rather than being malignant
in character. Thus, distinction of reversible lesions
from true neoplasms is necessary to elucidate relationship
between Hp infection and gastric carcinogenesis
in this animal model.
* 1 Department of Gastrointestinal Surgery, Postgraduate
School of Medicine, The University of Tokyo
* 2 First Department of Surgery, Shinshu University,
School of Medicine
18
2. Real-time observation of micrometastasis
formation in the living mouse liver
using a GFP gene-tagged rat tongue
carcinoma cell line
Nakanishi, H., Itoh, H.* 1 , Ikehara, Y., Kato, T.* 1 , Nakao,
A.* 2 and Tatematsu, M.
Initial arrest, attachment, extravasation, and
subsequent extravascular growth of tumor cells in
secondary organs are believed to be crucial events
for hematogenous metastasis, but the actual processes
in living animals remains unclear. For the
present study, we established green fluorescent
protein (GFP)-expressing rat tongue carcinoma cell
lines permiting real-time analysis of micrometastasis
formation in combination with intravital video
microscopy (IVVM). GFP expressing metastatic
(LM-EGFP) and non-metastatic (E2-EGFP) cell
lines could be visualized at the cellular level in live
mice for more than one month. Real-time IVVM
analysis of liver metastases after intraportal injection
of the cells via mesenteric vein revealed that
both LM-EGFP and E2-EGFP tumor cells arrest
similarly in sinusoidal vessels near terminal portal
venules within 0.4 sec, during which time no evidence
of “rolling” -like movement along endothelial
cell surface is observed. Quantitative analysis of
GFP positive foci showed that E2-EGFP cells were
completely sheared from the liver sinusoid within 3
days, with no solitary dormant cells, whereas a substantial
number of LM-EGFP cells remained in the
liver, probably due to stable attachment to the sinusoidal
wall. Confocal laser scanning microscopy
(CLSM), in combination with laminin immunohistochemistry,
revealed that only LM-EGFP cells
started growth at 3 to 4 days after inoculation and
that most of the growing foci were surrounded by a
subsinusoidal basement membrane (BM). Our results
suggest that micrometastasis formation by
LM-EGFP cells consists of initial tumor cell arrest
due to size constraints of the vessel, stable attachment
to subsinusoidal BM and subsequent intravascular
growth before extravasation. The difference
in metastatic potential between the 2 lines

1/11 (9.1%) p53 (+/+) mice developed SCCs. At
25 weeks, the incidences of SCCs were 7/16
(43.8%) and 8/14 (57.1%) in p53 (+/–) mice and
1/13 (7.7%) and 2/10 (20.0%) in p53 (+/+) mice
receiving 5 and 15 p.p.m., respectively. Of the
SCCs examined by PCR-single strand conformation
polymorphism analysis, 61% (14/23) from p53
(+/–) and 50% (6/12) from p53 (+/+) mice demonstrated
mutations in the p53 gene (exons 5-8).
These results suggest that p53 may not be a direct
target of MNU but rather play an important role as a
gatekeeper in mouse stomach carcinogenesis induced
by this direct acting agent. However, they
indicate the order of susceptibility to
MNAN-induced esophageal tumorigenesis to be as
follows: nullizygotes (–/–) > heterozygotes (+/–) >
wild type (+/+), and provide strong evidence of involvement
of p53 mutations in the development of
esophageal SCCs.
* 1 Department of Veterinary Pathology, Gifu University
* 2 Nagoya Laboratories, Pfizer Global Research & Development
* 3 Department of Molecular Virology and Microbiology,
Baylor College of Medicine, Houston
5. Polymorphisms of two fucosyltransferase
genes (Lewis and Secretor
genes) involving type I Lewis antigens
are associated with the presence of
anti-Helicobacter pylori IgG antibodies
Ikehara,Y., Nishihara, S.* 1 , Yasutomi, H., Kitamura, T.,
Matsuo, K., Shimizu, N., Inada, K., Kodera, Y.* 2 ,
Yamamura, Y.* 2 , Narimatsu, H.* 1 , Hamajima, N.* 3 and
Tatematsu, M.
Helicobacter pylori attach to the gastric mucosa
though adhesin, which binds to Lewis b (Le(b)) or
H type I carbohydrate structures. The products of
the Secretor (Se) and Lewis (Le) gene are involved
in type I Le antigen synthesis. The present study
was therefore performed to investigate the possibility
that Se and Le gene polymorphisms alter the risk
of H. pylori infection. Two hundred thirty-nine participants
were genotyped for Se and Le and tested
for the presence of anti-H. pylori IgG antibodies.
Using the normal gastric mucosa from 60 gastric
cancer patients, we then assessed immunohistochemically
whether type I Le antigen expression
depended on the Se and Le genotype. The H pylori
infection rate was positively associated with the
number of Se alleles (se/se group, 45.1%; Se/se
group, 64.6%; and Se/Se group, 73.3%) and nega-
20
tively associated with the number of Le alleles (le/le
group, 76.4%; Le/le group, 68.3%; and Le/Le group,
55.6%). When the subjects were classified into
three groups [low risk, (se/se, Le/Le) genotype;
high risk, (Se/Se, le/le), (Se/Se, Le/le), and (Se/se,
le/le) genotypes; moderate risk, other than low- or
high-risk group], the odds ratio relative to the
low-risk group was 3.30 (95% confidence interval,
1.40-7.78) for the moderate-risk group and 10.33
(95% confidence interval, 3.16-33.8) for the
high-risk group. Immunohistochemical analysis
supported the finding that Se and Le genotypes affected
the expression of H. pylori adhesin ligands.
We conclude that Se and Le genotypes strongly affect
susceptibility to H. pylori infection.
* 1 Division of Cell Biology, Institute of Life Science,
Soka University
* 2 Division of Gastroenterological Surgery, Aichi Cancer
Center Hospital
* 3 Epidemiology and Prevention, Aichi Cancer Center
Research Institute

Professor Youlin Qiao, from Cancer Institute and Hospital Chinese Academy of Medical Sciences and
Peking Union Medical College giving us the lecture entitled “ Helicobacter pylori Seropositivity
and Cardia Stomach Cancer: Positive Association in a Prospective, Nested Case-cohort Study from
Linxian, China“ in the 8th Aichi Cancer Center International Symposium held on February 16,
2002 (see p. 89).
21

22
From left to right
First row: Dr. T. Nakagawa, Mr. Y. Tatematsu, Dr. H. Nagai and Dr. S. Tomida.
Second row: Dr. A. Masuda, Dr. T. Takeuchi, Dr. H. Osada, Dr. K. Koshikawa, Dr. H. Konishi, Dr. Ta.
Takahashi, Dr. K. Mizuno, Ms. T. Harano and Dr. H. Saito.

Division of Molecular Oncology
________________________________________________________________________________
Takashi Takahashi, M.D., Ph.D., Chief
Hirotaka Osada, M.D., Ph.D., Section Head
Akira Masuda, Ph.D., Senior Researcher
Ken-ichi Kozaki, D.D.S., Ph.D., Senior Researcher (until Dec. 2000)
Hiroyuki Konishi, M.D., Ph.D., Senior Researcher (as of Jul. 2001)
Hiroko Saito, Ph.D., Research Assistant
Tomoko Harano, B.S., Research Assistant
Yoshio Tatematsu, B.S., Research Assistant
Postdoctoral Fellows
Hiroyuki Konishi, M.D., Ph.D. (until Jun. 2001)
Toshiyuki Takeuchi, Ph.D. (as of Jul. 2001)
Visiting Trainees
Kiyoshi Yanagisawa, M.D., Nagoya University School of Medicine (until Nov. 2000)
Nobuhiro Haruki, M.D., Nagoya City University School of Medicine (until Oct. 2000)
Katsumi Koshikawa, M.D., Nagoya University School of Medicine (as of May 2000)
Taku Nakagawa, M.D., Nagoya University School of Medicine (as of Apr. 2000)
Kotaro Mizuno, M.D., Nagoya City University School of Medicine (as of Sept. 2000)
General Summary
Lung cancer is soon expected to become the leading cause of cancer death in Japan, currently claiming
nearly 50,000 lives annually. Our goal is to understand the molecular pathogenesis of this fatal disease, as a
basis for designing novel strategies for better diagnosis, treatment and prevention.
Accumulating evidence indicates that lung cancer is a disease caused by accumulation of multiple genetic
defects and our previous studies identified multiple tumor suppressor genes and dominant oncogenes involved
in its pathogenesis. In addition, aiming at an in vitro model system suitable for analysis of various
aspects of lung carcinogenesis, we have established and extensively characterized peripheral lung epithelial
cell lines, termed HPL1A to E, which are, to our knowledge, the first immortalized human peripheral airway
cells. Metastasis is a major cause of cancer-related deaths and we are also devoting ourselves to investigate
the underlying mechanisms with the goal of control of this life-threatening process.
We are continuing to focus on molecular pathogenesis of lung cancer, employing a variety of strategies
for its better understanding in line with realization of our major objective, "from bench top to bedside".
1. Multi-faceted analyses of a highly metastatic
human lung cancer cell line
NCI-H460-LNM35 suggest mimicry of inflammatory
cells in metastasis
Kozaki, K., Koshikawa, K., Osada, H., Konishi, H.,
Tatematsu, Y., Miyaishi, O. *1 , Saito, H., Hida, T. *2 ,
Mitsudomi, T. *3 and Takahashi Ta.
Despite considerable advances in the understanding
of the molecular pathogenesis of lung
cancer, the majority of patients eventually die because
of widespread metastases. To date, various
molecules have been suggested as playing a role in
the underlying processes, but it is also evident that
we still need to learn more about how cancer cells
metastasize to distant organs. To propel studies on
the molecular mechanisms of lung cancer metasta-
sis, we have established a highly metastatic human
lung cancer cell line, NCI-H460-LNM35 (hereafter
referred to as LNM35), which is capable of spontaneous
metastasis, not only via hematogenous but
also lymphogenous routes, with a 100% incidence.
To gain insight into the molecular mechanisms of
the highly metastatic capability of LNM35, genome-wide
screening for genes differentially expressed
in LNM35 and a representative low metastatic
clone (NCI-H460-N15, hereafter referred to as
N15) of NCI-H460, the parental line for LNM35,
was performed with the aid of a microarray containing
9,844 cDNA fragments. This search consequently
identified 13 genes with a more than
2.5-fold up-regulation in LNM35, whereas a more
than 2.5-fold down-regulation was detectable for 15
genes. Our previous study showed increased
23

COX-2 expression in LNM35 and significant inhibition,
although in vitro, of motility and invasion as
a result of treatment with nimesulide, and we here
noted up-regulation of various other genes known
to be related to inflammation, including a proinflamatory
cytokine IL-1α, two C-X-C chemokines
(i.e., ENA-78 and NAP-2), complement component
3 and a complement-activating factor, CD55 in the
present microarray analysis. In addition, we
cloned and characterized a novel gene, CLCP1,
whose corresponding EST was identified in our expression
profiling analysis. CLCP1 expression
was confirmed by Northern blot analysis to be
up-regulated during in vivo selection of LNM35,
and increase was also identified in a significant
fraction of human lung cancer cases in vivo, especially
in lymphogenous metastatic sites. In fact,
our results indicate increased CLCP1 expression to
be present in 36% of primary sites, in 58% of
lymph node metastases and in 33% of distant metastases.
Taken together, these findings indicate
that future studies with our unique model system
are warranted in order to improve understanding of
the molecular mechanisms of lung cancer metastasis,
in the hope that this may ultimately provide
clues as to how to reduce the present, extremely
large number of lung cancer deaths.
*1
Department of Basic Gerontology, National Institute
for Longevity Sciences
*2
Department of Internal Medicine, Aichi Cancer Center
Hospital
*3
Department of Thoracic Surgery, Aichi Cancer Center
Hospital
2. Gene silencing by aberrant DNA methylation
and abnormalities in chromatin
configuration in human lung cancer
cells
Osada, H., Tatematsu, Y., Yatabe, Y. *1 , Masuda, A.,
Konishi, H., Harano, T., Nakagawa, T., Saito, T. *2 ,
Sugiyama, M. *2 , Yanagisawa, K., Takada, M. *3 and
Takahashi, Ta.
TGF-β strongly inhibits epithelial cell proliferation
and alterations of its signaling are thought to
play a role in tumorigenesis. We have found that
most lung cancer cell lines demonstrated loss of the
growth-inhibitory responses to TGF-β, and the affected
tumors being divided into two major groups,
TGFβRII (+)/Smad7(+) and TGFβRII (-)/Smad7(-),
suggesting heterogeneity in mechanisms underlying
TGF-β response. The mechanism of the loss of
24
TGFβRII expression in the latter group was further
studied, identifying aberrant DNA methylation of
the promoter region in a limited fraction of cell
lines. Interestingly, we found that alteration of
chromatin structure because of histone deacetylation
may also be involved, showing a good correlation
with loss of TGFβRII expression. This notion
was supported by the findings of a restriction enzyme
accessibility assay, of a chromatin immunoprecipitation
assay with anti-acetyl histone antibodies,
and of in vivo induction of TGFβRII expression
by histone deacetylase inhibitors including
trichostatin A (TSA) and sodium butyrate. In vitro
induction of TGFβRII promoter reporter activity by
TSA was also detected and found to require a
CCAAT box within the –127/-75 region. This is the
first study to demonstrate that, in addition to the
TSA-responsive region in the TGFβRII promoter,
the alteration of histone deacetylation may be involved
in loss of TGFβRII expression in lung cancer
cell lines.
We also identified another target for epigenetic
alteration in lung cancers, i.e, 14-3-3σ, one isoform
of the 14-3-3 family, which plays a role in the G2
checkpoint by sequestering Cdc2-cyclinB1 in the
cytoplasm. Loss of expression has been suggested
to cause a G2 checkpoint defect, resulting in chromosomal
aberrations. Our recent study on DNA
methylation status and expression level of the
14-3-3σ gene in lung cancer cell lines and primary
lung tumor specimens revealed small cell lung cancer
(SCLC) cell lines to frequently feature DNA
hypermethylation (~70%) and silencing of the
14-3-3σ gene. Among non-small cell lung cancers
(NSCLC), only large cell lung cancer cell lines
showed frequent hypermethylation and silencing of
14-3-3σ (~60%), whereas hypermethylation occurred
very rarely (6%) in other types including
squamous cell carcinomas and adenocarcinomas.
All eight primary SCLC specimens examined also
showed loss or significant reduction of 14-3-3σ expression
in vivo, while this was very rare in primary
NSCLC specimens (5%). This is the first description
that indicates lung cancers frequently show
significant inactivation of the 14-3-3σ gene, mainly
due to DNA hypermethylation, in SCLC, but rarely
in NSCLC, suggesting involvement of the 14-3-3σ
gene in lung tumorigenesis in a histological
type-specific manner. These epigenetic changes in
lung cancers may be of considerable clinical interest,
because their potential reversible nature may be
regarded as a rational target for development of
novel therapeutic measures.

*1
Department of Pathology and Molecular Diagnostics,
Aichi Cancer Center Hospital.
*2
Laboratory of Ultrastructural Research (until March
1999)
*3
Pulmonary Medicine, Rinku General Medical Center.
3. Persistent increase in chromosome instability
in lung cancers
Haruki, N., Masuda, A., Harano, T., Kiyono, T. *1 ,
Takahashi, Takao *2 , Tatematsu, Y., Shimizu, S. *3 ,
Mitsudomi, T. *3 , Konishi, H., Osada, H., Fujii, Y. *4 and
Takahashi, Ta.
Lung cancer cells have been shown to exhibit
frequent chromosomal abnormalities, from both
numerical (i.e. aneupoidy) and structural points of
view. These are believed to contribute to tumor development
and progression by facilitating loss of
heterozygosity and inactivation of tumor suppressor
genes, as well as favoring polysomy of chromosomes
that habor growth-promoting genes. However,
it has not been directly established whether
aneuploidy is in fact associated with a persistent increase
in the rate of chromosomal losses and gains
(i.e. chromosomal instability or CIN).
To clarify whether CIN is a common feature in
lung cancer cell lines in association with the presence
of significant aneuploidy, we examined the
rates of divergence of chromosome numbers in 10
lung cancer cell lines during cultivation by means
of fluorescence in situ hybridization with specific
centromere probes. Then, we examined the correlation
of degree of CIN with various other molecular
and cellular parameters in lung cancer cell lines,
such as the presence of mutations in p53, degree of
aneuploidy, morphology of centrosomes, and integrity
of the mitotic spindle checkpoint. As results,
we found that lung cancer cell lines exhibiting significant
aneuploidy showed a persistent CIN phenotype.
In addition, the CIN phenotype correlated
well with the presence of p53 mutations, and moderately
with centrosome abnormalities. However,
human papilloma virus 16-E6-directed inactivation
of p53 in a representative non-CIN lung cancer cell
line did not result in the induction of CIN, at least
up to the 25 th generation, suggesting that inactivation
of this tumor suppressor gene is itself unlikely
to directly induce CIN in lung cancer cells. Interestingly,
however, significant CIN could be induced
in p53-inactivated cells in conjunction with the
generation of aneuploid populations, when mitotic
spindle formation was transiently abrogated with a
microtubule interfering reagent.
In the present study, we provided clear evidence
for the first time that the majority of human lung
cancer cell lines, which exhibit significant aneuploidy,
show persistent CIN. The results suggest
that inactivation of p53 may allow lung cancer cells
to go through an inappropriate second division cycle
under certain forms of mitotic stress, which
would result in the induction of the CIN phenotype
in conjunction with generation of aneuploidy. Further
studies are now needed to identify and clarify
the underlying mechanisms directly responsible for
the induction of CIN. Their clarification should
provide important clues to the development of new
therapeutic approaches for this fatal cancer.
*1
Laboratory of Viral Oncology
*2
Laboratory of Ultrastructural Research (until March
1999)
*3
Department of Thoracic Surgery, Aichi Cancer Center
Hospital
*4
Department of Surgery II, Nagoya City University
School of Medicine
4. Identification of frequent G2 checkpoint
impairment and a homozygous deletion
of 14-3-3ε at 17p13.3 in small cell lung
cancers
Konishi, H., Nakagawa, T., Harano, T., Mizuno, K.,
Saito, H., Masuda, A., Osada, H. and Takahashi Ta.
It has been shown that the short arm of chromosome
17 (17p) is one of the most frequently affected
chromosomal regions in lung cancers, as well
as in a variety of other human neoplasias. Although
the p53 gene at 17p13.1 is well accepted as a genuine
molecular target in the frequent 17p deletions,
we previously performed a detailed LOH analysis
for 17p using 100 cases of primary lung cancers
representing all four major histologic subtypes, and
suggested that, in addition to the p53 gene, an as yet
unidentified tumor suppressor gene(s) residing at
17p13.3 might also play a role in lung carcinogenesis.
We therefore screened 65 lung cancer cell lines
with the aid of nine markers mapped within the
commonly deleted region of lung cancers at
17p13.3. A single STS marker corresponding to
the 14-3-3ε gene was consequently found to yield
no amplification products in either of two SCLC
cell lines on duplex PCR analysis. The two SCLC
cell lines, ACC-LC-48 and ACC-LC-52, had been
established from distinct metastases of the same pa-
25

tient after different treatment periods, suggesting
the occurrence of homozygous deletions before
metastasis in vivo. Northern blot analysis showed
complete absence of 14-3-3ε expression in the two
SCLC cell lines.
14-3-3ε is one of the seven 14-3-3 isoforms thus
far identified, which form a complex with a variety
of molecules. Among their binding partners,
Cdc25C has been shown to be involved in the G2
checkpoint response, and to bind mainly to 14-3-3ε
among the seven 14-3-3 isoforms, in Xenopus egg
extract. Although 14-3-3ε also forms a complex
with Cdc25C in human cells, it is not clear which
14-3-3 isoform actually plays a significant role in
the G2 checkpoint response. We therefore examined
the function of the G2 checkpoint using a
14-3-3ε-null ACC-LC-48 cell line. The mitotic
index in ACC-LC-48 after one Gy irradiation remained
at over 50% of the non-irradiated value,
suggesting an inefficient G2 arrest and G2 checkpoint
impairment. In addition, the G2 checkpoint
response could be restored to a significant extent by
both transient and stable introduction of exogenous
14-3-3ε into ACC-LC-48. These observations indicate
that the homozygous loss of 14-3-3ε perturbs
the G2 checkpoint response to x-ray-irradiation in
ACC-LC-48. Next we examined mitotic indices after
exposure to one Gy irradiation in an additional
seven SCLC cell lines, and found the G2 checkpoint
response to be frequently impaired to various
degrees. The G2 checkpoint is one of the most
highly conserved mechanisms that regulate the cell
cycle by preventing damaged cells from progression
through the cell cycle. Our finding that a significant
fraction of SCLCs, which are very sensitive to irradiation
and chemotherapy and at the same time the
most aggressive type of lung cancers, exhibit an
abnormal G2 checkpoint response is thus of great
interest not only from a biological point of view but
also in terms of clinical implications.
5. In vitro molecular analysis of carcinogenesis
of human lung adenocarcinomas
with the aim of clinical applications
Masuda, A., Konishi, H., Yatabe, Y. *1 , Hida, H. *2 , Saito,
T. *3 and Takahashi, Ta.
It is well known that oncogenes such as K-ras,
c-myc and c-jun are often activated or overexpressed
in human lung cancers. However, the precise
roles of such oncogene activation in the devel-
26
opment and progression of human lung cancers are
not yet fully understood. In the present study, we
examined effect of modulation of signaling using a
newly established immortalized human peripheral
lung epithelial cell line (HPL1D) and an immortalized
lung airway epithelial cell line (BEAS2B),
with the aim of identifying molecular targets for
future therapeutic modalities.
We observed that a transformed cell-like morphology
could be induced in HPL1D in response to
EGF under the anchorage-dependent conditions and
that apoptosis with anchorage-independence was
markedly reduced, consequently resulting in colony
formation in soft agar. Interestingly, although anchorage-dependent
proliferation of HPL1D was
markedly enhanced with HGF, which is known to
stimulate cell proliferation via signal transduction
pathways similar to those elicited with EGF, HGF
did not induce anchorage-independent cell growth
at all. We are currently investigating the mechanisms
underlying these differential responses.
In addition, the biological effects of oncogene
activation were examined by establishing stable
transfectants of BEAS2B, in which oncogenes were
placed under the regulation by tetracycline-off system.
Addition of either EGF or FCS failed to induce
endogenous c-jun under anchorage-independent
conditions and did not lead to
formation of soft agar colonies. In contrast, expression
of exogenously introduced c-jun considerably
stimulated anchorage-independent colony
formation. Interestingly, eight of 18 human lung
cancer cell lines exhibited loss of the anchorage-dependence
of c-jun induction. Furthermore,
we could show that introduction of activated K-ras
alone was insufficient to induce anchorage-independent
colony formation, but that anchorage-independent
cell growth could be stimulated
in conjunction with the forced expression of
c-jun, suggesting cooperative roles in transformation
of lung epithelial cells.
Taken together, these results point to the utility
of immortalized normal human lung epithelial cells
for understanding tumorigenesis in the lung.
*1
Department of Pathology and Molecular Diagnostics,
Aichi Cancer Center Hospital.
*2
Department of Internal Medicine, Aichi Cancer Center
Hospital
*3
Laboratory of Ultrastructural Research (until March
1999)

Dr. Roderich E. Schwarz, from University of Medicine and Dentistry of New Jersey Robert Wood Johnson
Medical School, U.S.A, giving us the lecture entitled “Surgery and Adjuvant Therapy for
Gatsric Carcinoma in the U.S.A.“ in the 8th Aichi Cancer Center International Symposium held on
February 16, 2002 (see p. 93).
27

28
From left to right
Front row: Dr. M. Suguro-Katayama, Ms. Y. Kasugai, Ms. H. Suzuki, Dr. M. Seto and Dr. Y. Hosokawa
Back row: Dr. R. Suzuki, Dr. H. Tagawa, Dr. Dr. K. Mayama, Dr. K. Izumiyama, Dr. S. Tsuzuki and Mr.
S. Karnan

Division of Molecular Medicine
________________________________________________________________________________
Masao Seto, M.D., Dr.M.Sc. Chief
Yoshitaka Hosokawa, M.D., Dr.M.Sc. Section Chief
Ryoji Ishida, Ph.D. Senior Researcher
Ritsuro Suzuki, M.D., Dr.M.Sc., Senior Researcher
Shinobu Tsuzuki, M.D., Ph.D., Senior Researcher (As of October, 2001)
Keiko Nishida, B.P. Senior Research Assistant (Until March, 2000)
Hiroko Suzuki, B.P. Senior Research Assistant
Yumiko Maeda, B.S. Research Assistant
Hiroyuki Tagawa, M.D., Ph.D. Research Resident (As of April, 2000)
Visiting Trainees
Yoshitoyo Kagami, M.D., Department of Hematology and Chemotherapy, Aichi Cancer Center Hospital
Hidenobu Takahashi, M.D., The First Department of Internal Medicine, Niigata University School of Medicine
(Until September, 2000)
Masakatsu Yonezumi, M.D., The Third Department of Internal Medicine, Hokkaido University School of
Medicine (Until March, 2001)
Ko Izumiyama, M.D., The Third Department of Internal Medicine, Hokkaido University School of Medicine
(As of April, 2001)
Toshihiro Nakanishi, B.P., Gifu Pharmaceutical University (Until March, 2000)
Karnan Sivasundaram, Nagoya University Graduate School of Science (As of June, 2001)
Miyuki Suguro-Katayama, M.D., The second Department of Internal Medicine, Mie University School of
Medicine (As of July, 2001)
Dr. Ko Mayama, M.D., The First Department of Internal Medicine, Hirosaki University School of Medicine (As
of October 2001)
General Summary
Research in this laboratory is aimed at generating a better understanding of the genetic and molecular
bases of human cancer, with eventual application of the acquired knowledge in the field of medical oncology.
Our work has been mainly focused on hematologic malignancies, in cooperation with researchers of the Department
of Hematology and Chemotherapy (Chief. Dr. Yasuo Morishima), and the Pathology and Clinical
Laboratories (Dr. Shigeo Nakamura). Hematologic malignancies have several advantages for studying the
molecular bases of neoplasia. Chromosomal abnormalities have been analyzed by a large number of researchers
and the observed strong association between specific chromosome changes and specific hematopoietic
tumors provides direct evidence that the resultant gene alterations play a pivotal role in the disease
development. Over the last two years we have concentrated attention on the 18q2l translocation (associated
with the mucosa-associated lymphoid tissue lymphoma), the 3q27 translocation (associated with diffuse large
cell lymphomas), and the 11q13 (associated with mantle cell lymphomas). The second advantage of
studying hematopoietic malignancies is that the various leukemias and lymphomas have been classified in
detail with respect to cell surface markers, developmental lineages and stages, so that they can be used for
studying important factors and signals for cell differentiation and proliferation. Indeed, we could identify
subsets of diffuse large B-cell lymphoma with cell surface markers. Finally, we are also trying to analyze
T-cell malignancies, most of whose chromosome aberrations are not well understood. We could identify
the TCBA1 gene, a candidate that may be involved in chromosome 6q aberrations.
1. Search for MALT1-associated proteins
using yeast two-hybrid strategy
Hosokawa, Y., Suzuki, H., and Seto, M.
The category of mucosa-associated lymphoid
tissue (MALT) lymphoma was first proposed by
Isaacson et al. and it is now clearly recognized and
categorized as extranodal marginal zone lymphoma
of MALT type in REAL classification. It often
originates from chronic inflammation, such as H.
pylori gastritis or autoimmune disease, tends to remain
localized for a long time and mostly exhibits
29

From left to right
First row: Ms. Y. Matsudaira, Dr. K. Kuzushima and Ms. Y. Nakao. Second row: Dr. E. Kondo, Dr. K. Tsujimura, Dr.
To. Takahashi, Ms. H. Tamaki, Dr. K. Tajima, Ms. K. Nishida, Dr. M. Miyazaki, Dr. Y. Akatsuka and Dr. T.
Nishida.
Insets: Dr. Y. Obata and Mr. S. Iwase.
32

Division of Immunology
________________________________________________________________________________
Toshitada Takahashi, M.D. Chief
Yuichi Obata, Ph.D. Section Head (until March 2001)
Kunio Tsujimura, M.D. Senior Researcher
Yoshiki Akatsuka, M.D. Senior Researcher (as of July 2000)
Yasue Matsudaira, B.S. Senior Research Assistant
Keiko Nishida, B.P. Senior Research Assistant (as of April 2001)
Satoshi Ozeki, D.V.M. Research Assistant (until March 2001)
Visiting Scientists
Kazuhiro Yoshikawa, B.M.T., M.D. Second Department of Pathology, Aichi Medical University
Yuichi Obata, Ph.D. Head, Department of Biological Systems, RIKEN BioResource Center (as of April 2001)
Visiting Trainees
Shigeru Iwase, B.P. Department of Hygienic Chemistry, Faculty of Pharmaceutical Sciences, Nagoya City
University
Keigo Mizutani, M.D. Department of Pediatrics, Nagoya City University School of Medicine
Eisei Kondo, M.D. Second Department of Internal Medicine, Okayama University School of Medicine
Kouhei Tajima, M.D. First Department of Surgery, Gunma University School of Medicine
Masahiro Yoshida, M.D. Department of Orthopedic Surgery, Nagoya University School of Medicine
Mikinori Miyazaki, M.D. Second Department of Internal Medicine, Nagoya City University School of Medicine
Tetsuya Nishida, M.D. First Department of Internal Medicine, Nagoya University School of Medicine
General Summary
The object of our research is to characterize the biological nature of cancer cells, with emphasis on immunogenetic
analysis of cell-surface molecules of both human and experimental tumors. The major projects
undertaken over the past two years are summarized below.
In the field of human tumor antigens, three projects are in progress. Firstly, production of single chain antibodies
(scFv) reactive with a truncated type of the epidermal growth factor receptor (EGFR) expressed on
glioblastomas was achieved and their application for tumor imaging has been investigated. Secondly, we
have carried out an antigen analysis of stomach, breast, and prostate cancers using the expression cloning
method (SEREX). Fifty to 100 cDNA clones encoding antigens detected by autologous IgG antibodies were
selected for each type of cancer and are now being analyzed. Thirdly, an attempt has been made to detect
minor histocompatibility antigens by generating cytotoxic T cells (CTL) from HLA identical bone marrow
transplantation patients. In addition, an efficient in vitro CTL generation method was established using retrovirally
transduced B cells as antigen-presenting cells.
In the second area of interest, studies on mouse tumor antigens have been conducted with the thymus-leukemia
(TL) antigen as a model. Immunization with dendritic cells engineered to express TL is able to
induce rejection of TL positive lymphoma cells. In addition, in order to monitor TL-specific CTL, TL
tetramers were prepared. Unexpectedly, these proved reactive not only with TL-specific CTL, but also with
normal intestinal intra epithelial lymphocytes and thymocytes.
1. Production of a single chain variable
fragment (scFv) antibody against type
III mutant EGFR
Yoshikawa, K. *1 , Nakayashiki, N. *1, 2 , Takasu, S. *1, 2 ,
Okamoto, K. *1, 2 , Nakamura K. *3 , Hanai, N. *3 , Okamoto,
S. *2 , Mizuno, M. *2 , Wakabayashi, T. *2 , Saga, S. *1 ,
Yoshida, J. *2 and Takahashi, To.
The type III deletion-mutant of the epidermal
growth factor receptor (EGFR) is a potential target
in diagnostic and therapeutic approaches for glioblastomas
characterized by its expression. In this
study, a single chain variable fragment (scFv)
antibody was produced, based on the mouse monoclonal
antibody, 3C10 (IgG2b) specifically recognizing
this mutant EGFR. (Nakayashiki, et al., Jpn.
J. Cancer Res., 91, 1035-1043, 2000). Partial determination
of its N-terminal amino acid sequence
and preparation of adequate primers for VH and VL
genes were assembled with a linker, (Gly4Ser)3 and
33

ligated into a bacterial expression vector to express
the scFv as cytoplasmic inclusion bodies. After appropriate
refolding, the scFv were purified using a
mutant peptide-conjugated column. On Biacore
analysis, the affinity (KA) of the parental 3C10 for
the mutant peptide was 9.7 x 10 7 M -1 , while that of
3C10 scFv was 2.45-2.48 x 10 7 M -1 , approximately
4 fold weaker. On ELISA, 3C10 scFv showed selective
reactivity with the mutant peptide, similarly
to the parental 3C10 antibody. Immunostaining
analysis revealed scFv staining in a glioblastoma
case with type III mutant EGFR, the biodistribution
of 99m Tc-labeled 3C10 scFv being evaluated in
athymic mice bearing transformants expressing the
mutant EGFR. 99m Tc-labeled 3C10 scFv accumulated
in tumors, with a tumor/blood radio of 3-5,
which was, however found to be lower than that
with the parental antibody (12-18). The results altogether
suggest that the scFv antibody still maintains
the antibody structure to detect a conformational
epitope, similarly to the parental antibody.
*1
Second Department of Pathology, Aichi Medical
University
*2
Department of Neurosurgery, Nagoya University
School of Medicine
*3
Tokyo Research Laboratories, Kyowa Hakko Kogyo
Co. Ltd.
2. Immunogenic gene products in cancer
patients
Obata, Y. *1 , Takahashi, To., Tamaki, H., Tajima, K.,
Yoshida, M., Miura, S. *2 , Iwase, T. *2 , Iwata, H *2 ,
Mitsudomi, T. *3 , Takahashi, M. *4 , Sakamoto, J. *5 , Chen,
Y.-T. *6 , Stockert, E. *6 and Old, L.J. *6
Molecular characterization of cancer antigens
recognized by cytotoxic T cells by Dr. Boon's group
has opened a new era of cancer immunology. The
search for cancer antigens that can be used for immunodiagnosis
and immunotherapy of cancer is
still at a very early stage. For their identification,
several methods using cytotoxic T cells have been
developed. In addition, a molecular technique using
antibodies produced in cancer patients was also devised
by Dr. Pfreundschuh and his colleagues. This
technique, named SEREX (serological analysis of
cancer antigens by recombinant cDNA expression
cloning), identifies protein antigens that have elicited
high titer IgG responses in cancer patients. Expression
cDNA phage libraries constructed using
mRNA from tumor specimens are screened with
autologous or allogeneic sera from cancer patients.
34
Positive clones are isolated and sequenced for identification
of genes encoding antigens. One of the
advantages of the SEREX method in identification
of cancer antigens is the lack of any requirement for
establishing cultured cell lines from tumor specimens,
a process that is very difficult for most
epithelial cancers. Another advantage is direct identification
of genes encoding antigens through DNA
sequencing. Since the introduction of the method,
SEREX analysis targeting various cancer types has
defined several categories of antigens with potential
use in the clinical setting: cancer-testis (CT) antigens,
mutational antigens, fusion antigens, differentiation
antigens, over-expressed antigens,
spliced-variant antigens and viral antigens.
In our own SEREX study, we have surveyed so
far five gastric, three prostate, two breast, one lung
and one colon cancer libraries. The immune systems
of the cancer patients were found to be surprising
extremely active, with antibodies production
against diverse sets of gene products. Nearly 500
antigens were identified. To select examples useful
for diagnosis and therapy, the genes and their products
were analyzed for; (1) “DNA alterations” such
as mutation, splicing abnormality and translocation;
(2) “cancer-restricted expression” by examining the
presence of transcripts in cancer and normal tissues;
and (3) “cancer-restricted immune recognition” by
examining the frequency of antibody production in
cancer patients and control individuals. Characterization
of these and future SEREX-defined antigens
promises identification of potential targets for immunodiagnosis
and immunotherapy of cancer, as
well as genes that are significant in cancer biology.
*1
RIKEN BioResource Center
*2
Department of Breast Surgery
*3
Department of Thoracic Surgery
*4
Department of Orthopedic Surgery
*5
Aichi Hospital
*6
Ludwig Institute for Cancer Research
3. Targeted cloning of cytotoxic T cells
specific for minor histocompatibility antigens
restricted by HLA class I molecules
of interest
Akatsuka, Y., Kondo, E., Nishida, T., Taji, H. *1 ,
Morishima, Y. *1 , Obata, Y. *2 , Kodera Y. *3 and Takahashi,
To.
Minor histocompatibility antigens (mHAs) are
MHC (HLA in human)-associated peptides originating
from polymorphisms in the genome that

trigger T cell responses between MHC identical allogeneic
individuals. Graft-versus-host disease
(GVHD) and graft-versus-tumor (GVT) effects in
hematopoietic stem cell transplant (HCT) recipients
are initiated by donor T cell recognition of mHAs
on recipient cells, and it has been suggested that
particular mHAs may be selectively involved in
GVHD or GVT reactions. Thus, identification of
novel mHAs, especially restricted by HLA-A24
which is common in Japanese, is important to specify
recipients at risk of severe GVHD and antigenic
targets for immunotherapy to augment GVT responses.
We have developed a novel approach to isolate
T cell clones restricted by HLA class I alleles of interest
directly from cytotoxic T lymphocyte (CTL)
cell lines using interferon (IFN)-γ-based techniques.
CTL lines were successfully generated from 3 of 4
patients entered in this study. Enzyme-linked immunospot
(ELISPOT) assays were first performed
to identify the HLA alleles presenting mHAs to the
CTL lines using panels of B-LCL that do not share
any HLA alleles with the recipients and engineered
to express each one of the recipient HLA-A, B and
C alleles. ELISPOT assays conducted on 2 CTL
lines generated from peripheral blood specimens of
2 patients early after HCT demonstrated almost all
T cells in these CTL lines to be restricted by
HLA-B44 and HLA-A24, respectively. Multiple
HLA alleles were used as restriction molecules in
the other 3 CTL lines generated relatively long after
HCT. IFN-γ secreting T cells were then positively
selected in response to stimulation with the B-LCL
expressing the HLA alleles selected based on the
ELISPOT results, and then directly cloned. We
have successfully cloned 2 distinct
HLA-B44-restricted CTL clones and 4 distinct
HLA-A24-restricted CTL clones. Of note is that
HLA-A24-restricted CTL clones were obtained
from all of 3 HLA-A24 positive patients with this
approach. All CTL clones showed hematopoietic
lineage-specific cytotoxicity, so it can be expected
that they will recognize mHAs useful for immunotherapy
of hematopoietic malignancies. We are
currently identifying genes encoding mHAs recognized
by these CTL clones. This new method could
potentially be applied to isolate T cell clones that
recognize any antigen in the context of a specific
HLA allele of interest.
*1
Department of Hematology and Chemotherapy
*2
RIKEN BioResource Center
*3
Department of Hematology, Japanese Red Cross
Nagoya First Hospital
4. Binding of thymus leukemia (TL) antigen
tetramers to normal intestinal
intraepithelial lymphocytes and thymocytes
Tsujimura, K., Obata, Y. *1 , Matsudaira, Y., Ozeki, S.,
Yoshikawa, K. *2 , Saga, S. *2 and Takahashi, To.
Thymus leukemia (TL) antigens belong to the
family of nonclassical MHC class I antigens and
can be recognized by both TCRαβ and TCRγδ CTL
with TL- but not H-2-restriction. We previously
reported that the CTL epitope is TAP-independent,
but antigenic molecules presented by TL have yet to
be determined. In the present study, TL tetramers
were prepared with T3 b -TL and murine
β2-microglobulin not including antigenic peptides,
to determine binding specificity. CTL clones
against TL antigens were stained with the T3 b -TL
tetramer, and the binding shown to be CD3- and
CD8-dependent. Normal lymphocytes from various
origins were also studied. Surprisingly, most CD8 +
intraepithelial lymphocytes (IEL) derived from the
small intestines (iIEL), as well as CD8 + and
CD4 + CD8 + thymocytes, were stained, while only
very minor populations of CD8 + cells derived from
other peripheral lymphoid tissues such as spleen
and lymph nodes were positive. The binding of
T3 b -TL tetramers to CD8 + iIEL and thymocytes was
CD8-dependent but CD3-independent, in contrast
to that to TL-restricted CTL. These results altogether
show that TL-restricted CTL can be monitored
by CD3-dependent binding of T3 b -TL tetramers.
In addition, CD3-independent T3 b -TL tetramer
binding to iIEL and thymocytes may imply that TL
expressed on intestinal epithelium and cortical
thymocytes has a physiological function, interacting
with these tetramer + CD8 + T lymphocytes.
*1
RIKEN BioResource Center
*2
Second Department of Pathology, Aichi Medical
University
35

36
From left to right
First row: Dr. T. Kiyono, Dr. T. Tsurumi, Dr. H. Nakamura, and Dr. K. Kuzushima.
Second row: Dr. M. Fujita, Ms. A. Kudoh, Ms. N. Hayashi, Dr. S. Nakasu, Ms. T. Yoshida, Dr. Y.
Sugaya, and Mr. Y. Nishikawa.

Division of Virology
________________________________________________________________________________
Tatsuya Tsurumi, M.D. Chief
Tohru Kiyono, M.D. Section Head (until March 2002)
Kiyotaka Kuzushima, M.D. Section Head (until March 2002)
Hiromu Nakamura , PhD. Section Head (from April 2000 to March 2002)
Masatoshi Fujita, M.D. Senior Researcher
Shou Nakasu, PhD. Senior Researcher (as of April 2000)
Naoaki Yokoyama, Vet. M.D. Researcher (until September 2000)
Yutaka Sugaya, PhD. Research Resident (as of April 2001)
Toyoko Yoshida. Research Assistant
Yasuhiro Nishikawa. Research Assistant (as of April 2000)
Visiting Trainees
Yo Hoshino. Department of Pediatrics, Nagoya University School of Medicine (until March 2002)
Ken Fujii. Division of Biological Science, Nagoya University Graduate School of Science (until March 2001)
Ayumi Kudoh. Graduate School of Science and Technology, Faculty of Science, Kumamoto University
Naomi Hayashi. Department of Pediatrics, Nagoya University School of Medicine (until March 2002)
Yoriko Yamashita. First Department of Pathology, Nagoya University School of Medicine (From March 2000
to March 2002)
General Summary
Approximately 15% of all human cancers have a viral etiology, but only six viruses have actually been
implicated in their development. Among these the Epstein-Barr virus (EBV) and human papillomavirus
(HPV) are the objects of our own studies. EBV is a ubiquitous gamma herpesvirus associated with several
malignant diseases, including Burkitt’s lymphoma, nasopharyngeal lymphoma, a subset of Hodgkin’s lymphomas,
some gastric cancers, and B cell lymphomas in immunosuppressed patients. HPV is causally
linked to cervical cancers and probably to other anogenital and also to some skin and oropharyngeal cancers.
Our research aims are to elucidate the molecular mechanisms of viral DNA replication and oncogenesis
of EBV and HPV as part of the world-wide effort to combat virus-infected cancers and to characterize cellular
immunity against EBV-associated tumors in order to contribute to clinical diagnosis and therapy. During
the period 2000-2001, our research interest was concentrated on the following issues: 1) protein-protein interactions
among EBV replication proteins; 2) purification and characterization of EBV replication proteins;
3) mechanisms of inhibiting re-replication during late S-G2-M phase in mammalian cells ; 4) immortalization
of human cells by HPV; 5) determination of Epstein-Barr virus-specific CD8 + T cell frequencies
by flow cytometry and related clinical applications; 6) identification of HLA A*2402-restricted
EBV or CMV-specific CD8 + T cell epitopes by a computer algorithm and an enzyme-linked immunospot
assay.
1. The Epstein-Barr Virus Pol catalytic
subunit physically interacts with the
BBLF4/BSLF1/BBLF2/3 complex
Fujii, K., Yokoyama, N. and Tsurumi, T.
At a replication fork, the following enzymatic
reactions play very important roles for the rapid and
accurate duplication of the genetic information: 1) a
DNA helicase unwinds the parental template; 2) a
primase manufactures RNA primers for Okazaki
fragment synthesis; 3) a DNA polymerase synthesizes
the nascent leading and lagging strands. During
these reactions, various protein-protein interac-
tions between the replication proteins have been
observed. For example, bacteriophage T7 gene 5
DNA polymerase interacts with the gene 4 helicase/primase,
while the catalytic subunit of the
HSV-1 DNA polymerase interacts with the carboxyl
terminus of the UL8 protein of the HSV-1
helicase/primase heterotrimeric complex.
In the case of EBV, the EBV DNA polymerase
holoenzyme consists of the BALF5 protein (Pol
catalytic subunit) and the BMRF1 protein (Pol accessory
subunit). Although enzymatic activities of
the EBV BBLF4/BSLF1/BBLF2/3 heterotrimeric
complex have yet to be demonstrated, we assume
37

that it may act as a helicase and primase like the
HSV-1 UL5/UL52/UL8 complex. In the present
study, we revealed by immunoprecipitation analyses
that the EBV DNA Pol catalytic subunit interacts
with the BBLF4/BSLF1/BBLF2/3 complex.
The same approach using anti-BSLF1 or
anti-BBLF2/3 antibodies with clarified lysates of
B95-8 cells in a viral productive cycle suggested
that the EBV Pol holoenzyme interacts with the
BBLF4/BSLF1/BBLF2/3 complex. By experiments
utilizing lysates from insect cells superinfected
with combinations of recombinant baculoviruses
capable of expressing each of the viral replication
proteins, it was shown that not the BMRF1
Pol accessory subunit but rather the BALF5 Pol
catalytic subunit directly interacts with the
BBLF4/BSLF1/BBLF2/3 complex. Furthermore,
double infection with pairs of recombinant viruses
revealed that each component of the BBLF4/
BSLF1/BBLF2/3 complex makes contact with the
BALF5 Pol catalytic subunit. The interactions of
the EBV DNA polymerase with the EBV putative
helicase-primase complex warrant particular attention
because they are thought to coordinate leading
and lagging strand DNA synthesis at the replication
fork.
2. Purification of the product of the Epstein-Barr
virus BZLF1 gene
Nakasu, S. and Tsurumi, T.
The product of the BZLF1 gene (pBZLF1) of
Epstein-Barr virus (EBV) encodes a nuclear protein
which is activators of the lytic cycle in cells latently
infected with EBV. pBZLF1 is suggested to activate
the genes required for the lytic cycle and induction
of viral DNA replication as a DNA binding
protein specific for the viral lytic origin of DNA
replication (ori lyt).
In order to understand the role of pBZLF1 in induction
of the lytic cycle, we have been trying to purify
the protein and to characterize its biochemical features.
This was first attempted with insect cells infected
with baculoviruses overproducing pBZLF1,
but the partially purified protein tended to form aggregates
and was eluted with a wide range of the
salt concentrations on ionic chromatography. The
results suggested the conformation of the pBZLF1
produced in the insect cells would not reflect the in
vivo case. Therefore, we next tried to purify
pBZLF1 from B95-8 cells, the cell line latently infected
with EBV. The lytic cycle was induced with
chemical agents such as TPA, sodium n-butyrate,
38
and calcium ionophore, and pBZLF1 could be extracted
with high salt buffer(0.6 to 1 M NaCl). The
pBZLF1 and purified more than 50 fold with the
hydrophobic column chromatography, DEAE
sephacel chromatography, phosphocellulose chromatography
and Heparin agarose chromatography.
However, the protein was not the main component
in the final fraction and further purification procedures
are therefore required.
3. Mechanisms by which Cdc2 kinase inhibits
re-replication during the late
S-G2-M phase in mammalian cells
Fujita, M. and Tsurumi, T.
Genomic DNA needs to be replicated completely
and only once during a single cell cycle and
inhibition of re-replication is one of most important
aspects of cell cycle control to maintain genome
integrity. We have demonstrated that, in mammalian
cells, Cdc2 kinase governs the inhibition of
re-replication during late S-G2-M phase through
prohibition of re-binding of the MCM heterohexameric
complex, an essential DNA replication
initiation factor, to chromatin. MCM complexes
are believed to be loaded onto chromatin by an origin
recognition complex (ORC) and CDC6 protein.
We have suggested that phosphorylation of the
MCM complex and CDC6 protein by Cdc2 kinase
may be one of mechanisms underlying prohibition
of re-binding. Recently, the Cdt1 protein has been
identified as another MCM-loading factor, its function
being proposed to be suppressed through binding
by a counteractive protein, geminin, thus prohibiting
MCM re-binding in the mitotic phase of the
Xenopus egg cell cycle. Interestingly, this geminin-mediated
regulation of Cdt1 function appears
to be independent of mitotic Cdc2 kinase activity.
However, considering our previous experimental
data, it is quite possible that the Cdt1 function is
regulated by Cdc2 kinase during the late phase of
the cell cycle in mammalian somatic cells (see below).
Therefore, we have been more precisely
analyzing mechanisms by which Cdc2 kinase inhibits
re-replication during the late phase of the mammalian
cell cycle, especially focusing on interrelationships
among Cdt1, geminin and Cdc2 kinase.
Using Cdc2 temperature-sensitive mutant murine
FT210 cells, we previously demonstrated that
inactivation of Cdc2 kinase leads to re-binding of
MCM complexes to chromatin during the G2/M
phase. We first investigated intranuclear behavior
of Cdt1 and geminin proteins when Cdc2 is inacti-

vated in FT210 cells. In G2/M phase-enriched
FT210 cells, MCM, Cdt1 and geminin were present
as soluble nucleoplasmic proteins. When Cdc2
kinase was inactivated, re-binding of MCM proteins
was observed, as we reported previously. Under
such conditions, most Cdt1 proteins also became
associated with chromatin, while geminin remained
soluble. It has been reported that the latter can
physically interact with Cdt1. One simple explanation
for the finding, therefore, might be that Cdc2
kinase activity enhances the Cdt1-geminin interaction,
and when Cdc2 is inactivated, Cdt1 becomes
free from geminin binding, then associating with
chromatin and functioning to load MCM. We found
that the Cdc2/cyclinA complex, but not Cdc2/cyclin
B complex, binds to Cdt1 through its Cy motif in
293T human kidney cells, this not interfering with
geminin binding to Cdt1. We also found that
when 293T cells are treated with purvalanol A, a
very specific Cdk2 and Cdc2 inhibitor, both
Cdt1-geminin and Cdt1-Cdc2/cyclin A interactions
are diminished. From these results, our current
working hypothesis is that Cdc2 inhibits Cdt1
re-loading MCM through (1) enhancing geminin
binding to Cdt1 by their phosphorylation and (2)
directly binding as a Cdc2/cyclin A complex to
Cdt1. For confirmation, we are now preparing recombinant
Cdt1, geminin and Cdk/cyclin complexes
to establish in vitro binding assay. We are
also examining whether wild-type or Cy-mutated
Cdt1 can induce re-replication in 293T cells.
4. Immortalization of human cells by HPV
Kiyono, T. and Tsurumi, T.
Normal human cells in culture undergo a limited
number of divisions and then enter a nondividing
state called replicative senescence. Most cancer
cells can divide indefinitely by escaping this senescence
program. The E6 and E7 genes of human
papillomavirus can cooperatively immortalize normal
human epithelial cells originating from skin or
mammal glands. Both inactivation of the RB
pathway by E7 and activation of telomerase by E6
are required for the immortalization. In the past two
years, we have tried to immortalize many cell types,
including: epithelial cells originated from bronchus,
small air ways, esophagus, stomach, mammal- and
prostate glands, endometrium, surface of the ovary,
tooth root, and hair follicle; endothelial cells from
umbilical veins, microvessels, and thoracic ducts;
mesothelial cells from omentum; mesenchymal
stem cells from bone marrow, T- and B lympho-
cytes; and fibroblasts from many tissues. Among
these cell types, only skin fibroblasts could be immortalized
by introduction of TERT, the catalytic
subunit of telomerase reverse transcriptase. Such
introduction proved sufficient for induction of telomerase
activity in all the cell types tested, but insufficient
for immortalization except in the skin fibroblasts.
We are trying to elucidate the underlying
mechanisms which could explain this anomaly.
So far, we found that all the above cell types other
than skin fibroblast showed gradual increase in
p16 Ink4a expression with increasing number of
population doublings, which in turn causes decreased
phosphorylation of RB resulting in cell cycle
arrest. Introduction of E6 and E7 extended the
life span of all the cell types tested, and in some
cases resulted in immortalization with active telomerase.
However, some cell types, including
mesenchymal stem cells from bone marrow,
showed an extended life span with no active telomerase,
and finally stopped growing. These cell
types were eventually immortalized by introduction
of TERT in addition to E6 and E7. These results
support our previous conclusion that both inactivation
of the RB pathway and activation of telomerase
are required for immortalization to many cell types.
However, inactivation of the RB pathway by E7
induced a significantly higher rate of apoptosis as
well as extending life span in cell types including
mesenchymal stem cells, and those introduced with
E7 together with TERT were difficult to propagate,
not because of senescence but because of apoptosis.
At least, practically, those cells required another
gene such as E6 to inhibit apoptosis induced by E7
for immortalization. E6 alone induces telomerase
in some cell types including skin keratinocytes and
mammary epithelial cells, but not in others. Introduction
of E7 together with TERT is a good strategy
to immortalize normal human cells, but is not ideal
for establishing normal human cell lines, because
E7 induces chromosomal instability and sometimes
apoptosis. We are looking for a better strategy to
establish normal human cell lines, which might be
useful not only for many research fields but also in
clinical areas such as regeneration medicine.
5. Longitudinal dynamics of Epstein-Barr
Virus-specific cytotoxic T lymphocytes
in the posttransplant lymphoproliferative
disorder
Kuzushima, K, Kimura, H.* 1 , Hoshino, Y.* 1 , Yoshimi,
A.* 1 , Tsuge, I.* 1 , Horibe, K.* 1 , Morishima, T.* 2 , Kojima,
S.* 1 and Tsurumi, T.
39

The Epstein-Barr virus (EBV)-associated lymphoproliferative
disorder (LPD) is a serious complication
after allogeneic bone marrow transplantation
(BMT). Dynamics of EBV-specific cytotoxic T
lymphocytes (CTL), which are important in controlling
EBV, during LPD have yet to be fully elucidated.
A patient with Wiskott-Aldrich syndrome
was diagnosed as suffering with the LPD on day 47
after BMT. Fluorescence-activated cell sorter
(FACS) analysis for interferon-γ production revealed
more than 70% of the patient’s CD8 + T cells
to be EBV-specific. They were directly cytotoxic to
donor-derived EBV + lymphoblastoid cells, thus being
blocked by an anti-class I antibody.
EBV-specific CD8 + T cell counts declined in parallel
with the EBV genome load and full recovery of
LPD was obtained with relaxation of immunosuppressive
drugs. The results illustrate longitudinal
dynamics of EBV-specific CTL during
post-transplant LPD and feature the advantages of
FACS analysis for EBV-specific CTL for treatment
decision making.
* 1 Departments of Pediatrics/Developmental Pediatrics
and * 2 Health Science, Nagoya University School of
Medicine, Nagoya Japan
6. Identification of HLA A*2402-restricted
cytomegalovirus-specific CD8 + T cell
epitopes by a computer algorithm and
an enzyme-linked immunospot assay
Kuzushima, K., Hayashi, N., Kimura, H.* 1 and Tsurumi,
T.
Antigenic peptides recognized by virus-specific
cytotoxic T lymphocytes (CTLs) are useful tools for
studying CTL responses specifically among those
40
who possess the presenting major histocompatibility
(MHC) class I molecules. For widening the application,
an efficient strategy to determine such
epitopes in the context of a given MHC is highly
desirable. We present here a rapid and effective
method for determination of CTL epitopes through
multiple screenings, consisting of a computer-assisted
algorithm, and MHC stabilization
and enzyme-linked immunospot assays. A major
cytomegalovirus (CMV)-specific CTL epitope,
QYDPVAALF in the amino acid sequence of its
lower matrix 65 kilo dalton phosphoprotein (pp65),
presented by HLA A*2402 molecules was identified
from 83 candidate peptides. The results indicate
that the CMV-specific CTL response is highly
focused on pp65 in the context of HLA A*2402.
Endogenous processing and presentation was confirmed
using a peptide-specific CD8 + T cell clone
as the effector and autologous fibroblast cells infected
with recombinant vaccinia virus expressing
pp65 gene or CMV as the antigen presenting cells.
Flow cytometric analysis of intracellular interferon-γ
production revealed between 0.04 and
0.27 % of CD8 + T cells in peripheral blood of HLA
A24-positive and CMV-seropositive donors to be
specific for the peptide. The tetrameric
MHC-peptide complexes specifically bound to the
reactive T cell clone and 0.79% of CD8 + T cells in
peripheral blood from a seropositive donor. The
peptide could thus be a useful reagent to study CTL
responses to CMV among populations positive for
HLA A*2402.
* 1 Departments of Pediatrics/Developmental Pediatrics,
Nagoya University School of Medicine, Nagoya Japan

Professor C.J.H. van de Velde, from Leiden University Medical Center, Netherlands, giving us the lecture
entitled “Ten Yeras Results of Prospective Randomized D1/D2 Gastric Cancer Trial Limited but
Definitive Benefits“ in the 8th Aichi Cancer Center International Symposium held on February 16,
2002 (see p. 93).
41

42
From left to right
First row: Ms. Keiko Miyazaki, Dr. Akiko Kanamori, Dr. Reiji Kannagi, Ms. Masumi Usui-Nozaki and
Dr. Nozomu Hiraiwa. Second row: Dr. Takaaki Hattori, Mr. Takunori Ogaeri, Dr. Osamu
Taguchi, Ms. Mineko Izawa and Dr. Akinari Watanabe.
Insets, Dr. Satoshi Saito, Dr. Kensuke Kumamoto, Ms. Yoshiko Goto, Dr. Kou Tei and Ms. Sasako
Eguchi.

Division of Molecular Pathology
________________________________________________________________________________
Reiji Kannagi, M.D., D.M.Sc., Chief
Osamu Taguchi, D.M.Sc., Section Head
Nozomu Hiraiwa, M.D., D.M.Sc., Senior Researcher
Akiko Kanamori, Ph.D., Researcher
Kumamoto Kensuke, Research Resident (as of April, 2001)
Mineko Izawa, B.A., Research Assistant
Chikako Mitsuoka, M.T., Research Assistant (until March, 2000)
Yoshiko Goto, D.V.M., Research Assistant
Keiko Miyazaki, M.T., Research Assistant (as of November, 2001)
Sasako Eguchi, Semi-regular Employee
Masumi Usui-Nozaki, Semi-regular Employee
Kayoko Kanda, M.T., Semi-regular Employee (until March, 2000)
Visiting Scientists
Hiroshi Ikeda, M.D., Aichi Medical University
Visiting Trainees
Katsuhiro Ohno, M.D., Nagoya University School of Medicine (until March, 2001)
Kensuke Kumamoto, M.D., Fukushima University School of Medicine (until March, 2001)
Chikako Mitsuoka, M.T., Shiroyama Hospital (as of April 2000, until April, 2001)
Kou Tei, M.D., Kyoto Prefectural University School of Medicine
Satoshi Saito, M.D., Nagoya University School of Medicine
Akinari Watanabe, M.D., Fukushima University School of Medicine (as of March, 2001)
General Summary
Cell adhesion molecules play important roles in infiltrative growth and distant metastasis of cancers, and
expression of functional carbohydrate determinants implicated in cell adhesion is remarkably enhanced upon
malignant transformation of cells. Especially, the carbohydrate determinants, sialyl Lewis a and sialyl
Lewis x, are frequently expressed on human malignant cells in patients with cancers or leukemia. These
determinants serve as ligands for selectins, cell adhesion proteins present on activated human endothelial
cells, and intimately involved in the process of hematogenous metastasis. During the period 2000-2001,
our research interest was concentrated on the following issues; 1) basic study on the ligand requirements of
three members of the selectin family of cell adhesion molecules; 2) expression and functional roles of a
newly found sulfated selectin ligand, sialyl 6-sulfo Lewis x, in solid tumors; 3) mechanisms of specific induction
of sialyl Lewis a and sialyl Lewis x expression upon malignant transformation of cells; and 4) experimental
trials for treatment of cancers by vaccination of MUC1 cDNA with dendritic cells.
1. Study of ligand specificity of three selectin
family cell adhesion molecules, E-,
P- and L-selectins, using genetically
engineered cells
Kanamori, A., Ohmori, K *1 , Goto, Y., Uchimura, K. *2 ,
Muramatsu, T. *2 , Kiso, M. *3 , Tamatani, T. *4 and
Kannagi, R.
Selectins, a family of cell adhesion molecules
with a C-type lectin domain at the outer terminus of
each molecule, have been shown to recognize specific
carbohydrate ligands such as sialyl Lewis X
and sialyl Lewis A. Cell adhesion mediated by
selectin and their carbohydrate ligands is implicated
in recruitment of leukocytes in inflammation, hematogenous
metastasis of cancer cells, and tissue
infiltration of leukemic cells. Recently we identified
sialyl 6-sulfo Lewis x as a major L-selectin
ligand on high endothelial venules of human peripheral
lymph nodes. We further investigated the
ligand activity of sialyl 6-sulfo Lewis x with E- and
P-selectins and made a comparison with the binding
activity of conventional sialyl Lewis x, using cultured
human lymphoid cells expressing both carbohydrate
determinants. The results of the recombinant
selectin binding studies and the non-static
43

monolayer cell adhesion assays indicated both sialyl
6-sulfo Lewis x and conventional sialyl Lewis x
to serve as ligands for E- and P-selectins, while
L-selectin appears quite specific for sialyl 6-sulfo
Lewis x. Treatment with anti-PSGL-1 antibodies
as well as O-sialoglycoprotein endopeptidase almost
completely abrogated the binding of P-selectin
but barely affected the binding of E-selectin. This
indicates that these carbohydrate determinants carried
by O-glycans of PSGL-1 selectively serve as
ligands for P-selectin, while the ligand for
E-selectin is not restricted to PSGL-1 nor to
O-sialoglycoprotein endopeptidase-sensitive glycans.
The binding of L-selectin was markedly reduced
by O-sialoglycoprotein endopeptidase treatment
but only minimally affected by anti-PSGL-1
antibodies, indicating O-glycans carrying sialyl
6-sulfo Lewis x to be the major L-selectin ligands,
while PSGL-1 is only a minor core protein for
L-selectin in these cells. These results indicated
that each member of the selectin family has a distinct
ligand binding specificity.
*1 Department of Biochemistry, Nagoya University,
School of Medicine.
*2 Department of Laboratory Medicine, Kyoto University,
School of Medicine.
*3 Department of Applied Bioorganic Chemistry, Gifu
University, School of Agriculture.
*4 Research Center for Advanced Science and Technology,
the University of Tokyo.
2. Expression of sialyl 6-sulfo Lewis X, a
new ligand for cell adhesion molecules
of the selectin family, in human colon
and cultured colon cancer cells
Izawa, M., Kumamoto, K., Kanamori, A., Kanda, K.,
Goto, Y., Ishida, H. *1 , Nakamura, S. *1 and Kannagi, R.
We recently identified sialyl 6-sulfo Lewis X
determinant as a major ligand for L-selectin on high
endothelial venules of human peripheral lymph
nodes. However, its expression is not limited to
endothelial cells. From our investigation of its
distribution in human colorectal cancer tissues and
cultured colon cancer cells, the sialyl 6-sulfo Lewis
X determinant is preferentially expressed in nonmalignant
colonic epithelium rather than cancer cells
(P < 0.001; n = 23). This is in contrast to the distribution
of conventional sialyl Lewis X, which is
preferentially expressed in cancer tissues rather than
nonmalignant epithelia (P = 0.007; n = 23), indicating
that 6-sulfation predominantly occurs in non-
44
malignant tissues and is suppressed upon malignant
transformation (Fig. 1). In confirmation of this, a
non-sialylated determinant 6-sulfo Lewis X was
also found to be preferentially localized in nonmalignant
epithelium. Significant expression of sialyl
6-sulfo Lewis X was observed in only 2 of 13
cultured colon cancer cell lines, whereas 8 were
positive for conventional sialyl Lewis X. Transfection
of cells with fucosyltransferase (Fuc-T) VI
induced expression of sialyl 6-sulfo Lewis X,
whereas transfection of Fuc-T III did not, suggesting
that the determinant was synthesized mainly by
Fuc-T VI in colonic epithelial cells. Members of
the sialic acid cyclase pathway, the de-N-acetyl
sialyl 6-sulfo Lewis X and cyclic sialyl 6-sulfo
Lewis X determinants, were also found to be preferentially
expressed in nonmalignant epithelium
rather than colonic cancer cells (P < 0.001; n = 23).
Stimulation of the sialyl 6-sulfo Lewis X-positive
colon cancer cell line with a calcium ionophore
ionomycin markedly reduced sialyl 6-sulfo Lewis X
and induced cyclic sialyl 6-sulfo Lewis X expres-
Fig. 1. Sialyl 6-sulfo sialyl Lewis X and conventional
sialyl Lewis X in a colon cancer. Note the
preferential expression of the 6-sulfo form on
non-malignant colonic epithelial cells (upper
panel), while non-sulfated sialyl Lewis X is
strongly expressed on cancer cells (lower panel).

sion. These results suggest that the metabolic
conversion of sialyl 6-sulfo Lewis X into cyclic
sialyl 6-sulfo Lewis X by a calcium-dependent enzyme,
sialic acid cyclase, as we hypothesized for
human leukocytes previously (C. Mitsuoka et al.,
Proc. Natl. Acad. Sci. USA, 96: 1597–1602, 1999),
also occurs in nonmalignant colonic epithelium.
*1 Central Clinical Laboratory, Aichi Cancer Center
Hospital.
3. Regulatory mechanisms for expression
of functional carbohydrate determinants
on malignant and non-malignant
cells:
3-1. Roles of sugar nucleotide transporters
in the enhanced expression of carbohydrate
ligands for selectins, sialyl
Lewis X and sialyl Lewis A, on solid
tumors
Kumamoto, K., Goto, Y., Ishida, N. *1 , Kawakita, M. *1
and Kannagi, R.
Extravasation of malignant cells involves interaction
of carbohydrate ligands on their surfaces
with selectins, cell adhesion molecules on endothelial
cells lining the blood vessels. Several molecular
species of carbohydrate ligands for selectins
are expressed on malignant cells, including sialyl
Lewis X and sialyl Lewis A, especially in solid tumors.
The molecular mechanisms underlying accelerated
expression of sialyl Lewis X/A in cancers
is not well understood.
Cancer-associated induction of some glycosyltransferases
has been assumed to influence expression
of determinants. Recent studies, however,
have indicated that cancer-associated alteration in
sugar transportation and intermediate carbohydrate
metabolism also play important roles in the induction
of sialyl Lewis X/A expression in cancer. A
series of human nucleotide sugar transporters in the
Golgi apparatus were recently cloned, including the
transporters for UDP-galactose (UDP-Gal),
UDP-N-acetylglucosamine (UDP-GlcNAc) and
CMP-sialic acid (CMP-SA). We have examined the
mRNA expression of these three transporters in
human colon cancer tissues by reverse transcription-PCR
analysis in comparison with that in nonmalignant
colonic mucosa prepared from the same
patients. The amount of mRNA for UDP-Gal transporter
was significantly increased in colon cancer
tissues compared with nonmalignant mucosa (P =
0.035; n = 20) (Fig. 2). The increase was more
prominent in patients with advanced colorectal cancer
of Dukes’ stages C and D, in which the amount
of UDP-Gal transporter mRNA showed on average
about a 3.6-fold increase over paired nonmalignant
samples (statistically significant at P = 0.004; n =
14). The mRNA content of the other two
transporters showed no significant difference between
the paired cancer and normal tissues. When
UDP-Gal transporter cDNA was stably transfected
into cultured human colon cancer cells, expression
of Thomsen-Friedenreich (TF) antigen and of sialyl
Lewis A (NeuAcα2→3Galβ1→3[Fucα1→4]Glc
NAcβ1→ R) and sialyl Lewis X (NeuAcα2→3Gal
β1→4[Fucα1→3]GlcNAcβ1→R) determinants was
significantly induced on transfectant cells, resulting
in markedly enhanced cell adhesion to vascular
E-selectin. These findings suggest that increase of
UDP-Gal transporter mRNA is involved in enhanced
expression of cancer-associated carbohydrate
determinants such as TF and sialyl Lewis A/X
Fig. 2. Typical examples of RT-PCR analyses of nucleotide sugar transporter gene expression in human colon
cancer tissues and non-malignant mucosa. After PCR reaction using the specific primers, the aliquots of
products were electrophoresed in 2% agarose gel and were stained with ethidium bromide. Sizes of each
band are indicated in basepairs (bp). Ca, cancer tissues; N, non-malignant mucosa prepared from the
same patient. UDP-Gal T, UDP-Gal transporter; G3PDH, glyceraldehyde 3-phosphate dehydrogenase.
45

antigens in colon cancers.
*1 Department of Physiological Chemistry, The Tokyo
Metropolitan Institute of Medical Science, Tokyo.
3. Regulatory mechanisms for expression
of functional carbohydrate determinants
on malignant and non-malignant
cells:
3-2. A T-box transcriptional factor that synergizes
with HTLV-1 Tax in transactivating
the selectin-ligand synthesizing
enzyme, fucosyltransferase VII
Hiraiwa, N. and Kannagi, R.
Sialyl Lewis x is reportedly to be expressed on
leukocytes as a ligand for selectins on vascular endothelium.
Molecular-biological studies have revealed
that its synthesis is critically regulated by the
key enzyme, fucosyltransferase VII (Fuc-T VII) in
leukocytes. We recently found that adult T-cell
leukemia malignant cells express sialyl Lewis x and
Fuc-T VII, increased expression of mRNA for the
latter actually being generated by Tax protein derived
from the human T-cell leukemic virus type 1
(HTLV-1) virus. We have demonstrated that Tax
activates the promoter of Fuc-T VII at a CRE-like
(cAMP-responsive element) site approx. 150-bp
away from the initiator of the gene. Detailed
studies revealed that the sequence of the CRE-like
site where CREB-1, other CREB/ATF transcriptional
factors can bind, resembles the 21-bp sequence
of the HTLV-1 long-terminal repeat (LTR)
and is critical for Tax-activation. We therefore
postulate the presence of unknown factor(s) with
the ability to bind to the site and associate with
other factors like members of CREB/ATF or Tax.
To clarify this possibility, we conducted a
one-hybrid experiment with a cDNA library from
ATL-related cells, cloned several binding-factors,
and demonstrated that a new T-box type transcriptional
factor, F7CAF-1 (Fuc-T VII CRE-associated
Factor), associates with CREB-1 and Tax at the
CRE-like site to facilitate Tax-induced Fuc-T VII
activation. The CRE-like site of the promoter of
the Fuc-T VII is akin to the semi-palindromic sequence
of the reported T-box binding site. The
exact sequence to which the F7CAF-1 binds remains
to be elucidated. Using JPX-9 cells, a derivative
of Jurkat T-cells carrying the transfected
Tax gene under the control of metallothionein promoter,
the induction of Tax was found to result in
appearance of mRNA of F7CAF-1 along with the
46
Fuc-T VII. A reporter assay with expression-plasmids
for the F7CAF-1, CREB-1 or Tax
showed F7CAF-1 to transactivate Tax-induced expression
of Fuc-T VII in cooperation with CREB-1.
Immunoprecipitation studies revealed association of
F7CAF-1 with Tax and CREB-1, indicating that the
complex formed with these factors might carry out
activation of Fuc-T VII in ATL cells. Northern
blots showed that F7CAF-1 is expressed in lung,
liver, and also in spleen, lymph nodes, and bone
marrow, closely related to expression of Fuc-T VII.
In ATL-related cell lines, F7CAF-1 was found to be
remarkably transcribed in association with Fuc- T
VII, but other kinds of T-cell leukemic/lymphoma
cell lines and B-cell leukemic/lymphoma cell lines
proved negative.
These results indicate that ATL cells overexpress
the Fuc-T VII gene that can be transactivated
by HTLV-1 Tax in concert with a T-box factor,
F7CAF-1. Cloning of F7CAF-1 should facilitate
research on Tax-induced transactivation of various
genes and elucidate underlying mechanisms
4. A murine model of tumor suppression
by vaccination with MUC1 DNA and
dendritic cells
Kontani, K. *1 and Taguchi, O.
Among specific approaches to cancer immunotherapy,
DNA vaccines are thought to be more applicable
for clinical use than other methods, such as
vaccines with peptides or autologous cancer cells,
or adoptive transfer of cytotoxic T lymphocytes
(CTLs), for the following reasons: 1) DNA vaccines
are inexpensive and simple to use once the DNA
vector is prepared; 2) adjuvants are usually unnecessary
for DNA vaccination; 3) high levels of antigen
expression can be maintained; 4) autologous
immune cells or cancer cells are not needed; and 5)
facilities and techniques for cell culture are not necessary.
Therefore, DNA vaccines targeting tumor
antigens have great potential for anti-cancer
immunotherapy.
MUC1 antigen is abundantly expressed on
breast, pancreas, lung, and colon cancer cells, eliciting
strong anti-tumor immunity in hosts. The
tandem repeat domain on its core protein contains
antigenic epitopes which are recognized by CTLs in
both mice and humans. Therefore, MUC1 is suitable
target for cancer immunotherapy.
In order to induce specific anti-tumor immunity
in mice, we attempted to immunize C57BL/6 mice
with a DNA vaccine encoding the MUC1 polypep-

tide. When mice immunized with MUC1 DNA
were challenged with EL4-muc, MUC1-transfected
syngeneic lymphoma cells, they completely prevented
tumor growth. In contrast, when the vaccine
was given to EL4-muc tumor-bearing mice,
suppression was not observed. However, activated,
but non-primed dendritic cells (DCs) obtained from
syngeneic mice, if applied simultaneously with the
MUC1 DNA vaccine to the same sites of EL4-muc
tumor-bearing mice, tumor growth was markedly
suppressed with prolongation of survival. MUC1
antigen could be detected on the dendritic cells at
the vaccination site and in regional nodes in the
targeted mice, which showed strongly enhanced
cellular immune responses specific for MUC1 as
compared to those in mice vaccinated with MUC1
DNA alone. No significant difference in titers of
antibodies to MUC1 between the two groups was
observed, suggesting that non-primed DCs inoculated
into DNA vaccine sites are essential for eliciting
strong anti-tumor cellular immunity to suppress
tumor growth efficiently. This animal model
should prove useful for developing DNA vaccines
for anti-cancer immunotherapy.
*1 Second Department of Surgery, Shiga University of
Medical Science.
47

48
From left to right
First row: Dr. F. Hirose, Dr. K. Nagata, Mr. Y. Yasui, Dr. I. Izawa, Dr. M. Inagaki and Ms. M.
Nishizawa.
Second row: Ms. T. Yuhara, Ms. N. Saitoh, Ms. A. Kawajiri, Ms. Y. Hayashi, Ms. N. Ohshima, Dr. N.
Hanai, Mr. T. Oguri, Dr. T. Yokoyama, Mr. T. Siromizu and Dr. K. Ohtakara.
Insets: Dr. K. Ohno, Dr. M. Yamaguchi, Dr. H. Yoshida and Dr. M. Kato.

Division of Biochemistry
________________________________________________________________________________
Masaki Inagaki, M.D. Chief
Koh-ichi Nagata, M.D. Section Head
Masamitsu Yamaguchi, Ph.D. Section Head (until June, 2001)
Ichiro Izawa, M.D. Senior Researcher
Hiroyasu Inada, M.D. Senior Researcher (until June,2001)
Fumiko Hirose, Ph.D. Senior Researcher
Miwako Nishizawa, B.P. Senior Research Assistant
Yoshihiro Yasui, M.P. Research Assistant
Noriko Saito, B.M.T. Research Assistant
Yoshio Nishimoto, B.S. Senior Research Assistant (until March 2001)
Yuko Hayashi, B.S. Research Assistant
Visiting Scientists
Hidemasa Goto, Domestic Research Fellow, Japan Science and Technology Corporation (until October,2001)
Visiting Trainees
Hideaki Togashi, M.S. Faculty of Science,Nagoya University,Graduate School of Science (until March,2001)
Mihoko Takagishi, B.P. Faculty of Pharmaceutical Science, Nagoya City University (until March,2001)
Seiaya Matsui, B.P. Faculty of Pharmaceutical Science, Nagoya City University (until March,2002)
Kazuhiro Ohtakara, M.D. Department of Pediatrics, Mie University School of Medicine
Aie Kawajiri, M.S. Department of Pathology, Nagoya University School of Medicine (as of April,2000)
Katsuhiko Ohno, Ph.D. Graduate School of Science, Nagoya University (until September, 2001)
Eun-Jeong Kwon, M.S. Graduate School of Science, Nagoya University
Hideki Yoshida, M. S. Faculty of Science and Technology, Tokyo Science University (until September, 2001)
Masaki Kato, M.S. Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology (until
September, 2001)
General Summary
Research in Division of Biochemistry is concerned with the regulation of tumor development, invasion
and metastasis, and our attention is focused on four specific areas: (1) Phosphorylation-dependent regulation
of elements of the cytoskeleton such as intermediate filaments, intermediate filament associated proteins and
septin proteins and cell adhesion molecules; (2) Identification and functional analysis of protein kinases involved
in cell division; (3) A search for intermediate filament binding proteins to elucidate mechanism of
regulation of the cell adhesion machinery, including hemidesmosome and desmosome functions; and (4)
Transcriptional regulation of genes related to cell proliferation.
1. Aurora B and Rho-kinase regulate
cleavage furrow-specific vimentin
phosphorylation in the cytokinetic process
Yasui, Y., Goto, H., Kawajiri, A. *1 , Nigg, E.A. *2 , Terada,
Y. *3 , Tatsuka, M. *4 , Matsui, S. *5 , Manser, E. *6 , Lim, L. *6,7 ,
Nagata, K. and Inagaki, M.
Vimentin, one of the type III intermediate filament
(IF) proteins, is expressed not only in mesenchymal
cells but also in most types of tumor cells.
We have introduced several types of vimentin mutated
at putative phosphorylation sites in its
amino-terminal head domain into type III
IF-negative T24 cells. Site-specific mutation induced
the formation of an unusually long
bridge-like IF structure between unseparated
daughter cells, although these mutants formed a
filament network similar to that in wild type interphase
cells. Among the phosphorylation sites
vimentin-Ser72 was one mutation site essential for
this phenotype. We further demonstrated that the
novel phosphorylation site, vimentin-Ser72, was
phosphorylated specifically at the cleavage furrow
during cytokinesis.
Aurora B is a kinase involved in cytokinesis, but
49

little is known about its target substrates. We provided
evidence that vimentin-Ser72 is phosphorylated
specifically at the border of the Aurora
B-localized area from anaphase to telophase during
mitosis. Expression of a dominant-negative mutant
of Aurora B leads to a reduction in cleavage furrow-specific
phosphorylation. We have identified
Ser6, Ser24, Ser38, Ser46, Ser64, Ser65, Ser72, and
Ser86 on vimentin as Aurora B phosphorylation
sites in vitro. Mutations in there were found to
induce the formation of the unusually long
bridge-like IF structure. Together with finding
that the cyclic AMP-dependent protein kinase active
form and p21-activated kinase, possible
vimentin-Ser72 kinases, are not localized at cleavage
furrow during mitotic phase, the results indicate
that Aurora B regulates cleavage furrow-specific
vimentin phosphorylation and that functional alteration
ensues.
We previously demonstrated vimentin-Ser71,
which is phosphorylated by Rho-kinase, to be also
phosphorylated specifically at the cleavage furrow.
Thus, the cleavage furrow-specific vimentin phosphorylation
may be regulated by at least two distinct
kinases (Aurora B and Rho-kinase). We
propose that continuous furrow ingression may increase
vimentin accessibility not only to Aurora B
in the spindle midzone but also to Rho-kinase in the
plasma membrane of the cleavage furrow.
*1
Department of Pathology, Nagoya University School
of Medicine, Nagoya, Aichi 466-8550, Japan.
*2
Max-Planck Institute for Biochemistry, Department
of Cell Biology, D-82152 Martinsried, Germany.
*3
Department of Genetics, Cell Biology and Development,
University of Minneapolis, MN55455, USA.
*4
RIRBM, Hiroshima University, Hiroshima 734-8553,
Japan.
*5
Department of Biological Chemistry, Faculty of
Pharmaceutical Science, Nagoya City University,
Nagoya 467-8603, Japan.
*6
Glaxo-IMCB Group, Institute of Molecular & Cell
Biology, Singapore 117609, Singapore.
*7
Institute of Neurology, University College London,
London WC1N 1PJ, UK.
2. Aurora B phosphorylation of histone H3
at serine28 prior to the mitotic chromosome
condensation
Goto, H. *1 , Yasui, Y., Nigg, E.A. *2 and Inagaki, M.
Histone H3 (H3) phosphorylation plays important
roles in mitotic chromosome condensation.
50
We have reported that H3 phosphorylation occurs
not only at Ser10 but also at Ser28 during mitosis,
at least in mammals. Aurora B was recently
demonstrated to be responsible for Ser10 phosphorylation
in S. cerevisiae, C. elegans, Drosophila,
and a Xenopus egg extract. To determine whether
Aurora B might phosphorylate H3 not only at serine10
but also at serine28 in mammals, we compared
the distribution of the enzyme with that of H3
phosphorylation. Aurora B was found to be primarily
localized in heterochromatin of late G2
phase cells where only Ser10 phosphorylation was
observed. Treatment of such cells with calyculin
A induced Ser28 phosphorylation in the Aurora
B-localized area. During prophase to metaphase,
Aurora B becomes distributed in condensing chromosomes
demonstrating Ser10 and Ser28 phosphorylation.
Aurora B can phosphorylate
H3-Ser10 and -Ser28 in nucleosomes in vitro,
transfection of a dominant-negative mutant resulting
in reduction of H3 phosphorylation not only at
Ser10 but also Ser28 during mitosis. This occurs
during mitotic chromosome condensation, and the
level of Ser28 phosphorylation is diminished to an
undetectable level by PP1 phosphatase prior to entry
into mitosis.
*1
Division of Signal Transduction, Nara Institute of
Science and Technology
*2
Max-Planck Institute for Biochemistry, Department
of Cell Biology, D-82152 Martinsried, Germany.
3. Keratin attenuates tumor necrosis factor-induced
cytotoxicity through association
with TRADD
Inada, H., Izawa, I., Nishizawa, M., Fujita, E. *1 , Kiyono,
T., Takahashi, T., Momoi, T. *1 and Inagaki, M.
Keratin 8 and 18 (K8/18) are the major components
of intermediate filament (IF) proteins of simple
or single-layered epithelia. Recent data show
that normal and malignant epithelial cells deficient
in K8/18 are nearly 100 times more sensitive to tumor
necrosis factor (TNF)-induced cell death. We
have now identified the human TNF receptor 1
(TNFR1)-associated death domain protein
(TRADD) to be a K18-interacting protein. Among
IF proteins tested in two-hybrid systems, TRADD
specifically bound to K18 and K14, type I (acidic)
keratins, the COOH-terminal region of TRADD interacting
with the coil Ia of the rod domain of K18.
Endogenous TRADD coimmunoprecipitated with
K18, and colocalized with K8/18 filaments in hu-

man mammary epithelial cells. Overexpression of
the NH2-terminus (aa 1-270) of K18 containing the
TRADD-binding domain as well as overexpression
of K8/18 in SW13 cells, which are devoid of keratins,
render the cells more resistant to killing by
TNF. We also showed that the overexpressed
NH2-termini of K18 and K8/18 associate with endogenous
TRADD in SW13 cells, resulting in inhibition
of activation of caspase-8. These results indicate
that K18 may sequester TRADD to attenuate
its interactions with activated TNFR1 and moderate
TNF-induced apoptosis in simple epithelial cells.
*1 Division of Development and Differentiation, National
Institute of Neuroscience, NCNP, Kodaira,
Tokyo 187-8502, Japan
4. ERBIN associates with p0071, an armadillo
protein, at cell-cell junctions of epithelial
cells
Izawa, I., Nishizawa, M., Tomono, Y. *1 , Ohtakara, K.,
Takahashi, T. and Inagaki, M.
ERBIN, an ErbB2 receptor-interacting protein,
belongs to a recently described family termed LAP
[leucine-rich repeats and PSD-95/dlg-A/ZO-1
(PDZ) domains], that plays essential roles in establishment
of cell polarity. To identify new ER-
BIN-binding proteins, we screened a yeast
two-hybrid library, using the carboxyl-terminal
fragment of ERBIN containing a PDZ domain as
bait, and isolated p0071 (also called plakophilin-4)
as an ERBIN-interacting protein. p0071 is a
member of the p120 catenin family, defined as proteins
with 10 armadillo repeats, and is localized
along cell-cell border. ERBIN PDZ domain binds
the COOH-terminus of p0071 containing the PDZ
domain-binding sequence, and endogenous ERBIN
was found to be co-immunoprecipitated with p0071.
In fully polarized Madin-Darby canine kidney
(MDCK) cells, ERBIN co-localized largely with
β-catenin and partly with desmoplakin along the
lateral plasma membrane. At these cell-cell contact
regions, ERBIN co-localizes with p0071.
Overexpression of the dominant active forms of
Cdc42, Rac1, or RhoA, Rho family small GTPases,
resulted in the marked accumulation of ERBIN at
cell-cell contacts of MDCK and HeLa cells.
These results show that ERBIN interacts in vivo
with p0071 and may be involved in the organization
of adherens junctions and desmosomes of epithelia.
In addition, we have demonstrated that the subcellular
localization of ERBIN might be regulated by
Rho family small GTPases. These observations
should pave the way toward further research on cell
polarity and adhesion as well as generating understanding
of pathological mechanisms of cancer.
*1 Division of Molecular and Cell Biology, Shigei
Medical Research Institute, Okayama 701-0202, Japan
5. Characterization of a mammalian septin
MSF-A
Nagata, K., Kawajiri, A. *1 , Saito, N., Togashi,H.,
Takagishi, M., Matsui, S., Hotani, H. *2 and Inagaki, M.
Septins are a family of conserved proteins implicated
in cell growth and cell cycle regulation, although
their properties and modes of action are largely
unknown. A septin termed MSF (MLL
septin-like fusion) has been identified as a fusion
partner gene in mixed lineage leukemia (MLL) in a
case of therapy-related acute myeloid leukemia with
a t(11;17)(q23;q25). Two alternative splicing
variants, MSF-A and MSF-B, have now been identified
while another report documented a complicated
transcriptional pattern of MSF. MSF has
been found to be mutated in some cases of breast
and ovarian cancers and is considered to be a candidate
tumor suppressor gene. The mutations may
be associated with allelic loss of the 17q25 region.
Although these findings have provided insights into
a possible role for MSF in leukemogenesis and oncogenesis,
and the molecular mechanism(s) linking
MSF function and tumorigenesis and also biochemical
and biological properties of MSF proteins
remain to be elucidated.
We therefore carried out an immunocytochemical
and biochemical characterization of MSF-A,
using an antibody specific for MSF subfamily proteins.
Expression was found to be predominantly
in mammary HMEC cells, in associated with microtubules
in interphase but the mitotic spindle and
bundle of microtubule in the midzone during mitosis.
Biochemical analysis revealed direct binding
of MSF-A with polymerized tubulin through its
central region containing guanine nucleotide-interactive
motifs, although GTPase activity
was not required for the association. Conditions
that disrupted the microtubule network also
disrupted the MSF-A-containing filament structure,
resulting in a punctate cytoplasmic pattern.
Unlike Nedd5, a septin thought to be involved in
cytokinesis, a MSF mutant deficient in GTPase activity
was found to form filament indistinguishable
51

from those of the wild type. These results indicate
that the interaction of MSF-A with microtubule
might be an important mechanism regulating a variety
of septin-dependent cellular processes.
*1
Department of Pathology, Nagoya University School
of Medicine
*2
Division of Biological Sciences, Graduate School of
Science, Nagoya University
6. Functional analysis of DREF using
transgenic flies
Hirose, F., Ohshima, N., Kwon, E-J. #1 , Yoshida, H. #2 ,
Inoue, Y.H. #3 , Matsukage, A. #4 and Yamaguchi, M. #2
The promoters of Drosophila genes encoding
DNA replication-related proteins contain transcription
regulatory elements DRE (5'-TATCGATA) in
addition to E2F recognition sites. A specific
DRE-binding factor, DREF, positively regulates
DRE-containing genes. In addition, it has been
reported that DREF can bind to a sequence in the
hsp70 scs' chromatin boundary element that is also
recognized by boundary element-associated factor
(BEAF) , and thus DREF may play a role in regulating
insulator activity. To examine DREF functions
in vivo, we have established transgenic flies in
which ectopic expression of DREF is targeted to the
eye imaginal discs. Adult flies expressing DREF
exhibited a severe rough eye phenotype, featuring
ectopic DNA and abolition of photoreceptor differentiation.
Furthermore, DREF expression caused
apoptosis in the imaginal disc cells. The DREF
induced rough eye phenotype could be suppressed
by a half dose reduction of the E2F gene, one of the
genes under DREF regulation, indicating that the
DREF overexpression phenotype is useful for
screening for modifiers of DREF activity. Among
Polycomb/trithorax-group genes, we found that half
dose reduction of some of trithorax-group genes
involved in determining chromatin structure or
chromatin-remodeling (brahma, moira and osa)
significantly suppressed while that of Distal-less
enhanced the DREF-induced rough eye phenotype.
The results suggest the possibility that DREF activity
might be regulated by protein complexes that
play roles in modulating chromatin structure.
To examine DREF functions in developing tissues,
overexpression was accomplished in wing
imaginal discs using a GAL4-UAS targeted expression
system in Drosophila. A notching wing
phenotype was induced, associated with ectopic
apoptosis. In addition, overexpression of the 32
52
kDa boundary element-associated factor (BEAF-32),
suggested to compete against DREF for common
binding sites in genomic regions, rescued the
DREF-induced notching wing phenotype, while a
half reduction of the genomic region, including the
BEAF-32 gene, exerted enhancing effects. To our
knowledge, this is the first evidence for any genetic
interaction between DREF and BEAF-32. The
DREF-induced notching wing phenotype is caused
by induction of apoptosis in the Drosophila wing
imaginal disc.
Current addresses
#1
Massachusetts General Hospital.
#2
Division of Biotechnology, Kyoto Institute of Technology.
#3
Drosophila Genetic Resource Center, Kyoto Institute
of Technology.
#4
Chemical and Biological Sciences, Japan Women’s
University.
7. Functional analysis of BEAF32A using
transgenic flies
Yamaguchi, M. #1 , Yoshida, H. #1 , Hirose, F., Inoue,
Y.H. #2 , Hayashi, Y., Yamagishi, M. #3 , Nishi Y., Tamai,
K. #4 , Sakaguchi, K. #5 and Matsukage, A. #6
Transgenic flies were established in which ectopic
expression of boundary element-associated
factor (BEAF) 32A was targeted to the Drosophila
eye imaginal disc. The adult fly eyes were found
to display a severe rough eye phenotype, most ommatidia
being fused with generation of irregularly
shaped rabdomeres. In the developing eye imaginal
disc, expression of BEAF32A inhibited differentiation
of photoreceptor cells and also induced
extensive apoptosis of eye imaginal disc cells.
Consistent with this, co-expression of baculovirus
P35 in the eye imaginal disc suppressed the
BEAF32A-induced rough eye phenotype. To investigate
the effects of BEAF32A on regulation of
chromatin structure, genetic crosses of
BEAF32A-overexpressing flies with loss-of-function
mutants for genes encoding other boundary
element-binding factors or regulators of chromatin
structure were conducted. Interestingly half-dose
reduction of the su(Hw) gene strongly enhanced the
rough eye phenotype induced by BEAF32A. Furthermore,
genetic crosses of the transgenic flies
with loss-of-function mutants for genes interacting
with Polycomb revealed specific links between
BEAF32A and genes such as Distal-less and kohtalo,
suggesting a relation to the chromatin insulator

chromatin insulator function of BEAF. In addition,
genetic crosses of transgenic flies expressing
BEAF32A with a collection of Drosophila deficiency
stocks allowed us to identify several genomic
regions, deletions of which caused enhancement
or suppression of the BEAF32A-induced
rough eye phenotype. The transgenic flies established
in this study should be useful to identify targets
of BEAF32A and its positive or negative regulators
in Drosophila.
Present addresses
#1 Division of Biotechnology, Kyoto Institute of Tech-
nology.
#2
Drosophila Genetic Resource Center, Kyoto Institute
of Technology.
#3
Neuroscience Research Institute, National Institute of
Advanced Industrial Science and Technology.
#4
Medical and Biological Laboratories Corporation
Limited.
#5
Department of Applied Biological Science, Science
University of Tokyo.
#6
Chemical and Biological Sciences, Japan Women’s
University.
53

54
From left to right
First row : Ms. Y. Hayashi, Ms. H. Fukami, Ms. Y. Iwata, Ms. S. Matsumoto and Dr. Y. Yamane.
Second row: Mr. H. Nakamura, Dr. K. Ishizaki, Ms. S. Abe, Mr. Y. Nishimoto and Dr. H. Kuminoto.

Division of Central Laboratory & Radiation Biology
________________________________________________________________________________
Kannji Ishizaki, Ph.D. Chief
Hiroshi Kumimoyo, Ph.D. Researcher
Yoshio Nishimoto, B.S. Senior Research Assistant
Hiroko Fukami, Research Assistant
Nobuhiro Uchida, Ph.D. Researcher (until February 2001)
Kennzou Ohtsuka, Ph.D. Section Head (until March 2001)
Mami Hata, B.P. Senior Research Assistant (until March 2001)
Visiting Trainees
Hideaki Nakamura, M.S. School of Medicine, Nagoya University
Yoshihiro Yamane, Ph.D. Faculty of Science , Nagoya University
General summary
Our main research project is analysis of the molecular genetics of human esophageal tumors. In resent
years, our effort has been focused on identification of a candidate tumor suppressor gene on 13q12-13, which
may be closely related to the prognosis of esophageal tumors. So far we have analyzed several candidate genes
located in or near this region, such as the Rb, BRCA2, AS3 and LATS2 genes. However, tumor specific
mutations were not detected. Now, we are intensively searching for a new gene in this region by constructing
a fine deletion map with esophageal tumors.
We are also studying interaction provide between genetic polymorphism and life-style factors with reference
to esophageal tumors to provide clue to effective prevention. We have found that a specific allele of the
L-myc gene is associated with induction of esophageal tumors in individual with smoking and drinking habits.
We are also now analyzing polymorphisms in other genes.
Another research project is to study genetic effects of low dose radiation on human cells. For this purpose
we have established immortal cell lines derived from normal controls and patients with radiation sensitive
genetic diseases by introducing the human telomerase gene. These cell lines are immortal but without any
change in p53 and other genes that are involved in cellular signal transduction, exhibiting normal responses
to low dose radiation. Using these cell lines we are now studying induced mutations.
1. Analysis of a candidate tumor suppressor
gene, LATS2, on 13q11-12 in
esophageal squamous cell carcinoma
Ishizaki, K., Fujimoto, J. *1 , Kumimoto, H., Nishimoto,
Y., Shimada, Y. *2 , Shinoda, M. *3 and Yamamoto, T. *1
We previously reported that loss of heterozygosity
on 13q12-13 is related to a poor prognosis
with esophageal cancer. We intensively screened
esophageal tumor tissues and cell lines for genetic
change in the Rb, BRCA2, and AS3 genes, located in
or near this region. But none of these genes was
found to exhibit a significant number of tumor specific
mutations, indicating the existence of another
tumor-suppressor gene in this region. Recently,
LATS2, a new human homologue of the Drosophila
tumor suppressor gene (lats/warts) was found on
13q11-12. We therefore screened esophageal tumor
cell lines and tumor tissues to detect tumor specific
mutations in this gene by PCR-SSCP and direct sequencing
with genomic DNA and cDNA. Although
we found 5 different polymorphisms (4 single-base
changes and a 6 bp insertion), a tumor specific mutation
was identified in only one out of 60 tumor
tissues. These results indicated that the LATS2 gene
is inactivated only rarely in esophageal tumors, if at
all, and that there is still another tumor suppressor
gene in this region. We are now constructing a fine
deletion map for its identification.
*1
Department of Oncology, Institute of Medical Science,
University of Tokyo
*2
Department of Surgery and Surgical Basic Science,
Graduate School of Medicine, Kyoto University
*3
Department of Thoracic Surgery, Aichi Cancer Center
Hospital
55

3. Establishment of immortal normal and
ataxia telangiectasia fibroblast cell lines
by introduction of the hTERT gene
Nakamura, H., Fukami, H., Kiyono, T. * 1 and Ishizaki,
K.
For analyzing effects of low doses and
low-dose-rate radiation on human cells, signal
transduction system including p53 responses are
important. However, they may not function normally
in cells by immortalized introduction of SV
40 or in cancer cells. Recently, however it has been
reported that cellular response may be maintained
in human cells immortalized by introduction of the
human catalytic subunit of telomerase (hTERT)
gene.
Therefore we introduced the hTERT gene with a
retrovirus vector into both normal (SuSa) and ataxia
telangiectasia (AT10S) fibroblast cells to establish
immortal cell lines (SuSa/T-n and AT10S/T-n). After
hTERT introduction, these cells continued to
grow beyond a population doubling number 300
with no indication of senescence, like flat shape and
400
350
300
250
200
150
100
50
0
0 100 200 300 400 500 600 700 800
Culture Day
Fig. 1
SuSa/neo
SuSa/T-n
AT1OS/T-n
reduced growth rate (Fig. 1). Induction of p53,
phosphorylation of Ser15 in p53, and induction of
p21 by X-ray irradiation in SuSa/T-n were not affected
by the hTERT introduction. Both SuSa/T-n
and AT10S/T-n cells exhibited an apparent inhibition
of growth, like the parental cells when they
reached confluence. Karyotype analysis has revealed
that they are in the diploid range. SuSa/T-n
cells maintained their original radiosensitivity (Fig.
2), while the AT10S/T-n line appears to be slightly
more resistant than the original cells. However, the
difference is very small, and AT cells are still much
more sensitive than normal cells.
These results suggest that cells immortalized by
hTERT introduction retain their original characteristics,
except for immortalization, and that they may
be useful for analyzing various effects of radiation.
We are now analyzing the influence of low dose
rate radiation on human cells using these immortal
cells as today.
*1
Division of Virology, Aichi Cancer Center Research
Institute
0.001
0 2 4 6 8 10
Dose (Gy)
Fig. 1. Growth curves after hTERT-introduction. Both hTERT-introduced cells (SuSa/T-n and AT1OS/T-n) demonstrate
growth far beyond the point at which control cells (SuSa) stopped growing.
Fig. 2. X-ray sensitivity of original (SuSa and AT1OS) and immortalized cells (SuSa/T-n and AT1OS/T-n).
Both immortalized lines show radiosensitivity similar to their parental cells. Bars in figure indicate standard
deviations (n = 3).
1
0.1
0.01
Fig. 2
SuSa/T-n
SuSa
AT1OS/T-n
AT1OS
57

58
From left to right
First row: Ms. Y. Kanie, Mr. M. Terashima and Ms. M. Yamamoto, second row: Dr. K. Tanabe, Dr. H.
Nakamura and Mr. Y. Minoura

Central Service Unit
________________________________________________________________________________
Kazushi Tanabe, M.S., D.M.Sc. Section Head (until March 2002)
Hiromu Nakamura, D.M.Sc. Section Head (as of April 2002)
Morio Terashima, B.A. Senior Research Assistant (Technical photography specialist)
Yasushi Minoura, B.P. Senior Research Assistant
Masami Yamamoto, D.V.M. Research Assistant (as of April 2002)
Mikio Hagino Research Assistant (Animal care specialist)
General Summary
The Central Service Unit was formerly known as the Biophysics Unit, but the Unit name was changed in
April 2000. In addition, the past section head of the Unit, Dr. K. Tanabe retired in March 2002, and Dr. H.
Nakamura has succeeded him.
Over the past several years, we concentrated on the planning and construction of our new Institute
building, which was finally completed in January 2002. On the opening of new building, the maintenance
and management of the fixtures and facilities of the new building, such as the air conditioning system,
security system, water purifying system, waste water treatment system, carbon dioxide gas supply system
and the experimental animal facilities became our duties.
The Central Service Unit fulfills many functions in assisting the investigations performed by the Institute,
and has responsibilities for the maintenance and operation of various instruments for biotechnology research.
These are the protein sequencer (ABI 476A), DNA sequencer (ABI 3100), flow-cytometer
(Becton-Dickinson FACSCalibur), high performance liquid chromatography equipment (Waters), imaging
analyzers (Fujix BAS2500Mac, Amersham-Pharmacia Imagemaster and FluorImager 595), X-ray machine
(Hitachi), electron microscopes (JEOL and Hitachi), confocal laser scanning microscope (Bio-Rad Radiance),
real-time PCR analyzer (Roche Light Cycler), ultracentrifuges (Beckman and Hitachi), and computer system
for image treatment. Furthermore, the Central Service Unit maintains and manages the radioisotope
experiment facilities, SPF and conventional animal facilities, the laboratory of translational research,
technical photographic work, hazardous chemical storage, ultra-low temperature freezers, cold rooms, liquid
nitrogen storage room, LAN system, and contributes to many other aspects of the Institute's functions. Our
activities provide background support to all of the investigations carried out by the Research Institute.
59

Librarians
________________________________________________________________________________
60
From left to right
Librarians, Ms. K. Adachi, Ms. S. Mori, Ms. M. Teratani and Ms. T. Yasuda, supporting
scientific and medical informations.

A transmission electron micrograph of an adult compound eye from a wild-type fly elaborated by Dr.
Fumiko Hirose, a senior researcher in the Division of Biochemistry, adopted as the cover photograph of
Molecular and Cellular Biology, volume 22, issue 8. Ectopic expression of transcription factor DREF (for
DNA replication-related element-binding factor) in Drosophila melanogaster imaginal discs induces DNA
synthesis, apoptosis, and unusual morphogenesis and results in a severe rough eye phenotype (see
Publication J043).
62

Researches Supported by Special Project Programme
________________________________________________________________________________
1. Identification of tumor-associated
antigens recognized by T cells
infiltrating Epstein-Barr virus-positive
gastric carcinomas
Kuzushima, K., Nakamura, S. *1 , Nakamura, T. *2 ,
Yamamura, Y. *3 , Hayashi, N. and Tsurumi, T.
Gastric adenocarcinomas carrying the
Epstein-Barr virus (EBV) are known to be
accompanied by massive lymphocyte infiltration.
To characterize the tumor infiltrating lymphocytes
(TILs), we isolated and cultured such cells from
surgically resected lesions. They were found to be
positive for CD3 and CD8 and consists of
HLA-class I-restricted CD8 + cytotoxic T
lymphocytes (CTLs) which killed autologous
EBV-transformed cells but not PHA blast cells and
recognized HLA-A24 as a restriction molecule.
However, the TILs did not recognize known EBV
antigenic peptides presented by HLA-A24 nor
HLA-A24 positive fibroblasts infected with
vaccinia recombinant virus expressing each of the
EBV latent proteins. EBV-positive gastric
carcinomas do not express conventional target
proteins of EBV-specific CTLs and the data suggest
that some cellular proteins may be involved in the
strong T cell response to EBV-associated gastric
carcinomas. Frequencies of CD8 + antigen-specific
T cells in TILs versus PBMCs were 1.9 % versus
0.013 % by limiting dilution assay, 7.0% versus
undetected by intracellular interferon (IFN)-γ
production assay and 22.8% versus 1.5 % by
enzyme-linked immunospot (ELISpot) assay. These
data demonstrate that class-I-restricted
antigen-specific CD8 + CTLs are specifically
expanded within EBV-positive gastric carcinoma
tissue.
To target EBV-positive gastric carcinomas,
where EBV protein expression is limited, utilization
of cytotoxic T lymphocytes recognizing latent
membrane protein (LMP2) could be an option. We
adopted an approach for determination of CTL
epitopes through multiple screenings, consisting of
a computer-assisted algorithm, a major
histocompatibility complex (MHC) stabilization
assay and ELISpot assay, an LMP2 epitope was
identified. Further analysis using a CTL clone
recognizing the epitope revealed that it is not
presented on the surface of HLA A24-positive
fibroblast cells infected with recombinant vaccinia
viruses expressing LMP2. ELISpot assays using the
CTL clone and various antigen presenting cells
demonstrated that the presentation was partially
restored by pretreatment of the fibroblast cells with
IFN-γ. The hydrophobic epitope, IYVLVMLVL,
was presented on transporters associated with
antigen processing-negative T2 cells transfected
with plasmids encoding HLA A*2402 and the
minimal epitope, indicating the presentation to be
TAP-independent. The identified peptide could be a
useful reagent to study and/or elicit CTL responses
to EBV-positive gastric carcinomas expressing the
LMP2
Departments of *1 Pathology and Clinical Laboratories,
*2 *3
Gastroenterology, Gastroenterological Surgery, Aichi
Cancer Center Hospital
61

catalytic subunit physically interacts with the
BBLF4/BSLF1/BBLF2/3 Complex. J. Virol. 74:
2550-2557, 2000.
J018. Fujita, M., Ishimi, Y., Nakamura, H.,
Kiyono, T. and Tsurumi, T.: Nuclear organization
of DNA replication initiation proteins in
mammalian cells. J. Biol. Chem. 277:
10354-10361, 2002.
J019. Fukumoto, M., Sugiyama, A., Ishida, K.,
Ikeno, T., Murakami, M., Kawasaki, S., Ota, H.,
Tatematsu, M. and Katsuyama, T.: Timing of
N-methyl-N-nitrosourea administration affects
gastric carcinogenesis in Mongolian gerbils infected
with Helicobacter pylori. Cancer Lett.,160, 99-105,
2000.
J020. Futamura, N., Nakamura, S., Tatematsu, M.,
Yamamura, Y., Kannagi, R. and Hirose, H.:
Clinicopathologic significance of sialyl Le x
expression in advanced gastric carcinoma. Br. J.
Cancer 83: 1681-1687, 2000.
J021. Gao, C., Li, Z., Ding, J., Wang, J., Hu, X.,
Liu, T., Xue, T., Li, H., Fujiyoshi, T., Takezaki, T.
and Tajima, K.: Study on the relation between
HLA-DRB1 alleles and Helicobacter pylori
infection. Chin. J. Epidemiol. 21: 417-419, 2000.
J022. Gohara, R., Tang, D., Inada, H., Inagaki,
M., Takasaki, Y. and Ando, S.: Phosphorylation of
vimentin head domain inhibits interaction with the
carboxyl-terminal end of alpha-helical rod domain
studied by surface plasmon resonance
measurements. FEBS Lett. 489:182-186, 2001.
J023. Hamada, T., Hirota, H., Yokoyama, S.,
Otsubo, N., Ishida, H., Kiso, M., Kanamori, A. and
Kannagi, R.: NMR analysis of novel ganglioside
GM4 analogues containing de-N-acetyl and
lactamized sialic acid: probes for searching new
ligand structures for human L-selectin. Magn.
Reson. Chem. 40: 517-523, 2002.
J024. Hamajima, N. and Matsuo, K.: Subtle
instruction to quit smoking may be efficacious for
certain smokers. Asian Pacific J. Cancer Prev. 1:
257-258, 2000.
J025. Hamajima, N., Fukumitsu, T., Odauchi, S.,
Akashi, T., Usui, T. and Ido, M.: A large-scale
follow-up study of smokers visiting medical
facilities in Japan. Asian Pacific J. Cancer Prev. 2:
185-191, 2001.
J026. Hamajima, N., Iwata, H., Obata, Y., Matsuo,
K., Mizutani, M., Iwase, T., Miura, S., Okuma, K.,
64
Ohashi, K. and Tajima, K.: No association of the 5’
promoter region polymorphism of CYP17 with
breast cancer risk in Japan. Jpn. J. Cancer Res. 91:
880-885, 2000.
J027. Hamajima, N., Katsuda, N., Matsuo, K.,
Saito, T., Ito, L.S., Ando, M., Inoue, M., Takezaki,
T. and Tajima, K.: Smoking habit and Interleukin
1B C-31T polymorphism. J. Epidemiol. 11:
120-125, 2001.
J028. Hamajima, N., Matsuo, K. and Yuasa, H.:
Adjustment of prognostic effects in prevalent
case-control studies on genotype. J. Epidemiol. 11:
204-210, 2001.
J029. Hamajima, N., Matsuo, K., Saito, T., Tajima,
K., Okuma, K., Yamao, K. and Tominaga, S.:
Interleukin 1 polymorphisms, lifestyle factors, and
Helicobacter pylori infection. Jpn. J. Cancer Res.
92: 383-389, 2001.
J030. Hamajima, N., Matsuo, K., Suzuki, T.,
Nakamura, T., Matsuura, A., Tajima, K. and
Tominaga, S.: Low expression myeloperoxidase
genotype negatively associated with Helicobacter
pylori infection. Jpn. J. Cancer Res. 92: 488-493,
2001.
J031. Hamajima, N., Matsuo, K., Tajima, K.,
Mizutani, M., Iwata, H., Iwase, T., Miura, S., Oya,
H. and Obata, Y.: Limited association between a
catechol-O-methyltransferase (COMT) poly-
morphism and breast cancer risk in Japan. Int. J.
Clin. Oncol. 6: 13-18, 2001.
J032. Hamajima, N., Saito, T., Matsuo, K., Kozaki,
K. Takahashi, Ta. and Tajima, K.: Polymerase
chain reaction with confronting two-pair primers
for polymorphism genotyping. Jpn. J. Cancer Res.
91: 865-868, 2000.
J033. Hamajima, N., Takezaki, T., Matsuo, K.,
Saito, T., Inoue, M., Hirai, T., Kato, T., Ozeki, J.
and Tajima, K.: Genotype frequencies of
cyclooxygenase 2 (COX2) rare polymorphisms for
Japanese with and without colorectal cancer. Asian
Pacific J. Cancer Prev. 2: 57-62, 2001.
J034. Hara, S., Kami, M., Miyakoshi, S., Suzuki,
R., Takeuchi, K., Seki, T., Oki, Y., Kishi, Y.,
Ueyama, J., Morinaga, S. and Muto, Y.: Central
nervous system involvement in pyothorax-
associated lymphoma: ring enhancement on CT
scan. Ann. Hematol. 80: 174-177, 2001
J035. Harada, H., Uchida, N., Shimada, Y.,
Kumimoto, H., Shinoda, M., Imamura, M. and

Ishizaki, K.: Polymorphism and allelic loss at the
AS3 locus on 13q12-13 in esophageal squamous
cell carcinoma. Int. J. Oncol. 18: 1003-1007,
2001.
J036. Haruki, N., Harano, T., Masuda, A., Kiyono,
T., Takahashi, T., Tatematsu, Y., Shimizu, S.,
Mitsudomi, T., Konishi, H., Osada, H., Fujii, Y.
and Takahashi, Ta.: Persistent increase in
chromosome instability in lung cancer : Possible
indirect involvement of p53. Am. J. Pathol. 159:
1345-1352, 2001.
J037. Haruki, N., Saito, H., Harano, T., Nomoto,
S., Takahashi, Takao, Osada, H., Fujii, Y. and
Takahashi, Ta.: Molecular analysis of the mitotic
checkpoint genes BUB1, BUBR1 and BUB3 in
human lung cancers. Cancer Lett. 162: 201-205,
2001.
J038. Haruki, N., Saito, H., Tatematsu, Y., Konishi,
H., Harano,T., Masuda, A., Fujii, Y. and
Takahashi, Ta.: Histological type-selective,
tumor-predominant expression of a novel CHK1
isoform and infrequent in vivo somatic CHK2
mutation in small cell lung cancer. Cancer Res. 60:
4689-92, 2000.
J039. Haruki, N., Yatabe, Y., Travis, W. D.,
Nomoto, S., Osada, H., Nakamura, S., Nakao, A.,
Fujii, Y. and Takahashi, Ta.: Characterization of
high-grade neuroendocrine tumors of the lung in
relation to menin mutations. Jpn. J. Cancer Res. 91:
317-323, 2000.
J040. Hata, M. and Ohtsuka, K.: Cloning and
expression of murine Hsp40 gene: Differences in
initiation sites between heat-induced and
constitutive transcript. DNA Seq. 11: 213-223,
2000.
J041. Hata, M. and Ohtsuka, K.: Murine cDNA
encoding a novel type I HSP40/DNAJ homolog,
mmDjA4. Biochim. Biophys. Acta 1493(1-2):
208-210, 2000.
J042. Hida, T., Kozaki, K., Muramatsu, H.,
Masuda, A., Shimizu, S., Mitsudomi, T., Sugiura,
T., Ogawa, M. and Takahashi, Ta.:
Cyclooxygenase (COX)-2 inhibitor induces
apoptosis and enhances cytotoxicity of various
anticancer agents in non-small cell lung cancer cell
lines. Clin. Cancer Res. 6: 2006-2011, 2000.
J043. Hirose, F., Ohshima, N., Shiraki, M., Inoue,
Y. H., Taguchi, O., Nishi, Y., Matsukage, A. and
Yamaguchi, M.: Ectopic expression of DREF
induces DNA synthesis, apoptosis and unusual
morphogenesis in the Drosophila eye imaginal disc:
possible interaction with Plycomb and trithorax
group proteins. Mol. Cell. Biol. 21: 7231-7242,
2001.
J044. Hirose, K., Tajima, K., Hamajima, N.,
Takezaki T., Inoue, M., Kuroishi T., Miura, S.,
and Tokudome, S.: Association of family history
and other risk factors with breast cancer risk among
Japanese premenopausal and postmenopausal
women. Cancer Causes Cont. 12: 349-358, 2001.
J045. Hirota, T., Morisaki, T., Nishiyama, Y.,
Marumoto, T., Tada, K., Hara, T., Masuko, N.,
Inagaki, M., Hatakeyama, K. and Saya, H.:
Zyxin, a regulator of actin filament assembly, plays
a role in cell division by interacting with
LATS1/h-warts tumor suppressor on mitotic
apparatus. J. Cell Biol. 149: 1073-1086, 2000.
J046. Hoshino, Y., Kimura, H., Kuzushima, K.,
Tsurumi, T., Nemoto, K., Kikuta, A., Nishiyama,
Y., Kojima, S., Matsuyama, T. and Morishima, T.:
Early intervention in post-transplant lympho-
proliferative disorders based on Epstein-Barr viral
load. Bone Marrow Transplant. 26:199-201, 2000.
J047. Hosokawa, Y., Maeda, Y. and Seto, M.: Low
frequency of expression of dominant-negative
Ikaros isoforms in human leukemia and lymphoma
cell lines. Leukemia Res. 24: 263-264, 2000.
J048. Hosokawa, Y., Maeda, Y. and Seto, M.:
Target genes downregulated by the BCL-6/LAZ3
oncoprotein in mouse Ba/F3 cells. Biochem.
Biophys. Res. Commun. 283: 563-568, 2001.
J049. Hosokawa, Y., Maeda, Y., Ishinohasama, R.,
Miura, I., Taniwaki, M. and Seto, M.: The Ikaros
gene, a central regulator of lymphoid differentiation,
fuses to the BCL6 gene as a result of
t(3;7)(q27;p12) translocation in a patient with
diffuse large B-cell lymphoma. Blood 95:
2719-2721, 2000.
J050. Hosokawa, Y., Nagai, E. and Seto, M.:
Truncated TSG101 transcripts in human leukemia
and lymphoma cell lines. J. Cancer Res. Clin.
Oncol. 126: 79-84, 2000.
J051. Hosokawa, Y., Papanikolaou, A., Cardiff,
R.D., Yoshimoto, K., Bernstein, M., Wang, T.C.,
Schmidt, E.V. and Arnold, A.: In vivo analysis of
mammary and non-mammary tumorigenesis in
MMTV-cyclin D1 transgenic mice deficient in p53.
Transgenic Research, 5: 471-478, 2001.
65

K. and Kato, M.: A new model population-based
cancer registration system in Aichi Prefecture,
Japan. Asian Pacific J. Cancer Prev. 1: 67-71, 2000.
J068. Inoue, M.: Information feedback by means of
electronic media. JACR Monogr. 5: 44-47, 2000 (in
Japanese).
J069. Inoue, T., Nakanishi, H., Inada, K., Hioki,
T., Tatematsu, M. and Sugimura, Y.: Real time
revwrse transcriptase polymerase chain reaction of
urinary cytokeratin 20 detects transitonal cell
carcinoma cells. J. Urol., 166, 2134-2141, 2001.
J070. Ishiguro, K., Kadomatsu, K., Kojima, T.,
Muramatsu, H., Tsuzuki, S., Nakamura, E.,
Kusugami, K., Saito, H. and Muramatsu, T.:
Syndecan-4 deficiency impairs focal adhesion
formation only under restricted conditions. J. Biol.
Chem. 25: 5249-5252, 2000.
J071. Ishii, K., Usui, S., Sugimura, Y., Yoshida, S.,
Hioki, T., Tatematsu, M., Yamamoto, H. and
Hirano, K.: Aminopeptidase N regulated by zinc in
human prostate participates in tumor cell invasion.
Int. J. Cancer, 92, 49-54, 2001.
J072. Ishii, K., Usui, S., Yamamoto, H., Sugimura,
Y., Tatematsu, M. and Hirao, K.: Decreases of
Metallothionein and Aminopeptidase N in Renal
Cancer Tissues. J. Biochem, 129, 253-258, 2001.
J073. Ishizaki, K., Nishizawa, K., Kato, T., Kitao,
H., Han, Z-B., Hirayama, J., Suzuki, F., Cannon
T. F., Kamigaichi, S., Tawarayama, Y., Masukawa,
M., Shimazu, T. and Ikenaga, M.: Genetic changes
induced in human cells in space shuttle experiment
(STS-95). Aviat. Space Environ. Med. 72: 794-798,
2001.
J074. Ito, H., Matsuo, K., Saito, T., Hirose, K.,
Inoue, M., Takezaki, T., Hamajima, N., Kuroishi,
T., and Tajima, K.: Valid responses to ABO blood
type question in self-reporting questionnaire. Asian
Pacific J. Cancer Prev. 2: 315-317, 2001.
J075. Ito, K., Ye, C.L., Hibi, K., Mitsuoka, C.,
Kannagi, R., Hidemura, K. ando, H., Kasai, Y.,
Akiyama, S. and Nakao, A.: Paired tumor marker
of soluble E-selectin and its ligand sialyl Lewis A
in colorectal cancer. J. Gastroenterol. 36: 823-829,
2001.
J076. Ito, L.S., Oba, S.M., Hamajima, N., Marie,
S.K.N., Uno, M., Shinjo, S.K., Kino, A., Lavilla, F.,
Inoue, M., Tajima, K. and Tominaga, S.:
Helicobacter pylori seropositivity among 963
Japanese Brazilians according to sex, age,
generation, and lifestyle factors. Jpn. J. Cancer. Res.
92: 1150-1156, 2001.
J077. Ito, S., Nakanishi, H., Ikehara, Y., Kato, T.,
Kasai, Y., Ito, K., Akiyama, S., Nakao, A. and
Tatematsu, M.: Real-time observation of
micrometastasis formation in the living mouse liver
using a green fluorescent protein gene-tagged rat
tongue carcinoma cell line. Int. J. Cancer, 93,
212-217, 2001.
J078. Iwase, S., Tsujimura, K., Matsudaira, Y.,
Ozeki, S., Onozaki, K., Obata, Y. and Takahashi,
To.: Comparison of anti-tumor responses against
TL positive lymphoma induced by skin grafting and
dendritic cell immunization. Microbiol. Immunol.
44: 609-618, 2000.
J079. Iwata, H., Yamamoto, M., Nemori, R.,
Mizutani, M., Iwase, T., Miura, S., Obata, Y.,
Hara, Y., Omoto, Y., Toyama, T., Yamashita, H.,
Iwase, H. and Kobayashi, S.: Localization of
gelatinolytic activity can be detected in breast
cancer tissues by film in situ zymography. Breast
Cancer 8: 111-115, 2001.
J080. Izawa, I., Nishizawa, M. Ohtakara, K.,
Ohtsuka, K., Inada, H. and Inagaki, M.:
Identification of Mrj, a DnaJ/Heat shock protein 40
family protein, as a keratin 8/18 filament-regulatory
protein. J. Biol. Chem. 275: 34521-34527, 2000.
J081. Izawa, M., Kumamoto, K., Mitsuoka, C.,
Kanamori, A., Ohmori, K., Ishida, H., Nakamura,
S., Kurata-Miura, K., Sasaki, K., Nishi, T. and
Kannagi, R.: Expression of sialyl 6-sulfo Lewis x is
inversely correlated with conventional sialyl Lewis
x expression in human colorectal cancer. Cancer
Res. 60: 1410-1416, 2000.
J082. Izumi, M., Yokoi, M., Nishikawa, N. S.,
Miyazawa, H., Sugino, A., Yamagishi, M.,
Yamaguchi, M., Matsukage, A., Yatagai, F. and
Hanaoka, F.: Transcription of the catalytic
180-kDa subunit gene of mouse DNA polymerase
αis controlled by E2F, an Ets-related transcription
factor, and Sp1. Biochem. Biophys. Acta. 1492 :
341-352, 2000.
J083. Kagami, Y., Jung, J., Choi, YS., Osumi, K.,
Nakamura, S., Morishima, Y. and Seto, M.:
Establishment of a follicular lymphoma cell line
(FLK-1) dependent on follicular dendritic cell-like
cell line HK. Leukemia 15: 148-156, 2001.
J084. Kanamori, A., Kojima, N., Uchimura, K.,
Muramatsu, T., Tamatani, T., Berndt, M.C.,
67

Kansas, G.S. and Kannagi, R.: Distinct sulfation
requirements of selectins disclosed using cells
which support rolling mediated by all three
selectins under shear flow: L-SELECTIN PREFERS
CARBOHYDRATE 6-SULFATION TO TYROSINE
SULFATION WHEREAS P-SELECTIN DOES NOT. J. Biol.
Chem. in press.
J085. Kanda, Y., Mineishi, S., Saito, T., Seo, S.,
Saito, A., Ohnishi, M., Suenaga, K., Niiya, H.,
Nakai, K., Takeuchi, T., Kawahigashi, Y., Shoji,
N., Ogasawara, T., Tanosaki, R., Kobayashi, Y.,
Tobinai, K., Kami, M., Mori, S., Suzuki, R.,
Kunitoh, H. and Takaue, Y.: Pre-emptive therapy
against cytomegalovirus (CMV) diseases guided by
CMV antigenemia assay after allogeneic
hematopoietic stem cell transplantation: a
single-center experience in Japan. Bone Marrow
Transplant. 27: 437-444, 2001.
J086. Kannagi, R.: Monoclonal anti-glycosphingo-
lipid antibodies in sphingolipid metabolism and cell
signaling. Method. Enzymol. 312: 160-179, 2000.
J087. Kannagi, R.: Use of liposomes containing
carbohydrates for production of monoclonal
antibodies. Method. Mol. Biol. 199: 203-218,
2002.
J088. Katsuda, N., Hamajima, N., Matsuo, K.,
Saito, T., Ito, S.L., Inoue, M., Takezaki, T., Tajima,
K. and Tominaga, S.: Association between the
interleukin 1B (C-31T) polymorphism and
Helicobacter pylori infection in health checkup
examinees. Jpn. J. Public Health 48: 604-611, 2001
(in Japanese).
J089. Kikuchi, Y., Hirano, M., Seto, M. and
Takatsu, K.: Identification and characterization of a
molecule, BAM11, that associates with the
pleckstrin homology domain of mouse Btk. Int.
Immunol. 12: 1397-1408, 2000.
J090. Kimura, H., Nagasaka, T., Hoshino, Y.,
Hayashi, N., Tanaka, N., Xu, J.L., Kuzushima, K.
and Morishima T.: Severe Hepatitis caused by
Epstein-Barr virus without infection of hepatocytes.
Hum. Pathol. 32:757-762,2001.
J091. Kishi, Y., Kami, M., Oki, Y., Kazuyama, Y.,
Kawabata, M., Miyakoshi, S., Morinaga, S.,
Suzuki, R., Mori, S. and Muto, Y.: Donor
lymphocyte infusion for treatment of
life-threatening respiratory syncytial virus infection
following bone marrow transplantation. Bone
Marrow Transplant. 26: 573-576, 2000.
68
J092. Kobayashi, Y., Kume, A., Li, M., Doyu, M.,
Hata, M., Ohtsuka, K. and Sobue, G.: Chaperones
Hsp70 and Hsp40 suppress aggregate formation and
apoptosis in cultured neuronal cells expressing
truncated androgen receptor protein with expanded
polyglutamine tract. J. Biol. Chem. 275: 8772-8778,
2000.
J093. Kohno, A., Tsuzuki, S., Kasai, M.,
Miyamura, K., Emi, N., Tanimoto, M. and Saito,
H.: Acute promyelocytic leukemia with apparently
normal karyotype: molecular findings and response
to all-trans retinoic acid. Leuk. Lymphoma, 42:
151-61, 2001.
J094. Koike, C., Luppi, P., Sharma, S.B., Kannagi,
R., Nakashima, I., Starzl, T.E. and Trucco, M.:
Molecular basis of evolutionary loss of
α1,3-galactosyltransferase gene in higher primates.
J. Biol. Chem. 277: 10114-10120, 2002.
J095. Kondo, E., Ogura, M., Kagami, Y., Taji, H.,
Miura, K., Takeuchi, T., Maeda, S., Asakura, S.,
Suzuki, R., Nakamura, S. and Morishima, Y.:
Assessment of prognostic factors in follicular
lymphoma patients. Int. J. Hematol. 73: 363-368,
2001.
J096. Kontani, K., Taguchi, O., Narita, T.,
Hiraiwa, N., Sawai, S., Hanaoka, J., Ichinose, M.,
Tezuka, N., Inoue, S., Fujino, S. and Kannagi, R.:
Autologous dendritic cells or cells expressing both
B7-1 and MUC1 can rescue tumor-specific
cytotoxic T lymphocytes from MUC1-mediated
apoptotic cell death. J. Leukocyte Biol. 68:
225-232, 2000.
J097. Kontani, K., Taguchi, O., Narita, T., Izawa,
M., Hiraiwa, N., Zenita, K., Takeuchi, T., Murai,
H., Miura, S. and Kannagi, R.: Modulation of
MUC1 mucin as an escape mechanism of breast
cancer cells from autologous cytotoxic
T-lymphocytes. Br. J. Cancer 84: 1258-1264,
2001.
J098. Kosako, H., Yoshida, T., Matsumura, F.,
Ishizaki, T., Narumiya, S. and Inagaki, M.:
Rho-kinase/ROCK is involved in cytokinesis
through the phosphorylation of myosin light chain
and not ezrin/radixin/moesin proteins at the
cleavage furrow. Oncogene 19: 6059-6064, 2000.
J099. Kozaki, K., Koshikawa, K., Tatematsu, Y.,
Miyaishi, O., Saito, H., Hida, T., Osada, H. and
Takahashi, Ta.: Multi-faceted analyses of a highly
metastatic human lung cancer cell line
NCI-H460-LNM35 suggest mimicry of

inflammatory cells in metastasis. Oncogene 20:
4228-4234, 2001.
J100. Kozaki, K., Miyaishi, O., Tsukamoto, T.,
Tatematsu, Y., Hida, T., Takahashi, To. and
Takahashi, Ta.: In vivo selected human lung
cancer cell line H460-LNM35 consistently exhibits
lymphogenous metastasis via both subcutaneous
and orthotopic propagation. Cancer Res. 69:
2535-2540, 2000.
J101. Kumamoto, K., Goto, Y., Sekikawa, K.,
Takenoshita, S., Ishida, N., Kawakita, M. and
Kannagi, R.: Increased expression of
UDP-galactose transporter mRNA in human colon
cancer tissues and its implication in synthesis of
Thomsen-Friedenreich antigen and sialyl Lewis
A/X determinants. Cancer Res. 61: 4620-4627,
2001.
J102. Kumimoto, H., Hamajima, N., Nishizawa, K.,
Nishimoto, Y., Matsuo, K., Harada, H., Shinoda,
M., Hatooka, S. and Ishizaki, K.: Different
susceptibility of each L-myc genotype to
esophageal cancer risk factors. Jpn. J. Cancer Res.
92: 735-739, 2001.
J103. Kumimoto, H., Hamajima, N., Nishizawa, K.,
Nishimoto, Y., Matsuo, K., Harada, H., Shimada,
Y., Imamura, M., Shinoda, M., Hatooka, S. and
Ishizaki, K.: Different susceptibility of each L-myc
genotype to esophageal cancer risk factors. Jap.
J. Cancer Res., 92:735-739, 2001.
J104. Kuroishi, T., Hirose, K., Suzuki, T. and
Tominaga, S.: Effectiveness of mass screening for
breast cancer in Japan. Breast Cancer 7: 1-8, 2000.
J105. Kuzushima, K., Hayashi, N., Kimura, H. and
Tsurumi, T.: Efficient identification of HLA
A*2402-restricted cytomegalovirus-specific CD8 +
T cell epitopes by a computer algorithm and an
enzyme-linked immunospot assay. Blood 98:
1872-1881, 2001.
J106. Kuzushima, K., Kimura, H., Hoshino, Y.,
Yoshimi , A., Tsuge, I., Horibe, K., Morishima,
T., Tsurumi, T. and Kojima, S.: Longitudinal
dynamics of Epstein-Barr virus-specific cytotoxic T
lymphocytes during the posttransplant lympho-
proliferative disorder. J. Infect. Dis. 182: 937-
940, 2000.
J107. Kwon, E.-J., Oh, E.-J., Kim, Y.-S., Hirose,
F., Ohno, K., Nishida, Y., Matsukage, A.,
Yamaguchi, M. and Yoo. M.-A.: Transcriptional
regulation of the Drosophila raf proto-oncogene
by the transcription factor dE2F. Nucleic Acids Res.
29: 1808-1814, 2001.
J108. Kwon, E.-J., Park, H.-S., Kim, Y.-S., Oh,
E.-J., Nishida, Y., Matsukage A., Yoo, M.-A. and
Yamaguchi, M.: Transcriptional regulation of the
Drosophila raf proto-oncogene by D-STAT during
development and immune response. J. Biol. Chem.
275:19824-19830, 2000.
J109. Lu, J., Landerholm, T.E., Wei, J.S., Dong,
X.-R., Wu, S.-P., Liu, X., Nagata, K., Inagaki, M.
and Majesky, M.W.: Coronary smooth muscle
differentiation from proepicardial cells requires
Rho-a-mediated actin reorganization and p160
Rho-kinase activity. Dev. Biol. 240: 404-418, 2001.
J110. Lucas, PC., Yonezumi, M., Inohara, N.,
McAllister-Lucas, L.M., Abazeed, M.E., Chen,
F.F., Yamaoka, S., Seto, M. and Nunez, G.: Bcl10
and MALT1, independent targets of chromosomal
translocation in malt lymphoma, cooperate in a
novel NF-kappa B signaling pathway. J. Biol.
Chem. 276: 19012-19019, 2001.
J111. Lunevicius, R., Nakanishi, H., Ito, S.,
Kozaki, K., Kato, T., Tatematsu, M. and Yasui, K.:
Clinicopathological significance of fibrotic capsule
formation around liver metastasis from colorectal
cancer. J. Cancer Res. Clin. Oncol., 127,193-199,
2001.
J112. Lymphoma Study Group of Japanese
Pathology: The World Health Organization
classification of malignant lymphomas in Japan:
Incidence of recently recognized entities. Pathol. Int.
50: 696-702, 2000.
J113. Maeda, S., Kagami, Y., Ogura, M., Taji, H.,
Suzuki, R., Kondo, E., Asakura, S., Takeuchi, T,
Miura K., Ando, M., Nakamura, S., Ito, T.,
Kinoshita, T., Ueda, R. and Morishima, Y.:
CD34+-selected autologous peripheral blood stem
cell transplantation conditioned with total body
irradiation for malignant lymphoma: increased risk
of infectious complications. Int. J. Hematol. 74:
214-221, 2001.
J114. Maruta, F., Ota, H., Genta, M., Sugiyama,
A., Tatematsu, M., Katsuyama, T. and Kawasaki,
S.: Role of N-methyl-N-nitrosourea in the induction
of intestinal metaplasia and gastric adenocarcinoma
in Mongolian Gerbils infected with Helicobacter
Pylori. Scand. J. Gastroenterol. , 3, 283-290, 2001.
J115. Masuda, A., Osada, H., Yatabe, Y., Kozaki,
K., Tatematsu, Y., Takahashi , T., Hida, T.,
69

Takahashi, To. and Takahashi, Ta.: Protective
function of p27 KIP1 against apoptosis in small cell
lung cancer cells in unfavorable microenvironments.
Am. J. Pathol. 158: 87-96, 2001.
J116. Matsuo, K., Hamajima, N., Hirose, K., Inoue,
M., Takezaki, T., Kuroishi, T. and Tajima, K.:
Alcohol, smoking, and dietary status and
susceptibility to malignant lymphoma in Japan:
Results of a hospital-based case-control study at
Aichi Cancer Center. Jpn. J. Cancer Res. 92:
1011-1017, 2001.
J117. Matsuo, K., Hamajima, N., Morishima, Y.
and Harada, M.: Hospital capacity and
post-transplant survival after allogeneic marrow
transplantation: analysis of data from Japana
Society for Hematopoietic Cell Transplantation.
Bone Marrow Transplant. 26: 1061-1067, 2000.
J118. Matsuo, K., Hamajima, N., Shinoda, M.,
Hatooka, S., Inoue, M., Takezaki, T. and Tajima,
K.: Gene-environment interaction between an
aldehyde dehydrogenase-2 (ALDH2) polymorph-
ism and alcohol consumption for the risk of
esophageal cancer. Carcinogenesis 22: 913-916,
2001.
J119. Matsuo, K., Hamajima, N., Shinoda, M.,
Hatooka, S., Inoue, M., Takezaki, T. and Tajima,
K.: Possible risk reduction in esophageal cancer
associated with MPO -463 A allele. J. Epidemiol.
11: 109-114, 2001.
J120. Matsuo, K., Hamajima, N., Suzuki, R.,
Nakamura, S., Seto, M., Morishima, Y. and
Tajima, K.: No substantial difference in genotype
frequencies of interleukin and myeloperoxidase
polymorphsisms between malignant lymphoma
patients and non-cancer controls. Haematologica
86: 602-608, 2001.
J121. Matsuo, K., Hamajima, N., Tominaga, S.,
Suzuki, T., Nakamura, T., Matsuura, A. and
Kitayama, K.: Helicobacter pylori IgG antibody test
established in the United States showed a
substantially lower sensitivity for Japanese
population. Am. J. Gastroenterol. 95: 1597-1598,
2000.
J122. Matsuo, K., Suzuki, R., Hamajima, N.,
Ogura, M., Kagami, Y., Taji, H., Kondoh, E.,
Maeda, S., Asakura, S., Kaba, S., Nakamura, S.,
Seto, M., Morishima, Y. and Tajima, K.:
Association between polymorphisms of folate- and
methionine-metabolizing enzymes and
susceptibility to malignant lymphoma. Blood 97:
70
3205-3209, 2001.
J123. Matsuoka, S., Nakagawa, T., Masuda, A.,
Haruki, N., Elledge, S. J. and Takahashi, Ta.:
Reduced expression and impaired kinase activity of
a Chk2 mutant identified in human lung cancer.
Cancer Res. 61: 5362-5365, 2001.
J124. Meloni, G., Capria, S., Vignetti, M., Alimena,
G., de Fabritiis, P., Montefusco, E., Mandelli, F.,
Matsuo, K, Hamajima, N., Suzuki, R., Nakamura,
S., Seto, M., Morishima, Y. and Tajima, K.:
Ten-year follow-up of a single center prospective
trial of unmanipulated peripheral blood stem cell
autograft and interferon-a in early phase chronic
myeloyd leukemia. Haematologica 86: 596-601,
2001.
J125. Mitsudomi, T., Hamajima, N., Ogawa, M.
and Takahashi, Ta.: Prognostic significance of p53
alterations in patients with non-small cell lung
cancer: a meta-analysis. Clin. Cancer Res. 6:
4055-4063, 2000.
J126. Mizoshita, T., Inada, K., Tsukamoto, T.,
Kodera, Y., Yamamura, Y., Hirai, T., Kato, T., Jou,
T., Itoh, M. and Tatematsu, M.: Expression of
Cdx1 and Cdx2 mRNAs and relevance of this
expression to differentiation in human
gastrointestinal mucosa-with special emphasis on
participation in intestinal metaplasia of the human
stomach. Gastric Cancer, 4, 185-191, 2001.
J127. Mizutani, K., Matsubayashi, T., Iwase, S.,
Doi, T.S., Kasai, K., Yazaki, M., Wada, Y.,
Takahashi, To. and Obata, Y.: Murine delta
homologue, mDelta1, expressed on feeder cells
controls cellular differentiation. Cell. Struct. Funct.
25: 21-31, 2000.
J128. Moore, M.A. and Tatematsu, M.: Are the
phenotypes of preneoplastic lesions of significance
for cancer prevention? 1. Liver. Asian Pacific J.
Cancer. Prev., 2, 27-42, 2001.
J129. Motegi, M., Yonezumi, M., Suzuki, H.,
Suzuki, R., Hosokawa, Y., Hosaka, S., Kodera, Y.,
Morishima, Y., Nakamura, S. and Seto, M.:
API2-MALT1 chimeric transcripts involved in
mucosa-associated lymphoid tissue type lymphoma
predict heterogenous products. Am. J. Pathol. 156:
807-812, 2000.
J130. Nakagawa, A., Hara, K., Takeuchi, K., Seto,
M. and Nakamura, S.: Sarcomatoid variant of
anaplastic large cell lymphoma with cytoplasmic
ALK and a-smooth muscle actin expression: a

mimic of inflammatory myofibroblastic tumor. Am.
J. Pathol. in press.
J131. Nakamura, S., Kato, M., Ichimura, K.,
Yatabe, Y., Kagami, Y., Suzuki, R., Taji, H.,
Kondo, E., Asakura, S., Kojima, M., Murakami, S.,
Yamao, K., Tsuzuki, T., Adachi, K., Miwa, A. and
Yoshida, T.: Peripheral T/natural killer-cell
lymphoma involving female genital tract: a
clinicopathologic study of 5 cases. Int. J. Hematol.
73: 108-114, 2001.
J132. Nakamura, T., Nakamura, S., Yonezumi, M.,
Seto, M. and Yokoi, T.: The t(11; 18)(q21; q21)
translocation in H. pylori-negative low-grade
gastric MALT lymphoma. Am. J. Gastroenterol. 95:
3314-3315, 2000.
J133. Nakamura, T., Nakamura, S., Yonezumi, M.,
Suzuki, T., Matsuura, A., Yatabe, Y., Yokoi, T.,
Ohashi, K. and Seto, M.: Helicobacter pylori and
the t(11;18)(q21;q21) translocation in gastric
low-grade B-cell lymphoma of mucosa-associated
lymphoid tissue type. Jpn. J. Cancer Res. 91:
301-309, 2000.
J134. Nakamura, Y., Hashimoto, R., Amano, M.,
Nagata, K., Matsumoto, N., Goto, H., Fukusho, E.,
Mori, H., Kashiwagi, Y., Kudo, T., Inagaki, M.
and Takeda, M.: Localized phosphorylation of
vimentin by Rho-kinase in neuroblastoma N2a cells.
Genes Cells 5:823-837, 2000.
J135. Nakanishi, H., Kodera, Y., Yamamura, Y.,
Ito, S., Kato, T., Ezaki, T. and Tatematsu, M.:
Rapid quantitative detection of carcinoembryonic
antigen-expressing free tumor cells in the peritoneal
cavity of gastric-cancer patients with real-time
RT-PCR on the lightcycler. Int. J. Cancer, 89:
411-417, 2000.
J136. Nakayashiki, N., Yoshikawa, K., Nakamura,
K., Hanai, N., Okamoto, K., Okamoto, S., Mizuno,
M., Wakabayashi, T., Sega, S., Yoshida, J. and
Takahashi, To.: Prodauction of a single chain
variable fragmanet recognizing type III mutant
epidermal growth factor receptor. Jpn. J. Cancer
Res. 91: 1035-1043, 2000.
J137. Naoe, T., Takeyama, K., Yokozawa, T., Kiyoi,
H., Seto, M., Uike, N., Ino, T., Utsunomiya, A.,
Maruta, A., Jin-nai, I., Kamada, N., Kubota, Y.,
Nakamura, H., Shimazaki, C., Horiike, S., Kodera,
Y., Saito, H., Ueda, R., Wiemels, J. and Ohno, R.:
Analysis of genetic polymorphism in NQO1,
GST-M1, GST-T1, and CYP3A4 in 469 Japanese
patients with therapy-related leukemia/
myelodysplastic syndrome and de novo acute
myeloid leukemia. Clin. Cancer Res. 6: 4091-4095,
2000.
J138. Naruse, T., Yuzawa, Y., Akahori, T., Mizuno,
M., Maruyama, S., Kannagi, R., Hotta, N., Matsuo,
S.: P-selectin-dependent macrophage migration into
the tubulointerstitium in unilateral ureteral
obstruction. Kidney Int. 62: 94-105, 2002.
J139. Natume, A., Tsujimura, K., Mizuno, M.,
Takahashi, To. and Yoshida, J.: IFN-β gene
therapy induces systemic antitumor immunity
against malignant glioma. J. Neurooncol. 47:
117-124, 2000.
J140. Nishida, K., Seto, M. and Ishida, R.:
Different susceptibilities of post-mitotic
checkpoint-proficient and -deficient BALB/ 3T3
cells to ICRF-193, a catalytic inhibitor of DNA
topoisomerase II. Jpn. J. Cancer Res. 92: 193-202,
2001.
J141. Nomoto, S., Haruki, N., Tatematsu, Y.,
Konishi, H., Mitsudomi, T., Takahashi, To. and
Takahashi, Ta.: Frequent allelic imbalance
suggests the involvement of a tumor suppressor
gene at 1p36 in the pathogenesis of human lung
cancers. Genes Chrom. Cancer 28: 342-346, 2000.
J142. Obata, Y., Takahashi, To., Sakamoto, J.,
Tamaki, H., Tominaga, S., Hamajima, N., Chen,
Y.-T. and Old, L.J.: SEREX analysis of gastric
cancer antigens. Cancer Chemother. Pharmacol. 46
(suppl): S37 - S42, 2000.
J143. Ogawa, K., Nakanishi, H., Takeshita, F.,
Futkuchi, M., Asamoto, M., Imaida, K., Tatematsu,
M. and Shirai, T.: Establishment of rat
hepatocellular carcinoma cell lines with differing
metastatic potential in nude mice. Int. J. Cancer,
91: 797-802, 2001.
J144. Ohgaki, H., Fukuda, M., Tohma, Y., Huang,
H., Stoica, G., Tatematsu, M. and Donehowr,
L.A.: Effect of intragastric application of
N-methylnitrosourea in p53 knockout mice.
Molecular Carcinogenesis. 28: 97-101, 2000.
J145. Ohmori, K., Kanda, K., Mitsuoka, C.,
Kanamori, A., Kurata-Miura, K., Sasaki, K., Nishi,
T., Tamatani, T. and Kannagi, R.: P- and
E-Selectins recognize sialyl 6-sulfo Lewis X, the
recently-identified L-selectin ligand. Biochem.
Biophys. Res. Commun. 278: 90-96, 2000.
J146. Ohno, K., Takahashi, Y., Hirose, F., Inoue,
Y. H., Taguchi, O., Nishida, Y., Matsukage, A. and
71

Yamaguchi, M.: Characterization of a Drosophila
homologue of the human myelodysplasia/myeloid
leukemia factor (MLF). Gene 260: 133-143, 2000.
J147. Ohshige, K., Morio, S., Mizushima, S.,
Kitamura, K., Tajima, K., Suyama, A., Usuku, S.,
Tia, P., Hor, L.B., Heng, S., Sapjonn, V.,
Tochikubo, O. and Soda, K.: Behavioral and
serological human immunodeficiency virus risk
factors among female commercial sex workers in
Cambodia. Int. J. Epidemiol. 29: 344-354, 2000.
J148. Ohshige, K., Morio, S., Mizushima, S.,
Kitamura, K., Tajima, K., Ito, A., Usuku, S.,
Suyama, A., Saphonn, V., Heng, S., Hor, L.B., Tia,
P. and Soda, K.: Cross-sectional analysis on risk
factors of HIV among female commercial sex
workers in Cambodia. Epidemiol. Infect. 124:
143-152, 2000.
J149. Ohta, T., Haga, H., Osada, H., Tanaka, K.,
Maeda, K., Takazaki, T., Seki, N., Ohyama, Y,
Nakanishi, Y. and Ishikawa, K.: Development and
evaluation of a QOL questionnaire for elderly
subject living in a community. Jpn. J. Public Health
48: 258-267, 2001 (in Japanese).
J150. Ohtakara, K., Inada, H., Goto, H., Taki, W.,
Manser., E., Lim, L., Izawa, I. and Inagaki, M.:
p21-activated kinase PAK phosphorylates desmin at
the different sites from those for Rho-associated
kinase. Biochem. Biophys. Res. Commun. 272:
712-716, 2000.
J151. Ohtsuka, K. and Hata, M.: Mammalian
HSP40/DNAJ homologs: Cloning of novel cDNAs
and a proposal for their classification and
nomenclature. Cell Stress & Chaperones 5: 98-112,
2000.
J152. Okino, T., Onda, M., Matsukura, N., Inada,
K., Tatematsu, M., Suzuki, S. and Shimada, T.:
Sequential Histopathological Changes in vivo after
suicide gene therapy of gastric cancer induced by
N-methyl-N’-nitro-N-nitrosoguanidine in Rats. Jpn.
J. Cancer Res, 92, 673-679, 2001.
J153. Osada, H., Tatematsu Y., Masuda, A., Saito,
T., Sugiyama, M., Yanagisawa, K. and Takahashi,
Ta.: Heterogeneous TGF-β unresponsiveness and
loss of TGFβRII expression caused by histone
deacetylation in lung cancer cell lines. Cancer Res.
61: 8331-8339, 2001.
J154. Oshige, M., Yoshida, H., Hirose, F., Takata,
K., Inoue, Y. H., Aoyagi, N., Yamaguchi, M.,
Koiwai, O., Matsukage, A. and Sakaguchi, K.:
72
Molecular cloning and expression during
development of the Drosophila gene for the
catalytic subunit of DNA polymerase ε. Gene 256:
93-100, 2000.
J155. Oyama, T., Kagami, Y., Seto, M. and
Morishima, Y.: Mechanism of action on B cell
lymphoma by chimeric anti-CD20 monoclonal
antibody. Leukemia 15: 1667, 2001.
J156. Ozawa, Y., Towatari, M., Tsuzuki, S.,
Hayakawa, F., Maeda, T., Miyata, Y., Tanimoto,
M. and Saito, H.: Histone deacetylase 3 associates
with and represses the transcription factor GATA-2.
Blood 98: 2116-23, 2001.
J157. Pei, J., Akatsuka, Y., Anasetti, C., Lin, M.-T.,
Petersdorf, E.W., Hansen, J.A. and Martin, P.J.:
Marrow graft rejection through recognition of an
HLA-C alloantigen: analysis of alloimmunity by T
cell cloning and testing of T cell repertoire
rearrangements. Biol. Blood Marrow Transplant. 7:
378-383, 2001.
J158. Sakai, H., Tsukamoto, T., Yamamoto, M.,
Shirai, N., Iidaka, T., Yanai, T., Masegi, T., and
Tatematsu, M.: Differential effects of partial
hepatectomy and carbon tetrachloride administra-
tion on induction of liver cell foci in a model for
detection of initiation activity. Jpn. J. Cancer Res,
92, 1018-1025, 2001.
J159. Sakai, H., Tsukamoto, T., Yamamoto, M.,
Yanai, T., Masegi, T., Inada, K., Nakanishi, H.
and Tatematsu, M.: Summation of initiation
activities of low doses of the non-hepatocarcinogen
1,2-dimethylhydrazine in the liver after carbon
tetrachloride administration. Cancer Lett.,148,
59-63, 2000.
J160. Sato, E., Yokoyama, N., Miyazawa, T,
Maeda, K, Ikeda, Y., Nishimura, Y ) , Fujita, K,
Kohmoto, M ., Takahashi, E., and Mikami, T.:
Efficient expression of the envelope protein of
feline immunodeficiency virus in a recombinant
feline herpesvirus type 1 (FHV-1) using the gC
promoter of FHV-1. Virus Res. 70: 13-23. 2000.
J161. Sawada, M., Moriya, S., Saito, S., Shineha,
R., Satomi, S., Yamori, T., Tsuruo, T., Kannagi, R.
and Miyagi, T.: Reduced sialidase expression in
highly metastatic variants of mouse colon
adenocarcinoma 26 and retardation of their
metastatic ability by sialidase overexpression. Int.
J. Cancer 97: 180-185, 2002.
J162. Sawada, M., Nakashima, S., Kiyono, T.,

Nakagawa, M., Yamada, J., Yamakawa, H.,
Banno, Y., Shinoda, J., Nishimura, Y., Nozawa, Y.
and Sakai, N.: p53 regulates ceramide formation by
neutral sphingomyelinase through reactive oxygen
species in human glioma cells. Oncogene 20:
1368-1378, 2001.
J163. Sawada, M., Nakashima, S., Kiyono, T.,
Yamada, J., Hara, S., Nakagawa, M., Shinoda, J.,
Sakai, N.: Acid sphingomyelinase activation
requires caspase-8 but not p53 nor reactive oxygen
species during Fas-induced apoptosis in human
gioma cels. Exp. Cell Res. 273:157-168, 2002.
J164. Sekine, M., Taya, C., Kikkawa, Y.,
Yonekawa, H., Takenaka, M., Matsuoka, Y., Imai,
E., Izawa, M., Kannagi, R. and Suzuki, A.:
Regulation of mouse kidney tubular epithelial
cell-specific expression of core 2 GlcNAc
transferase. Eur. J. Biochem. 268: 1129-1135,
2001.
J165. Shimizu, K., Hashimoto, T., Harihara, S.,
Tajima, K., Sonoda, S. and Zaninovic, V.:
B-globin gene haplotype characteristics of
Colombian Amerinds in South America. Human
Heredity 51: 54-63, 2001.
J166. Shimizu, N., Ikehara, Y., Inada, K.,
Nakanishi, H., Tsukamoto, T., Nozaki, K.,
Kaminishi, M., Kuramoto, S., Sugiyama, T.,
Katsuyama, T. and Tatematsu, M.: Eradication
diminishes enhancing effects Helicobacter pylori
infection on glandular stomach carcinogenesis in
mongolian gerbils. Cancer Res., 60, 1512-1514,
2000.
J167. Shimizu, N., Ikehara, Y., Nozaki, K.,
Kaminishi, M., Tsuda, H., Sugiyama, T.,
Katsuyama, T. and Tatematsu, M.: Effects of
lactoferrin administration in Helicobacter pylori
infection animal model. In: K. Shimazaki, H. Tsuda,
M. Tomita, T. Kuwata and Perraudin, J. P. (eds.),
Lactoferrin: Structure, Function and Applications,
pp. 209-215, Oxford, UK: Elsevier Science B.V.,
2000.
J168. Shimizu, S., Yatabe, Y., Koshikawa, T.,
Haruki, N., Hatooka, S., Shinoda, M., Suyama, M.,
Ogawa, M., Hamajima, N., Ueda, R., Takahashi,
T. and Mitsudomi, T.: High frequency of clonally
related tumors in cases of multiple synchronous
lung cancers as revealed by molecular diagnosis.
Clin. Cancer Res. 6: 3994-3999, 2000.
J169. Shimizu, S., Yatabe, Y., Koshikawa, T.,
Haruki, N., Hatooka, S., Shinode, M., Suyama, M.,
Ogawa, M., Hamajima, N., Ueda, R., Takahashi,
Ta. and Mitsudomi, T.: High frequency of clonally
related tumors in cases of multiple synchronous
lung cancers as revealed by molecular diagnosis.
Clin. Cancer Res. 6: 3994-3999, 2000.
J170. Song, J., Ugai, H., Ogawa, K., Wang, Y.,
Sarai, A., Obata, Y., Kanazawa, I., Sun, K.,
Itakura, K. and Yokoyama, K. K.: Two
consecutive zinc fingers in Sp1 and in MAZ are
essential for interactions with cis-elements. J. Biol.
Chem. 276: 30429-30434, 2001.
J171. Sonoda, S., Li, H.-C., Cartier, L., Nunez, L.
and Tajima, K.: Ancient HTLV type I provirus
DNA of Andean mummy. AIDS Res. Human
Retrovirus 16: 1753-1756, 2000.
J172. Stellman, S.D., Takezaki, T., Wang, L.,
Chen, Y., Citron, M.L., Djordjevic, M.V., Harlap,
S., Muscat, J.E., Neugut, A.I., Wynder, A.L.,
Ogawa, H., Tajima, K. and Aoki, K.: Smoking and
Lung Cancer Risk in American and Japanese Men:
An International Case-control Study. Cancer
Epidemiol. Biomarkers Prev, 10: 1193-1199, 2001.
J173. Sugaya, Y., Ihara, K., Masuda, Y., Ohtsubo,
E. and Maki, H.: Hyper-processive and slower
DNA chain elongation catalysed by DNA
polymerase III holoenzyme purified from the
dnaE173 mutator mutant of Escherichia coli. Genes
Cells 7: 385-99, 2002
J174. Sugiyama,T., Saka,K., Nakamura, S.,
Yonezumi, S. and Seto, M.: API2-MALT1
chimeric transcript is a predictive marker for the
responsiveness of H. pylori eradication treatment in
low-grade gastric MALT lymphoma.
Gastroenterology 120: 1884-1889, 2001.
J175. Suzuki, R., Kagami, Y., Takeuchi, K., Kami,
M., Okamoto, M., Ichinohasama, R., Mori, N.,
Kojima, M., Yoshino, T., Yamabe, H., Shiota, M.,
Mori, S., Ogura, M., Hamajima, N., Seto, M.,
Suchi, T., Morishima, Y. and Nakamura, S.:
Prognostic significance of CD56 expression for
ALK-positive and ALK-negative anaplastic
large-cell lymphoma of T/null cell phenotype.
Blood 96: 2993-3000, 2000.
J176. Suzuki, R., Seto, M. and Nakamura, S.:
Idiopathic eosinophilia. N. Engl. J. Med. 342:
660-661, 2000. (discussion)
J177. Suzuki, R., Seto, M., Nakamura, S.,
Nakagawa, A., Hara, K. and Takeuchi, K.:
Sarcomatoid variant of anaplastic large cell
73

lymphoma with cytoplasmic ALK and
alpha-smooth muscle actin expression: a mimic of
inflammatory myofibroblastic tumor. Am. J. Pathol.
159: 383-384, 2001.
J178. Suzuki, R., Takemura, K., Tsutsumi, M.,
Nakamura, S., Hamajima, N. and Seto, M.:
Detection of cyclin D1 overexpression by real-time
reverse-transcriptase-mediated quantitative poly-
merase chain reaction for the diagnosis of mantle
cell lymphoma. Am. J. Pathol. 159: 425-429, 2001.
J179. Tagawa, H., Miura, I., Suzuki, R., Suzuki,
H., Hosokawa, Y. and Seto, M.: Molecular
cytogenetic analysis of the breakpoint region at
6q21-22 in T-cell lymphoma/leukemia cell lines.
Genes Chrom. Cancer, 34: 175-185, 2002.
J180. Tajima, K. and Sonoda, S.: Origins of
HTLV-I with South America. Nat. Med. 6: 232-233,
2000.
J181. Takahashi, H., Maeda, Y., Seto, M. and
Hosokawa, Y.: Nucleotide insertions and deletions
within the homopolymeric runs of adenines and
thymidines of BCL10 cDNAs in normal peripheral
blood leukocytes. Blood 95: 2728-2729, 2000.
J182. Takeyama, K., Seto, M., Uike, N., Hamajima,
N., Ino, T., Mikuni, C., Kobayashi, T., Maruta, A.,
Muto, Y., Maseki, N., Sakamaki, N., Saitoh, H.,
Shimoyama, M. and Ueda, R.: Therapy-related
leukemia and myelodysplastic syndrome:
alarge-scale Japanese study of clinical and
cytogenetic features as well as prognostic factors.
Int. J. Hematol. 71: 144-152, 2000.
J183. Takezaki, T., Gao, C.M., Wu, J.Z., Ding,
J.H., Liu, Y.T., Zhang, Y., Li, S.P., Su, P., Liu,
T.K. and Tajima, K.: Dietary protective and risk
factors for esophageal and stomach cancers in a
low-epidemic area for stomach cancer in Jiangsu
Province, China; comparison with those in a
high-epidemic area. Jpn. J. Cancer Res. 92:
1157-1165, 2001.
J184. Takezaki, T., Hamajima, N., Matsuo, K.,
Tanaka, R., Hirai, T., Kato, T., Ohashi, K. and
Tajima, K.: Association of polymorphisms in the
beta-2 and beta-3 adrenoceptor genes with risk of
colorectal cancer in Japanese. Int. J. Clin. Oncol. 6:
117-122, 2001.
J185. Takezaki, T., Hirose, K., Inoue, M.,
Hamajima, N. Yatabe, Y., Mitsudomi, T., Sugiura,
T., Kuroishi, T., and Tajima, K.: Dietary factors
and lung cancer risk in Japanese: with special
74
reference to fish consumption and adenocarcinomas.
Br. J. Cancer 84: 1199-1206, 2001.
J186. Takezaki, T., Shinoda, M., Hatooka, S.,
Hasegawa, Y., Nakamura, S., Hirose, K., Inoue,
M., Hamajima, N., Kuroishi, T., Matsuura, H. and
Tajima, K.: Subsite-specific risk factors for
hypopharyngeal and esophageal cancer (Japan).
Cancer Causes Cont. 11: 597-608, 2000.
J187. Takubo, T., Kanashima, H., Terada, Y.,
Shibata, H., Aoyama, Y., Nakamae, H.,
Yamamura, R., Shima, M., Makita, K., Tanaka, K.,
Ohta, K., Yamane, T., Hino, M., Ohta, T.,
Hashimoto, S., Kamitani, T., Tatsumi, N.,
Kumamoto, K. and Kannagi, R.: Analysis of
E-selectin mRNA on leukemia cells: a report on
two patients with acute leukemia and one patient
with lymphoma leukemia accompanied by high
serum E-selectin levels. Med. Postgraduat. 40:
270-273, 2002.
J188. Tamaki, S., Ichinohe, T., Matsuo, K.,
Hamajima, N., Hirabayashi, N. and Dohy, H.:
Superior survival of blood and marrow stem cell
recipients given maternal grafts over recipients
given paternal grafts. Bone Marrow Transplant. 28:
375-380, 2001.
J189. Tamakoshi, A., Ohno, Y., Yamada, T., Aoki,
K., Hamajima, N., Wada, M., Kawamura, T.,
Wakai, K. and Lin, Y.S.: Depressive mood and
suicide among middle-aged workers: findings from
a prospective cohort study in Nagoya, Japan. J.
Epidemiol. 10: 173-178, 2000.
J190. Tamura, A., Miura, I., Iida, S., Yokota, S.,
Horiike, S., Nishida, K., Fuji, H, Nakamura, S.,
Seto, M., Ueda, R. and Taniwaki, M.: Interphase
detection of immunoglobulin heavy chain gene
translocations with specific oncogene loci in 173
patients with B-cell lymphoma. Cancer Genet.
Cytogenet. 129: 1-9, 2001.
J191. Tanaka, H., Shimada, Y., Harada, H.,
Shinoda, M., Hatooka, S., Imamura, M., and
Ishizaki, K.: Polymorphic variation of the ARP
gene on 3p21 in Japanese esophageal cancer
patients. Oncol. Rep. 7: 591-593, 2000.
J192. Tauchi, H., Komatsu, K., Ishizaki, K.,
Yatagai, F. and Kato, T.: Mutation spectrum of
MSH3-deficient HHUA/chr.2 cells reflects in vivo
activity of the MSH3 gene product in mismatch
repair. Mutation Res., 447: 155-164, 2000.
J193. Tei, K., Kawakami-Kimura, N., Taguchi, O.,

J222. Yonezumi, M., Suzuki, R., Suzuki, H.,
Yoshino, T., Oshima, K., Hosokawa, Y., Asaka, M.,
Morishima, Y., Nakamura, S. and Seto, M.:
Detection of AP12-MALT1 chimaeric gene in
extranodal and nodal marginal zone B-cell
lymphoma by reverse transcription polymerase
chain reaction (PCR) and genomic long and
accurate PCR analyses. Br. J. Haematol. 115:
588-594, 2001.
J223. Yoo, K.-Y., Tajima, K., Park, S.-K., Kang, D.,
Kim, S.-U., Hirose, K., Takeuchi, T. and Miura,
S.: Postmenopausal obesity as a breast cancer risk
factor according to estrogen and progesteron
receptor status (Japan). Cancer Letters, 167: 57-63,
2001.
J224. Yoshida, K., Hamajima, N., Kozaki, K., Saito,
H., Maeno, K., Sugiura, T., Ookuma, K. and
Takahashi, Ta.: Association between the dopamine
D2 receptor A2/A2 genotype and smoking behavior
in the Japanese. Cancer Epidemiol. Biomarkers
Prev. 10: 403-405, 2001.
J225. Yuasa, H., Hamajima, N. and Matsuo, K.:
Investigation for smoking cessation support on
internet survey of health and smoking awareness of
users. Jpn. J. Public Health 47: 820-827, 2000 (in
Japanese).
J226. Zhao, S.-M., Li, H.-C., Lou, H., Lu, X.-X.,
Yu, X.-F., Gao, D.-H., Hu, J., Chiba, H., Takezaki,
T., Yashiki, S., Fujiyoshi, T., Sonoda, S. and
Tajima, K.: High prevalence of HBV in Tibet,
China. Asian Pacific J. Cancer Prev. 2: 299-304,
2001.
J227. Zhong, S., Zhange, Y., Jansen, C., Goto, H.,
Inagaki, M. and Dong, Z.: MAP kinases mediate
UVB-induced phosphorylation of histone H3 at
Serine 28. J. Biol. Chem. 276: 12932-12937, 2001.
J228. Zhong, S., Jansen, C., She, Q.B., Goto, H.,
Inagaki, M., Bode, A.M., Ma, W.Y. and Dong, Z.:
Ultraviolet B-induced phosphorylation of histone
H3 at serine 28 is mediated by MSK1. J. Biol.
Chem. 276: 33213-33219, 2001.
77

Reviews and Books
R001. Akatsuka, Y.: Minor histocompatibility
antigens and their clinical significance. Saishin
Igaku 56: 186-191, 2001. (in Japanese)
R002. Ando, S. and Inagaki, M.: Protein kinase
domain structure and phosphorylation specificity.
Experimental Medicine 18(1): 7-12, 2000. (in
Japanese)
R003. Chen Y.-T., Scanlan, M.J., Obata, Y. and
Old, L.J.: Identification of human tumor antigens
by serological expression cloning. In Principles and
Practice of the Biologic Therapy of Cancer.
Philadelphia; Lippincott Williams & Wilkins,
Rosenberg, S.A., ed., pp. 557-570, 2000.
R004. Fujita, M.: Cell cycle regulation of DNA
replication initiation proteins bycyclin/CDK.
Experimental Medicine 20: 540-545, 2002. (in
Japanese)
R005. Fujita, M.: Function and cell cycle
regulation of DNA replication initiation proteins in
mammalian cells. Experimental Medicine 18:
933-939, 2000. (in Japanese)
R006. Goto, H., Kosako, H. and Inagaki, M.:
Regulation of intermediate filament organization
during cytokinesis: possible roles of Rho-associated
kinase. Microsc. Res. Tech. 49:173-182, 2000.
R007. Hamajima, N., Matsuo, K., Saito, T., Hirose,
K., Inoue, M., Takezaki, T., Kuroishi, T. and
Tajima, K.: Gene-environment interactions and
polymorphism studies of cancer risk in the
Hospital-based Epidemiologic Research Program at
Aichi Cancer Center II (HERPACC-II). Asian
Pacific J. Cancer Prev. 2: 99-107, 2001.
R008. Hamajima, N.: PCR-CTPP: a new
genotyping technique in the era of genetic
epidemiology. Expert Rev. Mol. Diagn. 1: 119-123,
2001.
R009. Hida, T. and Takahashi, Ta.: Development
of novel treatments of lung cancer 2: application of
COX-2 inhibitors. Chiryogaku, 35: 70-73, 2001. (in
Japanese)
R010. Ikenaga, M., Ishizaki, K., Nishizawa, K.,
Han, Z.-B., Kitao, H., Hirayama, J. and Kato, T.:
Genetic effects of space radiation and microgravity
on cultured human tumor cells. In: Exploring
Future Research Strategies in Space Radiation
78
Sciences, Eds. H. J. Majima and K. Fujitaka, pp.
86-91, Iryokagakusha, Co. Ltd., Tokyo, 2000.
R011. Inada, H., Nagata, K., Goto, H. and Inagaki,
M.: Regulation of intermediate filament dynamics:
A novel approach using site-and phosphorylation
state-specific antibodies. Cytoskeleton: Signalling
and Cell Regulation: A Practical Approach. (The
practial Approach Series), eds. Carraway, K.L. and
Carraway, C.A.C. Oxford University Press 183-207,
2000.
R012. Inoue, M.: Recent observations in the
epidemiology of gastric cancer in Japan. 4th
International Gastric Cancer Congress, eds., M.F.
Brennan, M.S. Karpeh, p. 63-67, Monduzzi Editore,
Italy, 2001.
R013. Kannagi, R. and Hakomori, S.: A guide to
monoclonal antibodies directed to glycotopes. Adv.
Exp. Med. Biol. 491: 587-630, 2001.
R014. Kannagi, R., Kumamoto, K., Tei, K., Saitou,
S., Izawa, M., Goto, Y. and Fukui, F.:
Carbohydrate determinants in cancer. Molecular
Medicine, 38: 1190-1199, 2001. (in Japanese)
R015. Kannagi, R., Mitsuoka, C., Kanamori, A.,
Uchimura, K., Muramatsu, T., Imai, T., Yoshie, O.,
Matsushima, K. and Ohmori, K.: Expression and
selectin-binding activity of sialyl 6-sulfo Lewis X
determinant on human helper memory T
lymphocytes. In: D.Y. Mason (ed.), Leukocyte
Typing VII, pp. 39-41, Oxford: Oxford University
Press, 2002.
R016. Kannagi, R.: Regulatory roles of
carbohydrate ligands for selectins in homing of
lymphocytes. Curr. Opin. Struct. Biol., in press.
R017. Kannagi, R.: Transcriptional regulation of
expression of carbohydrate ligands for cell adhesion
molecules in the selectin family. Adv. Exp. Med.
Biol. 491: 267-278, 2001.
R018. Kannagi, R.: Fucosyltransferase V. In: N.
Taniguchi, K. Honke and M. Fukuda (eds.),
Handbook of Glycosyltransferases and Their
Related Genes, pp. 232-236, Tokyo:
Springer-Verlag, 2001.
R019. Kannagi, R.: Fucosyltransferase VI. In: N.
Taniguchi, K. Honke and M. Fukuda (eds.),
Handbook of Glycosyltransferases and Their
Related Genes, pp. 237-245, Tokyo:
Springer-Verlag, 2001.
R020. Kannagi, R.: Selectins and their

Abstracts for International Conferences
A001. Akatsuka, Y., Warren, E.H., Brickner, A.G.,
Lin, M-T., Gooly, T., Martin, P.J., Hansen, J.A.,
Engelhard, V.H. and Riddell, S.R.: Effect of
disparity in the newly identified minor
histocompatibility antigen SKH13 on the
development of graft-versus-host disease after
marrow transplantation from an HLA-identical
sibling. The 42nd Annual Meeting of the American
Society of Hematology. abstract p.202, San
Francisco, California, 2000.
A002. Akatsuka, Y.: Identification of human new
histocompatibility antigens and their clinical
application. The fourth Nagoya International Blood
and Marrow Transplantation Symposium. abstract
pp.36-37, Nagoya, 2001.
A003. Das, K., Basu, M., Henshaw, J., Li, S-C.,
Kannagi, R. and Basu, S.: GalNAcT-1 activity
isolated from guinea pig bone marrow. XVIth
International Symposium on Glycoconjugates, The
Hague, Abstracts, pp. 71, Glycoconjugate J., 18: 92,
2001.
A004. Fujita, M. and Tsurumi, T.: Intranuclear
organization of DNA replication initiation proteins
in mammalian cells. Abstract of the Meeting on
Eukaryotic DNA Replication at The Salk Institute.
pp. 37, 2000.
A005. Fujita, M.: Cell Cycle regulation of DNA
replication initiation proteins in mammalian cells.
Abstract of Symposium on Molecular Network of
G1-S Regulation in Eukaryotic Cells: From G1
Regulaters to DNA Replication Machinery. pp. 13.
2001.
A006. Hamajima, N. and Tajima, K.: Estimation
of cancer susceptibility based on gene-environment
interaction. Aichi Cancer Center International
Symposium VII, p. 36-37, 2001.
A007. Hamajima, N., Katsuda, N., Matsuo, K.,
Saito, T., Ito, L.S., Ando, M., Inoue, M., Takezaki,
T. and Tajima, K.: Smoking habit and interleukin
1B C-31T polymorphism, Program & Abstract of
the 3rd Asian-Pacific Congress of Epidemiology,
2001, p. 48, 2001.
A008. Hamajima, N., Matsuo, K., Mizutani, M.,
Iwata, H., Iwase, T., Miura, S., Obata, Y. and
Tajima, K.: Catechol-O-methyltransferase
polymorphism and breast cancer risk in Japan.
Abstract of the Twentieth International Symposium
of the Sapporo Cancer Seminar Foundation: Gene
Environment Interaction and Cancer Prevention, p.
25, 2000.
A009. Hamajima, N., Tajima, K., Fukumitsu, T.,
Odauchi, S., Usui, T. and Akashi, T.: A large-scale
follow-up study of smokers who visited medical
facilities in Japan. The 22nd Annual Meeting of the
International Association of Cancer Registries,
Book of Abstract, p. 81, 2000.
A010. Haruki, N., Harano, T., Masuda, A.,
Kiyono, T., Tatematsu, Y., Shimizu, S., Mitsudomi,
T., Konishi, H., Osada, H., Fujii, Y. and
Takahashi, Ta.: Persistent increase of chromosome
instability in lung cancer: possible indirect
involvement of p53 inactivation. The 5 th Joint
Conference of the American Association for Cancer
Research and the Japanese Cancer Association. pp.
C-28, Maui, USA, 2001.
A011. Hirose, F., Yamaguchi, M., Taguchi, O.
and Matsukage, A.: Transcriptional regulatory
factor, DREF is involved in regulation of human
CMV US3 gene expression. A Keystone
Symposium Chromatin Structure and Function,
Abstract pp. 80, 2000.
A012. Ichinohe, T., Matsuo, K., Tamaki, S.,
Hamajima, N., Hirabahashi, N. and Dohi, H.:
Superior outcome of blood and marrow stem-cell
transplantations using maternal grafts over
transplants using paternal grafts. Blood, Vol 96
Abstract of the Annual Meeting of the 42 nd
American Society of Hematology, p. 208a, 2000.
A013. Ikehara, Y., Kojima, N., Kurosawa, N.,
Kono, M., Nishihara, S., Itzkowitz, S.H.,
Tatematsu, M., Tsuji, S., and Narimatsu, H.:
Cloning and Expression of the probable candidate
for the cancer associated Sialyl-Tn antigen synthase
(human ST6GalNAcI). XIII International Congress
International Academy of Pathology, p. 181, 2000.
A014. Inada, K., Tanaka, H., Nakanishi, H.,
Tsukamoto, T., Ikehara, Y., Tatematsu, K.,
Nakamura, S., Porter, E.M., and Tatematsu, M.:
Identification of Paneth cells in pyloric glands
associated with gastric and intestinal mixed type
intestinal metaplasia of the human stomach. XIII
International Congress International Academy of
Pathology, p85, 2000.
A015. Inoue, M., Tajima, K. and Tominaga, S.:
Probabilities of developing cancer over the whole
life span of a Japanese. The 22nd Annual Meeting
of the International Association of Cancer
Registries, Book of Abstract, p. 34, 2000.
81

A016. Inoue, M., Tajima, K. and Tominaga, S.:
The estimation of cancer incidence in Aichi
Prefecture, Japan: use of model area with good
quality registry data. The 23rd Annual Meeting of
the International Association of Cancer Registries,
Abstracts book, p. 54, 2001.
A017. Inoue, M., Tominaga, S., Tajima, K.,
Matsuura, A. and Nakamura, S.: Chronic atrophic
gastritis and subsequent gastric cancer: prospective
cohort study in Japan. The 22nd Annual Meeting of
the International Association of Cancer Registries,
Book of Abstract, p. 17, 2000.
A018. Inoue, M.: Recent observations in the
epidemiology of gastric cancer in Japan. 4th
International Gastric Cancer Congress, Abstract
Book, p. 8, 2001.
A019. Kannagi, R., Mitsuoka, C., Kanamori, A.,
Uchimura, K., Muramatsu, T., Imai, T., Yoshie, O.,
Matsushima, K. and Ohmori, K.: Expression and
selectin-binding activity of sialyl 6-sulfo Lewis X
determinant on human helper memory T-
lymphocytes. The 7th International Conference on
Human Leukocyte Differentiation Antigens,
Harrogate, Tissue Antigens 55(Suppl.): 8, 2000.
A020. Kannagi, R.: Glycoconjugate ligands and
regulation of selectin-mediated cell adhesion.
(Plenary Lecture). XVIth International Symposium
on Glycoconjugates, The Hague, Abstracts p. 7,
Glycoconjugate J., 18: 10, 2001.
A021. Kannagi, R.: Regulation of T-lymphocyte
traffic by carbohydrate ligands for selectin.
(Invited Speaker). International Symposium on
Protein Traffic, Glycosylation, and Human Health,
Interlaken, Abstracts p. 17, 2001.
A022. Kato, M., Hayashi, Y. and Yamaguchi, M.:
Identification of dRFX2 that binds to a novel
transcriptional regulatory element of the Drosophila
PCNA gene. Abstracts of Eukaryotic DNA
Replication pp.105, 2001.
A023. Kumamoto, K., Izawa, M., Mitsuoka, C.,
Takenoshita, S. and Kannagi, R.: Significant
alteration of carbohydrate 6-sulfotransferases in
colon cancer and modulation by histone deacetylase
inhibitors. The 5th Joint Conference of the
American Association for Research and the
Japanese Cancer Association "Molecular Biology
and New Therapeutic Strategies: Cancer Research
in the 21st Century," Hawaii, Abstract, pp. B-2,
2001.
82
A024. Kwon, E.-J., Hirose, F., Ohshima, N. and
Yamaguchi, M.: Transcriptional regulation of a
gene for DREF, a key regulatory factor for DNA
replication related genes in Drosophila. Abstracts
of Eukaryotic DNA Replication pp.118, 2001.
A025. Matsuo, K., Hamajima, N. and Tajima, K.:
Methylenetetrahydrofolate reductase (MTHFR)
polymorphsims and life style in malignant
lymphoma. Abstract of the Twentieth International
Symposium of the Sapporo Cancer Seminar
Foundation: Gene Environment Interaction and
Cancer Prevention, p. 26, 2000.
A026. Nagata, K. and Inagaki, M.: Char-
acterization of septin MSF: Possible involve- ment
in cytoskeletal reorganization. 41th American
Society of Cell Biology Annual Meeting.
Molecular Biology of the Cell, 12: 435a, 2001.
A027. Nakanishi, H., Ito, S., and Tatematsu, M.:
Sequential observation of peritoneal micro-
metastasis formation by LacZ gene and GFP
gene-tagged murine carcinoma cells. XIII
International Congress International Academy of
Pathology, 2000.
A028. Nakanishi, H.: Molecular Diagnostic
detection of free cancer cells in the body fluids of
gastrointestinal and bladder cancer patients with
real-time PCR. Program and abstracts of Aichi
Cancer Center International symposium VII, p.
26-27, 2001.
A029. Ohmori, K., Mitsuoka, C., Kanamori, A.,
Adachi, K., Kameyama, A., Takahashi, N.,
Nonoyama, S. and Kannagi, R.: Novel anti-CD15
antibodies with distinct specificity towards Lewis X
determinants carried by mucin core carbohydrates.
The 7th International Conference on Human
Leukocyte Differentiation Antigens, Harrogate,
Tissue Antigens 55(Suppl.):31, 2000.
A030. Ohmori, K., Tei, K., Kumamoto, K., Imai, T.,
Yoshie, O., Hasegawa, H., Matsushima, K. and
Kannagi, R.: Site-specific recruitment of human
helper T cells by synergistic action of sulfated
ligand for selectins and chemokines. The 5th Joint
Conference of the American Association for
Research and the Japanese Cancer Association
"Molecular Biology and New Therapeutic
Strategies: Cancer Research in the 21st Century,"
Hawaii, Abstract, pp. A-62, 2001.
A031. Ohno, K., Takahashi, Y., Hirose, F., Inoue,
Y. H., Taguchi, O., Nishida, Y., Matsukage, A. and
Yamaguchi, M.: Functional analysis of the

Drosophila MLF homologue isolated as a factor
interacting with DREF. A Keystone Symposium
Chromatin Structure and Function, Abstract pp.
69, 2000.
A032. Tajima, K. and Moore, M.: Forefront of
cancer epidemiology and prevention in the Asian
Pacific area, Program & Abstract of The 3rd
Asian-Pacific Congress of Epidemiology, 2001, p.
27, 2001; Proceeding of the 16th Asia Pacific
Cancer Conference, p. 27, 2001.
A033. Tajima, K., Hirose, K., Hamajima, N.,
Inoue, M., Takezaki, T. and Kuroishi, T.:
Lifestyles and cancer in Japan: with special
reference to results from the Hospital-based
Epidemiologic Research Program at Aichi Cancer
Center (HERPACC). Proceedings of the 3rd
Korea-Japan Joint Epidemiology Seminar, Diet and
Health, p. 5, 2001.
A034. Tajima, K., Takezaki, T., Li, H.-C. and
Sonoda, S.: Expansion of ethnoepidemiological
studies on HTLV in Japan and the world. Abstract
of the 8th Japanese-German Workshop on
Molecular and Cellular Aspects of Carcinogenesis,
p. 35-39, 2001.
A035. Tajima, K.: Adult T-cell leukemia/
lymphoma in Japan: Epidemiologic pattern and
prevention strategy. Proceeding of Asian Cancer
Conference in Shizuoka, p. 8, 2000.
A036. Takahashi, Ta., Haruki, N., Kozaki, K.,
Harano, T., Masuda, A., Konishi, H., Hida, T.,
Tatematsu, Y., Saito, H., Yatabe, Y., Yanagisawa,
K., Yoshida, K., Mitsudomi, T. and Osada, H.:
Multidirectional approach for studies on the
carcinogenesis and progression of human lung
cancers. Plenary session, The 9 th World Conference
of Lung Cancer. p. 35-36, Tokyo, Japan, 2000.
A037. Takahashi, Ta., Kozaki, K., Yatabe, Y.,
Achiwa, H. and Hida, T.: Increased expression of
COX-2 in the development of human lung cancers.
Cyclooxygenase-2, a Molecular Target for Cancer
Chemoprevention. p.3, Seoul, Korea, 2001.
A038. Takahashi, Ta.: Altered cell cycle control in
human lung cancer. Symposium on Cell Ceycle
Control & Lung Cancer. p. 5, Hong Kong, China,
2000.
A039. Takahashi, Ta.: Smoking behavior and its
consequences in the pathogenesis of lung cancer.
Molecular Epidemiology of Nicotine Addiction and
Lung Cancer. Maui, USA, 2001.
A040. Takahashi, Ta.: Updates on molecular
pathogenesis of human lung cancer.
Meet-the-Expert-Sunrise-Session. The 91 st Annual
Meeting of the American Association for Cancer
Research. p. ?, San Francisco, USA, 2000.
A041. Takahashi, Ta.: Smoking behavior and it’s
fingerprint in the precess of lung carcinogenesis.
The 20 th International Symposium of the Sapporo
Cancer Seminar Foundation. pp16-17, Sapporo,
Japan, 2001.
A042. Takezaki, T., Ikeda, S., Inoue, M., Tajima,
K. and Tominaga, S.: Cooking methods and risk of
stomach cancer incidence: a 14-year prospective
study in a rural Area of Japan. Program & Abstract
of the 3rd Asian-Pacific Congress of Epidemiology,
2001 -IEA Regional Scientific Meeting in Japan, p.
41, 2001.
A043. Tatematsu, M.: Analysis of the origin of
gastric cancers using animal models. XIII
International Congress International Academy of
Pathology, p.56-63, 2000.
A044. Tatematsu, M.: Reversibility of gastric
submucosal tumor-like lesions in Helicobacter
pylori infected mongolian gerbils with eradication.
U.S._Japan Cooperative Medical Science Program.
Environmental Mutagenesis and Carcinogenesis
Panel, p.10, 2001.
A045. Tsukamoto, T., Fukami, H., Yasutomi, H.,
Yamanaka, S., Nakanishi, H., Aoki, I., and
Tatematsu, M.: Transcriptional down regulation of
hexosaminidase alpha and beta subunits in abrrant
crypts in the 1, 2-dimethylhydrazine treated rat
colon. XIII International Congress International
Academy of Pathology, p103, 2000.
A046. Tsukamoto, T., Inada, K., Fukami, H.,
Yamamoto, M., Tanaka, H., Kusakabe, M, Bishop,
C.E., and Tatematsu, M.: Mouse stain
susceptibility to diethylnitrosamine induced
hepatocarcinogenesis is cell autonomous whereas
sex-susceptibility is due to the micro-environment.
The 5th Joint Conference of the American
Association for Research and the Japanese Cancer
Association "Molecular Biology and New
Therapeutic Strategies: Cancer Research in the 21st
Century," Hawaii, Abstract, pp. A-90, 2001.
A047. Tsurumi, T.: Epstein-Barr virus and human
cancer: Epstein-Barr virus replication proteins. The
21 st International Cancer Symposium in Sapporo.
pp. 52-53. 2001 (Sapporo, Japan)
83

A048. Uchimura, K., El-Fasakhany, F.M.,
Kannagi, R. and Muramatsu, T.: Molecular
cloning, characterization and its implications in
biosynthesis of 3'sulfo-Lewis a . XVIth International
Symposium on Glycoconjugates, The Hague,
Abstracts, pp. 941, Glycoconjugate J., 18: 120,
2001.
A049. Yamaguchi, M., Hayashi, Y. and Hirose, F.:
The homeodomain protein, Distal-less negatively
regulates the function of DREF, a master regulatory
factor for DNA replication-related genes in
Drosophila. Abstracts of Eukaryotic DNA
Replication pp. 226, 2001.
A050. Yamaguchi, M., Hayashi, Y., Ohno, K.,
Kwon, E.-J., Yoshida, H., Hirose, F., Inoue, Y. H.
and Matsukage, A.: Transcriptional regulatory
network for Drosophila. DNA replication-related
genes. Abstracts of Eukaryotic DNA Replication
Meeting pp.122, 2000.
A051. Yoshida, H., Inoue, Y. H., Hirose, F.,
Sakaguchi, K., Matsukage, A. and Yamaguchi,
M.: Ectopic DREF expression induces S phase
and apoptosis in Drosophila wing imaginal discs.
Abstracts of Eukaryotic DNA Replication Meeting.
pp.125, 2000.
A052. Yoshioka, S., Sekine, M., Kannagi, R. and
Suzuki, A.: Mouse kidney tubular epithelial
cell-specific regulation of glycochains and megalin.
XVIth International Symposium on Glyco-
conjugates, The Hague, Abstracts, pp. 95,
Glycoconjugate J., 18: 121, 2001.
84

Abstracts
Epigenetic Alteration in Human Stomach
Cancers
Toshikazu Ushijima
Carcinogenesis Division, National Cancer Center Research
Institute, Tokyo, Japan
CpG methylation plays important roles in carcinogenesis.
To search for tumor suppressor genes
and genes with altered expressions using aberrant
CpG methylation as a marker, we previously developed
a comprehensive genome-scanning method
for differential methylations, methylation-sensitive-
representational difference analysis (MS-RDA)
[Ushijima et al., PNAS, 94; 2284, 1997].
A pair of a gastric cancer and its surrounding
normal tissue was analyzed by MSRDA. Six DNA
fragments were isolated as being flanked by a CpG
island (CGI) and possibly hypermethylated. Three
of the six flanking CGIs were confirmed to be hypermethylated
in the cancer, and two of them had
known genes in their vicinities.
DNA fragment 3A1 was derived from a CGI in
the 5’ region of the Insulin-induced protein 1 (IN-
SIG1/CL-6) gene. Hypermethylation of the CGI
was present in 50% of the primary gastric cancers
(11 of 22), and the hypermethylation was associated
with reduced expression of INSIG1. When a cell
line with hypermethylation and reduced expression
of INSIG1 was treated with 5-aza-2’-deoxycytidine
(aza-dC), a demethylating agent, demethylation of
the CGI and re-expression of INSIG1 were observed.
INSIG1 is known to be expressed when a
fibroblast differentiates into an adipocyte, and it
was suggested that the silencing of INSIG1 was related
to malignant phenotypes in gastric cancers.
DNA fragment 3B4 was derived from intron 7,
near exon 8, of the p41-Arc gene. A CGI spanning
exon 8 was found to be hypermethylated in 10 of
the 22 gastric cancers. p41-Arc expression was
markedly reduced in seven cancers, five of which
contained signet-ring cancer cells. A CGI in the
p41-Arc 5’ upstream region was hypermethylated in
one of these five cancers. p41-Arc is known to be
essential in actin polymerization and cell-shape
control. It was suggested that its decreased expression
was involved in gastric cancer cell morphology,
especially in signet-ring cell cancers.
The role of these new players in gastric carcinogenesis
is being studied by their introduction
88
into gastric cancer cell lines.
E-cadherin Germline Mutations in Familial
Gastric Cancer
Parry J. Guilford
Cancer Genetics Laboratory, Department of Biochemistry,
University of Otago, Dunedin, New Zealand
Hereditary diffuse gastric cancer (HDGC) is a
cancer syndrome caused by inactivating germline
mutations in the gene for the homophilic cell-to-cell
adhesion protein E-cadherin (CDH-1). This syndrome
is typified by early-onset, histologically diffuse
gastric cancer. HDGC families also have a
six-fold increased risk of developing breast cancer,
but do not present with the intestinal form of gastric
cancer. The penetrance of HDGC is higher in females
than males (83% and 67% respectively) and
the age of cancer onset ranges from 14 years upwards.
There is no evidence for any phenotype
variation associated with mutations at different locations
in the CDH-1 gene.
To date, about 25 families from a broad range
of ethnic groups have been identified with inactivating
CDH-1 germline mutations. However, in
Asian populations, the only germline CDH-1 mutations
found in gastric cancer families have been
substitution mutations. It is probable that the high
rate of sporadic gastric cancer in Asia is masking
the “true” inherited gastric cancer families. For reasons
which are not yet clear, HDGC is
over-represented in the New Zealand Maori population.
We speculate that the CDH-1 mutations may
have led to a heterozygote advantage, perhaps related
to E-cadherin’s role as a receptor for the bacterial
pathogen Listeria monocytogenes. Unlike
other familial cancers, LOH is not a common
mechanism for inactivation of the second allele. Instead,
the dominant mechanism for the 2nd hit on
CDH-1 appears to be promoter hypermethylation.
The demonstration that the 2nd hit on CDH-1 need
not be an irreversible event suggests that environmental
or physiological factors which lead to sustained
downregulation of E-cadherin can influence
the genetics of tumor progression. Therefore, the
maintenance of E-cadherin expression must be regarded
as a critical target for chemopreventative
strategies.
Recent detailed histological analyses of HDGC

gastrectomies have shown that stomachs from
HDGC patients develop multifocal clusters of signet
ring cells at a relatively early age. One patient
we analysed recently had >45 independent lesions.
The existence of these multiple foci, which have
lost all E-cadherin expression, suggests that the
second CDH-1 hit has occurred in multiple cells at
a similar moment in time. This broad 2 nd hit argues
strongly for the involvement of an environmental
trigger such as a carcinogen, or a general physiological
event such as inflammation, in the progression
of HDGC. The multifocal disease also suggests
that few irreversible genetic hits are required for the
development of signet ring cells. These lesions are,
however, likely to be slow growing and may constitute
precursor lesions that are not yet fully invasive.
Molecular Classification of Stomach Cancer
by Gene Expression Profiling
Hiroyuki Aburatani
Genome Science Division, Research Center for Advanced
Science and Technology, University of Tokyo,
Tokyo, Japan
Recent molecular analyses have clarified
many genetic alterations in gastric carcinogenesis,
such as p53, α-catenin, E-cadherin, TFF1 and
c-met, but it is still hardly enough to understand
common pathway of carcinogenesis and progression
of gastric cancer. Furthermore, gastric cancer
shows diverse clinical properties such as histological
type, metastatic status, invasiveness and responsiveness
to chemotherapy. Only a little is known
about genes associated with these characteristics.
To gain molecular understanding of carcinogenesis,
progression and diversity of gastric cancer,
22 primary human advanced gastric cancer and 8
non-cancerous gastric tissues were analyzed by
high-density oligonucleotide microarray in this
study. Based on expression analysis of approximately
6800 genes on HuFL array, a two-way clustering
algorithm distinguished cancer tissues from
non-cancerous tissues. Subsequently, differentially
expressed genes between cancer and non-cancerous
tissues were identified with Mann-Whitney's U-test;
162 and 129 genes highly expressed (P

pylori seropositivity were 1.87 (95% confidence interval
[CI] = 1.10 to 3.17) for cardia stomach cancer,
2.29 (95% CI = 1.26 to 4.14) for non-cardia stomach
cancer, and 2.04 (95% CI =1.31 to 3.18) for
both stomach cancer sub-sites combined. Conclusions:
H. pylori seropositivity was associated with
similarly increased risks for both cardia and
non-cardia stomach cancer in this well-characterized
cohort.
Implications: Contrary to most earlier reports, these
data suggest that the procarcinogenic potential of
H. pylori carriage is whole stomach, rather than
limited to the cardia stomach. Future studies should
address whether or not H. pylori eradication represents
a logical stomach cancer prevention strategy
in this high-risk population.
Polymorphisms of Fucosyltransferase Genes
and Helicobacter pylori Infection
Risk
Nobuyuki Hamajima
Division of Epidemiology and Prevention, Aichi Cancer
Center Research Institute, Nagoya, Japan
Helicobacter pylori (HP) infection increases
the risk of diseases including peptic ulcer and
stomach cancer. Although the infection largely depends
on the environmental factors, especially
sanitary conditions in childhood, the genetic factors
play a role in the infection, as a twin study shows.
To date, as well as HLA types, polymorphisms of
TNF-A, Lewis (Le, fucosyltransferase 3), secretor
(Se, fucosyltransferase 2), IL-1B, and myeloperoxidase
have been reported to be possible genetic
factors associated with the infection. The latter four
genes were found out of fifty polymorphisms
screened in Aichi Cancer Center. In this presentation,
I focus on the association of polymorphisms of
Le and Se with the persistent HP infection.
HP with babA2 gene encoding blood-group
antigen-binding adhesin (BabA) has binding activity
to Leb and H type I antigens. Se and Le enzymes
metabolize Type I precursor into H type I and Lea
antigens, respectively. H type I is further metabolized
into Leb by Le enzyme. Commonly observed
in Japanese are Se1, Se2, Le, and le3 for functional
alleles (Se and Le) and sej, se5, le1, and le2 for reduced/no-functional
allele (se and le), respectively.
We examined the associations between
anti-HP IgG antibody and the above genotypes for
241 non-cancer outpatients, and found that individuals
with se/se & Le/Le genotypes had the lowest
seropositivity (33.3%, 9/27) and those with
90
Se/Se & le/le, Se/Se & Le/le, or Se/se & le/le genotypes
the highest (83.8%, 31/37), and the rest intermediate
(62.3%, 109/175 excluding two not
genotyped). Sex-age-adjusted OR of being infected
relative to the lowest group was 3.34 for the intermediate
and 10.21 for the highest.
The expression of Leb and Lea antigens depending
on the genotypes was confirmed in gastric
foveolar epithelium. These findings suggest that Se
and Le genotypes influence the risk of the continued
HP infection through the expression of ligands
for BabA.
Helicobacter pylori Is a Promoter of Stomach
Cancer rather than an Initiator
Masae Tatematsu
Division of Oncological Pathology, Aichi Cancer Center
Research Institute, Nagoya, Japan
In 1994, the World Health Organization/International
Agency for Research on Cancer
concluded that Helicobacter pylori (Hp) is a definite
carcinogen based on the epidemiological evidence.
For detailed analysis of the role of Hp in
stomach carcinogenesis, it is essential to establish a
small animal model. We have established experimental
models of stomach carcinogenesis in Mongolian
gerbils (MGs) using the chemical carcinogens,
N-methyl-N'-nitro-N-nitrosoguanidine (MN-
NG) and N-Methyl-N-nitrosourea (MNU). The lesions
were generally well differentiated, although
poorly differentiated adenocarcinomas were also
found. Hp infection enhances glandular stomach
carcinogenesis in MGs treated with MNNG or
MNU. Animals with high titers of anti-Hp antibodies
are at greatest risk of developing neoplasms. Hp
infection and high-salt diet administration are both
considered being important factors for gastric carcinogenesis
in man. Hp infection exerts stronger
promoting effects than a high-salt diet on gastric
carcinogenesis, and that the two factors act as synergistically
to enhance development of stomach
cancer.
Eradication diminishes enhancing effects of
Hp infection on glandular stomach carcinogenesis
in MGs. Hp eradication may be useful as a prevention
approach. On the other hand, submucosal proliferative
tumor-like lesions are also induced in the
glandular stomach with Hp infection alone, and often
they are similar to carcinomas. To explore if the
role of Hp infection is promotion or initiation, we
established an experimental model of long term Hp
infection and eradication in MGs, without chemical

Current Status and Future Perspective of
Laparoscopic Operation for Early Gastric
Cancer
Michitaka Fujiwara
Department of Surgery II, School of Medicine, Nagoya
University, Nagoya, Japan
Background: Endoscopic mucosal resection
(EMR) and laparoscopic wedge resection offer improved
quality of life after treatment for early gastric
cancer (EGC), but the degree of curability that
can be obtained through these procedures in terms
of systemic lymph node dissection is severely limited.
In 1993, laparoscopy-assisted distal gastrectomy
(LADG) with lymphadenectomy emerged as a
novel option for treating the EGCs of the middle to
lower stomach with potential lymph node involvement.
Due to recent refinements in the technique
and instruments for laparoscopic surgery, some investigators
suggest that LADG can be applied to
more advanced disease.
Where we are today: Between 1995 and 2001,
we have performed 120 laparoscopic operations,
including 7 laparoscopic proximal gastrectomies,
and one laparoscopic total gastrectomy, for gastric
carcinoma that has been diagnosed as confined to
the mucosa or the submucosa through endoscopic
ultrasonography. The number of patients treated
with laparoscopic wedge resection was relatively
small at 20, because EMR is primarily indicated for
a subset of EGC that is estimated not to have lymph
node metastasis. LADG, the most frequently performed
procedure under the laparoscopy, was performed
in 92 patients. LADG was converted to
open surgery in 4 of 92 cases because of uncontrollable
bleeding, positive proximal margin, and macroscopic
finding of lymph node metastasis (confirmed
by frozen section during surgery), but there
was no mortality associated with this procedure.
When the outcome of these patients was compared
with that of the historical control consisting of 80
patients treated with conventional open surgery
(1992~1997), no significant differences in the blood
loss, morbidity, duration of postoperative fever
elevation, and maximum value of CRP were observed.
On the other hand, a smaller amount of analgesics
was required for the LADG patients who
also had better postoperative recovery in terms of
the duration before passing of the flatus and ambulation.
LADG requires use of costly equipments,
but has nevertheless proved less expensive in terms
of the total cost required per a patient, due primarily
to the shorter hospital stay. These patients have
92
been followed for a mean of 24.8 months (range:
2~58 months). One case from each group has so far
died of the recurrent disease. There was no significant
difference in the number of resected lymph
nodes. We believe that D2 lymphadenectomy as defined
by the Japanese Classification for Gastric
Carcinoma can be performed adequately under the
laparoscopy, although a longer follow-up time is
needed for confirmation of the long-term consequences
obtained through this approach.
Future perspective: Advanced laparoscopic
surgery requires intensive training as well as the use
of costly surgical equipments, and can currently be
performed only in specialized institutions. Besides
constructing an adequate training program, further
improvements in surgical devices and navigation
systems may facilitate such operation. From this
viewpoint, we have started the use of
three-dimensional CT angiography for preoperative
simulation as well as for navigation during surgery.
Further progress in the field of optical technology is
warranted.
Update of JCOG 9501 Study; a Randomized
Controlled Trial to Evaluate Para-aortic
Lymphadenectomy for Gastric Carcinoma
Yasuhiro Kodera
Department of Surgery II, School of Medicine, Nagoya
University, Nagoya, Japan
Background: Radical gastrectomy with D2
lymphadenectomy has been a standard procedure
for treatment of gastric carcinoma in Japan and is
considered responsible for the excellent
stage-by-stage survival of these patients. Some patients
nevertheless have recurrences in the
para-aortic lymph nodes. Long-term survivors have
been reported among the population treated with
systemic resection of these nodes in several pilot
studies.
Study design: A multi-institutional randomized
controlled trial was performed to compare treatment
results of para-aortic lymphadenectomy with those
of standard Japanese-style D2 resection. Patients
with histologically proven gastric adenocarcinoma
who at laparotomy was found to have cancer invasion
as far as or beyond the subserosa (T2), negative
cytology (CY0), and no distant or extensive
node metastases (N0~2, M0) were randomized to
receive either the standard D2 resection (Group A)
or D2 plus extensive para-aortic lymph node dissection
(Group B). Primary endpoint of this trial is the

overall survival that is to be evaluated after 5 years
of follow-up. Secondary endpoints include recurrence-free
survival, morbidity, operative mortality,
postoperative hospital stay, and QOL.
Results: The patient accrual started in June
1995 and was completed in April 2001. A total of
523 patients were randomized into Groups A
(n=263) and B (n=260). Cumulative 4-year survival
rate was 69.7%. Difference in survival between the
2 groups has not been evaluated at this time. Two
deaths due to postoperative complications and another
two due to rapid disease progression were
observed, hence the overall hospital mortality of
0.8%. Operative time was longer (300 mins versus
237 mins) and bleeding amount greater (660 mL
versus 430 mL) for Group B, as has been expected.
There was no difference between the groups in the
incidence of major surgical complications such as
leakage, pancreatic fistula, and intra-abdominal abscess,
although complication as a whole was more
frequent among Group B. Complications specific to
Group B were paralytic ileus and prolonged lymphorrhea.
The mean number of lymph nodes retrieved
was 54 (range: 14~161) for Group A and 74
(range: 30~235) for Group B. The mean number of
para-aortic lymph nodes resected by the super-extended
lymphadenectomy was 25 (range:
4~75).
Conclusion: Extended lymphadenectomy with
and without para-aortic lymph node dissection is
safe and feasible when performed at specialized
centers in Japan. Super-extended lymphadenectomy
was associated with longer operating time, greater
blood loss, and higher incidence of surgical complications.
Final survival analyses to assess whether
these shortcomings can be compensated for by a
significant survival benefit is eagerly awaited.
Surgery and Adjuvant Therapy for Gastric
Carcinoma in the U.S.A.
Roderich E. Schwarz
Cancer Institute of New Jersey and Department of Surgery,
University of Medicine and Dentistry of New Jersey,
Robert Wood Johnson Medical School, New Brunswick,
U.S.A.
Gastric cancer, still the predominant cause of
cancer death in the U.S. 60 years ago, has significantly
decreased in incidence and mortality. Physicians
diagnosing or treating gastric cancer in the
U.S. are facing specific characteristics and challenges:
significant social and ethnic patient heterogeneity,
geographic variations in incidence, ad-
vanced stages at diagnosis, significant comorbidity,
a growing number of elderly patients, and a continuing
trend in the prevalence of cardia or proximal
disease location. Treatment is increasingly influenced
by managed care organizations, and access to
specialized cancer- centers can be limited. Thus, the
majority of patients continue to be treated in a
low-volume setting.
Gastrectomy remains the mainstay of therapy
for potentially curable gastric cancer. Although
radical regional resections, including extended
lymph node dissection (ELND), had been utilized
here since the middle of the 20th century, ELND is
still not widely practiced throughout the country. In
a nationwide survey, survival after gastrectomy in
the U.S. remains inferior to that obtained in Japan
or Western Europe. Specialized centers, however,
in which ELND has been routinely applied, are
generating stage-adjusted survival after gastrectomy
which approaches that achieved in Japanese centers
or series. Morbidity and length of hospital stay have
continued to decrease during the past decade. Patterns
of first recurrence after gastrectomy and
ELND suggest that transserosal and hematogenous
dissemination are operational in the vast majority of
clinical relapses, but isolated regional (nodal) recurrences
remain sparse.
A recent U.S. Intergroup trial of postoperative
adjuvant chemoradiation followed by chemotherapy
(INT 116) has resulted in a significant overall survival
and relapse-free survival benefit. While the
chemoradiation therapy (CRT) components were
well quality-controlled, the operative treatment was
not; only 10% of patients underwent formal ELND.
CRT appeared to primarily reduce “local” and “regional”
recurrences, and was associated with an increase
in the relative frequency of distant relapses.
It is unclear whether the radiation component could
functionally substitute in part for the limited regional
resection extent.
In light of the specific characteristics of gastric
cancer treatment in the U.S., one can conclude that:
there is room for standardization of quality assurance
of operative treatment; postoperative adjuvant
chemoradiation therapy has shown a measurable
benefit; CRT has not been validated for patients
having undergone ELND; and routine use of CRT
in different settings (outside the U.S., different recurrence
patterns) appears not warranted at this
time. Adjuvant therapy strategies should be tested
for disease specific challenges as indicated by
dominant relapse patterns.
93

Author index for research reports and publications
________________________________________________________________________________
Akatsuka, Y. 34, J001, J008, J157, R001, R043,
R044, A001, A002
Aoki, K. 7, J067, J189
Ariyoshi, Y. 7, J197
Doi, T. J055, J127
Fujii, K. 37, 37
Fujita, M. 37, 38, J004, J017, J018, J221,
R004, R005, R056, A004, A005
Fukami, H. 57, J201, J202, J203, A045, A046
Gao, C. J021, J183
Goto, H. 49, 50, J134, J150, J215, J227,
J228, R006, R011
Goto, Y. 43, 44, 45, J101, R014
Hagino, M. 59
Hamajima, N. 7, 10, 12, 20, 30, 56, J024, J025,
J026, J027, J028, J029, J030, J031,
J032, J033, J044, J052, J053, J054,
J057, J066, J074, J076, J088, J102,
J103, J116, J117, J118, J119, J120,
J121, J122, J124, J125, J142, J168,
J169, J175, J178, J182, J184, J185,
J186, J188, J189, J207, J224, J225,
R007, R008, R032, R033, R036,
R046, R047, A006, A007, A008,
A009, A012, A025, A033
Harada, H. J035, J102, J103, J191, J205
Harano, T. 24, 25, J036, J037, A010, A036
Haruki, N. 25, J005, J036, J037, J038, J039,
J123, J141, J168, J169, A010,
A036
Hasegawa, Y. J186
Hata, M. J040, J041, J092, J151, R038,
R039
Hatooka, S. J102, J103, J118, J119, J168, J169,
J186, J191, J196
Hayashi, N. 40, 61, J090, J105
Hayashi, Y. 52, 52, J211, A022, A049, A050
Hayashi, Y. J211, A022, A049, A050
Hida, T. 23, J042, J099, J100, J115, R009,
A036, A037
Hirai, T. J033, J126, J184
Hiraiwa, N. 46, J096, J097
Hirata, M. J221
Hirose, F. 52, J043, J107, J146, J154, J211,
J216, A011, A024, A031, A049,
A050, A051
Hirose, K. 5, 10, 12, J044, J054, J066, J074,
J104, J116, J185, J186, J223,
R007, R046, R047, R055, A033
Hoshino, Y. 39, J046, J090, J106
Hosokawa, Y. 29, 30, 31, J047, J048, J049, J050,
J051, J058, J129, J179, J181, J222
Huang, X. J053, J054
Ichinose, M. J096
Iidaka, T. 19, J158
Ikehara, Y. 18, J057, J060, J077, J166, J167,
A013, A014
Imai, T. R015, A019, A030
Inada, H. 50, J022, J059, J080, J150, R011
Inada, K. 18, 19, 20, J057, J060, J069, J126,
J152, J159, J166, J201, A014,
A046
Inagaki, M. 49, 50, 51, J022, J045, J059, J062,
J080, J098, J109, J134, J150, J194,
J208, J215, J227, J228, R002,
R006, R011, R035, A026
Inagaki, N. J062
Inoue, M. 5, 7, 12, J027, J033, J044, J054,
J064, J065, J066, J067, J068, J074,
J076, J088, J116, J118, J119, J185,
J186, J202, J203, R007, R012,
R046, R047, A007, A015, A016,
A017, A018, A033, A042
Inoue, Y. 52, J043, J063, J146, J154, J211,
J216, A031, A050, A051
Inoue, Y.H. 52
Ishida, H. 44, J023, J081
Ishida, R. J140
Ishizaki, K. 55, 56, 57, J035, J073, J102, J103,
J191, J192, J205, R010
Ito, H. J074
Ito, S. J077, J088, J111, J135, A027
Iwase, S. J078, J127, J199
Iwase, T. 34, J026, J031, J053, J066, J079,
A008
Iwata, H. J026, J031, J053, J066, J079,
A008
Izawa, I. 50, 51, J059, J080, J150, R035
Izawa, M. 44, J081, J097, J164, R014, A023
Kagami, Y. J083, J095, J113, J122, J131, J155,
J175, J210
Kanamori, A. 43, 44, J006, J007, J023, J081,
95

J084, J145, J206, R015, R037,
A019, A029
Kanda, K. 44, J145
Kannagi, R. 43, 44, 45, 46, J013, J016, J020,
J023, J056, J075, J081, J084, J086,
J087, J094, J096, J097, J101, J138,
J145, J161, J164, J187, J193, J206,
R013, R014, R015, R016, R017,
R018, R019, R020, R037, A003,
A019, A020, A021, A023, A029,
A030, A048, A052
Kato, M. J067, J131, A022
Kato, T. 18, J033, J073, J077, J111, J126,
J135, J184, J192, R010
Kawajiri, A. 49, 51
Kawakami-
Kimura, N. J193
Kitamura, T. 20, J057
Kiyono, T. 25, 37, 39, 50, 57, J017, J018,
J036, J059, J162, J163, J221,
R021, R022, R023, R024, A010
Kobayashi, S. J079, J196
Kobayashi, T. J182
Kodera, Y. 20, J052, J057, J126, J129, J135,
J137, J212
Koiwai, O. J154
Kondo, E. 34, J095, J113, J131
Konishi, H. 23, 24, 25, 26, J036, J038, J141,
J217, A010, A036
Kosako, H. J098, R006
Koshikawa, K. 23, J099
Koshikawa, T. J168, J169
Kozaki, K. 23, J032, J042, J099, J100, J111,
J115, J224, A036, A037
Kumamoto, K. 44, 45, J081, J101, J187, J193,
J206, R014, A023, A030
Kumimoto, H. 55, 56, J035, J102, J103, J205
Kuroishi, T. 5, 7, 11, J054, J074, J104, J116,
J185, J186, R007, R046, R047,
A033
Kuzushima, K. 37, 39, 40, 61, J017, J046, J090,
J105, J106, J198, J221, R025,
R026, R027, R028, R029, R030,
R031
Kuzuya, K. 7
Kwon, E. -J. 52, J107, J108, A024, A050
Maeda, Y. J047, J048, J049, J181
Masuda, A. 24, 25, 26, J002, J036, J038, J042,
J115, J123, J153, J213, A010,
96
A036
Matsudaira, Y. 35, J078, J199, J200
Matsui, S. 49, 51, J215
Matsukage, A. 52, J043, J063, J082, J107, J146,
J154, J211, J216, A011, A031,
A050, A051
Matsuo, K. 20, 56, J024, J026, J027, J028,
J029, J030, J031, J032, J033, J053,
J057, J074, J088, J102, J103, J116,
J117, J118, J119, J120, J121, J122,
J184, J188, J225, R007, R032,
R033, A007, A008, A012, A025
Matsuura, A. J030, J065, J121, J133, R032,
A017
Matsuura, H. J186
Minoura, Y. 59
Mitsudomi, T. 7, 23, 25, 34, J005, J014, J036,
J042, J125, J141, J168, J169, J185,
J217, R034, R052, A010, A036
Mitsuoka, C. J075, J081, J145, R015, R037,
A019, A023, A029
Miura, K. J081, J095, J145
Miura, S. 7, 34, J026, J031, J044, J053,
J066, J079, J097, J223, A008
Mizoshita, T. 19, J126
Mizuno, K. 25
Mizutani, K. J127
Mizutani, M. J026, J031, J053, J066, J079,
A008
Moore, M. J128, R045, A032
Morishima, Y. 34, J083, J095, J113, J117, J120,
J122, J124, J129, J155, J175, J210,
J218, J219, J222, R033
Motegi, M. J129
Murai, H. J097
Nagata, K. 49, 51, J109, J134, J194, J208,
J215, R011, R035, A026
Nagatake, M. J213
Nakagawa, T. 24, 25, J123
Nakamura, H. 57, 59, J018, J137
Nakamura, S. 30, 44, 61, J020, J039, J060, J061,
J065, J081, J083, J095, J113, J120,
J122, J124, J129, J130, J131, J132,
J133, J174, J175, J176, J177, J178,
J186, J190, J196, J210, J217, J218,
J219, J222, A014, A017
Nakamura, T. 30, 61, J030, J065, J121, J132,
J133, J196, R032
Nakanishi, H. 18, 19, J002, J060, J069, J077,

J111, J135, J143, J159, J166,
A014, A027, A028, A045
Nakasu, S. 38
Nakayashiki, N. 33, J136
Narita, T. J096, J097
Natsume, A. J003
Nishi, Y. J043, J211
Nishida, K. J140, J190
Nishida, T. 34
Nishimoto, Y. 55, J063, J102, J103
Nishizawa, K. J073, J102, J103, J205, R010
Nishizawa, M. 50, 51, J059, J062, J080
Nomoto, S. J037, J039, J141
Nozaki, K. 18, J166, J167
Obata, Y. 34, 35, J026, J031, J055, J078,
J079, J127, J142, J170, J199, J200,
R003, R036, A008
Ogura, M. J095, J113, J122, J175
Ohashi, K. J026, J065, J133, J184, J196
Ohno, K. J010, J107, J146, A031, A050
Ohshima, N. 52, J043, A024
Ohtakara, K. 51, J080, J150
Ohtsuka, K. J040, J041, J080, J092, J151, J209,
J221, R038, R039, R040
Okuma, K. 7, J026, J029
Osada, H. 23, 24, 25, J036, J037, J039, J099,
J115, J149, J153, J213, R041,
R042, A010, A036
Ozeki, S. 35, J078, J199, J200
Saito, H. 23, 25, J037, J038, J070, J093,
J099, J137, J156, J195, J204, J212,
J224, A036
Saito, N. 51
Saito, S. J161
Saito, T. 7, J027, J029, J032, J033, J053,
J074, J088, R007, A007
Saito, T. 24, 26, J153, J213
Sakai, H. 19, J158, J159
Seto, M. 29, 30, 31, J009, J015, J047, J048,
J049, J050, J061, J083, J089, J110,
J120, J122, J124, J129, J130, J132,
J133, J137, J140, J155, J174, J175,
J176, J177, J178, J179, J181, J182,
J190, J196, J210, J218, J219, J222
Shimizu, N. 18, 20, J057, J166, J167
Shimizu, S. 25, J005, J036, J042, J168, J169,
A010
Shinoda, M. 55, J035, J102, J103, J118, J119,
J168, J186, J191, J196
Shirai, N. 19, J158
Shiraki, M. J043, J063
Suchi, T. J175, J218, J219
Sugaya, Y. J173
Sugiura, T. 7, J042, J185, J224
Sugiyama, M. 24, J153, J213
Suyama, M. J168, J169
Suzuki, H. 29, 30, J129, J179, J222
Suzuki, R. 30, 31, J034, J085, J091, J095,
J113, J120, J122, J124, J129, J131,
J175, J176, J177, J178, J179, J210,
J212, J218, J219, J222, R033
Suzuki, S. J152
Suzuki, T. 11, J030, J065, J104, J121, J133,
J196, R032, R040
Taguchi, O. 46, J043, J096, J097, J146, J193,
A011, A031
Taji, H. 34, J095, J113, J122, J131
Tajima, K. 5, 7, 10, 12, 34, J021, J026, J027,
J029, J030, J031, J032, J033, J044,
J052, J053, J054, J064, J065, J066,
J067, J074, J076, J088, J116, J118,
J119, J120, J122, J124, J147, J148,
J165, J171, J172, J180, J183, J184,
J185, J186, J214, J223, J226,
R007, R045, R046, R047, R048,
R049, R050, R051, A006, A007,
A008, A009, A015, A016, A017,
A025, A032, A033, A034, A035,
A042
Takagishi, M. 51, J194
Takahashi, H. J181
Takahashi, M. 34, J202, J203
Takahashi, Ta. 23, 24, 25, 26, J005, J012, J014,
J032, J036, J037, J038, J039, J042,
J099, J100, J115, J123, J125, J141,
J153, J169, J197, J213, J217, J224,
R009, R034, R041, R042, R052,
R053, A010, A036, A037, A038,
A039, A040, A041
Takahashi, To. 33, 34, 35, J003, J055, J059, J078,
J100, J115, J127, J136, J139, J141,
J142, J199, J200, J220, R036
Takahashi, Y. J146, A031
Takeuchi, T. J085, J095, J097, J223
Takezaki, T. 7, 12, J021, J027, J033, J054,
J074, J088, J116, J118, J119, J172,
J183, J184, J185, J186, J226,
R007, R046, R047, R049, R054,
97

A007, A033, A034, A042
Tamaki, H. 34, J142, R036
Tanabe, K. 59
Tanaka, H. J060, J191, J201, J202, J203,
A014, A046
Tatematsu, M. 18, 19, 20, J019, J020, J057, J060,
J069, J071, J072, J077, J111, J114,
J126, J128, J135, J143, J144, J152,
J158, J159, J166, J167, J201, J202,
J203, A013, A014, A027, A043,
A044, A045, A046
Tatematsu, Y. 23, 24, 25, J036, J038, J099, J100,
J115, J141, A010, A036
Tei, K. J193, R014, A030
Terashima, M. 59
Togashi, H. J194
Tokumasu, S. J205
Tominaga, S. 5, 11, J029, J030, J052, J054,
J064, J065, J066, J067, J076, J088,
J104, J121, J142, R032, R036,
A015, A016, A017, A042
Tsujimura, K. 35, J003, J078, J139, J199, J200
Tsukamoto, T. 18, 19, J060, J100, J126, J158,
J159, J166, J201, J202, J203,
A014, A045, A046
Tsurumi, T. 37, 38, 39, 40, 61, J017, J018,
J046, J105, J106, J221, R029,
R056, R057, R058, R059, A004,
A047
98
Uchida, K. J213
Uchida, N. J035, J205
Ueda, R. J015, J061, J113, J137, J168,
J169, J182, J190, J210, J212
Yamagishi, M. 52, J082, J211
Yamaguchi, M. 52, J010, J043, J063, J082, J107,
J108, J146, J154, J210, J211, J216,
J218, J219, A011, A022, A024,
A031, A049, A050, A051
Yamamoto, M. 19, J079, J158, J159, J201, A046
Yamamura, Y. 20, 61, J020, J052, J057, J126,
J135
Yanagisawa, K. 24, J153, J213, A036
Yasui, K. J111
Yasui, Y. 49, 50, J215
Yasutomi, H. 20, J057, A045
Yatabe, Y. 24, 26, J039, J115, J131, J133,
J168, J169, J185, J217, J218, J219,
A036, A037
Yokoyama, N. 37, J011, J017, J160, J221
Yonezumi, M. J110, J129, J132, J133, J222
Yoshida, H. 52, J154, J211, J216, A050, A051
Yoshida, M. 34
Yoshida, T. J098, J131
Yoshikawa, K. 33, 35, J136, J200
Yuasa, H. J028, J225
Zenita, K. J097