118

Advanced Clinical Research Center

Division of Clinical Genome Research
臨床ゲノム腫瘍学分野

Professor
Yoichi Furukawa, M.D., Ph.D.
Associate Professor Kiyoshi Yamaguchi, Ph.D.
Assistant Professor Kiyoko Takane, M.D., Ph.D.

教 授准教授助 教

博士（医学）
博士（薬学）
博士（医学）

古山高川口根洋 一
貴世志希世子Research Projects
The aim of our research is the application of findings in basic cancer research toclinics. Currently, we are working on the following six projects: 1) Identification ofnovel molecular targets for the treatment of colorectal cancer, 2) understanding therole of Wnt/β-catenin signaling pathway in human carcinogenesis, 3) discovery ofWnt inhibitors through a screening of large-scale chemical libraries, 4)establishment of intrahepatic cholangiocarcinoma mouse model by orthotopic
transplantation of syngeneic tumor cells, 5) elucidation of the genetic features ofrare cancers and the mechanisms of their development, and 6) clinical sequencing
for the implementation of genomic medicine.

1. Identification of novel molecular targets for thetreatment of colorectal cancer
Kiyoshi Yamaguchi, Yoichi Furukawa
Epigenetic modifications such as DNA methylation and histone modification change in gene expression with global dynamics of chromatin structure.
Accumulated evidence has demonstrated that aberrant epigenetic modifications are involved in carcinogenesis. Bromodomain has been known as a protein-interaction module that recognizes acetylated
lysine residues. Through this interaction, protein containing a bromodomain(s) directs the assembly of nuclear factor complexes to their target sites on chromatin, resulting in the transcriptional activation.
Recently, we found that bromodomain containing 8(BRD8) was frequently accumulated in colorectal cancer. Transcriptome analysis coupled with genome-wide mapping of BRD8-binding sites disclosedthat BRD8 regulates the expression of multiple subunits of the pre-replicative complex in concert with theactivator protein-1. Depletion of BRD8 induced
cell-cycle arrest at the G1 phase and suppressed cellproliferation. We also showed that the bromodomainof BRD8 is indispensable for not only the interaction
with histone H4 or transcriptional regulation but also
its own protein stability. These findings highlight theimportance of bromodomain as a therapeutic target.
2. Understanding the role of Wnt/β-catenin signaling pathway in human carcinogenesis
Kiyoshi Yamaguchi, Yoichi Furukawa
Aberrant Wnt/β-catenin signaling has been foundin the various types of cancer, particularly colon cancer. This activation leads to the accumulation of β-catenin in the nucleus, where it functions as a transcriptional co-activator of the TCF/LEF family. Therefore,
comprehensive understanding of genes directlytransactivated by the heterodimeric β-catenin/TCF
transcriptional complex will lead to the better understanding of the role of this pathway in human carcinogenesis. Previously, our gene expression analysis
using β-catenin-depleted colorectal cancer cells identified a total of 64 candidate genes whose expression
was regulated by the Wnt/β-catenin signaling. One of119
these genes was visinin-like 1 (VSNL1), which belongs
to the neuronal calcium-sensor gene family. The expression of VSNL1 was regulated by β-catenin/TCF7L2 via two Wnt-responsive elements in intron 1.
Knockdown of VSNL1 induced apoptosis in colorectal cancer cells. In contrast, forced expression of wildtype VSNL1, but not a myristoylation, Ca2+-binding,or dimerization-defective mutant, suppressed the apoptosis induced by camptothecin and doxorubicin.
Our data suggest that VSNL1, a novel target gene of
the Wnt/β-catenin signaling pathway, is associatedwith apoptosis resistance in colorectal cancer cells.
3. Discovery of Wnt inhibitors through a screening of large-scale chemical libraries
Kiyoshi Yamaguchi, Yoichi Furukawa, Yoshitaka
Ohishi, Satoru Nagatoishi1, Kouhei Tsumoto1,2,3: 1Project Division of Advanced Biopharmaceutical Science,2
Medical Proteomics Laboratory, IMSUT, 3Department of Bioengineering, School of Engineering, The
University of Tokyo
A variety of cell-based assays have contributed tothe discovery of small molecules that modulate Wntsignaling. Previously, we developed a sensitive andspecific cell-based reporter assay for the detection of
the Wnt/β-catenin signaling activity. Using this assay,
we established a high-throughput screening system,and performed a screening of small molecule and natural product libraries. As a result, several compounds
that inhibit Wnt/β-catenin signaling activity wereidentified, and their target(s) are currently under investigation. To elucidate the detailed mode of actionof these hit compounds, we have adopted an approach based on the estimation of gene network fromgene expression data.
4. Establishment of intrahepatic cholangiocarcinoma mouse model by orthotopic transplantation of syngeneic tumor cells
Kiyoko Takane, Yoichi Furukawa
Genetically engineered mice are useful tools forstudying human diseases including cancer. In thisproject, we previously generated a novel mouse model of intrahepatic cholangiocarcinoma (ICC) using liver-specific expression of oncogenic Kras and homozygous Pten deletion (AKPP: Alb-Cre+; LSL-KrasG12D/+;Ptenflox/flox). Subsequently, a cell line was establishedfrom the ICC of AKPP mice and named AKPP cells.
First, we confirmed that conventional ICC markers
such as cytokeratin-19 and pan-cytokeratin wereabundantly expressed in AKPP cells. In contrast, thecells expressed low levels of hepatocyte nuclear factor
4 alpha (HNF4α), a marker of hepatocellular carcinoma. To investigate the downstream effects expectedfrom the loss of Pten and activation of Kras, the phosphorylation levels of Akt (Ser473) and Erk1/2 wereanalyzed by immunoblotting. As a result, increased
phosphorylation of these effectors was observed in
AKPP cells, compared to mouse fibroblast L-cells. We
are currently investigating the tumorigenic potentialof AKPP cells using syngeneic immunocompetentmice.
5. Elucidation of the genetic features of rare cancers and the mechanisms of their development
Kiyoko Takane, Kiyoshi Yamaguchi, Yoichi Furukawa, Seiya Imoto1, Satoru Miyano2, Hideaki Yano3,Atsushi Kaneda4: 1Division of Health Medical Intelligence, Human Genome Center, IMSUT, 2Systems
Biology for Intractable Diseases, Medical ResearchInstitute, Tokyo Medical and Dental University, 3Department of Surgery, National Center for Global
Health and Medicine, 4Department of MolecularOncology, Graduate School of Medicine, Chiba University
Pseudomyxoma peritonei (PMP) is a rare disease
with an incidence of 1 – 2 cases per million and characterized by the presence of mucin-producing tumors
in the abdominal cavity. Primary tumors of PMP develop most frequently in the appendix and occasionally in other organs including the ovary, colorectum,
gallbladder, stomach, pancreas, fallopian tube, urachus, lung, and breast. To elucidate the molecularmechanisms underlying PMP, we previously analyzed 18 appendiceal PMPs by targeted sequencing using the Cancer Hotspot Panel. Consequently, we
found that KRAS and/or GNAS mutations are common genetic features of PMP. In addition, we suggested that mutations in TP53 and/or genes related to thePI3K-AKT pathway may confer malignant properties
to PMP.
We further performed genome-wide DNA methylome analysis of 15 appendiceal PMP samples using
Infinium 850K BeadChip. As a result, the 15 PMPswere classified into at least two epigenotypes, uniquemethylation epigenotype and normal-like methylation epigenotype. In addition, we have found that
HOXD1 and TSPYL5 showed high methylation andlow expression in all PMP samples tested. We are currently investigating the molecular mechanisms ofPMP involved in the low expression of HOXD1 andTSPYL5. These data may contribute to the better understanding of molecular mechanisms underlying
PMP, and help in the diagnosis, treatment, and prevention of this life-threatening disease.
6.  Clinical sequencing for the implementation ofgenomic medicine
Kiyoko Takane, Kiyoshi Yamaguchi, Yoichi Furukawa, Eigo Shimizu1, Rika Kasajima1, Kotoe Katayama1, Seiya Imoto1, Tetsuo Shibuya2, Kazuaki
Yokoyama3, Yasuhito Nanya3, Koichiro Yuji4, Rui

120Yamaguchi5, Satoru Miyano6: 1Division of Health
Medical Intelligence, 2Division of Medical Data Informatics, Human Genome Center, 3Department ofHematology/Oncology, 4Project Division of International Advanced Medical Research, Advanced Clinical Research Center, IMSUT, 5Division of Cancer
Systems Biology, Aichi Cancer Center Research Institute, 6Systems Biology for Intractable Diseases,Medical Research Institute, Tokyo Medical andDental University
The application of Next-Generation Sequencing(NGS) technology in clinical medicine has revolutionized molecular diagnostics by enabling multiple genetesting, or analysis of the entire exon or whole genome with a limited amount of DNA. In collaboration
with Human Genome Center and Advanced ClinicalResearch Center, we have been working on two projects: 1) genetic diagnosis of patients with suspected
hereditary cancer predisposition and 2) implementation of precision medicine for patients with rare or
intractable cancer.
In the first project, we applied NGS technology formolecular diagnostics of hereditary colon cancer syndromes such as familial adenomatous polyposis(FAP), Lynch syndrome (LS), and polymerase proofreading-associated polyposis (PPAP). Long-read sequencing has technical advantages over short-readsequencing for the detection of structural variants.
Therefore, we tested the ability of nanopore sequenc-ing and adaptive sampling for the enrichment of 91
genes involved in hereditary cancer predisposition
using DNA from a patient with LS carrying a characterized SV in our previous study. Application of theadaptive sampling using peripheral blood DNA resulted in approximately six times higher target composition than the nonadaptive sampling. Consequently, we successfully identified the breakpoints of apathogenic SV that was difficult to identify by shortread sequencing technology.
In the second project, we have been working on
the implementation of genomic data in clinics. We offered consultation of genetic analysis to patients withrare or intractable cancer as an outpatient clinic service in IMSUT hospital. Patients with colorectal, esophageal, cervical, pancreatic, and breast cancer gave
written informed consent for genetic analysis andprediction of treatment using artificial intelligence
were enrolled in this study. Genetic alterations intheir tumors were determined by NGS, and the data
were subsequently analyzed by QIAGEN Clinical Insights (QCI). The results of QCI including predicted
driver mutations and suggested actionable drugs
were discussed in the IMSUT Tumor Board. The Tumor Board is composed of members from a variety ofspecialties, including physicians, medical oncologists,
genetic counsellors, geneticists, bioinformaticians,and experts of ethics, and meets online every twoweeks.

Publications
1.		Hiranuma, R., Sato, R., Yamaguchi, K., Nakamizo,S., Asano, K., Shibata, T., Fukui, R., Furukawa, Y.,Kabashima, K. and Miyake, K. Aberrant monocytopoiesis drives granuloma development in sarcoidosis. Int Immunol. 2023 in press
2.		Nanamiya, T., Takane, K., Yamaguchi, K., Okawara, Y., Arakawa, M., Saku, A., Ikenoue, T., Fujiyuki, T., Yoneda, M., Kai, C. and Furukawa, Y. Expression of PVRL4, a molecular target for cancer
treatment, is transcriptionally regulated by FOS.
Oncol Rep. 51(1):17, 2024.
3.		Tanaka, N., Okada, H., Yamaguchi, K., Seki, M.,
Matsubara, D., Gotoh, N., Suzuki, Y., Furukawa,Y., Yamashita, T., Inoue, J.I., Kaneko, S. and Sakamoto, T. Mint3-depletion-induced energy stresssensitizes triple-negative breast cancer to chemotherapy via HSF1 inactivation. Cell Death Dis.
14(12):815, 2023.
4.		Kawachi, K., Tang, X., Kasajima, R., Yamanaka, T.,
Shimizu, E., Katayama, K., Yamaguchi, R.,
Yokoyama, K., Yamaguchi, K., Furukawa, Y., Miyano, S., Imoto, S., Yoshioka, E., Washimi, K., Okubo, Y., Sato, S., Yokose, T. and Miyagi, Y. Geneticanalysis of low-grade adenosquamous carcinomaof the breast progressing to high-grade metaplas-tic carcinoma. Breast Cancer Res Treat. 202(3):563573, 2023.
5.		Takeuchi, K., Tabe, S., Takahashi, K., Aoshima, K.,Matsuo, M., Ueno, Y., Furukawa, Y., Yamaguchi,
K., Ohtsuka, M., Morinaga, S., Miyagi, Y., Yamaguchi, T., Tanimizu, N. and Taniguchi, H. Incorporation of human iPSC-derived stromal cells creates a pancreatic cancer organoid withheterogeneous cancer-associated fibroblasts. Cell
Rep. 42(11):113420, 2023.
6.		Matsuura, M., Takane, K., Yamaguchi, K., Ikenoue,T., Hatakeyama, S., Kurokawa, S., Tamate, M.,
Akimoto, T., Iwasaki, M., Sugita, S., Hasegawa, T.,
Ota, Y., Saito, T. and Furukawa, Y. Identification ofcancer driver mutations in liquid-based cytology
samples for the screening of endometrial diseases.
BJC Rep. 1:18, 2023.
7.		Rizq, O., Mimura, N., Oshima, M., Momose, S.,
Takayama, N., Itokawa, N., Koide, S., Shibamiya,A., Miyamoto-Nagai, Y., Rizk, M., Nakajima-Takagi, Y., Aoyama, K., Wang, C., Saraya, A., Seimiya,M., Watanabe, M., Yamasaki, S., Shibata, T., Yamaguchi, K., Furukawa, Y., Chiba, T., Sakaida, E., Nakaseko, C., Tamaru, J.I., Tai, Y.T., Anderson, K.C.,
Honda, H. and Iwama, A. UTX inactivation in ger-121
minal center B cells promotes the development ofmultiple myeloma with extramedullary disease.
Leukemia. 37(9):1895-1907, 2023.
8.		Ikushima, H., Yamaguchi, K., Furukawa, Y., Imoto, S., Koda, H., Mizukami, T., Morikawa, T. andUchino, K. Transdifferentiation of cervical squamous cell carcinoma with ERBB2 amplification toadenocarcinoma: whole genome sequence analysis and successful control by anti-HER2 therapy.
BJC Rep. 1:12, 2023.
9.		Shibata, T., Sato, R., Taoka, M., Saitoh, S.I., Komine,M., Yamaguchi, K., Goyama, S., Motoi, Y., Kitaura,J., Izawa, K., Yamauchi, Y., Tsukamoto, Y., Ichinohe, T., Fujita, E., Hiranuma, R., Fukui, R., Furukawa, Y., Kitamura, T., Takai, T., Tojo, A., Ohtsuki,M., Ohto, U., Shimizu, T., Ozawa, M., Yoshida, N.,Isobe, T., Latz, E., Mukai, K., Taguchi, T., Hemmi,
H., Akira, S. and Miyake, K. TLR7/8 stress response drives histiocytosis in SLC29A3 disorders.
J Exp Med. 220(9):e20230054, 2023.
10.	Suzuki, M., Kasajima, R., Yokose, T., Shimizu, E.,
Hatakeyama, S., Yamaguchi, K., Yokoyama, K.,
Katayama, K., Yamaguchi, R., Furukawa, Y., Miyano, S., Imoto, S., Shinozaki-Ushiku, A., Ushiku,
T. and Miyagi, Y. KMT2C expression and DNA
homologous recombination repair factors in lungcancers with a high-grade fetal adenocarcinoma
component. Transl Lung Cancer Res. 12(8):17381751, 2023.
11.	Li, D., Johmura, Y., Morimoto, S., Doi, M., Nakanishi, K., Ozawa, M., Tsunekawa, Y., Inoue-Yamauchi, A., Naruse, H., Matsukawa, T.,
Takeshita, Y., Suzuki, N., Aoki, M., Nishiyama, A.,
Zeng, X., Konishi, C., Suzuki, N., Nishiyama, A.,
Harris, A.S., Morita, M., Yamaguchi, K., Furukawa, Y., Nakai, K., Tsuji, S., Yamazaki, S., Yamanashi,Y., Shimada, S., Okada, T., Okano, H., Toda, T. andNakanishi, M. LONRF2 is a protein quality control ubiquitin ligase whose deficiency causes
late-onset neurological deficits. Nat Aging.
3(8):1001-1019, 2023.
12.	Iwamoto, R., Yamaguchi, K., Katayama, K., Ando,H., Setsukinai, K.I., Kobayashi, H., Okabe, S., Imoto, S. and Kitajima, M. Identification of SARSCoV-2 variants in wastewater using targeted amplicon sequencing during a low COVID-19prevalence period in Japan. Sci Total Environ.
887:163706, 2023.

13.	Nakajima-Takagi, Y., Oshima, M., Takano, J.,
Koide, S., Itokawa, N., Uemura, S., Yamashita, M.,Andoh, S., Aoyama, K., Isshiki, Y., Shinoda, D.,Saraya, A., Arai, F., Yamaguchi, K., Furukawa, Y.,
Koseki, H., Ikawa, T. and Iwama, A. Polycomb repressive complex 1.1 coordinates homeostatic andemergency myelopoiesis. Elife. 12:e83004, 2023.
14.	Tage, H., Yamaguchi, K., Nakagawa, S., Kasuga,S., Takane, K., Furukawa, Y. and Ikenoue, T. Visinin-like 1, a novel target gene of the Wnt/β-cateninsignaling pathway, is involved in apoptosis resistance in colorectal cancer. Cancer Med. 12(12):1342613437, 2023.
15.	Takaori, A., Hashimoto, D., Ikeura, T., Ito, T., Nakamaru, K., Masuda, M., Nakayama, S., Yamaki,S., Yamamoto, T., Fujimoto, K., Matsuo, Y., Akagawa, S., Ishida, M., Yamaguchi, K., Imoto, S., Hirota, K., Uematsu, S., Satoi, S., Sekimoto, M. and Naganuma, M. Impact of neoadjuvant therapy on gutmicrobiome in patients with resectable/borderline
resectable pancreatic ductal adenocarcinoma.
Pancreatology. 23(4):367-376, 2023.
16.	Washimi, K., Kasajima, R., Shimizu, E., Sato, S.,
Okubo, Y., Yoshioka, E., Narimatsu, H., Hiruma,
T., Katayama, K., Yamaguchi, R., Yamaguchi, K.,
Furukawa, Y., Miyano, S., Imoto, S., Yokose, T. andMiyagi, Y. Histological markers, sickle-shaped
blood vessels, myxoid area, and infiltrating
growth pattern help stratify the prognosis of patients with myxofibrosarcoma/undifferentiatedsarcoma. Sci Rep. 13(1):6744, 2023.
17.	Yamaguchi, K., Nakagawa, S., Saku, A., Isobe, Y.,
Yamaguchi, R., Sheridan, P., Takane, K., Ikenoue,T., Zhu, C., Miura, M., Okawara, Y., Nagatoishi, S.,
Kozuka-Hata, H., Oyama, M., Aikou, S., Ahiko, Y.,Shida, D., Tsumoto, K., Miyano, S., Imoto, S. andFurukawa, Y. Bromodomain protein BRD8 regulates cell cycle progression in colorectal cancer
cells through a TIP60-independent regulation of
the pre-RC complex. iScience. 26(4):106563, 2023.
18.	Yamaguchi, K., Horie, C., Takane, K., Ikenoue, T.,Nakagawa, S., Isobe, Y., Ota, Y., Ushiku, T., Tanaka, M., Fujishiro, J., Hoshino, N., Arisue, A., Nishizuka, S., Aikou, S., Shida, D. and Furukawa, Y.
Identification of odontogenic ameloblast associated (ODAM) as a novel target gene of the Wnt/β-catenin signaling pathway. Cancer Sci.
114(3):948-960, 2023.