Updated on 2021/10/19

写真a

 
TSUBOTA Shoma
 
Organization
Graduate School of Medicine Program in Integrated Medicine Biological Chemistry Assistant Professor
Graduate School
Graduate School of Medicine
Undergraduate School
School of Medicine Department of Medicine
Title
Assistant Professor

Degree 2

  1. Doctor of Philosophy (Medical Science) ( 2017.10   Nagoya University ) 

  2. Master of Medical Science ( 2013.3   Nagoya University ) 

Research Interests 3

  1. Neuroblastoma

  2. Cancer biology

  3. Molecular Biology

Research Areas 1

  1. Life Science / Tumor biology

Research History 1

  1. Nagoya University   Graduate School of Medicine Program in Integrated Medicine Biological Chemistry   Assistant Professor

    2019.4

 

Papers 11

  1. Combination of tumor necrosis factor-α and epidermal growth factor induces the adrenergic-to-mesenchymal transdifferentiation in SH-SY5Y neuroblastoma cells. Reviewed

    Huang Y, Tsubota S, Nishio N, Takahashi Y, Kadomatsu K

    Cancer science   Vol. 112 ( 2 ) page: 715 - 724   2021.2

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

    Neuroblastoma, a type of cancer that is common in children, is composed of two genetically clonal but epigenetically distinct cell types: mesenchymal (MES) and adrenergic (ADRN) types, controlled by super-enhancer-associated lineage-specific transcription factor networks. Mesenchymal-type cells are more migratory, resistant to chemotherapy, and prevalent in relapse tumors. Importantly, both cell types spontaneously transdifferentiate into one another, and this interconversion can be induced by genetic manipulations. However, the mechanisms of their spontaneous transdifferentiation and extracellular factors inducing this phenomenon have not yet been elucidated. Using a unique approach involving gene set enrichment analysis, we selected six ADRN and 10 MES candidate factors, possibly inducing ADRN and MES phenotypes, respectively. Treatment with a combination of 10 MES factors clearly induced the MES gene expression profile in ADRN-type SH-SY5Y neuroblastoma cells. Considering the effects on gene expression profile, migration ability, and chemoresistance, a combination of tumor necrosis factor alpha (TNF-α) and epidermal growth factor (EGF) was sufficient to synergistically induce the ADRN-to-MES transdifferentiation in SH-SY5Y cells. In addition, human neuroblastoma cohort analysis revealed that the expression of TNF and EGF receptors was strongly associated with MES gene expression signatures, supporting their important roles in transdifferentiation in vivo. Collectively, we propose a mechanism of neuroblastoma transdifferentiation induced by extracellular growth factors, which can be controlled in clinical situations, providing a new therapeutic possibility.

    DOI: 10.1111/cas.14760

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  2. Origin and initiation mechanisms of neuroblastoma Reviewed International journal

    Shoma Tsubota, Kenji Kadomatsu

    Cell and Tissue Research   Vol. 372 ( 2 ) page: 211 - 221   2018.5

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    Language:Japanese   Publishing type:Research paper (scientific journal)   Publisher:Springer Verlag  

    Neuroblastoma is an embryonal malignancy that affects normal development of the adrenal medulla and paravertebral sympathetic ganglia in early childhood. Extensive studies have revealed the molecular characteristics of human neuroblastomas, including abnormalities at genome, epigenome and transcriptome levels. However, neuroblastoma initiation mechanisms and even its origin are long-standing mysteries. In this review article, we summarize the current knowledge about normal development of putative neuroblastoma sources, namely sympathoadrenal lineage of neural crest cells and Schwann cell precursors that were recently identified as the source of adrenal chromaffin cells. A plausible origin of enigmatic stage 4S neuroblastoma is also discussed. With regard to the initiation mechanisms, we review genetic abnormalities in neuroblastomas and their possible association to initiation mechanisms. We also summarize evidences of neuroblastoma initiation observed in genetically engineered animal models, in which epigenetic alterations were involved, including transcriptomic upregulation by N-Myc and downregulation by polycomb repressive complex 2. Finally, several in vitro experimental methods are proposed that hopefully will accelerate our comprehension of neuroblastoma initiation. Thus, this review summarizes the state-of-the-art knowledge about the mechanisms of neuroblastoma initiation, which is critical for developing new strategies to cure children with neuroblastoma.

    DOI: 10.1007/s00441-018-2796-z

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  3. PRC2-Mediated Transcriptomic Alterations at the Embryonic Stage Govern Tumorigenesis and Clinical Outcome in MYCN-Driven Neuroblastoma Reviewed International journal

    Shoma Tsubota, Satoshi Kishida, Teppei Shimamura, Miki Ohira, Satoshi Yamashita, Dongliang Cao, Shinichi Kiyonari, Toshikazu Ushijima, Kenji Kadomatsu

    CANCER RESEARCH   Vol. 77 ( 19 ) page: 5259 - 5271   2017.10

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    Authorship:Lead author   Language:Japanese   Publishing type:Research paper (scientific journal)   Publisher:AMER ASSOC CANCER RESEARCH  

    Pediatric cancers such as neuroblastoma are thought to involve a dysregulation of embryonic development. However, it has been difficult to identify the critical events that trigger tumorigenesis and differentiate them from normal development. In this study, we report the establishment of a spheroid culture method that enriches early-stage tumor cells from TH-MYCN mice, a preclinical model of neuroblastoma. Using this method, we found that tumorigenic cells were evident as early as day E13.5 during embryo development, when the MYC and PRC2 transcriptomes were significantly altered. Ezh2, an essential component of PRC2, was expressed in embryonic and postnatal tumor lesions and physically associated with N-MYC and we observed that H3K27me3 was increased at PRC2 target genes. PRC2 inhibition suppressed in vitro sphere formation, derepressed its target genes, and suppressed in situ tumor growth. In clinical specimens, expression of MYC and PRC2 target genes correlated strongly and predicted survival outcomes. Together, our findings highlighted PRC2-mediated transcriptional control during embryogenesis as a critical step in the development and clinical outcome of neuroblastoma. (C) 2017 AACR.

    DOI: 10.1158/0008-5472.CAN-16-3144

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  4. Screening of novel Midkine binding protein by BioID2-based proximity labeling. Reviewed

    Komata Y, Tsubota S, Sakamoto K, Ikematsu S, Kadomatsu K

    Nagoya journal of medical science   Vol. 83 ( 3 ) page: 495 - 508   2021.8

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    Language:English   Publisher:Nagoya Journal of Medical Science  

    Midkine (MK), a heparin-binding growth factor, is associated with the poor prognosis of the pediatric tumor, neuroblastoma. MK would be a druggable target as many studies showed inhibition of its function in various cancers suppressed tumor developments. To establish the therapy targeting MK, identification of its binding partners, and elucidation of its intracellular signaling are needed. It was reported that exogenous MK induced phosphorylation of ribosomal protein S6 (RPS6) downstream of mTOR signaling. Using RPS6 phosphorylation as a marker of MK response, we searched for MK reactive cell lines. We found that MK cell lines expressing less MK tended to respond better to MK. Next, using an MK reactive neuroblastoma cell line, MK-knocked down SH-SY5Y cells, we employed a proximity-dependent biotin identification method, which was invented to evaluate protein-protein interactions by biotinylation. We confirmed that secreted MK fused to the biotin ligase BioID2 (MK-BioID2) was able to biotinylate proteins from the cells. Biotinylated proteins were identified by liquid chromatography-mass spectrometry analyses. Twenty five proteins were found to be overlapped after three independent experiments, among which insulin-like growth binding protein 2 (IGFBP2) was further analyzed. IGFBP2 was indeed detected with immunoblotting after streptavidin pull down of MK-BioID2 labeled cell extract of MK-knocked down SH-SY5Y cells. Our study suggests that the BioID2 method is useful to identify binding partners of growth factors.

    DOI: 10.18999/nagjms.83.3.495

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  5. Thymidylate synthase inhibitor raltitrexed can induce high levels of DNA damage in MYCN-amplified neuroblastoma cells. Reviewed International journal

    Ken Yamashita, Shinichi Kiyonari, Shoma Tsubota, Satoshi Kishida, Ryuichi Sakai, Kenji Kadomatsu

    Cancer science   Vol. 111 ( 7 ) page: 2431 - 2439   2020.7

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

    MYCN gene amplification is consistently associated with poor prognosis in patients with neuroblastoma, a pediatric tumor arising from the sympathetic nervous system. Conventional anticancer drugs, such as alkylating agents and platinum compounds, have been used for the treatment of high-risk patients with MYCN-amplified neuroblastoma, whereas molecule-targeting drugs have not yet been approved. Therefore, the development of a safe and effective therapeutic approach is highly desired. Although thymidylate synthase inhibitors are widely used for colorectal and gastric cancers, their usefulness in neuroblastoma has not been well studied. Here, we investigated the efficacies of approved antifolates, methotrexate, pemetrexed, and raltitrexed (RTX), on MYCN-amplified and nonamplified neuroblastoma cell lines. Cell growth-inhibitory assay revealed that RTX showed a superior inhibitory activity against MYCN-amplified cell lines. We found no significant differences in the protein expression levels of the antifolate transporter or thymidylate synthase, a primary target of RTX, among the cell lines. Because thymidine supplementation could rescue the RTX-induced cell growth suppression, the effect of RTX was mainly due to the reduction in dTTP synthesis. Interestingly, RTX treatments induced single-stranded DNA damage response in MYCN-amplified cells to a greater extent than in the nonamplified cells. We propose that the high DNA replication stress and elevated levels of DNA damage, which are a result of deregulated expression of MYCN target genes, could be the cause of increased sensitivity to RTX.

    DOI: 10.1111/cas.14485

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  6. 新規tagmentation-assisted PCR法を用いたCD19キメラ抗原受容体遺伝子導入T細胞の遺伝子挿入部位解析(Integration mapping of CD19 chimeric antigen receptor T cells by novel tagmentation-assisted PCR) Reviewed International journal

    濱田 太立, 奥野 友介, 西尾 信博, 鈴木 哲, 川島 希, 村松 秀城, 坪田 庄真, Wilson Matthew H., 盛田 大介, 片岡 伸介, 市川 大輔, 村上 典寛, 谷口 理恵子, 鈴木 喬悟, 小島 大英, 関屋 由子, 西川 英里, 成田 敦, 濱 麻人, 小島 勢二, 中沢 洋三, 高橋 義行

    臨床血液   Vol. 59 ( 9 ) page: 1586 - 1586   2018.9

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    Language:English   Publishing type:Research paper (scientific journal)   Publisher:(一社)日本血液学会-東京事務局  

  7. Integration Mapping of piggyBac-Mediated CD19 Chimeric Antigen Receptor T Cells Analyzed by Novel Tagmentation-Assisted PCR. Reviewed International journal

    Motoharu Hamada, Nobuhiro Nishio, Yusuke Okuno, Satoshi Suzuki, Nozomu Kawashima, Hideki Muramatsu, Shoma Tsubota, Matthew H Wilson, Daisuke Morita, Shinsuke Kataoka, Daisuke Ichikawa, Norihiro Murakami, Rieko Taniguchi, Kyogo Suzuki, Daiei Kojima, Yuko Sekiya, Eri Nishikawa, Atsushi Narita, Asahito Hama, Seiji Kojima, Yozo Nakazawa, Yoshiyuki Takahashi

    EBioMedicine   Vol. 34   page: 18 - 26   2018.8

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

    Insertional mutagenesis is an important risk with all genetically modified cell therapies, including chimeric antigen receptor (CAR)-T cell therapy used for hematological malignancies. Here we describe a new tagmentation-assisted PCR (tag-PCR) system that can determine the integration sites of transgenes without using restriction enzyme digestion (which can potentially bias the detection) and allows library preparation in fewer steps than with other methods. Using this system, we compared the integration sites of CD19-specific CAR genes in final T cell products generated by retrovirus-based and lentivirus-based gene transfer and by the piggyBac transposon system. The piggyBac system demonstrated lower preference than the retroviral system for integration near transcriptional start sites and CpG islands and higher preference than the lentiviral system for integration into genomic safe harbors. Integration into or near proto-oncogenes was similar in all three systems. Tag-PCR mapping is a useful technique for assessing the risk of insertional mutagenesis.

    DOI: 10.1016/j.ebiom.2018.07.008

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  8. Development of the sympathoadrenal lineage from neural crest and neuroblastoma Reviewed International journal

    Tsubota Shoma, Kadomatsu Kenji

    CANCER SCIENCE   Vol. 109   page: 22 - 22   2018.1

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    Publishing type:Research paper (scientific journal)  

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  9. Neurocan, an extracellular chondroitin sulfate proteoglycan, stimulates neuroblastoma cells to promote malignant phenotypes. Reviewed International journal

    Zhendong Su, Satoshi Kishida, Shoma Tsubota, Kazuma Sakamoto, Dongliang Cao, Shinichi Kiyonari, Miki Ohira, Takehiko Kamijo, Atsushi Narita, Yinyan Xu, Yoshiyuki Takahashi, Kenji Kadomatsu

    Oncotarget   Vol. 8 ( 63 ) page: 106296 - 106310   2017.12

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

    Neurocan (NCAN), a secreted chondroitin sulfate proteoglycan, is one of the major inhibitory molecules for axon regeneration in nervous injury. However, its role in cancer is not clear. Here we observed that high NCAN expression was closely associated with the unfavorable outcome of neuroblastoma (NB). NCAN was also highly and ubiquitously expressed in the early lesions and terminal tumor of TH-MYCN mice, a NB model. Interestingly, exogenous NCAN (i.e., overexpression, recombinant protein and conditioned medium) transformed adherent NB cells into spheres whose malignancies in vitro (anchorage-independent growth and chemoresistance) and in vivo (xenograft tumor growth) were potentiated. Both chondroitin sulfate sugar chains and NCAN's core protein were essential for the sphere formation. The CSG3 domain was essential in the moiety of NCAN. Our comprehensive microarray analysis and RT-qPCR of mRNA expression suggested that NCAN treatment promoted cell division, and urged cells to undifferentiated state. The knockdown of NCAN in tumor sphere cells cultured from TH-MYCN mice resulted in growth suppression in vitro and in vivo. Our findings suggest that NCAN, which stimulates NB cells to promote malignant phenotypes, is an extracellular molecule providing a growth advantage to cancer cells.

    DOI: 10.18632/oncotarget.22435

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  10. Neuroblastoma stem cells and CFC1 Reviewed International journal

    Shoma Tsubota, Kenji Kadomatsu

    ONCOTARGET   Vol. 8 ( 28 ) page: 45032 - 45033   2017.7

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

    DOI: 10.18632/oncotarget.18464

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  11. Origin and mechanism of neuroblastoma Reviewed International journal

    Shoma Tsubota, Kenji Kadomatsu

    Oncoscience   Vol. 4 ( 7-8 ) page: 70 - 72   2017.7

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    Language:English   Publishing type:Research paper (scientific journal)   Publisher:Impact Journals LLC  

    Neuroblastoma is the most common extracranial solid tumor in childhood, and develops mainly in the adrenal medulla but also in the sympathetic ganglia. Some cases of neuroblastoma show spontaneous regression. Particularly, stage 4S, where S stands for “special”, exhibits spontaneous regression despite multifocal tumors. Another interesting feature is that, with few exceptions, driver gene mutations are rare in neuroblastoma, while copy number alterations are common, including MYCN amplification, 17q gain, and others, suggesting the involvement of epigenetic regulation [1]. These features suggest that neuroblastoma tumorigenesis is closely related to the development of the sympathoadrenal lineage. However, the underlying mechanisms, even the origin of neuroblastoma, remain largely obscure. To elucidate those mechanisms, three important papers have been recently published [2-4].

    DOI: 10.18632/oncoscience.360

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KAKENHI (Grants-in-Aid for Scientific Research) 2

  1. MYCNとEZH2が引き起こすエピゲノム異常による神経芽腫発生機構の解明

    Grant number:20K16355  2020.4 - 2022.3

    若手研究

    坪田 庄真

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    Authorship:Principal investigator 

    Grant amount:\4290000 ( Direct Cost: \3300000 、 Indirect Cost:\990000 )

    本研究では、神経芽腫のがん遺伝子として知られるMYCNによる発がんにおいて、MYCNとエピゲノム制御複合体PRC2の責任分子であるEZH2が物理的に結合する意義・その機構を解明する。具体的に、MYCNとEZH2の結合様式を明らかにし、EZH2と結合しないMYCN変異体を用いて神経芽腫の発生を検証する。また、それぞれのゲノム上での局在やエピゲノム修飾、遺伝子発現を網羅的に解析する。

  2. Mechanism of neuroblastoma development by MYCN-PRC2 mediated epigenomic aberrations

    Grant number:18K15208  2018.4 - 2020.3

    Grant-in-Aid for Early-Career Scientists

    Tsubota Shoma

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    Authorship:Principal investigator 

    Grant amount:\4160000 ( Direct Cost: \3200000 、 Indirect Cost:\960000 )

    Neuroblastoma is a childhood cancer, and is caused by the well-known oncogene, MYCN. The applicant previously reported that EZH2, an epigenomic regulator, is important for MYCN-induced neuroblastoma development, that MYCN binds to EZH2, and that inhibition EZH2 function can prevent neuroblastoma growth. In this study, we investigated whether the binding of MYCN to EZH2 is essential for the development of neuroblastoma. Unfortunately, we could not reproduce the previous report that a part of MYCN is required for the binding of EZH2 in the first place, so we did not achieve our goal within the study period.