内皮細胞特異的スプライシングバリアントからアプローチする癌微小環境の分子病態解析

2015 Fiscal Year Research-status Report

• Project/Area Number: 15K08973
• Research Institution: Sapporo Medical University
• Principal Investigator: 山本 英一郎 札幌医科大学, 医学部, 助教 (60567915)
• Project Period (FY): 2015-04-01 – 2018-03-31
• Keywords: 腫瘍血管 / 遺伝子発現 / 大腸がん

Outline of Annual Research Achievements

大腸がんの増殖・浸潤・転移に関わる血管新生メカニズムの解明を目的としている。大腸がんおよび対照非がん部臨床検体より内皮細胞特異的抗原を用いて内皮細胞を分離してRNAを抽出後、Illumina Hi-Seqを用いたRNA-シークエンス法によるトランスクリプトーム解析から新規TEC関連遺伝子、スプライスバリアント(SV)を多数同定した。がん間質内皮細胞（Tumor associated Endothelial Cells, TECs）特異的に発現亢進する遺伝子の一つである転写因子Aは定量PCR解析および免疫組織学的染色でTECに特異的に発現することを明らかにし、さらにヒト正常血管内皮細胞(HUVEC)を大腸がん細胞培養上清で刺激することで発現誘導されることが確認できた。またヒト血管内皮細胞(HUVEC)を用いた予備実験の結果、転写因子AをノックダウンすることでHUVECの増殖およびチューブ形成能を阻害すること、また細胞周期関連遺伝子の発現が顕著に変動することが明らかとなった。TECsに特異的なスプライシングバリアント候補遺伝子を絞り込み、現在定量PCR解析で多数の臨床検体で検証している。

Current Status of Research Progress

2: Research has progressed on the whole more than it was originally planned.

Reason

本年度の目標としていた、臨床検体からの内皮細胞の分離、新規TEC関連遺伝子、スプライスバリアント(SV)の絞り込みができた。

Strategy for Future Research Activity

新規TEC関連遺伝子、スプライスバリアント(SV)の機能解析を予定通り進める。in vitro実験モデルを作成し候補遺伝子の過剰発現、ノックダウンによる血管新生・細胞増殖への影響を検討する。次にin vivoにおける機能解析を計画する。

Causes of Carryover

順調に研究計画が進んだ

Expenditure Plan for Carryover Budget

次年度の機能解析により使用することで研究成果が得られることが期待できる

Research Products

• [Journal Article] Clinicopathological and molecular alterations in early gastric cancers with the microsatellite instability-high phenotype (2016)
  • Author(s): Sugimoto R, Sugai T, Habano W, Endoh M, Eizuka M, Yamamoto E, Uesugi N, Ishida K, Kawasaki T, Matsumoto T, Suzuki H.
  • Journal Title: Int J Cancer Volume: 1 Pages: 1689-97
  • DOI: 10.1002/ijc.29916
  • Peer Reviewed
• [Journal Article] Epigenetic silencing of NTSR1 is associated with lateral and noninvasive growth of colorectal tumors (2015)
  • Author(s): Kamimae S, Yamamoto E, Kai M, Niinuma T, Yamano HO, Nojima M, Yoshikawa K, Kimura T, Takagi R, Harada E, Harada T, Maruyama R, Sasaki Y, Tokino T, Shinomura Y, Sugai T, Imai K, Suzuki H
  • Journal Title: Oncotarget Volume: 6 Pages: 29975-90
  • DOI: 10.18632/oncotarget.5034
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Low-Frequency IL23R Coding Variant Associated with Crohn's Disease Susceptibility in Japanese Subjects Identified by Personal Genomics Analysis (2015)
  • Author(s): Onodera K, Arimura Y, Isshiki H, Kawakami K, Nagaishi K, Yamashita K, Yamamoto E, Niinuma T, Naishiro Y, Suzuki H, Imai K, Shinomura Y
  • Journal Title: PLoS One Volume: 10 Pages: e0137801
  • DOI: 10.1371/journal.pone.0137801
  • Peer Reviewed / Open Access
• [Journal Article] A Screen for Epigenetically Silenced microRNA Genes in Gastrointestinal Stromal Tumors. (2015)
  • Author(s): Isosaka M, Niinuma T, Nojima M, Kai M, Yamamoto E, Maruyama R, Nobuoka T, Nishida T, Kanda T, Taguchi T, Hasegawa T, Tokino T, Hirata K, Suzuki H, Shinomura Y
  • Journal Title: PLoS One Volume: 10 Pages: e0133754
  • DOI: 10.1371/journal.pone.0133754
  • Peer Reviewed / Open Access
• [Journal Article] Aberrant TET1 Methylation Closely Associated with CpG Island Methylator Phenotype in Colorectal Cancer. (2015)
  • Author(s): 5.Ichimura N, Shinjo K, An B, Shimizu Y, Yamao K, Ohka F, Katsushima K, Hatanaka A, Tojo M, Yamamoto E, Suzuki H, Ueda M, Kondo Y
  • Journal Title: Cancer Prev Res (Phila) Volume: 10 Pages: 702-11
  • DOI: 10.1158/1940-6207.CAPR-14-0306.
  • Peer Reviewed
• [Journal Article] Comparative analysis of methods to determine DNA methylation levels of a tumor-related microRNA gene. (2015)
  • Author(s): Konishi Y, Hayashi H, Suzuki H, Yamamoto E, Sugisaki H, Higashimoto H.
  • Journal Title: Anal Biochem Volume: 484 Pages: 66-71
  • DOI: 10.1016/j.ab.2015.05.003.
  • Peer Reviewed
• [Presentation] Identification of tumor endothelium-related genes in colorectal cancer (2016)
  • Author(s): Akira Yorozu, Yamamoto Eiichiro , Reo Maruyama, Masahiro Kai, Takeshi Niinuma, Tomohisa Furuhata, Tomohiko Nishidate, Tamotsu Sugai, Hiromu Suzuki.
  • Organizer: Tenth AACR-JCA Joint Conference on Breakthroughs in Cancer Research: From Biology to Therapeutics
  • Place of Presentation: Maui, Hawaii, USA
  • Year and Date: 2016-02-16 – 2016-02-20
  • Int'l Joint Research
• [Presentation] Identification of aberrant DNA methylation associated with the development of traditional serrated adenoma. (2016)
  • Author(s): Hironori Aoki, Eiichiro Yamamoto, Hiro-o Yamano, Takeshi Niinuma, Masahiro Kai, Reo Maruyama, Yasushi Adachi, Takao Endo, Tamotsu Sugai, Hiromu Suzuki.
  • Organizer: Tenth AACR-JCA Joint Conference on Breakthroughs in Cancer Research: From Biology to Therapeutics
  • Place of Presentation: Maui, Hawaii, USA
  • Year and Date: 2016-02-16 – 2016-02-20
  • Int'l Joint Research
• [Presentation] Possible role of TET1 dysregulation in inducing aberrant DNA methylation in colorectal cancer. (2016)
  • Author(s): Hiromu Suzuki, Masahiro Kai, Takeshi Niinuma, Eiichiro Yamamoto, Reo Maruyama, Akira Yorozu, Kohei Kumegawa, Hironori Aoki, Tetsuya Shindo, Takashi Tokino, Kohzoh Imai.
  • Organizer: Tenth AACR-JCA Joint Conference on Breakthroughs in Cancer Research: From Biology to Therapeutics
  • Place of Presentation: Maui, Hawaii, USA
  • Year and Date: 2016-02-16 – 2016-02-20
  • Int'l Joint Research
• [Presentation] Histone H3K27 trimethylation in normal colon is associated with development of colorectal tumor with CpG island methylator phenotype (2015)
  • Author(s): Eiichiro Yamamoto, Hiroo Yamano, Tamotsu Sugai, Hiromu Suzuki
  • Organizer: United European Gastroenterology Week 2015
  • Place of Presentation: Barcelona
  • Year and Date: 2015-10-24 – 2015-10-29
  • Int'l Joint Research
• [Presentation] ENDOSCOPIC AND MOLECULAR FEATURES REVEAL PROGRESSION OF SERRATED LESIONS (2015)
  • Author(s): Hironori Aoki, Eiichiro Yamamoto, Hiroo Yamano, Takao Endo, Tamotsu Sugai, Hiromu Suzuki
  • Organizer: United European Gastroenterology Week 2015
  • Place of Presentation: Barcelona
  • Year and Date: 2015-10-24 – 2015-10-29
  • Int'l Joint Research