抗菌ペプチドの抗腫瘍活性分子メカニズムおよび化学療法の効果増強作用の解明

• Project/Area Number: 14J05415
• Research Category: Grant-in-Aid for JSPS Fellows
• Allocation Type: Single-year Grants
• Section: 国内
• Research Field: Tumor therapeutics
• Research Institution: Tohoku University
• Principal Investigator: 黒田 健吾 東北大学, 農学研究科, 特別研究員(DC1)
• Project Period (FY): 2014-04-25 – 2017-03-31
• Project Status: Completed (Fiscal Year 2016)
• Budget Amount: ¥3,200,000 (Direct Cost: ¥3,200,000); Fiscal Year 2016: ¥1,000,000 (Direct Cost: ¥1,000,000); Fiscal Year 2015: ¥1,000,000 (Direct Cost: ¥1,000,000); Fiscal Year 2014: ¥1,200,000 (Direct Cost: ¥1,200,000)
• Keywords: 抗菌ペプチド / 大腸癌 / miRNA / microRNA / カセリシジン / 大腸癌細胞

Outline of Annual Research Achievements

前年度までに、大腸癌細胞であるHCT116は抗菌ペプチドで処理した際にmicroRNA (miRNA) であるmiR-663aが濃度依存的に増加する事を明らかにした。さらに、miR-663aの大腸癌細胞内での役割を明らかにするためにレンチウイルスベクターによるmiR-663a過剰発現株を作成した。本年度はmiR-663aがHCT116細胞に及ぼす影響を明らかにするための実験を行った。miR-663aを過剰に発現したHCT116細胞はベクター未導入の細胞およびコントロールベクターを導入した細胞に比べ、細胞老化の際に見られる細胞質の膨化が引き起こされていた。さらにmiR-663a強制発現によってHCT116の増殖は抑制され、その効果は免疫不全のヌードマウスによる異種移植モデルにおいても観察できた。
増殖抑制効果の詳細なメカニズムを明らかにする為に細胞周期を確認したところ、miR-663aの強制発現によってG2/M期における停止している細胞が多く見られた。細胞周期の調節は、非常に多くの因子によって複雑に制御されているが、G2/M期の調節に関与するp21の増加と非活性型のcdc2の割合の増加が観察された。更に、miR-663aの標的として考えられているケモカインレセプターCXCR4の発現を確認したところ、発現量が減少し、下流の活性型Aktの割合も減少していることが分かった。
これら一連の挙動の多くはmiR-663aを強制発現していないHCT116を抗菌ペプチドで処理していた際にも共通していたことから、本研究において抗菌ペプチドの大腸癌細胞に対する増殖抑制効果のメカニズムの一端にmiR-663aが関与する事を明らかに出来た。

Research Progress Status

28年度が最終年度であるため、記入しない。

Strategy for Future Research Activity

28年度が最終年度であるため、記入しない。

Research Products

• [Journal Article] miR-663a regulates growth of colon cancer cells, after administration of antimicrobial peptides, by targeting CXCR4-p21 pathway (2017)
  • Author(s): Kuroda K, Fukuda T, Krstic-Demonacos M, Demonacos C, Okumura K, Isogai H, Hayashi M, Saito K, Isogai E
  • Journal Title: BMC Cancer Volume: Jan 7;17(1):33 Issue: 1 Pages: 33-33
  • DOI: 10.1186/s12885-016-3003-9
  • Peer Reviewed / Open Access / Int'l Joint Research / Acknowledgement Compliant
• [Journal Article] Proliferation and Cell Cycle of Swine Fibroblast after Infection with Salmonella enterica (2016)
  • Author(s): Masuda M, Guo Y, Kuroda K, Xu J, Yoneyama H, Fukuda T, Hango’mbe BM, Okuno K, Isogai E
  • Journal Title: Advances in Microbiology Volume: 6 Issue: 13 Pages: 942-952
  • DOI: 10.4236/aim.2016.613088
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Oxidative Stress and Breast Cancer Biomarkers: The case of the cytochrome P450 2E1 (2016)
  • Author(s): Singh S, Rajendran R, Kuroda K, Isogai E, Krstic-Demonacos M, Demonacos C
  • Journal Title: Journal of Cancer Metastasis and Treatment Volume: 2 Issue: 7 Pages: 268-276
  • DOI: 10.20517/2394-4722.2016.42
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Establishment of an immortalized cell line derived from the prairie vole via lentivirus-mediated transduction of mutant cyclin-dependent kinase 4, cyclin D, and telomerase reverse transcriptase (2016)
  • Author(s): Katayama M, Kiyono T, Horie K, Hirayama T, Eitsuka T, Kuroda K, Donai K, Hidema S, Nishimori K and Fukuda T
  • Journal Title: Experimental Animals Volume: 65 Issue: 1 Pages: 87-96
  • DOI: 10.1538/expanim.15-0061
  • NAID: 130005125854
  • ISSN: 0007-5124, 1341-1357, 1881-7122
  • Peer Reviewed / Open Access
• [Journal Article] Functional structure and antimicrobial activity of persulcatusin, an antimicrobial peptide from the hard tick Ixodes persulcatus. (2016)
  • Author(s): Miyoshi N, Saito T, Ohmura T, Kuroda K, Suita K, Ihara K, Isogai E.
  • Journal Title: Parasit Vectors. Volume: - Issue: 1
  • DOI: 10.1186/s13071-016-1360-5
  • Peer Reviewed / Open Access
• [Journal Article] The human cathelicidin antimicrobial peptide LL-37 and mimics are potential anticancer drugs. (2015)
  • Author(s): Kuroda, K., OKumura, K., Isogai, H., Isogai, E.
  • Journal Title: Front Oncol Volume: 5 Pages: 1-10
  • DOI: 10.3389/fonc.2015.00144
  • Peer Reviewed / Open Access
• [Journal Article] Immortalization of fetal bovine colon epithelial cells by expression of human cyclin D1, mutant cyclin dependent kinase 4, and telomerase reverse transcriptase: An in vitro model for bacterial infection. (2015)
  • Author(s): Kuroda K, Kiyono T, Isogai E, Masuda M, Narita M, Fukuda T
  • Journal Title: PLoS One. Volume: - Issue: 12 Pages: e0143473-e0143473
  • DOI: 10.1371/journal.pone.0143473
  • Peer Reviewed / Open Access
• [Journal Article] Leptospiral lipopolysaccharide stimulates the expression of toll-like receptor 2 and cytokines in pig fibroblasts (2015)
  • Author(s): Guo Y, Fukuda T, Donai K, Kuroda K, Masuda M, Nakamura S, Yoneyama H, Isogai E
  • Journal Title: Animal Science Journal Volume: 86(2) Issue: 2 Pages: 238-244
  • DOI: 10.1111/asj.12254
  • Peer Reviewed / Open Access
• [Journal Article] The effect of antibody against leptospiral lipopolysaccharide (L-LPS) on inhibition the interaction between L-LPS and toll-like receptor 2 in pig fibroblast cell line (2015)
  • Author(s): Guo Y, Fukuda T, Nakamura S, Bai L, Kuroda K, Yoneyama H, Xu J, Tomioka R, Isogai E
  • Journal Title: Asian-Australia J Anim Sci. Volume: 28(2) Issue: 2 Pages: 273-279
  • DOI: 10.5713/ajas.14.0440
  • Peer Reviewed / Open Access
• [Journal Article] Establishment of cell lines derived from the genus Macaca through controlled expression of cell cycle regulators (2015)
  • Author(s): Kuroda K, Kiyono T, Eitsuka T, Isogai H, Takahashi K, Donai K, Isogai E, Fukuda T
  • Journal Title: J Cell Biochem. Volume: 116 Issue: 2 Pages: 205-211
  • DOI: 10.1002/jcb.24963
  • Peer Reviewed / Open Access
• [Journal Article] Antimicrobial peptide FF/CAP18 induces apoptotic cell death in HCT116 colon cancer cells via changes in the metabolic profile (2015)
  • Author(s): Kuroda K, Fukuda T, Isogai H, Kazuhiko Okumura, Krstic-Demonacos M, Isogai E
  • Journal Title: Int J Oncol. Volume: 46(4) Issue: 4 Pages: 1516-1526
  • DOI: 10.3892/ijo.2015.2887
  • Peer Reviewed / Open Access
• [Presentation] 腸内フローラ構成細菌を中心とした細菌による生体内除染 (2017)
  • Author(s): 齋藤和輝、木野康志、黒田健吾、西村順子、関根 勉、福田智一、福本 学、山城秀昭、小林 仁、篠田 壽、西田典永、河野麻実子、磯貝恵美子
  • Organizer: 第122回日本畜産学会総会
  • Place of Presentation: 神戸大学（兵庫県神戸市）
  • Year and Date: 2017-03-19
• [Presentation] 抗菌ペプチドによって誘導される大腸癌細胞由来エクソソームの解析 (2017)
  • Author(s): 林美和 黒田健吾 伊原航平 磯貝恵美子
  • Organizer: 日本農芸化学会2017年度京都大会
  • Place of Presentation: 京都女子大学（京都府京都市）
  • Year and Date: 2017-03-17
• [Presentation] Uptake of radioactive cesium by intestinal and probiotics bacteria (2016)
  • Author(s): Saito K, Kuroda K, Sekine T, Fukumoto M, Kino Y, Fukuda T, Yamashiro H, Shinoda H, Isogai H, Kobayashi J, Nishimura J, Abe Y, Nishida T, Kouno M, Isogai E
  • Organizer: The 14th International Symposium of Integrated Field Science
  • Place of Presentation: 東北大学（宮城県仙台市）
  • Year and Date: 2016-11-26
  • Int'l Joint Research
• [Presentation] The analogue human cathelicidin antimicrobial peptide is a potential anticancer drug (2016)
  • Author(s): Kuroda K, Fukuda T, Isogai H, Okumura K, Demonacos KM, Isogai E
  • Organizer: 21th World Congress on Advances in Oncology and 18th International Symposium on Molecular Medicine Athene
  • Place of Presentation: アテネ（ギリシャ）
  • Year and Date: 2016-10-06
  • Int'l Joint Research
• [Presentation] Anti-cancer and anti-microbial activities of ellagitannin oligomers from Punica granatum (2016)
  • Author(s): Ide A, Kuroda K, Hayashi M, Saito K, Iwaya T, Nishida N, Kohno M, Ito H, Isogai E
  • Organizer: 21th World Congress on Advances in Oncology and 18th International Symposium on Molecular Medicine Athene
  • Place of Presentation: アテネ（ギリシャ）
  • Year and Date: 2016-10-06
  • Int'l Joint Research
• [Presentation] Effective activity of synthetic antimicrobial peptides to antibiotic resistant Staphylococcus aureus (2016)
  • Author(s): Isogai E, Miyoshi N, Morozumi I, Kuroda K, Ihara K, Narita M, Isogai H, Hiramatsu K, Sasaki T
  • Organizer: 21th World Congress on Advances in Oncology and 18th International Symposium on Molecular Medicine Athene
  • Place of Presentation: アテネ（ギリシャ）
  • Year and Date: 2016-10-06
  • Int'l Joint Research
• [Presentation] プロバイオティクス細菌による新たな生体内除染方法の探索 (2016)
  • Author(s): 齋藤和輝、黒田健吾、関根 勉、福田智一、木野康志、福本 学、山城秀昭、小林 仁、篠田 壽、西村順子、西田典永、河野麻実子、磯貝恵美子
  • Organizer: 第70回日本細菌学会東北支部総会
  • Place of Presentation: 北里大学（青森県十和田市）
  • Year and Date: 2016-08-18
• [Presentation] Immortalization of Fetal Bovine Colon Epithelial Cells by Expression of Human Cyclin D1, Mutant CDK4, and TERT (2016)
  • Author(s): 黒田 健吾、清野 透、磯貝 恵美子、増田 みずき、成田 萌、奥野 克哉、小柳 友佳子、福田 智一
  • Organizer: 日本農芸化学会2016大会
  • Place of Presentation: 札幌コンベンションセンター(札幌市白石区)
  • Year and Date: 2016-03-27
• [Presentation] MicroRNA array analysis in colon cancer cells treated with antimicrobial peptides (2015)
  • Author(s): Kuroda K, Fukuda T, Isogai H, Okumura K, Isogai E
  • Organizer: 20th World Congress on Advances in Oncology and 18th International Symposium on Molecular Medicine
  • Place of Presentation: Metropolitan Hotel, Athens, Greece
  • Year and Date: 2015-10-08
  • Int'l Joint Research
• [Presentation] Uptake of radioactive cesium by the major member of intestinal flora, genus Bifidobacterium, Lactobacillus, Clostridium, and Bacteroides. (2015)
  • Author(s): Kuroda K, Suzuki R, Sekine T, Fukumoto M, Fukuda T, Isogai E.
  • Organizer: 15th International Congress of Radiation Research (ICRR2015)
  • Place of Presentation: 京都国際会館 (京都市左京区)
  • Year and Date: 2015-05-25
  • Int'l Joint Research
• [Presentation] Cell proliferation and inhibition of apoptosis by Salmonella infection on swine fibroblast (2014)
  • Author(s): Masuda M, Guo Y, Kuroda K, Fukuda T, Yoneyama Y, Isogai E
  • Organizer: 12th Epidemiology and Evolutionary Genetics of Infectious Diseases
  • Place of Presentation: Bangkok, Thailand
  • Year and Date: 2014-12-11 – 2014-12-13
• [Presentation] Antimicrobial activity of synthetic antimicrobial peptide from taiga tick, Ixodes persulcatus (persulcatusin) against Staphylococcus aureus (2014)
  • Author(s): Miyoshi N, Kuroda K, Mizuki M, Suita K, Yoneyama H, Ando T, Isogai E
  • Organizer: 12th Epidemiology and Evolutionary Genetics of Infectious Diseases
  • Place of Presentation: Bangkok, Thailand
  • Year and Date: 2014-12-11 – 2014-12-13
• [Presentation] Metabolites variation in colon cancer cell line HCT116 treated with antimicrobial peptide FF/CAP18 (2014)
  • Author(s): Kuroda K, Fukuda T, Isogai H, Okumura K, Isogai E
  • Organizer: 19th World Congress on Advances in Oncology and 17th International Symposium on Molecular Medicine
  • Place of Presentation: Athens, Greece
  • Year and Date: 2014-10-09 – 2014-10-11
• [Presentation] マダニ由来抗菌ペプチドpersulcatusinの構造と活性 (2014)
  • Author(s): 三好就英、斉藤 丈、大村忠弘、黒田健吾、増田みづ紀、吹田一将、米山 裕、安藤太助、磯貝 浩、磯貝恵美子
  • Organizer: 日本細菌学会東北支部会
  • Place of Presentation: 宮城県仙台市
  • Year and Date: 2014-08-20 – 2014-08-21
• [Presentation] Elucidation of metabolic pathways for the colon cancer cell line HCT116 treated with antimicrobial peptide FF/CAP18 (2014)
  • Author(s): Kuroda K, Fukuda T, Isogai H, Okumura K, Isogai E
  • Organizer: FOURTH INTERNATIONAL SYMPOSIUM ON ANTIMICROBIAL PEPTIDES
  • Place of Presentation: Lorient, France
  • Year and Date: 2014-06-04 – 2014-06-06