Mechanistic analysis of lymph node metastasis using gastric and colonic cancer cell lines tagged with GFP

• Project/Area Number: 18590394
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Experimental pathology
• Research Institution: Aichi Cancer Center Research Institute
• Principal Investigator: NAKANISHI Hayao Aichi Cancer Center Research Institute, 腫瘍病理学部, 室長 (20207830)
• Project Period (FY): 2006 – 2008
• Project Status: Completed (Fiscal Year 2008)
• Budget Amount: ¥4,230,000 (Direct Cost: ¥3,600,000、Indirect Cost: ¥630,000); Fiscal Year 2008: ¥910,000 (Direct Cost: ¥700,000、Indirect Cost: ¥210,000); Fiscal Year 2007: ¥1,820,000 (Direct Cost: ¥1,400,000、Indirect Cost: ¥420,000); Fiscal Year 2006: ¥1,500,000 (Direct Cost: ¥1,500,000)
• Keywords: 腫瘍 / 転移 / リンパ節転移 / 胃癌 / 大腸癌 / リンパ管新生 / リンパ管内皮細胞 / VEGF-C / Podoplanin / 大腸がん / リンパ管内皮細 / 胃がん

Research Abstract

胃がん、大腸がんの予後を規定する最も重要な因子であるリンパ節転移の機構を解明するため、臨床病理学的ならびに実験的に腫瘍リンパ管新生について検討し、以下の諸点を明らかにした。1)消化器がんにおける腫瘍リンパ管新生のin vivoにおける実態を明らかにするためにリンパ管内皮特異抗体D2-40を用いて胃癌50例、大腸癌61例のリンパ管新生を免疫組織学的に検討した。その結果、いずれの腫瘍においてもリンパ管新生は血管新生のように腫瘍組織内では殆ど起こらず、主として腫瘍周囲で起こることを明らかにした。VEGF-Cの発現はこれとよく一致して周囲正常上皮に比べ腫瘍細胞でむしろ減少傾向が認められたことから、リンパ管新生はVEGF-Cが既存のリンパ管に作用して誘導される可能性が示唆された。2)次にこれら臨床例の腫瘍リンパ管新生に関する観察結果を実験的に検証するためにGFP遺伝子を導入した胃癌、大腸癌リンパ節転移細胞株とmicrolymphangiographyを組み合わせて腫瘍リンパ管新生をヌードマウス皮下移植系で実験的に解析した。その結果、上記臨床例の観察結果とよく一致して、リンパ管新生は腫瘍組織内ではなくむしろ腫瘍周囲の既存のリンパ管で起こり、腫瘍細胞のリンパ管侵襲も腫瘍辺縁部で起きている可能性が示唆された。3)VEGF-Cのリンパ節転移における役割を明らかにするため、リンパ節非転移性で、VEGF-C低発現ヒト大腸癌細胞株にVEGF-Cを強制発現させた安定発現株ではリンパ節転移の有意な促進は見られなかった。一方、リンパ節転移能を有し、VEGF-Cを高発現するヒト大腸癌細胞株にVEGFC ShRNA plasmidベクターを導入した安定発現株ではリンパ管密度ならびにリンパ節転移能が軽度ながら抑制されたことから、リンパ節転移能の獲得にはVEGF-Cを介したリンパ管新生だけでは十分ではなく、腫瘍のリンパ管侵襲能も関与している可能性が示唆された。4)腫瘍細胞とリンパ管との相互作用を解析するためにHPVE6,E7およびhTERT遺伝子を導入して作成したヒト不死化リンパ管内皮細胞株(HuTLEC)および上記大腸癌リンパ節転移細胞株(COLM-5)を組み合わせて構築したin vitro解析システムを用いて、COLM-5細胞の培地中にVEGF-C刺激に比べ数倍高い転写活性をluciferase assayで検出し、VEGF-C以外の新しい管腔形成促進因子が存在する可能性を示唆した。

Research Products

• [Journal Article] Characterization of a novel lymph node metastasis model from human colonic cancer and its preclinical use for comparison of anti-metastatic efficacy between oral S-1 and UFT/LV. (2010)
  • Author(s): Ito Y, *Nakanishi H, Kodera Y, Hirai T, Nakao A, Kato, T
  • Journal Title: (in press)
  • NAID: 10027830674
• [Journal Article] Targeted delivery of oligomannose-coated liposome to the omental micrometastasis by peritoneal macrophages from patients with gastric cancer (2010)
  • Author(s): Matsui M, Shimizu, Y Ikehara Y, Kondo E, Kodera Y, Nakanishi H.
  • Journal Title: (in press)
  • NAID: 10026594802
• [Journal Article] Bone Marrow and Peripheral Blood Expression of ID1 in Human Gastric Carcinoma Patients Is a Bona Fide Indicator of Lymph Node and Peritoneal Metastasis (2009)
  • Author(s): Iwatsuki, M., Fukagawa, T., Mimori, K., Nakanishi, H., Ito, S., Ishii, H., Yokobori, T., Sasako, M., Baba, H., Mori, M.
  • Journal Title: Br J cancer 100 Pages: 1937-1942
• [Journal Article] Expression of L1 Cell Adhesion Molecule is a Significant Prognostic Factor in pT3-stage Gastric Cancer (2009)
  • Author(s): Kodera, Y., Nakanishi, H., Ito, S., Mochizuki, Y., Nakayama, G., Koike, M., Fujiwara, M., Yamamura, Y., Nakao, A.
  • Journal Title: AntiCancer Res. 29(10) Pages: 4033-9
• [Journal Article] Interleukin-2 potentiation of cetuximab anti-tumor activity for EGFR overexpressing gastric cancer xenografts via antibody-dependent cellular cytotoxicity (2008)
  • Author(s): Hara M, Nakanishi, H., Tsujimura, K., Matsui, M., Yatabe, Y., Manabe, T. Tatematsu, M.
  • Journal Title: Cancer Sci. 99(7) Pages: 1471-8
• [Journal Article] The effect of chemotherapy against micrometastases and isolated tumor cells in lymph nodes : an in vivo study (2008)
  • Author(s): Eguchi, T., Kodera, Y., Nakanishi, H., Yokoyama, H., Ohashi, N., Ito, Y., Nakayama, G., Koike, M., Fujiwara, M., Nakao, A.
  • Journal Title: In Vivo 22(6) Pages: 707-12
• [Journal Article] Matrix Metalloproteinase -9 Associated with Heparan Sulp hate Chains of GPI-Anchored Cell Surface Proteoglycans Mediates Motility of Murine Colon Adenocarcinoma Cells (2008)
  • Author(s): Koyama Y., et al.
  • Journal Title: J Biochem 143(5) Pages: 581-592
  • Peer Reviewed
• [Journal Article] The effect of chemotherapy a gainst micrometastases and isolated tumor cells in lymph nodes : an in vivo study (2008)
  • Author(s): Eguchi T., et al.
  • Journal Title: In Vivo 22(6) Pages: 707-12
  • Peer Reviewed
• [Journal Article] Immunohistochemical analysis of intarcellular dynamics of ClGalTassociated with molecular chaperone, Cosmc (2008)
  • Author(s): Narimatsu Y., et al.
  • Journal Title: Biochem Biophys Res Commun 366 Pages: 199-205
  • Peer Reviewed
• [Journal Article] Non-invasive and real-time monitoring of molecular targeting therapy for lymph node and peritoneal metastasis in nude mice bearing xenografts of human colorectal cancer cells tagged with GFP and DsRed (2007)
  • Author(s): Nakanishi, H., Hara, H., Ikehara, Y., Tatematsu, M.
  • Journal Title: Proceeding of SPIE 6449
• [Journal Article] Isolated tumor cell in lateral lymph node has no influences on the prognosis of rectal cancer patients (2007)
  • Author(s): Hara, M., Hirai, T., Nakanishi, H., Kanemitsu, K., Komori, K., Tatematsu, M., Kato, T.
  • Journal Title: Int. J. Colorectal Dis. 22(8) Pages: 911-7
• [Journal Article] Regional difference in intratumoral lymphangiogenesis of oral squamous cell carcinomas evaluated by immunohistochemistry using D2-40 and podoplanin antibody. An analysis in comparison with angiogenesis (2007)
  • Author(s): Ohno, F., Nakanishi, H., Abe, A., Seki, Y., Kinoshita, A.. Hasegawa, Y., Tatematsu, M., Kurita, K.
  • Journal Title: J. Oral Pathol. Med. 36(5) Pages: 281-9
• [Journal Article] Comparative analysis of intraperitoneal minimal free cancer cells between colorectal and gastric cancer patients using quantitative RT-PCR Possible reason for rare peritoneal recurrence in colorectal cancer (2007)
  • Author(s): Hara, M., Nakanishi, H., Jun, Q., Hirai, T., Kanemitsu, K., Ito, S., Mochizuki, Y., Kodera, Y., Tatematsu, M., Yamamura, Y., Kato, T.
  • Journal Title: Clin. Exp. Metastasis 24(3) Pages: 179-189
• [Journal Article] Non-invasive and real-time monitoring of molecular targeting therapy for lymph node and peritoneal metastasis in nude mice bearing xenografts of human colorectal cancer cells tagged with GFP and DsRed (2007)
  • Author(s): Nakanishi H, et. al.
  • Journal Title: Proceeding of SPIE 6449
  • Peer Reviewed
• [Journal Article] Isolated tumor cell in lateral lymph node has no influences on the prognosis of rectal cancer patients (2007)
  • Author(s): Hara M, et. al.
  • Journal Title: Int. J. Colorectal Dis. 22(8) Pages: 911-917
  • Peer Reviewed
• [Journal Article] Regional difference in intratumoral lymphangiogenesis of oral squamous cell carcinomas evaluated by immunohistochemistry using D2-40 and podoplanin antibody. An analysis in comparison with angiogenesis. (2007)
  • Author(s): Ohno, F, et. al.
  • Journal Title: J. Oral Pathol. Med. 36(5) Pages: 281-289
  • Peer Reviewed
• [Journal Article] Non-invasive and real-time monitoring; of molecular targeting therapy for lymph node and peritoneal metastasis in nude mice bearing xenografts of human colorectal cancer cells tagged with GFP and DsRed (2007)
  • Author(s): Nakanishi H, Hara H, Ikehara Y Y et al.
  • Journal Title: Proceeding of SPIE (in press)
• [Journal Article] Comparative analysis of intraperitoneal minimal free cancer cells between colorectal and gastric cancer patients using quantitative RT-PCR. Possible reason for rare peritoneal recurrence in colorectal cancer (2007)
  • Author(s): Hara M, Nakanishi H (Corresponding author), Jun Q, et al.
  • Journal Title: Clin.Exp.Metastasis (in press)
• [Journal Article] Isolated tumor cell in lateral lymph node has no influences on the prognosis of rectal cancer patients (2007)
  • Author(s): Hara M, Hirai T, Nakanishi H (Third author) et al.
  • Journal Title: Int.J.Colorectal Dis. (in press)
• [Journal Article] Intraoperative quantitative detection of CEA mRNA in the peritoneal lavage of gastric cancer patients with transcription reverse-transcription concerted (TRC) method.A comparative study with real-time quantitative RT-PCR. (2007)
  • Author(s): Ohashi N, Nakanishi E (Corresponding author), Kodera Y, et al.
  • Journal Title: Anticancer Res. (in press)
• [Journal Article] Regional difference in intratumoral lymphangiogenesis of oral squamous cell carcinomas evaluated by immunohistochemistry using D2-40 and podoplanin antibody. An analysis in comparison with angiogenesis (2007)
  • Author(s): Ohno F., Nakanishi H (Corresponding author), Abe A.et al.
  • Journal Title: J.Oral Pathol.Med. (in press)
• [Journal Article] Detection of minimal gastric cancer cells in peritoneal washings by focused microarray analysis eith multiple markers : Clinical implications. (2007)
  • Author(s): Mori K, Suzuki T, Uozaki H, Nakanishi H (Forth author), et al.
  • Journal Title: Ann.Surg.Oncol. (in press)
• [Journal Article] Peritoneal carcinomatosis; A new diagnostic and therapeutic strategy (2007)
  • Author(s): Nakanishi H, Ito S, Mochizuki Y et al.
  • Journal Title: Gastric Cancer Research Horizons (Nova Scientific Publishers, Inc.) (in press)
• [Journal Article] Biological significance of isolated tumor cells and micrometastasis in lymph nodes evaluated using a green fluorescent protein (GFP)-tagged human gastric cancer cell line (2006)
  • Author(s): Yokoyama, H., Nakanishi, H., Kodera, Y., Ikehara, Y., Ohashi, N., Ito, Y., Koike, M., Fujiwara, M., Tatematsu, M., Nakao, A.
  • Journal Title: Clin. Cancer Res. 12(2) Pages: 361-68
• [Journal Article] Molecular basis for sensitivity and acquired resistance to gefitinib in HER2 overexpressing human gastric cancer cell lines derivedfrom liver metastasis (2006)
  • Author(s): Yokoyama, H., Ikehara, Y., Kodera, Y., Ikehara, S., Yatabe, Y., Mochizuki, Y., Koike, M., Fujiwara, M., Nakao, A., Tatematsu, M., Nakanishi, H
  • Journal Title: Br J Cancer 95 Pages: 1504-1513
• [Journal Article] Molecular basis for sensitivity and acquired resistance to gefitinib in HER2-overexpressing human gastric cancer cell lines derived from liver metastasis (2006)
  • Author(s): Yokoyama H., Ikehara Y, Nakankshi H (Last author)
  • Journal Title: Br.J.Cancer 95 Pages: 1504-13
• [Journal Article] Biological significance of isolated tumor cells and micrometastasis in lymph nodes evaluated using a green fluorescent protein (GFP)-tagged human gastric cancer cell line. (2006)
  • Author(s): Yokoyama H., Nakanishi H (Corresponding author), Kodera Y et al.
  • Journal Title: Clin.Cancer Res. 12(2) Pages: 361-68
• [Journal Article] A polymorphism of C-to-T substitution at -31 ILIB is associated with the risk of advanced gastric adenocarcinoma in a Japanese population. (2006)
  • Author(s): Ikehara Y, Niwa T, Nakankshi H (last author)
  • Journal Title: Cancer Res. 66(17) Pages: 8740-8748
• [Journal Article] A carbohydrate recognition-based drug delivery and controlled release system using intraperitoneal macrophages as a cellular vehicle (2006)
  • Author(s): Ikehara SK, Ikehara Y, Nakanishi H, et al.
  • Journal Title: J.Hum.Genet. 51(11) Pages: 927-33
• [Journal Article] Apical Golgi localization of N,N'-diacetyllactosediamine synthase, {beta} 4GalNAc-T3, is responsible for LacdiNAc expression on gastric mucosa. (2006)
  • Author(s): Ikehara Y., Sato T., Nakanishi H (Tenth author)
  • Journal Title: Glycobiology 16(9) Pages: 777-785
• [Journal Article] Prognostic significance of intraperitoneal cancer cells in gastric carcinoma : Analysis of real time RT-PCR after 5 years of follow-up (2006)
  • Author(s): Kodera Y., Nakanishi H.(Second author), Ito S, et al.
  • Journal Title: J.Am.Coll.Surg. 202 Pages: 231-236
• [Presentation] HER family as potential molecular targets for anti-cancer therapy against gastrointestinai malignancies (2007)
  • Author(s): Nakanishi, H.,
  • Organizer: The 12th Aichi Cancer Center International Symposium
  • Place of Presentation: Nagoya, Japan
• [Presentation] Non-invasive and real-time monitoring of molececular targeting therapy for lymph node and peritoneal metastasis in nude mice bearing xenografts of human colorectal cancer cells tagged with GFP and DsRed, Genetically Engineered and Opticai Probes for Biomedical Applications IV. (2007)
  • Author(s): Nakanishi, H., Hara, M., Ikehara, Y., Tatematsu, M.
  • Organizer: The International Society for Opticai Engineering (SPIE), Photonics West 2007. Symposium
  • Place of Presentation: San Jose, USA.
• [Presentation] A new diagnostic and therapeutic strategy targeting micrometastasis for prevention of peritoneal recurrence in gastric cancer patients (2007)
  • Author(s): Nakanishi, H., Ito, S.,
  • Organizer: The 11th German-Japanese Cancer Workshop
  • Place of Presentation: Kyoto, Japan
• [Presentation] Non-invasive and real-time monitoring of molececular targeting therapy for lymph node and peritoneal metastasis in nude mice bearing xenografts of human colorectal cancer cells tagged with GFP and DsRed (2007)
  • Author(s): Nakanishi H, et. al. [Oral]
  • Organizer: Genetically Engineered and Opticai Probes for Biomedical Applications IV, The International Society for Opticai Engineering(SPIE)
  • Place of Presentation: San Jose, USA
• [Presentation] ヒト舌癌および口腔領域扁平上皮癌のリンパ管新生, リンパ管侵襲およびリンパ節転移に関する免疫組織化学的検討 : 第65回 (2006)
  • Author(s): 大野ふみ, 中西速夫, 阿部厚, 木下篤敬, 関泰, 安井昭夫, 栗田賢一, 立松正衞
  • Organizer: 日本癌学会学術総会
• [Presentation] GFP遺伝子導入ラット舌癌リンパ節微小転移モデルの樹立とリンパ節転移に対するS-1の転移抑制効果の検討 : 第65回 (2006)
  • Author(s): 関泰, 中西速夫, 木下篤敬, 大野ふみ, 今井隆夫, 栗田賢一, 立松正衞
  • Organizer: 日本癌学会学術総会