基礎研究

  • 分子血液グループ
  • 細胞免疫グループ
  • リンパ系腫瘍グループ

リンパ系腫瘍グループ (Lymphoid Malignancies Group)

中川雅夫 助教 グループリーダー・後藤秀樹 助教
吉岡康介 大学院1年・下埜城嗣 大学院4年・石尾崇 大学院2年

写真:血液がんの中でも特に診断及び治療が難しいT細胞リンパ腫を中心に、網羅的機能的遺伝子スクリーニングから、新しい治療法・診断法の創出を目指します。(右から石尾、後藤、中川、下埜、吉岡)

血液がんの中でも特に診断及び治療が難しい病気の一つが、悪性リンパ腫です。特にT細胞性リンパ腫は既存の治療法に抵抗性で、あるいは、一度は治療が奏功しても高頻度に再発をきたします。この現状を変えて行くために、新しい治療標的分子の同定、分子標的薬の開発が望まれていますが、その基盤となるべき分子病態の理解は遅れています。

近年は次世代シークエンサーの登場により網羅的な体細胞遺伝子変異解析が可能になりました。T細胞性リンパ腫の分野でも腫瘍形成に重要と考えられる遺伝子変異が多数報告されてきています(Nakagawa et al. JEM 211:2497-505, 2014.など)。

bcr-abl依存性耐性機序については、bcr-ablに変異が生じることによりimatinibの効果が無くなる場合、また、bcr-abl遺伝子が増幅し過剰発現することでimatinibの効果が不十分になる場合があります。
一方、bcr-abl非依存性耐性機序については、bcr-ablとは別に、他の癌遺伝子が活性化することでbcr-abl非依存的に細胞増殖が誘導される場合や、imatinibの細胞内濃度が低下するような細胞表面のトランスポーターの異常が示されています。

それらの遺伝子変異の発見が新しい治療法開発につながると良いのですが、実際はそう簡単ではないこともわかってきています。その遺伝子変異はリンパ腫ができ始めたきっかけだったかもしれませんが、病気が発見され治療が必要な時にはすでに不必要になっているかもしれません。また、がんは一つの遺伝子異常ではなく、たくさんの遺伝子群によるnet effectで腫瘍形質を維持しています。一つの遺伝子を標的としても、不十分な効果しか得られないこともあります。では、どの遺伝子を標的にすれば本当に治療効果をあげられるのでしょうか?これは全ての悪性リンパ腫患者さん、そして、その治療に関わる人々にとって、最も重要な問題ではないでしょうか?

この重要問題に正面から取り組むために、我々はCRISPR/Cas9網羅的遺伝子ノックアウトスクリーニングを基盤とした仕事を進めています。CRISPR/Cas9は近年開発されたばかりの革新的ゲノム編集技術であり、これを用いることで従来法より飛躍的に高い効率・特異性で標的遺伝子をノックアウトすることが可能になってきました。我々はこのシステムをヒトT細胞性リンパ腫細胞株に導入、約 20000遺伝子を網羅的に解析し、どの遺伝子を標的とすることでT細胞リンパ腫細胞を根絶できるのかを探っています(図)。我々の研究成果が、創薬へのシーズ、治療反応性バイオマーカの発見や薬剤耐性機序解明などに発展し、実医療へ貢献できることを目指しています

図:CRISPR Cas9 sgRNA KO Library screening technology

Lymphoid Malignancies Group:
Principal Investigator:
Masao Nakagawa, M.D., Ph.D.
Assistant Professor
Department of Hematology, Hokkaido University Graduate School of Medicine
Kita 15, Nishi 7, Kita-ku, Sapporo 060-8638, Japan
Tel : +81-(0)11-706-7214, Fax :+81-(0)11-706-7823
E-mail: nakagawam@med.hokudai.ac.jp

Team: (Alphabetical order)
Hideki Goto M.D., Ph.D. Assistant Professor
Takashi Ishio M.D. Graduate student
Joji Shimono M.D. Graduate student
Kousuke Yoshioka M.D. Graduate student

RESEARCH INTERESTS:

Functional genomics for discovering the therapeutic molecular targets in PTCL

> Identifying the "Achilles' heel" genes in peripheral T-cell lymphomas as molecular targets for therapeutic intervention.
> Defining the molecular mechanisms of frequently mutated CCR4 gene in PTCL.

Overview
Human peripheral T-cell lymphomas (PTCL) is approximately 10 % of non-Hodgkin Lymphomas in worldwide. PTCL comprises heterogeneous diseases, where most are aggressive with less than 40% of 5-year overall survival with current treatment strategy. A discovery for new therapeutic modalities is urgent need.

Recently, the Next generation sequencing technology enabled us to find frequently mutated genes in PTCL. In my previous work as a postdoctoral fellow in Lou Staudt Lab and Thomas Waldmann lab, we discovered frequent gain-of-function mutation in chemokine receptor CCR4 gene in Adult T-cell leukemia/lymphoma (ATLL) and provided functional evidences that inhibition of CCR4 signaling might have therapeutic potential for patients with ATLL. However most of other somatically mutated genes in PTCL were not fully characterized in terms of therapeutic targets.
On the other hand, it has been recently known that the malignant cells potentially acquire the dependency on un-mutated genes which is the phenomenon called as "non-oncogene addiction". These observations indicate that non-biased and comprehensive functional investigation should be conducted in order to discover the therapeutic molecular targets in PTCL.

The goal of our laboratory is to understand the oncogenic molecular networks in PTCL which can be exploited for therapeutic intervention. To directly address this issue, our lab applies revolutionizing CRISPR/Cas9/sgRNA library screening technology to the following functional investigation.

1) Identifying indispensable pathways/molecules for the cellular proliferation/survival of PTCL cells, especially ATLL.
2) Discovering the pathways/molecules associated with drug-resistance or drug-sensitivity in PTCL cells.

PUBLICATION LIST:
  1. Perera, LP., Zhang, M., Nakagawa, M., Petrus, MN., Maeda, M., Kadin, ME., Waldmann, TA., Perera, PY. Chimeric antigen receptor modified T cells that target chemokine receptor CCR4 as a therapeutic modality for T-cell malignancies. American Journal of Hematology, 2017. In Press.
  2. Shimono, J., Miyoshi, H., Kiyasu, J., Sato, K., Kamimura, T., Eto, T., Miyagishima, T., Nagafuji, K., Teshima, T., Ohshima, K. Clinicopathologic analysis of primary splenic diffuse large B-cell lymphoma. Br J Haemtol 2017 In Press.
  3. Shimono, J., Miyoshi, H., Seto, M., Ohshima, K. Clinical features of diffuse large B-cell lymphoma with polyploidy. Pathol Int, 67:17-23, 2017.
  4. Chen, J., Zhang. Y,, Petrus, MN., Xiao, W., Nicolae, A., Raffeld, M., Pittaluga, S., Bamford, RN., Nakagawa, M., Ouyanga, S., Epsteine, AL., Kadin, ME., Del Mistro, A., Woessnerh, R., Jaffe, ES., Waldmann, TA. Cytokine receptor signaling is required for the survival of ALK- anaplastic large cell lymphoma even in the presence of JAK1/STAT3 mutations. Proc. Natl. Acad. Sci. U S A, 2017. In Press.
  5. Yang, Y., Kelly, P., Shaffer, AL., Schmitz, R., Yoo, HM., Liu, X., Huang, da W., Webster, D., Young, RM., Nakagawa, M., Ceribelli, M., Wright, GW., Yang, Y., Zhao, H., Yu, X., Xu, W., Chan, WC., Jaffe, ES., Gascoyne, RD., Campo, E., Rosenwald, A, Ott, G., Delabie, J., Rimsza, L., Staudt, LM.  Targeting Non-proteolytic Protein Ubiquitination for the Treatment of Diffuse Large B Cell Lymphoma.  Cancer Cell, 29:494-507, 2016.
  6. Hodson, DJ., Shaffer, AL., Xiao, W., Wright, GW., Schmitz, R., Phelan, JD., Yang, Y., Webster, DE., Rui, L., Kohlhammer, H., Nakagawa, M., Waldmann, TA., Staudt, LM. Regulation of normal B-cell differentiation and malignant B-cell survival by OCT2. Proc. Natl. Acad. Sci. U S A, 113:E2039-46, 2016
  7. Miyoshi, H., Kiyasu, J., Kato, T., Yoshida, N., Shimono, J., Yokoyama, S., Taniguchi, H., Sasaki, Y., Kurita, D., Kawamoto, K., Kato, K., Imaizumi, Y., Seto, M., Ohshima, K. PD-L1 expression on neoplastic or stromal cells is respectively a poor or good prognostic factor for adult T-cell leukemia/lymphoma. Blood, 128:1374-81, 2016.
  8. Yokoyama, S., Miyoshi, H., Nakashima, K., Shimono, J., Hashiguchi, T., Mitsuoka, M., Takamori, S., Akagi, Y., Ohshima, K. Prognostic Value of Programmed Death Ligand 1 and Programmed Death 1 Expression in Thymic Carcinoma. Clin Cancer Res, 22:4727-34, 2016
  9. Yoshida, N., Miyoshi, H., Kato, T., Sakata-Yanagimoto, M., Niino, D., Taniguchi, H., Moriuchi, Y., Miyahara, M., Kurita, D., Sasaki, Y., Shimono, J., Kawamoto, K., Utsunomiya, A., Imaizumi, Y., Seto, M., Ohshima, K. CCR4 frameshift mutation identifies a distinct group of adult T cell leukaemia/lymphoma with poor prognosis. J Pathol, 238:621-6, 2016.
  10. Nakagawa, M., Schmitz, R., Xiao, W., Goldman, CK., Yang, Y., Xu, W., Yu, X., Waldmann, TA., Staudt, LM. Gain-of-Function CCR4 Mutations in Adult T-cell Leukemia/Lymphoma. J. Exp. Med., 211:2497-505, 2014.
  11. Yu, P., Petrus, MN., Ju, W., Zhang, M., Conlon, KC., Nakagawa, M., Maeda, M., Bamford, RN., Waldmann, TA. Augmented efficacy with the combination of blockade of the Notch-1 pathway, bortezomib and romidepsin in a murine MT-1 adult T-cell leukemia model. Leukemia, 29:556-66, 2014.
  12. Suguro, M., Yoshida, N., Umino, A., Kato, H., Tagawa, H., Nakagawa, M., Fukuhara, N., Karnan, S., Takeuchi, I., Hocking, TD., Arita, K., Karube, K., Tsuzuki, S., Nakamura, S., Kinoshita, T., Seto, M. Clonal heterogeneity of lymphoid malignancies correlates with poor prognosis. Cancer Sci., 105:897-904, 2014.
  13. Karube, K., Nakagawa, M., Tsuzuki, S., Takeuchi, I., Honma, K., Nakashima, Y., Shimizu, N., Ko, YH., Morishima, Y., Ohshima, K., Nakamura, S., Seto, M. Identification of FOXO3 and PRDM1 as tumor-suppressor gene candidates in NK-cell neoplasms by genomic and functional analyses. Blood, 118: 3195-204, 2011.
  14. Nakagawa, M., Tsuzuki, S, Honma, K., Taguchi, O., Seto, M. Synergistic effect of Bcl2, Myc and Ccnd1 transforms mouse primary B-cells into malignant cells. Haematologica, 96: 1318-1326, 2011.
  15. Umino, A., Nakagawa, M., Utsunomiya, A., Tsukasaki, K., Taira, N., Katayama, N., Seto, M. Clonal evolution of adult T-cell leukemia/lymphoma takes place in lymph node. Blood, 117: 5473-5478, 2011.
  16. Kato, H., Kagami, Y., Nakagawa, M., Karnan, S., Yatabe, Y., Nakamura, S., Morishima, Y., Seto, M. IL-4/CD40L Co-Stimulation Induces Long-Term Proliferation for CD10-Positive Germinal Center B Cell-Like Diffuse Large B-Cell Lymphoma. The Open Leukemia Journal, 3: 60-68, 2010.
  17. Seto, M., Honma, K., Nakagawa, M. Diversity of genome profiles in malignant lymphoma. Cancer Sci., 101: 573-578, 2010.
  18. Honma, K. Tsuzuki, S. Nakagawa, M., Tagawa, H., Nakamura, S., Morishima, Y., Seto, M. TNFAIP3/A20 functions as a novel tumor suppressor gene in several subtypes of non-Hodgkin lymphomas. Blood, 114: 2467-2475, 2009.
  19. Nakagawa, M., Nakagawa-Oshiro, A., Karnan, S., Tagawa, H., Utsunomiya, A., Nakamura, S., Takeuchi, I., Ohshima, K., Seto, M. Array CGH analysis of PTCL-U reveals a distinct subgroup with genetic alterations similar to lymphoma-type ATLL. Clin. Cancer Res., 15: 30-38, 2009.
  20. Takeuchi, I., Tagawa, H., Tsujikawa, A., Nakagawa, M., Katayama-Suguro, M., Guo, Y., Seto, M. The potential of copy number gains and losses, detected by array-based comparative genomic hybridization, for computational differential diagnosis of B-cell lymphomas and genetic regions involved in lymphomagenesis. Haematologica, 94: 61-69, 2009.
  21. Hashino, S., Kobayashi, S., Takahata, M., Onozawa, M., Nakagawa, M., Kawamura, T., Fujisawa, F., Izumiyama, K., Kahata, K., Kondo, T., Asaka, M. Graft-versus-tumor effect after reduced-intensity allogeneic hematopoietic stem cell transplantation in a patient with advanced colon cancer. Int. J. Clin. Oncol., 13: 176-180, 2008.
  22. Honma, K., Tsuzuki, S., Nakagawa, M., Karnan, S., Aizawa, Y., Kim, WS., Kim, YD., Ko, YH., Seto, M. TNFAIP3 is the target gene of chromosome band 6q23.3-q24.1 loss in ocular adnexal marginal zone B cell lymphoma. Genes Chrom. Cancer, 47: 1-7, 2008.
  23. Hashino, S., Morita, L., Takahata, M., Onozawa, M., Nakagawa, M., Kawamura, T., Fujisawa, F., Kahata, K., Izumiyama, K., Yonezumi, M., Chiba, K., Kondo, T., Asaka, M. Administration of micafungin as prophylactic antifungal therapy in patients undergoing allogeneic stem cell transplantation. Int. J. Hematol., 87: 91-97, 2008.
  24. Nakagawa, M., Hashino, S., Takahata, M., Kawamura, T., Fujisawa, F., Kahata, K., Kondo, T., Imamura, M., Ando, S., Asaka, M. Successful reduced-intensity stem cell transplantation with cord blood for a poor-prognosis adult with refractory chronic active epstein-barr virus infection. Int. J. Hematol., 85: 443-445, 2007.
  25. Fukuhara, N., Nakamura, T., Nakagawa, M., Tagawa, H., Takeuchi, I., Yatabe, Y., Morishima, Y., Nakamura, S., Seto, M. Chromosomal imbalances are associated with outcome of Helicobacter pylori eradication in t(11;18)(q21;q21) negative gastric MALT lymphomas. Genes Chrom. Cancer, 46: 784-790, 2007.
  26. Onozawa, M., Hashino, S., Takahata, M., Fujisaw, F., Kawamura, T., Nakagawa, M., Kahata, K., Kondo, T., Ota, S., Tanaka, J., Imamura, M., Asaka, M. Relationship between preexisting anti-varicella-zoster virus (VZV) antibody and clinical VZV reactivation in hematopoietic stem cell transplantation recipients. J. Clin. Microbiol., 44: 4441-4443, 2006.
  27. Nakagawa, M., Seto, M., Hosokawa, Y. Molecular pathogenesis of MALT lymphoma: two signaling pathways underlying anti-apoptotic effect of API2-MALT1 fusion protein. Leukemia, 20: 929-936, 2006.
  28. Nakagawa, M., Seto, M., Hosokawa, Y. Molecular pathogenesis of MALT lymphoma: two signaling pathways underlying anti-apoptotic effect of API2-MALT1 fusion protein. Leukemia, 20: 929-936, 2006.
  29. Hosokawa, Y., Suzuki, H., Nakagawa, M., Lee, T.H., Seto, M. API2-MALT1 fusion protein induces transcriptional activation of the API2 gene through NF-kappaB binding elements: Evidence for a positive feed-back loop pathway resulting in unremitting NF-kappaB activation. Biochem. Biophys. Res. Commun., 334: 51-60, 2005.
  30. Nakagawa, M., Kameoka, Y., Suzuki, R. Nucleophosmin in acute myelogenous leukemia. N. Engl. J. Med., 352: 1819-1820, 2005.
  31. Izumiyama, K., Nakagawa, M., Yonezumi, M., Kasugai, Y., Suzuki, R., Suzuki, H., Tsuzuki, S., Hosokawa, Y., Asaka, M., Seto, M. Stability and subcellular localization of API2-MALT1 chimeric protein involved in t(11;18) (q21;q21) MALT lymphoma. Oncogene, 22: 8085-8092, 2003.
  32. Nakagawa, M., Miyagishima, T., Kamata, T., Arai, S., Miura, Y., Onishi, S., Kishimoto, A., Kamishima, Y., Choi, G.H., Kudo, M., Okabe, M. Refractory idiopathic cold agglutinin disease successfully treated with intermittent high-dose cyclophosphamide. Rinsho Ketsueki, 42: 713-715, 2001.