Developments of HE, fluorescent immunostaining, and WSI analysis by FISH and applications to Hodgkin lymphoma

• Project/Area Number: 17K08746
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Human pathology
• Research Institution: Nagoya City University
• Principal Investigator: Murase Takayuki 名古屋市立大学, 医薬学総合研究院(医学), 准教授 (40315875)
• Co-Investigator(Kenkyū-buntansha): 稲垣 宏 名古屋市立大学, 医薬学総合研究院(医学), 教授 (30232507)
• Project Period (FY): 2017-04-01 – 2020-03-31
• Project Status: Completed (Fiscal Year 2019)
• Budget Amount: ¥4,810,000 (Direct Cost: ¥3,700,000、Indirect Cost: ¥1,110,000); Fiscal Year 2019: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000); Fiscal Year 2018: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000); Fiscal Year 2017: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
• Keywords: 蛍光 in situ hybridization / Sequential FICTION-WSI / 免疫染色 / IN-Cell Analyzer / 顕微鏡画像解析 / 染色体転座 / 多形腺腫 / 多発性骨髄腫 / 病理学 / 癌 / 細胞・組織 / 遺伝子 / トランスレーショナルリサーチ

Outline of Final Research Achievements

The CCND1-IGH, NSD2-IGH, IGH-MAF gene rearrangements by FISH are used for risk stratification in multiple myeloma (MM) cases. Compared with fresh cell-FISH, immunohistochemistry (IHC) is more cost-/time-efficient and can be readily applied to routinely prepared FFPE materials. We performed FFPE-FISH and FFPE-IHC, and examined the usefulness of IHC as a tool for detecting CCND1, NSD2, and MAF gene rearrangements. With cohort n=70, we performed FFPE-FISH and FFPE-IHC, and determined IHC cut-off points. The sensitivity and specificity for the molecules were >.90 and >.96, respectively. With cohort n=120 using unknown gene status cases, we performed IHC, and the gene status was estimated using the cut-off points determined with cohort n=70. The subsequent FISH analysis showed that the sensitivity and specificity for the molecules were >.92 and >.98, respectively. The MM gene rearrangements were estimated accurately by IHC, suggesting that fresh cell-FISH assays can be replaced by IHC.

Academic Significance and Societal Importance of the Research Achievements

次世代シーケンサーにより網羅的に遺伝子を解析し、腫瘍遺伝子異常をビッグデータ化することは腫瘍発生研究に重要であるが、腫瘍組織のビッグデータとの比較の報告はない。我々は1枚のパラフィン組織切片から蛍光免疫染色とFISH法を連続的に行い、スライド全体をビッグデータ化し、細胞単位で組織全体を解析する技術(Sequential FICTION-WSI法)を開発したが、基礎医学・臨床医学分野での応用が可能である。以上から遺伝子ビッグデータと細胞単位の形態・蛋白発現・遺伝子解析から得られた組織ビッグデータとを比較することは画期的であり、腫瘍組織を対象とした分子病理学の応用範囲を飛躍的に広げるものである。

Research Products

• [Journal Article] Mixed-type primary germ cell tumor of the mediastinum in a young adult male with a sudden life threatening condition: A case report. (2020)
  • Author(s): Sakane T, Okuda K, Murase T, Watanabe T, Oda R, Tatematsu T, Yokota K, Haneda H, Inagaki H, Nakanishi R.
  • Journal Title: Thorac Cancer. 11(1):166-169;2020 Volume: 11(1) Pages: 166-169
  • Peer Reviewed
• [Journal Article] Central pathology review of salivary gland adenoid cystic carcinoma. (2020)
  • Author(s): Ueda K, Murase T, Nagao T, Kusafuka K, Urano M, Yamamoto H, Nakaguro M, Taguchi KI, Masaki A, Hirai H, Kawakita D, Tsukahara K, Hato N, Nagao T, Fujimoto Y, Sakurai K, Hanai N, Kano S, Onitsuka T, Okami K, Nibu KI, Tada Y, Kawata R, Inagaki H.
  • Journal Title: Head Neck. Volume: In press
  • Peer Reviewed
• [Journal Article] Clinicopathological significance of EGFR pathway gene mutations and CRTC1/3-MAML2 fusions in salivary gland mucoepidermoid carcinoma. (2020)
  • Author(s): *Morita M, *Murase T, Okumura Y, Ueda K, Sakamoto Y, Masaki A, Kawakita D, Tada Y, Nibu K, Shibuya Y, Inagaki H.
  • Journal Title: Histopathol Volume: In press
  • Peer Reviewed
• [Journal Article] A mutation analysis of the EGFR pathway genes, RAS, EGFR, PIK3CA, AKT1, and BRAF, and TP53 gene in thymic carcinoma and thymoma type A/B3. (2019)
  • Author(s): Sakane T, Murase T, Okuda K, Saida K, Masaki A, Yamada T, Saito Y, Nakanishi R, Inagaki H.
  • Journal Title: Histopathology Volume: 75(5) Pages: 755-766
  • Peer Reviewed
• [Journal Article] Immunohistochemistry for identification of CCND1, NSD2, and MAF gene rearrangements in plasma cell myeloma. (2019)
  • Author(s): Murase T, Ri M, Narita T, Fujii K, Masaki A, Iida S, Inagaki H.
  • Journal Title: Cancer Sci Volume: 110(8) Pages: 2600-2606
  • Peer Reviewed
• [Journal Article] Plasma cell myeloma positive for t(14;20) with relapse in the central nervous system. (2019)
  • Author(s): Murase T, Inagaki A, Masaki A, Fujii K, Narita T, Ri M, Hanamura I, Iida S, Inagaki H.
  • Journal Title: J Clin Exp Hematop Volume: 59(3) Pages: 135-139
  • NAID: 130007720089
  • Peer Reviewed
• [Journal Article] CCR4 is rarely expressed in CCR4-mutated T/NK-cell lymphomas other than adult T-cell leukemia/lymphoma. (2019)
  • Author(s): Sakamoto Y, Fujii K, Murase S, Nakano S, Masaki A, Murase T, Kusumoto S, Iida S, Utsunomiya A, Ueda R, Ishida T, Inagaki H.
  • Journal Title: Int J Hematol. Volume: 110(4) Pages: 389-392
  • Peer Reviewed
• [Journal Article] Postoperative radiotherapy for T1/2N0M0 mucoepidermoid carcinoma positive for CRTC1/3‐MAML2 fusions (2018)
  • Author(s): Okumura Yoshihide、Murase Takayuki、Saida Kosuke、Fujii Kana、Takino Hisashi、Masaki Ayako、Ijichi Kei、Shimozato Kazuo、Tada Yuichiro、Nibu Ken‐ichi、Inagaki Hiroshi
  • Journal Title: Head & Neck Volume: 40 Issue: 12 Pages: 2565-2573
  • DOI: 10.1002/hed.24856
  • Peer Reviewed
• [Journal Article] CCR4 mutations associated with superior outcome of adult T-cell leukemia/lymphoma under mogamulizumab treatment (2018)
  • Author(s): Sakamoto Yuma、Ishida Takashi、Masaki Ayako、Murase Takayuki、Yonekura Kentaro、Tashiro Yukie、Tokunaga Masahito、Utsunomiya Atae、Ito Asahi、Kusumoto Shigeru、Iida Shinsuke、Ueda Ryuzo、Inagaki Hiroshi
  • Journal Title: Blood Volume: 132 Issue: 7 Pages: 758-761
  • DOI: 10.1182/blood-2018-02-835991
  • Peer Reviewed / Open Access
• [Journal Article] Mutation analysis of the EGFR pathway genes, EGFR, RAS, PIK3CA, BRAF, and AKT1, in salivary gland adenoid cystic carcinoma. (2018)
  • Author(s): Saida K, Murase T, Ito M, Fujii K, Takino H, Masaki A, Kawakita D, Ijichi K, Tada Y, Kusafuka K,Iida Y, Onitsuka T, Yatabe Y, Hanai N, Hasegawa Y
  • Journal Title: Oncotarget Volume: 9 Issue: 24 Pages: 17043-17055
  • DOI: 10.18632/oncotarget.24818
  • Peer Reviewed / Open Access
• [Journal Article] Four immunohistochemical assays to measure the PD-L1 expression in malignant pleural mesothelioma. (2018)
  • Author(s): Watanabe T, Okuda K, Murase T, Moriyama S, Haneda H, Kawano O, Yokota K, Sakane T, Oda R, Inagaki H, Nakanishi R.
  • Journal Title: Oncotarget. Volume: 9(29) Issue: 29 Pages: 20769-20780
  • DOI: 10.18632/oncotarget.25100
  • Peer Reviewed / Open Access
• [Journal Article] Improved clonality detection in Hodgkin lymphoma using a semi-nested modification of the BIOMED-2 PCR assay for IGH and IGK rearrangements: A paraffin-embedded tissue study. (2018)
  • Author(s): Han S, Masaki A, Sakamoto Y, Takino H, Murase T, Iida S, Inagaki H.
  • Journal Title: Pathol Int Volume: Epub Issue: 5 Pages: 12660-12660
  • DOI: 10.1111/pin.12660
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] A comparative study of PD-L1 immunohistochemical assays with four reliable antibodies in thymic carcinoma. (2018)
  • Author(s): Sakane T, Murase T, Okuda K, Takino H, Masaki A, Oda R, Watanabe T, Kawano O, Haneda H, Moriyama S, Saito Y, Yamada T, Nakanishi R, Inagaki H.
  • Journal Title: Oncotarget Volume: 9(6) Issue: 6 Pages: 6993-7009
  • DOI: 10.18632/oncotarget.24075
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] <i>Achromobacter</i> Infection Is Rare in Japanese Patients with Pulmonary B-cell Lymphoma (2018)
  • Author(s): Aoyama S, Masaki A, Sakamoto Y, Takino H, Murase T, Ohshima K, Yoshino T, Kato S, Inagaki H.
  • Journal Title: Internal Medicine Volume: 57 Issue: 6 Pages: 789-794
  • DOI: 10.2169/internalmedicine.9430-17
  • NAID: 130006507388
  • ISSN: 0918-2918, 1349-7235
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Improved clonality detection in B-cell lymphoma using a semi-nested modification of the BIOMED-2 PCR assay for IGH rearrangement: A paraffin-embedded tissue study. (2017)
  • Author(s): Sakamoto Y, Masaki A, Aoyama S, Han S, Saida K, Fujii K, Takino H, Murase T, Iida S, Inagaki H.
  • Journal Title: Pathol Int Volume: 67(9) Issue: 9 Pages: 453-460
  • DOI: 10.1111/pin.12566
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] MYB, MYBL1, MYBL2 and NFIB gene alterations and MYC overexpression in salivary gland adenoid cystic carcinoma. (2017)
  • Author(s): Fujii K, Murase T, Beppu S, Saida K, Takino H, Masaki A, Ijichi K, Kusafuka K, Iida Y, Onitsuka T, Yatabe Y, Hanai N, Hasegawa Y
  • Journal Title: Histopathology Volume: 7 Issue: 5 Pages: 823-834
  • DOI: 10.1111/his.13281
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Presentation] FFPE標本を用いた免疫染色は多発性骨髄腫におけるIGH関連遺伝子再構成の有用なスクリーニング法である． (2019)
  • Author(s): 村瀬貴幸、正木彩子、藤井慶一郎、上田佳緒璃、李政樹、飯田真介、稲垣宏
  • Organizer: 第108回 日本病理学会総会(2019.5.東京)
• [Presentation] 多発性骨髄腫におけるCCND1、MMSET、MAF遺伝子再構成の検出：パラフィン標本を用いた組織FISH法および免疫組織染色への応用 (2018)
  • Author(s): 村瀬貴幸、李政樹、成田朋子、吉田嵩、正木彩子、飯田真介、稲垣宏
  • Organizer: 第43回日本骨髄腫学会 (2018.05.)
• [Presentation] ETV6-RET融合遺伝子を有した唾液腺分泌癌の2例 (2018)
  • Author(s): 村瀬貴幸、奥村嘉英、齋田昂佑、坂本祐真、津田香那、滝野寿、正木彩子、稲垣宏
  • Organizer: 第107回日本病理学会総会 (2018.06.)
• [Presentation] MAFB遺伝子再構成陽性を示す多発性骨髄腫の1例 (2018)
  • Author(s): 村瀬貴幸、李政樹、稲垣淳、成田朋子、吉田嵩、正木彩子、飯田真介、稲垣宏
  • Organizer: 第58回日本リンパ・網内系学会 (2018.06.)
• [Presentation] Sequential FICTION-WSI法を用いた多形腺腫細胞のclonality解析 (2017)
  • Author(s): 村瀬 貴幸、石橋 謙一郎、藤井 香那、齋田 昂佑、滝野 寿、正木 彩子、稲垣 宏
  • Organizer: 第106回日本病理学会総会
• [Presentation] t(4;14)(p16;q32)/IGH-FGFR3とt(14;16)(q32;q23)/IGH-MAFに対するパラフィン検体による組織 FISH法の有用性 (2017)
  • Author(s): 村瀬 貴幸、成田 朋子、李 政樹、正木 彩子、吉田 嵩、飯田 真介、稲垣 宏
  • Organizer: 第42回日本骨髄腫学会
• [Presentation] 免疫染色により確定診断された唾液腺intercalated duct lesionの1例 (2017)
  • Author(s): 村瀬 貴幸、坂本 祐真、藤井 香那、齋田 昂佑、滝野 寿、正木 彩子、稲垣 宏
  • Organizer: 第62回日本唾液腺学会