Elucidation of the mechanism of intractability of pancreatobiliary cancers using patients-derived organoids

2022 Fiscal Year Annual Research Report

• Project/Area Number: 22H02839
• Allocation Type: Single-year Grants
• Research Institution: Tohoku University
• Principal Investigator: 古川 徹 東北大学, 医学系研究科, 教授 (30282122)
• Co-Investigator(Kenkyū-buntansha): 木下 賢吾 東北大学, 情報科学研究科, 教授 (60332293) ; 海野 倫明 東北大学, 医学系研究科, 教授 (70282043)
• Project Period (FY): 2022-04-01 – 2026-03-31
• Keywords: 膵臓がん / 胆道がん / オルガノイド / ゲノム / 化学療法

Outline of Annual Research Achievements

術前化学療法後切除膵臓がん臨床検体159例及び入手可能なそれら切除検体の術前生検標本120例においてGATA6, CK5, vimentinの発現を解析し，その特徴及び予後を含む臨床病理学的徴候との関連を明らかにした。膵臓がんはGATA6, CK5発現様式により，GATA6高CK5低のclassical群，GATA6高CK5高のhybrid群，GATA6低CK5高のbasal-like群，GATA6低CK5低のnull群に分けられ，予後との関連において，classical群が最も良く，null群は最も不良であった。Hybrid群とbasal-like群はそれらの中間の予後を示した。化学療法前の生検検体において，vimentin高の膵臓がんは予後不良であった。化学療法前と後でGATA6，CK5発現様式が変化した例が多数あり，化学療法効果判定で比較的よく奏功したとみなされたGrade 2/3例では予後不良の発現様式の膵臓がんに変化した例が有意に多く認められ，これらが膵臓がん再発に関与している可能性が示唆された。KRAS遺伝子変異はこれら発現様式とは関連していなかった。GATA6とCK5の発現様式は膵臓がん術前化学療法施行例の予後予測に有用であり，また，化学療法後の発現様式変化に関連することが示され，膵臓がんにおける術前化学療法施行の高精度化に寄与する情報を明らかにすることができた。また、予後不良であった肉腫様胆嚢がんの遺伝子・分子発現解析を施行し，TP53，ARID1A，SMAD4の異常が寄与していることを明らかにした。さらに，膵臓がん細胞においいて活性化しているMAPKの下流にあるlong non-coding RNAを検索し，LINC00941が特異的にupregulateし、予後と関連することを明らかにした。

Current Status of Research Progress

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

Reason

当初計画で予定していた、術前化学療法後切除膵臓がん臨床検体の術前生検組織と術後切除組織の臨床病理学的解析において論文発表に至る成果を得ることができた。膵臓がん、胆道がんにおけるゲノム解析についてもデータを得ることができ、また、患者組織由来オルガノイド培養についても継続的に行なうことができた。

Strategy for Future Research Activity

次年度も引き続き下記を継続して行う。
①術前化学療法後切除膵臓がん臨床検体の術前生検組織と術後切除組織の臨床病理学的解析：術前化学療法施行後切除組織について、術前病理組織像、術後組織における残存がん細胞の定性・定量的評価による化学療法効果のスコア化を行い、臨床データとの比較を含め、臨床病理学的特徴を抽出する。
②組織採取と核酸抽出：化学療法前生検組織はホルマリン固定パラフィン包埋（FFPE）標本であり、術後化学療法後切除組織はFFPEおよび一部は凍結組織が保存されている。それらから、がん細胞部分、がん細胞に隣接する間質組織部分、非腫瘍部分をマイクロダイセクションにて採取し、DNA、RNAを抽出する。
③ゲノム、トランスクリプトーム解析と統計解析：抽出したDNA、RNAを用い、次世代型シーケンサー（NGS）により全ゲノムまたはエクソームシーケンス，トランスクリプトームシーケンスを行う。塩基置換、挿入欠失、構造異常、コピー数異常、遺伝子発現変化を網羅的にリストアップする。ゲノム変異データについてはがんの体細胞性変異とホストのゲノム多型を解析する。分子異常は必要に応じ，サンガー法，ディジタルPCR法、Real time PCR法によりvalidationする。タンパク質発現を免疫組織化学法、免疫ブロット法により解析する。遺伝子変異、発現動態と化学療法感受性との関連性の統計解析を行い、化学療法感受性、抵抗性に関連する分子候補を見出す。
④オルガノイド培養と機能解析：新たに得られる検体について切除組織からがん細胞、間質細胞を分離しオルガノイド培養を行う。オルガノイド培養は申請者ラボの既報（Eur J Cancer 148:239-50, 2021）による。

Research Products

• [Int'l Joint Research] Johns Hopkins University/Memorial Sloan Kettering Cancer Center/Massachusetts General Hospital(米国)
  • Country Name: U.S.A.
  • Counterpart Institution: Johns Hopkins University/Memorial Sloan Kettering Cancer Center/Massachusetts General Hospital
• [Int'l Joint Research] University of Verona(イタリア)
  • Country Name: ITALY
  • Counterpart Institution: University of Verona
• [Int'l Joint Research] University of Korc(トルコ)
  • Country Name: TURKEY
  • Counterpart Institution: University of Korc
• [Int'l Joint Research] Asan Medical Center(韓国)
  • Country Name: KOREA (REP. OF KOREA)
  • Counterpart Institution: Asan Medical Center
• [Int'l Joint Research] University of Utrecht(オランダ)
  • Country Name: NETHERLANDS
  • Counterpart Institution: University of Utrecht
• [Journal Article] GATA6 and CK5 stratify the survival of patients with pancreatic cancer undergoing neoadjuvant chemotherapy. (2023)
  • Author(s): Kokumai T, Omori Y, Ishida M, Ohtsuka H, Mizuma M, Nakagawa K, Maeda C, Ono Y, Mizukami Y, Miura S, Kume K, Masamune A, Morikawa T, Unno M, Furukawa T.
  • Journal Title: Modern Pathology Volume: 36 Pages: 100102
  • DOI: 10.1016/j.modpat.2023.100102
  • Peer Reviewed
• [Journal Article] Molecular comparison of concurrent components of high-grade dysplasia, adenocarcinoma, and sarcomatoid carcinoma in a case of sarcomatoid carcinoma of the gallbladder. (2023)
  • Author(s): Hirose K, Omori Y, Ono Y, Mizukami Y, Kaneko Y, Maruyama T, Ohtani H, Furukawa T.
  • Journal Title: Virchows Archiv Volume: - Pages: -
  • DOI: 10.1007/s00428-023-03524-7
  • Peer Reviewed
• [Journal Article] Long non-coding RNAs associated with mitogen-activated protein kinase in human pancreatic cancer. (2023)
  • Author(s): Ishikawa T, Fukushige S, Saiki Y, Hirose K, Hiyoshi T, Ogawa T, Katori Y, Furukawa T.
  • Journal Title: Cancers Volume: 15 Pages: 303
  • DOI: 10.3390/cancers15010303
  • Peer Reviewed / Open Access
• [Journal Article] Mechanisms of development and progression of pancreatic neoplasms. (2022)
  • Author(s): Furukawa T.
  • Journal Title: Pathology International Volume: 72 Pages: 529-540
  • DOI: 10.1111/pin.13272
  • Peer Reviewed / Open Access
• [Journal Article] Application of patient-derived cancer organoids to personalized medicine. (2022)
  • Author(s): Shiihara M, Furukawa T.
  • Journal Title: Journal of Personalized Medicine Volume: 12 Pages: 789
  • DOI: 10.3390/jpm12050789
  • Peer Reviewed / Open Access
• [Presentation] Lessons Learned from Mistakes Made In the Interpretation of Small Biopsies and Cytologic Samples from the Pancreatobiliary Tract and Ampulla. (2023)
  • Author(s): Furukawa T.
  • Organizer: 112th Annual Meeting of the United States and Canadian Academy of Pathology.
  • Int'l Joint Research / Invited
• [Presentation] Molecular Phenotyping and the Combinatorial System by the Number of Accumulated Gene Alterations of Intraductal Papillary Mucinous Neoplasms Stratifies the Risk of Invasion and Outcome: New Insight for Management Strategy. (2023)
  • Author(s): Omori Y, Ono Y, Kokumai T, Date F, Higuchi R, Honda G, Mizukami Y, Morikawa T, Unno M, Furukawa T.
  • Organizer: 112th Annual Meeting of the United States and Canadian Academy of Pathology.
  • Int'l Joint Research
• [Presentation] Prognostic Performance of Microscopic Size Measurements in Small Invasive Carcinomas Arising in Intraductal Papillary Mucinous Neoplasms (IPMNs) of the Pancreas. (2023)
  • Author(s): Zhang ML, Omori Y, Hong SM, Ideno N, Date K, Missael Rocha Castellanous D, Shroff S, Furukawa T, Fernadez-del Castillo C, Mino-Kenudson M.
  • Organizer: 112th Annual Meeting of the United States and Canadian Academy of Pathology. 2023年3月11-17日 New Orleans, LO, USA Poster
  • Int'l Joint Research
• [Presentation] Whole exome sequencing of familial pancreatic cancer cases in Japan. (2022)
  • Author(s): Furukawa T.
  • Organizer: 第81回日本癌学会総会
• [Presentation] Pathological aspects. International Consensus Meeting Management of IPMN and MCN. (2022)
  • Author(s): Furukawa T.
  • Organizer: International Association of Pancreatology and Japan Pancreas Society Meeting 2022
  • Int'l Joint Research
• [Presentation] Three molecular development pathways of remnant pancreatic cancer impact survival after resection. (2022)
  • Author(s): Suzuki S, Omori Y, Hirose K, Ono Y, Karasaki H, Shimoda M, Nagakawa Y, Tsuchida A, Mizukami Y, Furukawa T.
  • Organizer: nternational Association of Pancreatology and Japan Pancreas Society Meeting 2022
  • Int'l Joint Research
• [Presentation] Interobserver agreement on biopsy-based diagnosis of autoimmune pancreatitis. (2022)
  • Author(s): Notohara K, Kamisawa T, Furukawa T, Fukushima N, Ikeura T, Kawa S, Okazaki K.
  • Organizer: International Association of Pancreatology and Japan Pancreas Society Meeting 2022
  • Int'l Joint Research
• [Presentation] Molecular Phenotypic Risk Stratification of Intraductal Papillary Mucinous Neoplasms. (2022)
  • Author(s): Omori Y, Ono Y, Morikawa T, Date F, Higuchi R, Honda G, Karasaki H, Mizukami Y, Unno M, Furukawa T.
  • Organizer: International Association of Pancreatology and Japan Pancreas Society Meeting 2022
  • Int'l Joint Research
• [Presentation] STK11-aletered intraductal papillary mucinous neoplasms develop characteristic clinico-patho-biology. (2022)
  • Author(s): Omori Y, Ono Y, Itoh T, Date F, Morikawa T, Motoi F, Higuchi R, Honda G, Karasaki H, Mizukami Y, Unno M, Furukawa T.
  • Organizer: International Association of Pancreatology and Japan Pancreas Society Meeting 2022
  • Int'l Joint Research
• [Presentation] Characteristics of growth and mucus secretion of patient-derived organoids may substantiate invasive phenotypes of IPMN. (2022)
  • Author(s): Shiihara M, Ishikawa T, Yoshida T, Omori Y, Hirose K, Morikawa T, Nakagawa K, Mizuma M, Otsuka H, Motoi F, Unno M, Okamura Y, Kinoshita K, Furukawa T.
  • Organizer: International Association of Pancreatology and Japan Pancreas Society Meeting 2022
  • Int'l Joint Research
• [Presentation] Rearrangement and expression of PRKACA/B governs phenotypes of pancreato-biliary oncocytic neoplasms. (2022)
  • Author(s): Itoh T, Omori Y, Seino M, Hirose K, Date F, Ono Y, Mizukami Y, Mizuma M, Morikawa T, Higuchi R, Honda G, Unno M, Furukawa T.
  • Organizer: International Association of Pancreatology and Japan Pancreas Society Meeting 2022
  • Int'l Joint Research
• [Presentation] Expression phenotypes of GATA6 and CK5 stratify the survival of pancreatic cancer. (2022)
  • Author(s): Kokumai T, Omori Y, Ishida M, Date F, Inoue K, Douchi D, Nakayama S, Miura T, Maeda S, Masuda K, Takikawa T, Miura S, Kume K, Ohtsuka H, Mizuma M, Nakagawa K, Morikawa T, Masamune A, Unno M, Furukawa T.
  • Organizer: International Association of Pancreatology and Japan Pancreas Society Meeting 2022
  • Int'l Joint Research
• [Presentation] Dual specificity phosphatase 6 deficiency improves insulin sensitivity and protects against obesity in mice. (2022)
  • Author(s): Jiang C, Saiki Y, Furukawa T.
  • Organizer: International Association of Pancreatology and Japan Pancreas Society Meeting 2022
  • Int'l Joint Research
• [Presentation] Intraductal neoplasms of the pancreas are precursor lesions of the pancreatic cancer. (2022)
  • Author(s): Furukawa T.
  • Organizer: 7th Japanese Cancer Association and American Association for Cancer Research Special Joint Conference. The Latest Advances in Pancreatic Cancer Research: From Basic Science to Therapeutics.
  • Int'l Joint Research / Invited