Identification of drug resistance-related proteins in pancreatic cancer by SILAC quantitative proteomic approach.

• Project/Area Number: 24501338
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Tumor diagnosis
• Research Institution: Chiba University
• Principal Investigator: SATOH Mamoru 千葉大学, 医学部附属病院, 寄附研究部門教員 (20401002)
• Co-Investigator(Kenkyū-buntansha): NOMURA Fumio 千葉大学, 大学院医学研究院, 教授 (80164739) ; MIYAZAKI Masaru 千葉大学, 大学院医学研究院, 教授 (70166156) ; YOSHITOMI Hideyuki 千葉大学, 医学部附属病院, 講師 (60375631) ; SOGAWA Kazuyuki 麻布大学, 生命・環境化学部, 講師 (50436440)
• Co-Investigator(Renkei-kenkyūsha): KODERA Yoshio 北里大学, 理学部, 准教授 (60265733) ; TAKANO Shigetsugu 千葉大学, 医学部附属病院, 助教 (20436380)
• Research Collaborator: KAWASHIMA Yusuke 理化学研究所, 統合生命医科学研究センター, 研究員
• Project Period (FY): 2012-04-01 – 2015-03-31
• Project Status: Completed (Fiscal Year 2014)
• Budget Amount: ¥5,330,000 (Direct Cost: ¥4,100,000、Indirect Cost: ¥1,230,000); Fiscal Year 2014: ¥1,690,000 (Direct Cost: ¥1,300,000、Indirect Cost: ¥390,000); Fiscal Year 2013: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000); Fiscal Year 2012: ¥2,080,000 (Direct Cost: ¥1,600,000、Indirect Cost: ¥480,000)
• Keywords: プロテオーム解析 / プロテオミクス解析 / 抗癌剤耐性 / 膵臓癌 / 培養細胞 / 培養上清 / SILAC

Outline of Final Research Achievements

In this study, we employed a quantitative proteomics approach using stable isotope labeling with amino acids in cell culture (SILAC) followed by mass spectrometry to quantify changes in protein levels between gemcitabine resistant and sensitive pancreatic cancer cells. Proteins were extracted with cell fractionation and difference of solubilization. 344 (cell fractionation) and 118 (difference of solubilization) proteins were shown more than 2-fold change between the resistant and sensitive cell lines. Expression ratio of 40 proteins were confirmed by WB. 35 proteins were indicated more than 2-fold change by secretome analysis.

Research Products

• [Journal Article] Urinary albumin and transferrin as early diagnostic markers of chronic kidney disease. (2015)
  • Author(s): Maeda H, Sogawa K, Sakaguchi K, Abe S, Sagizaka W, Mochizuki S, Horie W, Watanabe T, Shibata Y, Satoh M, Sanda A, Nomura F, Suzuki J.
  • Journal Title: J Vet Med Sci. Volume: -
  • NAID: 130005094672
  • Peer Reviewed / Open Access
• [Journal Article] Haploinsufficiency of the c-myc transcriptional repressor FIR, as a dominant negative-alternative splicing model, promoted p53-dependent T-cell acute lymphoblastic leukemia progression by activating Notch1. (2015)
  • Author(s): Matsushita K, Kitamura K, Rahmutulla B, Tanaka N, Ishige T, Satoh M, Hoshino T, Miyagi S, Mori T, Itoga S, Shimada H, Tomonaga T, Kito M, Nakajima-Takagi Y, Kubo S, Nakaseko C, Hatano M, Miki T, Matsuo M, Fukuyo M, Kaneda A, Iwama A, Nomura F.
  • Journal Title: Oncotarget. Volume: 6 Pages: 5102-5117
  • Peer Reviewed / Open Access
• [Journal Article] Serum cytokine and chemokine profiles in patients with chronic inflammatory demyelinating polyneuropathy. (2015)
  • Author(s): Beppu M, Sawai S, Misawa S, Sogawa K, Mori M, Ishige T, Satoh M, Nomura F, Kuwabara S.
  • Journal Title: J Neuroimmunol. Volume: 279 Pages: 7-10
  • Peer Reviewed
• [Journal Article] Increased cell proliferation and differential protein expression induced by low-level Er:YAG laser irradiation in human gingival fibroblasts: proteomic analysis. (2014)
  • Author(s): Ogita M, Tsuchida S, Aoki A, Satoh M, Kado S, Sawabe M, Nanbara H, Kobayashi H, Takeuchi Y, Mizutani K, Sasaki Y, Nomura F, Izumi Y.
  • Journal Title: Lasers Med Sci. Volume: -
  • Peer Reviewed
• [Journal Article] Nuclear accumulation of annexin A2 contributes to chromosomal instability by coilin-mediated centromere damage. (2014)
  • Author(s): Kazami T, Nie H, Satoh M, Kuga T, Matsushita K, Kawasaki N, Tomonaga T, Nomura F.
  • Journal Title: Oncogene. Volume: -
  • Peer Reviewed
• [Journal Article] How proteomic ApoE serotyping could impact Alzheimer's disease risk assessment: genetic testing by proteomics. (2014)
  • Author(s): Nishimura M, Satoh M, Matsushita K, Nomura F.
  • Journal Title: Expert Rev Proteomics. Volume: 11 Pages: 405-407
  • Peer Reviewed
• [Journal Article] Moesin is a possible target molecule for cytomegalovirus-related Guillain-Barré syndrome. (2014)
  • Author(s): Sawai S, Satoh M, Mori M, Misawa S, Sogawa K, Kazami T, Ishibashi M, Beppu M, Shibuya K, Ishige T, Sekiguchi Y, Noda K, Sato K, Matsushita K, Kodera Y, Nomura F, Kuwabara S.
  • Journal Title: Neurology. Volume: 83 Pages: 113-117
  • Peer Reviewed
• [Journal Article] Alternative splicing of FBP-interacting repressor coordinates c-Myc, P27Kip1/cyclinE and Ku86/XRCC5 expression as a molecular sensor for bleomycin-induced DNA damage pathway. (2014)
  • Author(s): Rahmutulla B, Matsushita K, Satoh M, Seimiya M, Tsuchida S, Kubo S, Shimada H, Ohtsuka M, Miyazaki M, Nomura F.
  • Journal Title: Oncotarget. Volume: 5 Pages: 2404-2417
  • Peer Reviewed / Open Access
• [Journal Article] Direct application of MALDI-TOF mass spectrometry to cerebrospinal fluid for rapid pathogen identification in a patient with bacterial meningitis. (2014)
  • Author(s): Segawa S, Sawai S, Murata S, Nishimura M, Beppu M, Sogawa K, Watanabe M, Satoh M, Matsutani T, Kobayashi M, Iwadate Y, Kuwabara S, Saeki N, Nomura F.
  • Journal Title: Clin Chim Acta. Volume: 435 Pages: 59-61
  • Peer Reviewed
• [Journal Article] Human apolipoprotein e resequencing by proteomic analysis and its application to serotyping. (2014)
  • Author(s): Nishimura M, Satoh M, Nishimura S, Kakinuma S, Sato K, Sawai S, Tsuchida S, Kazama T, Matsushita K, Kado S, Kodera Y, Nomura F.
  • Journal Title: PLoS One Volume: 9(1)
  • Peer Reviewed
• [Journal Article] Glucoamylase is a major allergen of Schizophyllum commune. (2014)
  • Author(s): Toyotome T, Satoh M, Yahiro M, Watanabe A, Nomura F, Kamei K.
  • Journal Title: Clin Exp Allergy Volume: 44(3) Pages: 450-457
  • Peer Reviewed
• [Journal Article] Increased concentrations of apo A-I and apo A-II fragments in the serum of patients with hepatocellular carcinoma by magnetic beads-assisted MALDI-TOF mass spectrometry. (2014)
  • Author(s): Liu Y, Sogawa K, Sunaga M, Umemura H, Satoh M, Kazami T, Yoshikawa M, Tomonaga T, Yokosuka O, Nomura F.
  • Journal Title: Am J Clin Pathol Volume: 1(1) Pages: 52-61
  • Peer Reviewed
• [Journal Article] The FLS (fatty liver Shionogi) mouse reveals local expressions of lipocalin-2, CXCL1 and CXCL9 in the liver with non-alcoholic steatohepatitis. (2013)
  • Author(s): Semba T, Nishimura M, Nishimura S, Ohara O, Ishige T, Ohno S, Nonaka K, Sogawa K, Satoh M, Sawai S, Matsushita K, Imazeki F, Yokosuka O, Nomura F.
  • Journal Title: BMC Gastroenterol Volume: 13 Pages: 120-120
  • Peer Reviewed
• [Journal Article] Application of quantitative proteomic analysis using tandem mass tags for discovery and identification of novel biomarkers in periodontal disease. (2013)
  • Author(s): Tsuchida S, Satoh M, Kawashima Y, Sogawa K, Kado S, Sawai S, Nishimura M, Ogita M, Takeuchi Y, Kobyashi H, Aoki A, Kodera Y, Matsushita K, Izumi Y, Nomura F.
  • Journal Title: Proteomics Volume: 13(15) Pages: 2339-50
  • Peer Reviewed
• [Journal Article] Prohibitin and its rapidly emerging role as a biomarker of systemic malignancies-reply. (2013)
  • Author(s): Guo F, Hiroshima K, Wu D, Satoh M, Abulazi M, Nomura F, Yoshino I, Tomonaga T, Nakatani Y
  • Journal Title: Hum Pathol Volume: 44(4) Pages: 679-80
  • Peer Reviewed
• [Journal Article] Overexpression of hydroxymethylglutaryl CoA synthase 2 and 2,4-dienoyl-CoA reductase in rat pancreas following chronic alcohol consumption. (2013)
  • Author(s): Satoh M, Haruta-Satoh E, Yamada M, Kado S, Nomura F
  • Journal Title: Pancreas. Volume: 42(3) Pages: 475-82
  • Peer Reviewed
• [Journal Article] Establishment and application of a high-quality comparative analysis strategy for the discovery and small-scale validation of low-abundance biomarker peptides in serum based on an optimized novel peptide extraction method. (2013)
  • Author(s): Sasito T, Kawashima Y, Minamida S, Matsumoto K, Araki K, Matsui T, Satoh M, Nomura F, Iwamura M, Maeda T, Kodera Y
  • Journal Title: Journal of Electrophoresis. Volume: 57 Pages: 1-9
  • NAID: 130003364779
  • Peer Reviewed
• [Journal Article] Cyclic sample pooling using two-dimensional liquid chromatography system enhances coverage in shotgun proteomics (2013)
  • Author(s): Kawashiraa Y, Satoh M, Saito T, Matsui T, Nomura F, Matsumoto H, Kodera Y
  • Journal Title: Biomedical Chromatography Volume: 27 Issue: 6 Pages: 691-694
  • DOI: 10.1002/bmc.2864
  • Peer Reviewed
• [Journal Article] Enhanced recovery of lyophilized peptides in shotgun proteomics by using an LC-ESI-MS compatible surfactant (2013)
  • Author(s): Kawashima Y, Takahashi N, Satoh M, Saito T, Kado S, Nomura F, Matsumoto H, Kodera Y
  • Journal Title: Proteomics Volume: 13 Issue: 5 Pages: 751-755
  • DOI: 10.1002/pmic.201200462
  • Peer Reviewed
• [Journal Article] Combined proteomic analysis of liver tissue and serum in chronically alcohol-fed rats (2013)
  • Author(s): Yamada M, Satoh M, Seimiya M, Sogawa K, Itoga S, Tomonaga T, Nomura F
  • Journal Title: Alcohol Clin Exp Res Volume: 37 Issue: s1
  • DOI: 10.1111/j.1530-0277.2012.01883.x
  • Peer Reviewed
• [Journal Article] Preanalytical evaluation of serum 25-hydroxyvitamin D3 and 25-hydroxyvitamin D2 measurements using LC-MS/MS. (2013)
  • Author(s): Mochizuki A, Kodera Y, Saito T, Satoh M, Sogawa K, Nishimura M, Seimiya M,Kubota M, Nomura F.
  • Journal Title: Clin Chim Acta. Volume: 420 Pages: 114-120
  • DOI: 10.1016/j.cca.2012.10.041
  • Peer Reviewed
• [Journal Article] The application of a three-step serum proteome analysis for the discovery and identification of novel biomarkers of hepatocellular carcinoma. (2012)
  • Author(s): Kimura A, Sogawa K, Satoh M, Kodera Y, Yokosuka O, Tomonaga T, Nomura F.
  • Journal Title: Int J Proteomics. Volume: 2012: Pages: 623190-623190
  • DOI: 10.1155/2012/623190
  • Peer Reviewed
• [Journal Article] Identification of a Novel Biomarker for Biliary Tract Cancer Using Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry. (2012)
  • Author(s): Kikkawa S, Sogawa K, Satoh M, Umemura H, Kodera Y, Matsushita K, Tomonaga T, Miyazaki M, Yokosuka O, Nomura F.
  • Journal Title: Int J Proteomics Volume: 108 Pages: 609-609
  • DOI: 10.1155/2012/108609
  • Peer Reviewed
• [Journal Article] Low-density lipoprotein receptorrelated protein-1 (LRP-1) mediates autophagy and apoptosis caused by Helicobacter pyloriVacA (2012)
  • Author(s): Yahiro, K., Suzuki, H., et al.
  • Journal Title: J. Biol. Chem Volume: 287(37) Issue: 37 Pages: 31104-15
  • DOI: 10.1074/jbc.m112.387498
  • Peer Reviewed
• [Journal Article] Akt/mTOR signaling pathway is crucial for gemcitabine resistance induced by Annexin II in pancreatic cancer cells (2012)
  • Author(s): Kagawa S, Takano S, Yoshitomi H, Kimura F, Satoh M, Shimizu H, Yoshidome H, Ohtsuka M, Kato A, Furukawa K, Matsushita K, Nomura F, Miyazaki M
  • Journal Title: T Surg Res Volume: 178 Issue: 2 Pages: 758-767
  • DOI: 10.1016/j.jss.2012.05.065
  • Peer Reviewed
• [Journal Article] Proteomic analysis of gingival crevicular fluid for discovery of novel periodontal disease markers. (2012)
  • Author(s): Tsuchida S
  • Journal Title: Proteomics. Volume: 12(13) Issue: 13 Pages: 2190-202
  • DOI: 10.1002/pmic.201100655
  • Peer Reviewed
• [Journal Article] SAP155-mediated splicing of FUSE-binding protein-interacting repressor serves as a molecular switch for c-myc gene expression (2012)
  • Author(s): Matsushita, K., Kajiwara, T., Tamura, M., Satoh, M., Tanaka, N., Tomonaga, T., Matsubara, H., Shimada, H., Yoshimoto, R., Ito, A., Kubo, S., Natsume, T., Levens, D., Yoshida, M., and Nomura, F
  • Journal Title: Mol. Cancer Res Volume: 10 Issue: 6 Pages: 787-799
  • DOI: 10.1158/1541-7786.mcr-11-0462
  • Peer Reviewed
• [Journal Article] Prohibitin in squamous cell carcinoma of the lung: its expression and possible clinical significance. (2012)
  • Author(s): Guo F, Hiroshima K, Wu D, Satoh M, Abulazi M, Yoshino I, Tomonaga T, Nomura F, Nakatani Y.
  • Journal Title: Hum Pathol. Volume: 43(8) Issue: 8 Pages: 1282-1288
  • DOI: 10.1016/j.humpath.2011.10.006
  • Peer Reviewed
• [Presentation] 質量分析の基礎知識 (2015)
  • Author(s): 佐藤守
  • Organizer: 平成26 年度 高度先端的医学セミナー 質量分析に関する研修会
  • Place of Presentation: 東京都 日本臨床検査技師会館
  • Year and Date: 2015-03-14 – 2015-03-15
  • Invited
• [Presentation] DAPTAD誘導体化を用いたLC-MS/MSによる血清ビタミンD代謝物の４項目同時定量 (2014)
  • Author(s): 佐藤守、石毛崇之、小川祥二郎、西村基、松下一之、東達也、野村文夫
  • Organizer: 第39 回 日本医用マススペクトル学会年会
  • Place of Presentation: 千葉県 三井ガーデンホテル千葉
  • Year and Date: 2014-10-16 – 2014-10-17
• [Presentation] イムノアフィニティー抽出DAPTAD誘導体化LC-MS/MSによる血清25OH-ビタミンDおよび1,25（OH）2-ビタミン Dの同時定量 (2014)
  • Author(s): 石毛崇之、佐藤守、小川祥二郎、西村基、土田祥央、澤井摂、松下一之、東達也、野村文夫
  • Organizer: 第39 回 日本医用マススペクトル学会年会
  • Place of Presentation: 千葉県 三井ガーデンホテル千葉
  • Year and Date: 2014-10-16 – 2014-10-17
• [Presentation] 前田浩人、前川武大、柴田悠衣、阿部沙織、堀江和香、望月俊輔、渡辺俊文、佐藤守、三田明弘、野村文夫、曽川一幸 (2014)
  • Author(s): ネコ慢性腎不全における尿を用いた早期診断マーカーの探索
  • Organizer: 第39 回 日本医用マススペクトル学会年会
  • Place of Presentation: 千葉県 三井ガーデンホテル千葉
  • Year and Date: 2014-10-16 – 2014-10-17
• [Presentation] キウイフルーツアレルギーにおけるアレルゲンタンパク質の解析 (2014)
  • Author(s): 河原彩花、曽川一幸、佐藤守、三田明弘、下条直樹、野村文夫
  • Organizer: 第39 回 日本医用マススペクトル学会年会
  • Place of Presentation: 千葉県 三井ガーデンホテル千葉
  • Year and Date: 2014-10-16 – 2014-10-17
• [Presentation] ナッツアレルギーにおけるアレルゲンタンパク質の解析 (2014)
  • Author(s): 安東圭奈美、曽川一幸、佐藤守、三田明弘、下条直樹、野村文夫
  • Organizer: 第39 回 日本医用マススペクトル学会年会
  • Place of Presentation: 千葉県 三井ガーデンホテル千葉
  • Year and Date: 2014-10-16 – 2014-10-17
• [Presentation] プロテオーム解析を用いた歯周病疾患マーカー探索 (2014)
  • Author(s): 土田祥央、佐藤守、曽川一幸、川島祐介、石毛崇之､別府美奈子､澤井摂、西村基、小寺義男、松下一之、野村文夫
  • Organizer: 第39 回 日本医用マススペクトル学会年会
  • Place of Presentation: 千葉県 三井ガーデンホテル千葉
  • Year and Date: 2014-10-16 – 2014-10-17
• [Presentation] 質量分析計を活用したがん培養細胞の分泌物解析とサイトケラチン断片の探索 (2014)
  • Author(s): 陳健嘉、西村基、西村里美、石毛崇之、佐藤守、松下一之、野村文夫
  • Organizer: 第39 回 日本医用マススペクトル学会年会
  • Place of Presentation: 千葉県 三井ガーデンホテル千葉
  • Year and Date: 2014-10-16 – 2014-10-17
• [Presentation] MALDI-TOF MSを用いた細菌性髄膜炎の起因菌迅速同定 (2014)
  • Author(s): 別府美奈子、澤井摂、瀬川俊介、村田正太、曽川一幸、渡邊正治、西村基、佐藤守、松下一之、野村文夫
  • Organizer: 第39 回 日本医用マススペクトル学会年会
  • Place of Presentation: 千葉県 三井ガーデンホテル千葉
  • Year and Date: 2014-10-16 – 2014-10-17
• [Presentation] Interactions between SF3b1(SAP155) and FUSE-binding protein-interacting repressor (FIR) links c-Myc and P27Kip1 expression through GeLC-MS analysis (2014)
  • Author(s): 松下一之、佐藤守、朝長毅、野村文夫
  • Organizer: 日本プロテオーム学会2014年会
  • Place of Presentation: 茨城県 つくば国際会議場
  • Year and Date: 2014-07-17 – 2014-07-18
• [Presentation] Differential solublization method to extract low-molecular-weight proteins/peptides for successful serum SRM analyses. (2013)
  • Author(s): Satoh M, Kawashima Y, Saito T, Ishibashi M, Sogawa K, Kado S, Kodera Y, Nomura F
  • Organizer: HUPO
  • Place of Presentation: Yokohama
• [Presentation] プロテオーム解析から臨床検査の現場を目指して (2013)
  • Author(s): 佐藤守、川島祐介、齋藤達也、石橋真澄、曽川一幸、荷堂清香、小寺義男、野村文夫
  • Organizer: 日本医用マススペクトル学会
  • Place of Presentation: 神戸市産業振興センター
• [Presentation] Differential solublization method to extract low-molecular-weight proteins/peptides for successful serum SRM analyses (2013)
  • Author(s): Mamoru Satoh, Yusuke Kawashima, Tatsuya Saito, Kazuyuki Sogawa, Sayaka Kado, Yoshio Kodera, Fumio Nomura
  • Organizer: The 5th Annual Conference on Mass Spectometry Applications to the Clinical Lab 2013
  • Place of Presentation: San Diego, USA
• [Presentation] ApoE typing from small amounts of blood samples (2013)
  • Author(s): Motoi Nishimura, Satomi Nishimura, Mamoru Satoh, Syouko Kakinuma, Kenichi Satoh, Setsu Sawai, Sayaka Kado, Yoshio Kodera, Fumio Nomura
  • Organizer: The 5th Annual Conference on Mass Spectometry Applications to the Clinical Lab 2013
  • Place of Presentation: San Diego, USA
• [Presentation] Effects of preanalytical factors on serum 25-hydroxyvitamin D3 and 25-hydroxyvitamin D2 measurements using LC-MS/MS for the clinical laboratory testing (2013)
  • Author(s): Ishige T, Mochizuki A, Kodera Y, Saito T, Satoh M, Sogawa K, Nishimura M, Sawai S, Seimiya M, Kubota M, Matsushita K, Nomura F
  • Organizer: The 5th Annual Conference on Mass Spectometry Applications to the Clinical Lab 2013
  • Place of Presentation: San Diego, USA
• [Presentation] 診断応用を目指した血清の定量的プロテオミクス (2012)
  • Author(s): 佐藤守、川島祐介、曽川一幸、齋藤達也、荷堂清香、小寺義男、野村文夫
  • Organizer: 第37回日本医用マススペクトル学会年会
  • Place of Presentation: 名古屋
  • Invited
• [Presentation] 血清よりの Apolipoprotein E (ApoE) タイピング (2012)
  • Author(s): 西村基、西村里美、佐藤守、柿沼翔子、佐藤謙一、澤井摂、 荷堂清香、小寺義男、野村文夫
  • Organizer: 第37回日本医用マススペクトル学会年会
  • Place of Presentation: 名古屋
• [Presentation] ジェムシタビン耐性ヒト膵癌細胞株のプロテオーム解析 (2012)
  • Author(s): 佐藤守
  • Organizer: 第63回日本電気泳動学会総会
  • Place of Presentation: 沖縄
  • Invited
• [Presentation] 慢性的アルコール投与ラット膵臓のプロテオーム解析 (2012)
  • Author(s): 佐藤守、佐藤（春田）恵里、山田真子、荷堂清香、野村文夫
  • Organizer: 第19 回日本遺伝子診療学会
  • Place of Presentation: 千葉
• [Book] 質量分析の消化器疾患の診療・研究への応用 第2回 MS解析に用いられる機器 (2012)
  • Author(s): 佐藤守、曽川一幸、野村文夫
  • Total Pages: 6
  • Publisher: 先端医学社
• [Book] 総説 定量的質量分析法の最近の進歩 (2012)
  • Author(s): 佐藤守、曽川一幸、野村文夫
  • Total Pages: 6
  • Publisher: 日本臨床自動化学会