title: Choline Kinase α Mediates Interactions Between the Epidermal Growth Factor Receptor and Mechanistic Target of Rapamycin Complex 2 in Hepatocellular Carcinoma Cells to Promote Drug Resistance and Xenograft Tumor Progression.
abstract: Choline kinase α (CHKA) catalyzes conversion of choline to phosphocholine and can contribute to carcinogenesis. Little is known about the role of CHKA in the pathogenesis of hepatocellular carcinoma (HCC).
  
  
  
  We performed whole-exome and transcriptome sequence analyses of 9 paired HCC and non-tumor-adjacent tissues. We performed tissue chip analyses of 120 primary HCC and non-tumor-adjacent tissues from patients who received surgery in Shanghai, China from January 2006 through December 2009; 48 sets of specimens (HCC and non-tumor-adjacent tissues) were also analyzed. CHKA gene copy number was quantified and findings were validated by quantitative reverse transcription polymerase chain reaction analysis. CHKA messenger RNA and protein levels were determined by polymerase chain reaction, immunohistochemical, and immunoblot analyses. CHKA was examined in 2 hepatocyte cell lines and 7 HCC-derived cell lines, and knocked down with small interfering RNAs in 3 HCC cell lines. Cells were analyzed in proliferation, wound healing, migration, and invasion assays. Cells were injected into tail veins of mice and tumor growth and metastasis were quantified. Immunoprecipitation and immunofluorescence assays were conducted to determine interactions between CHKA and the epidermal growth factor receptor (EGFR) and the mechanistic target of rapamycin complex 2.
  
  
  
  Levels of CHKA messenger RNA were frequently increased in HCC tissues compared with nontumor tissues; increased expression was associated with amplification at the CHKA loci. Tumors that expressed high levels of CHKA had more aggressive phenotypes, and patients with these tumors had shorter survival times after surgery compared to patients whose tumors expressed low levels of CHKA. HCC cell lines that stably overexpressed CHKA had higher levels of migration and invasion than control HCC cells, and formed larger xenograft tumors with more metastases in mice compared to HCC cells that did not overexpress CHKA. CHKA was required for physical interaction between EGFR and mechanistic target of rapamycin complex 2. This complex was required for HCC cells to form metastatic xenograft tumors in mice and to become resistant to EGFR inhibitors.
  
  
  
  We found levels of CHKA to be increased in human HCCs compared to nontumor tissues, and increased expression to be associated with tumor aggressiveness and reduced survival times of patients. Overexpression of CHKA in HCC cell lines increased their invasiveness, resistance to EGFR inhibitors, and ability to form metastatic tumors in mice by promoting interaction of EGFR with mechanistic target of rapamycin complex 2.
authors.affiliation: International Co-operation Laboratory on Signal Transduction, Eastern Hepatobiliary Surgery Institute, Second Military Medical University, Shanghai, China; Department of Hepatic Surgery, Eastern Hepatobiliary Surgery Hospital, Shanghai, China.
authors.forename: Xi-Meng
authors.initials: XM
authors.lastname: Lin
authors.affiliation: International Co-operation Laboratory on Signal Transduction, Eastern Hepatobiliary Surgery Institute, Second Military Medical University, Shanghai, China; Anal-Colorectal Surgery Institute, 150th Hospital of PLA, Luoyang, China.
authors.forename: Liang
authors.initials: L
authors.lastname: Hu
authors.affiliation: Tsinghua National Laboratory for Information Science and Technology, Bioinformatics Division, Synthetic and Systems Biology Center, Department of Automation, Tsinghua University, Beijing, China.
authors.forename: Jin
authors.initials: J
authors.lastname: Gu
authors.affiliation: Department of Hepatic Surgery, Eastern Hepatobiliary Surgery Hospital, Shanghai, China.
authors.forename: Ruo-Yu
authors.initials: RY
authors.lastname: Wang
authors.affiliation: International Co-operation Laboratory on Signal Transduction, Eastern Hepatobiliary Surgery Institute, Second Military Medical University, Shanghai, China; National Center for Liver Cancer, Shanghai, China.
authors.forename: Liang
authors.initials: L
authors.lastname: Li
authors.affiliation: Department of Neurosurgery, Wuhan General Hospital of Guangzhou Command, Wuhan, China.
authors.forename: Jing
authors.initials: J
authors.lastname: Tang
authors.affiliation: Department of Hepatic Surgery, Eastern Hepatobiliary Surgery Hospital, Shanghai, China.
authors.forename: Bao-Hua
authors.initials: BH
authors.lastname: Zhang
authors.affiliation: Department of Hepatic Surgery, Eastern Hepatobiliary Surgery Hospital, Shanghai, China.
authors.forename: Xing-Zhou
authors.initials: XZ
authors.lastname: Yan
authors.affiliation: International Co-operation Laboratory on Signal Transduction, Eastern Hepatobiliary Surgery Institute, Second Military Medical University, Shanghai, China; National Center for Liver Cancer, Shanghai, China.
authors.forename: Yan-Jing
authors.initials: YJ
authors.lastname: Zhu
authors.affiliation: International Co-operation Laboratory on Signal Transduction, Eastern Hepatobiliary Surgery Institute, Second Military Medical University, Shanghai, China.
authors.forename: Cong-Li
authors.initials: CL
authors.lastname: Hu
authors.affiliation: Department of Hepatic Surgery, Eastern Hepatobiliary Surgery Hospital, Shanghai, China.
authors.forename: Wei-Ping
authors.initials: WP
authors.lastname: Zhou
authors.affiliation: MOE Key Laboratory of Bioinformatics and Bioinformatics Division, Tsinghua National Laboratory for Information Science and Technology, Department of Automation, Tsinghua University, Beijing, China.
authors.forename: Shao
authors.initials: S
authors.lastname: Li
authors.affiliation: Mengchao Hepatobiliary Hospital, Fujian Medical University, Fuzhou, China.
authors.forename: Jing-Feng
authors.initials: JF
authors.lastname: Liu
authors.affiliation: Laboratory of Metabolism, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland.
authors.forename: Frank J
authors.initials: FJ
authors.lastname: Gonzalez
authors.affiliation: Department of Hepatic Surgery, Eastern Hepatobiliary Surgery Hospital, Shanghai, China.
authors.forename: Meng-Chao
authors.initials: MC
authors.lastname: Wu
authors.affiliation: International Co-operation Laboratory on Signal Transduction, Eastern Hepatobiliary Surgery Institute, Second Military Medical University, Shanghai, China; National Center for Liver Cancer, Shanghai, China. Electronic address: hywangk@vip.sina.com.
authors.forename: Hong-Yang
authors.initials: HY
authors.lastname: Wang
authors.affiliation: International Co-operation Laboratory on Signal Transduction, Eastern Hepatobiliary Surgery Institute, Second Military Medical University, Shanghai, China; National Center for Liver Cancer, Shanghai, China; Mengchao Hepatobiliary Hospital, Fujian Medical University, Fuzhou, China; Laboratory of Metabolism, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland. Electronic address: chenlei@smmu.edu.cn.
authors.forename: Lei
authors.initials: L
authors.lastname: Chen
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy / Q000378:metabolism* / Q000473:pathology; D002460:Cell Line; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000235:genetics; D002797:Choline Kinase / Q000235:genetics* / Q000378:metabolism; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D066246:ErbB Receptors / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000069347:Erlotinib Hydrochloride / Q000494:pharmacology; D000077156:Gefitinib; D056945:Hep G2 Cells; D006801:Humans; D015151:Immunoblotting; D007150:Immunohistochemistry; D008113:Liver Neoplasms / Q000188:drug therapy / Q000378:metabolism* / Q000473:pathology; D000076225:Mechanistic Target of Rapamycin Complex 2; D051379:Mice; D046912:Multiprotein Complexes / Q000378:metabolism*; D009361:Neoplasm Invasiveness / Q000235:genetics; D009368:Neoplasm Transplantation; D011799:Quinazolines / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism*; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism*; D014945:Wound Healing / Q000235:genetics; D023041:Xenograft Model Antitumor Assays
chemical_list: D000970:Antineoplastic Agents; D046912:Multiprotein Complexes; D011799:Quinazolines; D012333:RNA, Messenger; D000069347:Erlotinib Hydrochloride; C517212:CHKA protein, human; D002797:Choline Kinase; D066246:ErbB Receptors; D000076225:Mechanistic Target of Rapamycin Complex 2; D058570:TOR Serine-Threonine Kinases; D000077156:Gefitinib
keywords: Liver Cancer; Oncogene; Signal Transduction; Tumorigenesis