title: Stabilization of Notch1 by the Hsp90 Chaperone Is Crucial for T-Cell Leukemogenesis.
abstract: Purpose:Notch1 deregulation is assuming a focal role in T-cell acute lymphoblastic leukemia (T-ALL). Despite tremendous advances in our understanding of Notch1 transcriptional programs, the mechanisms by which Notch1 stability and turnover are regulated remain obscure. The goal of the current study is to identify intracellular Notch1 (ICN1, the activated form of Notch1) binding partner(s) regulating its stability and activity.Experimental Design:We employed immunoaffinity purification to identify ICN1-associating partner(s) and used coimmunoprecipitation to verify the endogenous protein interaction. Pharmacologic or short hairpin RNA-mediated inhibition was applied in loss-of-function assays to assess the role of tentative binding partner(s) in modulating ICN1 protein stability as well as affecting T-ALL cell expansionin vitroandin vivoMechanistic analysis involved protein degradation and polyubiquitination assays.Results:We identify the Hsp90 chaperone as a direct ICN1-binding partner essential for its stabilization and transcriptional activity. T-ALL cells exhibit constitutive endogenous ICN1-Hsp90 interaction and Hsp90 depletion markedly decreases ICN1 levels. The Hsp90-associated E3 ubiquitin ligase Stub1 mediates the ensuring proteasome-dependent ICN1 degradation. Administration of 17-AAG or PU-H71, two distinct Hsp90 inhibitors, depletes ICN1, inhibits T-ALL cell proliferation, and triggers dramatic apoptotic cell death. Systemic treatment with PU-H71 reduces ICN1 expression and profoundly inhibits murine T-ALL allografts as well as human T-ALL xenografts.Conclusions:Our findings demonstrate Hsp90 blockade leads to ICN1 destabilization, providing an alternative strategy to antagonize oncogenic Notch1 signaling with Hsp90-selective inhibitors.Clin Cancer Res; 23(14); 3834-46. ©2017 AACR.
authors.affiliation: Zhongnan Hospital of Wuhan University, Wuhan, PR China.|Medical Research Institute, Wuhan University, Wuhan, PR China. qingguoliang@whu.edu.cn.|School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, PR China.
authors.forename: Zhaojing
authors.initials: Z
authors.lastname: Wang
authors.affiliation: School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, PR China.
authors.forename: Yufeng
authors.initials: Y
authors.lastname: Hu
authors.affiliation: Medical Research Institute, Wuhan University, Wuhan, PR China. qingguoliang@whu.edu.cn.
authors.forename: Daibiao
authors.initials: D
authors.lastname: Xiao
authors.affiliation: Medical Research Institute, Wuhan University, Wuhan, PR China. qingguoliang@whu.edu.cn.
authors.forename: Jingchao
authors.initials: J
authors.lastname: Wang
authors.affiliation: School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, PR China.
authors.forename: Chuntao
authors.initials: C
authors.lastname: Liu
authors.affiliation: Protein Facility, Center of Biomedical Analysis, Tsinghua University, Beijing, PR China.
authors.forename: Yisheng
authors.initials: Y
authors.lastname: Xu
authors.affiliation: Protein Facility, Center of Biomedical Analysis, Tsinghua University, Beijing, PR China.
authors.forename: Xiaomeng
authors.initials: X
authors.lastname: Shi
authors.affiliation: School of Life Sciences, Tsinghua University, Beijing, PR China.
authors.forename: Peng
authors.initials: P
authors.lastname: Jiang
authors.affiliation: Department of Hematology, Affiliated Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, PR China.
authors.forename: Liang
authors.initials: L
authors.lastname: Huang
authors.affiliation: South China Institute for Stem Cell Biology and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, PR China.
authors.forename: Peng
authors.initials: P
authors.lastname: Li
authors.affiliation: Zhongnan Hospital of Wuhan University, Wuhan, PR China.|Medical Research Institute, Wuhan University, Wuhan, PR China. hudanliu@whu.edu.cn.
authors.forename: Hudan
authors.initials: H
authors.lastname: Liu
authors.affiliation: Zhongnan Hospital of Wuhan University, Wuhan, PR China.|Medical Research Institute, Wuhan University, Wuhan, PR China. qingguoliang@whu.edu.cn.
authors.forename: Guoliang
authors.initials: G
authors.lastname: Qing
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D016227:Benzoquinones / Q000008:administration & dosage; D063646:Carcinogenesis / Q000187:drug effects*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D018841:HSP90 Heat-Shock Proteins / Q000037:antagonists & inhibitors / Q000235:genetics*; D006801:Humans; D047029:Lactams, Macrocyclic / Q000008:administration & dosage; D051379:Mice; D018832:Molecular Chaperones / Q000235:genetics; D054218:Precursor T-Cell Lymphoblastic Leukemia-Lymphoma / Q000188:drug therapy* / Q000235:genetics / Q000473:pathology; D011485:Protein Binding; D060066:Protein Interaction Maps / Q000187:drug effects / Q000235:genetics; D059748:Proteolysis / Q000187:drug effects; D051881:Receptor, Notch1 / Q000037:antagonists & inhibitors / Q000235:genetics*; D015398:Signal Transduction / Q000187:drug effects; D023041:Xenograft Model Antitumor Assays
chemical_list: D016227:Benzoquinones; D018841:HSP90 Heat-Shock Proteins; D047029:Lactams, Macrocyclic; D018832:Molecular Chaperones; C497761:NOTCH1 protein, human; D051881:Receptor, Notch1; C112765:tanespimycin