title: Alpha-secretase inhibition reduces human glioblastoma stem cell growth in vitro and in vivo by inhibiting Notch.
abstract: The Notch pathway is dysregulated and a potential target in glioblastoma multiforme (GBM). Currently available Notch inhibitors block γ-secretase, which is necessary for Notch processing. However, Notch is first cleaved by α-secretase outside the plasma membrane, via a disintegrin and metalloproteinase-10 and -17. In this work, we used a potent α-secretase inhibitor (ASI) to test inhibition of glioblastoma growth and inhibition of Notch and of both novel and known Notch targets. Featured in this study are luciferase reporter assays and immunoblot, microarray analysis, chromatin immunoprecipitation (ChIP), quantitative real-time PCR, cell number assay, bromodeoxyuridine incorporation, plasmid rescue, orthotopic xenograft model, and local delivery of treatment with convection-enhanced delivery using nanoparticles, as well as survival, MRI, and ex vivo luciferase assay. A CBF1-luciferase reporter assay as well as an immunoblot of endogenous Notch revealed Notch inhibition by the ASI. Microarray analysis, quantitative real-time PCR, and ChIP of ASI and γ-secretase inhibitor (GSI) treatment of GBM cells identified known Notch pathway targets, as well as novel Notch targets, including YKL-40 and leukemia inhibitory factor. Finally, we found that local nanoparticle delivery of ASIs but not GSIs increased survival time significantly in a GBM stem cell xenograft treatment model, and ASI treatment resulted in decreased tumor size and Notch activity. This work indicates α-secretase as an alternative to γ-secretase for inhibition of Notch in GBM and possibly other cancers as well, and it identifies novel Notch targets with biologic relevance and potential as biomarkers.
authors.affiliation: Division of Neuro-Oncology, Neurology Department, University of Virginia Health System, Old Medical School-Room 4814, 21 Hospital Drive, Charlottesville, VA 22908, USA.
authors.forename: Desiree H
authors.initials: DH
authors.lastname: Floyd
authors.forename: Benjamin
authors.initials: B
authors.lastname: Kefas
authors.forename: Oleksandr
authors.initials: O
authors.lastname: Seleverstov
authors.forename: Olga
authors.initials: O
authors.lastname: Mykhaylyk
authors.forename: Charli
authors.initials: C
authors.lastname: Dominguez
authors.forename: Laurey
authors.initials: L
authors.lastname: Comeau
authors.forename: Christian
authors.initials: C
authors.lastname: Plank
authors.forename: Benjamin
authors.initials: B
authors.lastname: Purow
mesh_terms: D053829:Amyloid Precursor Protein Secretases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D000818:Animals; D017209:Apoptosis; D014408:Biomarkers, Tumor / Q000235:genetics / Q000378:metabolism; D015153:Blotting, Western; D001932:Brain Neoplasms / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology*; D002453:Cell Cycle; D049109:Cell Proliferation; D047369:Chromatin Immunoprecipitation; D020869:Gene Expression Profiling; D005909:Glioblastoma / Q000188:drug therapy / Q000378:metabolism / Q000473:pathology*; D006801:Humans; D066298:In Vitro Techniques; D008156:Luciferases / Q000378:metabolism; D008280:Magnetics; D051379:Mice; D008807:Mice, Inbred BALB C; D053758:Nanoparticles; D014411:Neoplastic Stem Cells / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D020411:Oligonucleotide Array Sequence Analysis; D010880:Piperidines / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D051880:Receptors, Notch / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013141:Spiro Compounds / Q000494:pharmacology; D014407:Tumor Cells, Cultured; D023041:Xenograft Model Antitumor Assays
chemical_list: D014408:Biomarkers, Tumor; C513090:INCB3619; D010880:Piperidines; D012333:RNA, Messenger; D051880:Receptors, Notch; D013141:Spiro Compounds; D008156:Luciferases; D053829:Amyloid Precursor Protein Secretases