title: Novel anti-apoptotic microRNAs 582-5p and 363 promote human glioblastoma stem cell survival via direct inhibition of caspase 3, caspase 9, and Bim.
abstract: Glioblastoma is the most common and lethal primary brain tumor. Tumor initiation and recurrence are likely caused by a sub-population of glioblastoma stem cells, which may derive from mutated neural stem and precursor cells. Since CD133 is a stem cell marker for both normal brain and glioblastoma, and to better understand glioblastoma formation and recurrence, we looked for dys-regulated microRNAs in human CD133+ glioblastoma stem cells as opposed to CD133+ neural stem cells isolated from normal human brain. Using FACS sorting of low-passage cell samples followed by microRNA microarray analysis, we found 43 microRNAs that were dys-regulated in common in three separate CD133+ human glioblastomas compared to CD133+ normal neural stem cells. Among these were several microRNAs not previously associated with cancer. We then verified the microRNAs dys-regulated in glioblastoma using quantitative real time PCR and Taqman analysis of the original samples, as well as human GBM stem cell and established cell lines and many human specimens. We show that two candidate oncogenic microRNAs, miR-363 and miR-582-5p, can positively influence glioblastoma survival, as shown by forced expression of the microRNAs and their inhibitors followed by cell number assay, Caspase 3/7 assay, Annexin V apoptosis/fluorescence activated cell sorting, siRNA rescue of microRNA inhibitor treatment, as well as 3'UTR mutagenesis to show luciferase reporter rescue of the most successful targets. miR-582-5p and miR-363 are shown to directly target Caspase 3, Caspase 9, and Bim.
authors.affiliation: Division of Neuro-Oncology, Departments of Neurology, Microbiology, and Biochemistry and Molecular Genetics, University of Virginia Health System, Charlottesville, Virginia, United States of America.
authors.forename: Desiree Hunt
authors.initials: DH
authors.lastname: Floyd
authors.affiliation: Division of Neuro-Oncology, Departments of Neurology, Microbiology, and Biochemistry and Molecular Genetics, University of Virginia Health System, Charlottesville, Virginia, United States of America.
authors.forename: Ying
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Division of Neuro-Oncology, Departments of Neurology, Microbiology, and Biochemistry and Molecular Genetics, University of Virginia Health System, Charlottesville, Virginia, United States of America.
authors.forename: Bijan K
authors.initials: BK
authors.lastname: Dey
authors.affiliation: Division of Neuro-Oncology, Departments of Neurology, Microbiology, and Biochemistry and Molecular Genetics, University of Virginia Health System, Charlottesville, Virginia, United States of America.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Kefas
authors.affiliation: Division of Neuro-Oncology, Departments of Neurology, Microbiology, and Biochemistry and Molecular Genetics, University of Virginia Health System, Charlottesville, Virginia, United States of America.
authors.forename: Hannah
authors.initials: H
authors.lastname: Breit
authors.affiliation: Division of Neuro-Oncology, Departments of Neurology, Microbiology, and Biochemistry and Molecular Genetics, University of Virginia Health System, Charlottesville, Virginia, United States of America.
authors.forename: Kaitlyn
authors.initials: K
authors.lastname: Marks
authors.affiliation: Division of Neuro-Oncology, Departments of Neurology, Microbiology, and Biochemistry and Molecular Genetics, University of Virginia Health System, Charlottesville, Virginia, United States of America.
authors.forename: Anindya
authors.initials: A
authors.lastname: Dutta
authors.affiliation: Division of Neurosurgical Research, Department of Neurosurgery, University of Heidelberg, Heidelberg, Germany.
authors.forename: Christel
authors.initials: C
authors.lastname: Herold-Mende
authors.affiliation: Charité University Clinics, Clinic for Neurosurgery, Berlin, Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Synowitz
authors.affiliation: Neurosurgical Research, University Clinics Munich, München, Germany.
authors.forename: Rainer
authors.initials: R
authors.lastname: Glass
authors.affiliation: Division of Neuro-Oncology, Departments of Neurology, Microbiology, and Biochemistry and Molecular Genetics, University of Virginia Health System, Charlottesville, Virginia, United States of America.
authors.forename: Roger
authors.initials: R
authors.lastname: Abounader
authors.affiliation: Division of Neuro-Oncology, Departments of Neurology, Microbiology, and Biochemistry and Molecular Genetics, University of Virginia Health System, Charlottesville, Virginia, United States of America.
authors.forename: Benjamin W
authors.initials: BW
authors.lastname: Purow
mesh_terms: D020413:3' Untranslated Regions / Q000235:genetics*; D000071916:AC133 Antigen; D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism; D051017:Apoptosis Regulatory Proteins / Q000235:genetics / Q000378:metabolism*; D000072224:Bcl-2-Like Protein 11; D053148:Caspase 3 / Q000235:genetics / Q000378:metabolism*; D053453:Caspase 9 / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000235:genetics* / Q000502:physiology; D002478:Cells, Cultured; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic; D005909:Glioblastoma / Q000235:genetics* / Q000378:metabolism; D006023:Glycoproteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D014411:Neoplastic Stem Cells / Q000166:cytology* / Q000378:metabolism*; D010455:Peptides / Q000235:genetics / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D020413:3' Untranslated Regions; D000071916:AC133 Antigen; D015703:Antigens, CD; D051017:Apoptosis Regulatory Proteins; C000606775:BCL2L11 protein, human; D000072224:Bcl-2-Like Protein 11; D006023:Glycoproteins; D008565:Membrane Proteins; D035683:MicroRNAs; C000606023:PROM1 protein, human; D010455:Peptides; D011518:Proto-Oncogene Proteins; D053148:Caspase 3; D053453:Caspase 9