Micro RNA Research. Ken Kosik. Harriman Professor, Department of Molecular, Cellular & Developmental Biology and Biomolecular Sciences & Engr.
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1 Ken Kosik Harriman Professor, Department of Molecular, Cellular & Developmental Biology and Biomolecular Sciences & Engr. Program Co-Director, Neurosciences Research Institute Micro RNA Research
2 Neuroscience Research Institute Cancer Cell cycle Wilson, Jordan, Lew Cell fusion Rothman micrornas-kosik Synaptic Biology K channels Vandenberg Protein trafficking Ma, Weimbs, Parsons Synaptic adherens junction Kosik, Szumlinski Neurodevelopment & Evolution Visual system Clegg, Reese Neurogenesis Kippen, Smith Stem cells University consortium Programmed cell death Rothman Evolution of the visual system--oakley Neurodegeneration Retinal degeneration--fisher Macular Degeneration Johnson, Anderson Alzheimer s Feinstein, Wilson, Jordan, Kosik, Lew, Lal
3 The Central Dogma DNA RNA mirnas Protein
4 genome genome prokaryote eukaryote
5 microrna nucleotide non-coding transcripts derived from a ~70 nt stem-loop precursor 5 terminal monophosphate 3 -terminal hydroxyl
6 lin-4 Negatively Regulates lin-14 m7gppp lin-14 mrna AAAAAAAAAA STOP 3' UTR lin-14 lin-4 5' UAAGGACUCAAAACUUACA3' 3' GUUCCUGAG UGAGUGU5' C A G C U G U C A A
7 Initiation Execution He and Hannon, 2004
8 Oligo Array II: expression profiling of 180 mirnas brain development neuronal differentiation dendritic translation apoptosis cell cycle stem cell differentiation plasticity brain disorders/brain tumors
9
10
11 MicroRNA is an Anti-Apoptotic Factor in Human Brain Tumors
12 Glioblastoma Multiforme (GBM) background Most common primary brain malignancy: 5-7 new cases per 100,000 per year Most malignant (highest-grade) infiltrative astrocytoma No cure; current therapy: surgery + RT + chemotherapy Mean survival: 11 months Histologic parameters have limited value in predicting survival among patients Multiple genetic alterations in oncogenes (EGFR and PDGFR), tumor suppressors (TP53 and PTEN) but no common mrna or protein markers, or genetic predictors of survival
13 mir-21 is Overexpressed in Glioblastoma Tissues and Cells Array of 181 mirnas in GBMs mir-21
14 mir-21 Knock-down Induces Apoptosis in Glioma Cells CellTiter Assay 48 hours post-transfection Caspase 3/7 Activation 48 hours post-transfection Fluorescence at 535 nm (fold induction) 0 Mock 2'OMe-scrambled 2'OMe-miR-21 LNA-scrambled LNA-miR-21 Mock 2'OMe-scrambled 2'OMe-miR-21 LNA-scrambled LNA-miR Cell Number (luminescence fold)
15 Conclusions mir-21 is strongly over-expressed in high-grade brain tumors and in established glioblastoma cell lines mir-21 inhibition triggers activation of caspases and induces apoptosis of glioma cells aberrantly expressed mir-21 may contribute to the malignant phenotype by blocking expression of critical apoptosis-related genes
16 DNA Onco-mirs Over-expressed micrornas whose suppression limits tumor cell growth RNA Protein micrornas Dysregulated mirna----mir-21 microrna expression profile Target mrnas greatly expand the repertoire for intervention in cellular processes Opens discovery to multiple targets in carcinogeneis Inhibits apoptotic genes
17
18 Laser Capture Day 15 Hippocampal culture Fixation with Ethanol GFAP staining Laser capture ion exchange column multi-plex
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20 mir-34a mir-124a mir-124a mir-124a let-7i let-7b
21 Kosik lab Min Jeong Kye I-Fan Wang Onur Sakarya Parul Kapil Thales Papagiannakopoulos Snigda Bannerjee Ben Groves Sourav Banerjee Asano Shoichi Robert Graf Fernando Santiago Sherry Hikita Kawther Abu-ElNeel Former Contributing Lab Members: Anna Krichevsky, Jennifer Chan, Sasha Levy Collaborators Silvia Monticell, Mark Ansel, Changchun Xiao, To-Ha Thai, Klaus Rajewsky, Anjana Rao Department of Pathology, Harvard Medical School, and CBR Nicholas D. Socci, Chris Sander, Computational Biology Center, Memorial Sloan-Kettering Cancer Center Nikolaus Rajewsky, Center for Functional Comparative Genomics, NYU Debora S. Marks, Dept of Systems Biology, HMS Kai Q. Lao, Applied Biosystems
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