BACKGROUND: Glioblastoma multiforme is the most common form of primary brain tumor, often characterized by poor survival. Glioblastoma initiating cells (GICs) regulate self-renewal, differentiation, and tumor initiation properties and are involved in tumor growth, recurrence, and resistance to conventional treatments. The sonic hedgehog (SHH) signaling pathway is essential for normal development and embryonic morphogenesis. The objectives of this study were to examine the molecular mechanisms by which GIC characteristics are regulated by NPV-LDE-225 (Smoothened inhibitor; (2,2'-[[dihydro-2-(4-pyridinyl)-1,3(2H,4H)-pyrimidinediyl]bis(methylene)]bis[N,N-dimethylbenzenamine). METHODS: Cell viability and apoptosis were measured by XTT and annexin V-propidium iodide assay, respectively. Gli translocation and transcriptional activities were measured by immunofluorescence and luciferase assay, respectively. Gene and protein expressions were measured by quantitative real-time PCR and Western blot analyses, respectively. RESULTS AND CONCLUSION: NPV-LDE-225 inhibited cell viability, neurosphere formation, and Gli transcriptional activity and induced apoptosis by activation of caspase-3 and cleavage of poly(ADP-ribose) polymerase. NPV-LDE-225 increased the expression of tumor necrosis factor-related apoptosis inducing ligand (TRAIL)-R1/DR4, TRAIL-R2/DR5, and Fas and decreased the expression of platelet derived growth factor receptor-α and Bcl2, and these effects were abrogated by Gli1 plus Gli2 short hairpin RNAs. NPV-LDE-225 enhanced the therapeutic potential of FasL and TRAIL by upregulating Fas and DR4/5, respectively. Interestingly, NPV-LDE-225 induced expression of programmed cell death 4 and apoptosis and inhibited cell viability by suppressing micro RNA (miR)-21. Furthermore, NPV-LDE-225 inhibited pluripotency-maintaining factors Nanog, Oct4, Sox2, and cMyc. The inhibition of Bmi1 by NPV-LDE-225 was regulated by induction of miR-128. Finally, NPV-LDE-225 suppressed epithelial-mesenchymal transition by upregulating E-cadherin and inhibiting N-cadherin, Snail, Slug, and Zeb1 through modulating the miR-200 family. Our data highlight the importance of the SHH pathway for self-renewal and early metastasis of GICs.
BACKGROUND:Glioblastoma multiforme is the most common form of primary brain tumor, often characterized by poor survival. Glioblastoma initiating cells (GICs) regulate self-renewal, differentiation, and tumor initiation properties and are involved in tumor growth, recurrence, and resistance to conventional treatments. The sonic hedgehog (SHH) signaling pathway is essential for normal development and embryonic morphogenesis. The objectives of this study were to examine the molecular mechanisms by which GIC characteristics are regulated by NPV-LDE-225 (Smoothened inhibitor; (2,2'-[[dihydro-2-(4-pyridinyl)-1,3(2H,4H)-pyrimidinediyl]bis(methylene)]bis[N,N-dimethylbenzenamine). METHODS: Cell viability and apoptosis were measured by XTT and annexin V-propidium iodide assay, respectively. Gli translocation and transcriptional activities were measured by immunofluorescence and luciferase assay, respectively. Gene and protein expressions were measured by quantitative real-time PCR and Western blot analyses, respectively. RESULTS AND CONCLUSION: NPV-LDE-225 inhibited cell viability, neurosphere formation, and Gli transcriptional activity and induced apoptosis by activation of caspase-3 and cleavage of poly(ADP-ribose) polymerase. NPV-LDE-225 increased the expression of tumor necrosis factor-related apoptosis inducing ligand (TRAIL)-R1/DR4, TRAIL-R2/DR5, and Fas and decreased the expression of platelet derived growth factor receptor-α and Bcl2, and these effects were abrogated by Gli1 plus Gli2 short hairpin RNAs. NPV-LDE-225 enhanced the therapeutic potential of FasL and TRAIL by upregulating Fas and DR4/5, respectively. Interestingly, NPV-LDE-225 induced expression of programmed cell death 4 and apoptosis and inhibited cell viability by suppressing micro RNA (miR)-21. Furthermore, NPV-LDE-225 inhibited pluripotency-maintaining factors Nanog, Oct4, Sox2, and cMyc. The inhibition of Bmi1 by NPV-LDE-225 was regulated by induction of miR-128. Finally, NPV-LDE-225 suppressed epithelial-mesenchymal transition by upregulating E-cadherin and inhibiting N-cadherin, Snail, Slug, and Zeb1 through modulating the miR-200 family. Our data highlight the importance of the SHH pathway for self-renewal and early metastasis of GICs.
Entities:
Keywords:
Gli transcription factors; NPV-LDE-225; brain cancer stem cells; cell survival; glioblastoma initiating cells; sonic hedgehog signaling
Authors: Silvia Buonamici; Juliet Williams; Michael Morrissey; Anlai Wang; Ribo Guo; Anthony Vattay; Kathy Hsiao; Jing Yuan; John Green; Beatriz Ospina; Qunyan Yu; Lance Ostrom; Paul Fordjour; Dustin L Anderson; John E Monahan; Joseph F Kelleher; Stefan Peukert; Shifeng Pan; Xu Wu; Sauveur-Michel Maira; Carlos García-Echeverría; Kimberly J Briggs; D Neil Watkins; Yung-mae Yao; Christoph Lengauer; Markus Warmuth; William R Sellers; Marion Dorsch Journal: Sci Transl Med Date: 2010-09-29 Impact factor: 17.956
Authors: Young A Yoo; Myoung Hee Kang; Hyun Joo Lee; Baek-hui Kim; Jong Kuk Park; Hyun Koo Kim; Jun Suk Kim; Sang Cheul Oh Journal: Cancer Res Date: 2011-10-05 Impact factor: 12.701
Authors: S A Ciafrè; S Galardi; A Mangiola; M Ferracin; C-G Liu; G Sabatino; M Negrini; G Maira; C M Croce; M G Farace Journal: Biochem Biophys Res Commun Date: 2005-09-09 Impact factor: 3.575