title: The indolinone MAZ51 induces cell rounding and G2/M cell cycle arrest in glioma cells without the inhibition of VEGFR-3 phosphorylation: involvement of the RhoA and Akt/GSK3β signaling pathways.
abstract: MAZ51 is an indolinone-based molecule originally synthesized as a selective inhibitor of vascular endothelial growth factor receptor (VEGFR)-3 tyrosine kinase. This study shows that exposure of two glioma cell lines, rat C6 and human U251MG, to MAZ51 caused dramatic shape changes, including the retraction of cellular protrusions and cell rounding. These changes were caused by the clustering and aggregation of actin filaments and microtubules. MAZ51 also induced G2/M phase cell cycle arrest. This led to an inhibition of cellular proliferation, without triggering significant cell death. These alterations induced by MAZ51 occurred with similar dose- and time-dependent patterns. Treatment of glioma cells with MAZ51 resulted in increased levels of phosphorylated GSK3β through the activation of Akt, as well as increased levels of active RhoA. Interestingly, MAZ51 did not affect the morphology and cell cycle patterns of rat primary cortical astrocytes, suggesting it selectively targeted transformed cells. Immunoprecipitation-western blot analyses indicated that MAZ51 did not decrease, but rather increased, tyrosine phosphorylation of VEGFR-3. To confirm this unanticipated result, several additional experiments were conducted. Enhancing VEGFR-3 phosphorylation by treatment of glioma cells with VEGF-C affected neither cytoskeleton arrangements nor cell cycle patterns. In addition, the knockdown of VEGFR-3 in glioma cells did not cause morphological or cytoskeletal alterations. Furthermore, treatment of VEGFR-3-silenced cells with MAZ51 caused the same alterations of cell shape and cytoskeletal arrangements as that observed in control cells. These data indicate that MAZ51 causes cytoskeletal alterations and G2/M cell cycle arrest in glioma cells. These effects are mediated through phosphorylation of Akt/GSK3β and activation of RhoA. The anti-proliferative activity of MAZ51 does not require the inhibition of VEGFR-3 phosphorylation, suggesting that it is a potential candidate for further clinical investigation for treatment of gliomas, although the precise mechanism(s) underlying its effects remain to be determined.
authors.affiliation: Department of Anatomy, Catholic Neuroscience Institute, College of Medicine, The Catholic University of Korea, Seoul, Korea.
authors.forename: Joo-Hee
authors.initials: JH
authors.lastname: Park
authors.affiliation: Department of Anatomy, Catholic Neuroscience Institute, College of Medicine, The Catholic University of Korea, Seoul, Korea.
authors.forename: Yoo-Jin
authors.initials: YJ
authors.lastname: Shin
authors.affiliation: Department of Anatomy, Catholic Neuroscience Institute, College of Medicine, The Catholic University of Korea, Seoul, Korea.
authors.forename: Tae-Ryong
authors.initials: TR
authors.lastname: Riew
authors.affiliation: Department of Anatomy, Catholic Neuroscience Institute, College of Medicine, The Catholic University of Korea, Seoul, Korea.
authors.forename: Mun-Yong
authors.initials: MY
authors.lastname: Lee
mesh_terms: D008841:Actin Cytoskeleton / Q000187:drug effects / Q000648:ultrastructure; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D001253:Astrocytes / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D048430:Cell Shape / Q000187:drug effects; D059565:G2 Phase Cell Cycle Checkpoints / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic; D038362:Glycogen Synthase Kinase 3 / Q000235:genetics* / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta; D006801:Humans; D007211:Indoles / Q000494:pharmacology*; D008870:Microtubules / Q000187:drug effects / Q000648:ultrastructure; D009281:Naphthalenes / Q000494:pharmacology*; D009457:Neuroglia / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology / Q000648:ultrastructure; D010766:Phosphorylation; D051057:Proto-Oncogene Proteins c-akt / Q000819:agonists / Q000235:genetics* / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D051381:Rats; D015398:Signal Transduction; D042582:Vascular Endothelial Growth Factor C / Q000494:pharmacology; D040321:Vascular Endothelial Growth Factor Receptor-3 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D020742:rhoA GTP-Binding Protein / Q000819:agonists / Q000235:genetics* / Q000378:metabolism
chemical_list: C441602:3-(4-dimethylamino-naphthalen-1-ylmethylene)-1,3-dihydro-indol-2-one; D000970:Antineoplastic Agents; D007211:Indoles; D009281:Naphthalenes; D034741:RNA, Small Interfering; D042582:Vascular Endothelial Growth Factor C; C066737:RHOA protein, human; C581309:FLT4 protein, human; D040321:Vascular Endothelial Growth Factor Receptor-3; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; C000605508:Gsk3b protein, rat; D051057:Proto-Oncogene Proteins c-akt; D038362:Glycogen Synthase Kinase 3; D020742:rhoA GTP-Binding Protein