title: Cyclin G-associated kinase is necessary for osteosarcoma cell proliferation and receptor trafficking.
abstract: Osteosarcoma is the most frequent primary malignant bone tumor among the children. The advent of neoadjuvant chemotherapy significantly improved the prognosis of patients with osteosarcoma in the 1980s, but it has since plateaued in the past decades. Recently, one of the most researched areas in sarcoma treatment is tyrosine kinases. Here, we describe research on a serine/threonine kinase, cyclin G-associated kinase (GAK), which has not been reported in osteosarcoma previously. In this study, a lentiviral based human shRNA library was utilized to screen for kinases in KHOS and U-2OS osteosarcoma cells. The expression of GAK was examined in osteosarcoma and the effect on cell proliferation was analyzed by GAK siRNA knockdown. The level of GAK expression and its correlation to prognosis was analyzed in osteosarcoma tissue microarray. The effect of GAK depletion on insulin-like growth factor and epidermal growth factor receptor-mediated signal transduction was analyzed by Western blot. We observed that GAK was overexpressed in both osteosarcoma cell lines and tissue samples when compared with human osteoblasts. GAK knockdown by siRNA decreased cell proliferation in both drug-sensitive and multidrug-resistant osteosarcoma cell lines. Immunohistochemistry of osteosarcoma tissue microarray revealed that overexpression of GAK was associated with poor prognosis. Finally, knockdown of GAK resulted in alterations of receptor trafficking and several downstream proteins. In conclusion, our results suggest that osteosarcoma cell proliferation and survival are dependent on GAK. These findings may lead to the development of new therapeutic options for osteosarcoma.
authors.affiliation: Department of Orthopaedic Surgery, and SarcomaBiology Laboratory, Center for Sarcoma and Connective Tissue Oncology, Massachusetts General Hospital, Boston, MA 02114, USA.
authors.forename: Michiro
authors.initials: M
authors.lastname: Susa
authors.forename: Edwin
authors.initials: E
authors.lastname: Choy
authors.forename: Xianzhe
authors.initials: X
authors.lastname: Liu
authors.forename: Joseph
authors.initials: J
authors.lastname: Schwab
authors.forename: Francis J
authors.initials: FJ
authors.lastname: Hornicek
authors.forename: Henry
authors.initials: H
authors.lastname: Mankin
authors.forename: Zhenfeng
authors.initials: Z
authors.lastname: Duan
mesh_terms: D015153:Blotting, Western; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D002470:Cell Survival; D055785:Gene Knockdown Techniques; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000172:deficiency / Q000378:metabolism*; D016086:Lentivirus / Q000235:genetics; D012516:Osteosarcoma / Q000201:enzymology* / Q000473:pathology*; D017346:Protein Serine-Threonine Kinases / Q000172:deficiency / Q000378:metabolism*; D021381:Protein Transport; D034741:RNA, Small Interfering / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000378:metabolism*; D015203:Reproducibility of Results; D016019:Survival Analysis
chemical_list: D047908:Intracellular Signaling Peptides and Proteins; D034741:RNA, Small Interfering; D011956:Receptors, Cell Surface; C486325:GAK protein, human; D017346:Protein Serine-Threonine Kinases