Methods: Tissue samples of 37 OS cases retrieved from Hiroshima University Hospital were used for correlating immunoexpression of AMBN and prognosis of patients . In vivo experiments, 143-B cells were injected into five-week-old male BALB/cAnNCrlCrlj mice, and tumor growth and metastasis were analyzed by bioluminescent imaging. In vitroexperiments, western blot, wound healing assay and immunofluorescence were used for molecular analyses of AMBN bioactivities as a tumor suppressor in OS.
Results: (1) In immunohistochemical analysis of 37 clinical OS cases, AMBN expression showed lower frequency of pulmonary metastasis (chi-square test, P＜0.05) and better prognosis. (2) In vivo, AMBN significantly inhibited tumor growth and pulmonary metastasis of OS by using mouse xenograft model. 3) In vitro, AMBN induced cell cycle arrest in G1 phase and suppressed proliferation of OS cells through the reduction of c-myc expression triggered by Src inactivation. In addition, AMBN suppressed the migration activity with focal adhesion and stress fiber formation via RhoA activation induced by Src inactivation. (t-test, P＜0.05).
Conclusions: We demonstrate that AMBN has a new role as a tumor suppressor gene via Src inactivation in OS. Our findings indicate that AMBN has a potential to be used as a new prognostic marker and a therapeutic target of OS. (This study was supported by JSPS.)