Arisa Tsuru

GLI2 is a novel therapeutic target for metastasis of osteosarcoma.

Aberrant activation of the Hedgehog (Hh) pathway has been reported in several malignancies. We previously demonstrated that knockdown of GLI2 inhibited proliferation of osteosarcoma cells through regulation of the cell cycle. In this study, we analyzed the function of GLI2 in the pathogenesis of osteosarcoma metastasis. Immunohistochemical studies showed that GLI2 was overexpressed in patient osteosarcoma specimens. Knockdown of GLI2 inhibited migration and invasion of osteosarcoma cells. In contrast, the forced expression of constitutively active GLI2 in mesenchymal stem cells promoted invasion. In addition, xenograft models showed that knockdown of GLI2 decreased lung metastasis of osteosarcomas. To examine clinical applications, we evaluated the efficacy of arsenic trioxide (ATO), which is a Food and Drug Administration‐approved antitumor drug, on osteosarcoma cells. ATO treatment suppressed the invasiveness of osteosarcoma cells by inhibiting the transcriptional activity of GLI2. In addition, the combination of Hh inhibitors including ATO, vismodegib and GANT61 prevented migration and metastasis of osteosarcoma cells. Consequently, our findings suggested that GLI2 regulated metastasis as well as the progression of osteosarcomas. Inhibition of the GLI2 transcription may be an effective therapeutic method for preventing osteosarcoma metastasis.

Ribosomal protein S3 regulates GLI2-mediated osteosarcoma invasion

It has been reported that GLI2 promotes proliferation, migration, and invasion of mesenchymal stem cell and osteosarcoma cells. To examine the molecular mechanisms of GLI2-mediated osteosarcoma metastasis, we performed a microarray analysis. The gene encoding ribosomal protein S3 (RPS3) was identified as a target of GLI2. Real-time PCR revealed that RPS3 was upregulated in osteosarcoma cell lines compared with normal osteoblast cells. Knockdown of GLI2 decreased RPS3 expression, whereas forced expression of a constitutively active form of GLI2 upregulated the expression of RPS3. RPS3 knockdown by siRNA decreased the migration and invasion of osteosarcoma cells. Although forced expression of constitutively active GLI2 increased the migration of human mesenchymal stem cells, knockdown of RPS3 reduced the up-regulated migration. In contrast, forced expression of RPS3 increased migration and invasion of osteosarcoma cells. Moreover, reduction of migration by GLI2 knockdown was rescued by forced expression of RPS3. Immunohistochemical analysis showed that RPS3 expression was increased in primary osteosarcoma lesions with lung metastases compared with those without. These findings indicate that GLI2-RPS3 signaling may be a marker of invasive osteosarcoma and a therapeutic target for patients with osteosarcoma.

Hairy/enhancer-of-split related with YRPW motif protein 1 promotes osteosarcoma metastasis via matrix metallopeptidase 9 expression

Background:Activation of the Notch pathway has been reported in various types of cancers. However, the role of the hairy/enhancer-of-split related with YRPW motif protein 1 (HEY1) in osteosarcoma is unknown. We examined the function of HEY1 in osteosarcoma.Methods:Expression of HEY1 was studied in human osteosarcoma. The effects of HEY1 in osteosarcoma were evaluated in vitro and in a xenograft model. Moreover, we examined the function of matrix metallopeptidase 9 (MMP9) as a downstream effector of HEY1.Results:HEY1 was upregulated in human osteosarcoma. Knockdown of HEY1 inhibited the invasion of osteosarcoma cell lines. In contrast, the forced expression of HEY1 increased the invasion of mesenchymal stem cell. In addition, lung metastases were significantly inhibited by the knockdown of HEY1. We found that MMP9 was a downstream effector of HEY1 that promotes the invasion of osteosarcoma cells. Knockdown of HEY1 decreased the expression of MMP9. Addition of MMP9 rescued the invasion of osteosarcoma cells that had been rendered less invasive by knockdown of HEY1 expression.Conclusions:Our findings suggested that HEY1 augmented the metastasis of osteosarcoma via upregulation of MMP9 expression. Therefore, inhibition of HEY1 may be a novel therapeutic strategy for preventing osteosarcoma metastasis.