miR-487b and TRAK2 that form an axis to regulate the aggressiveness of osteosarcoma, are potential therapeutic targets and prognostic biomarkers.

To investigate the effect of microRNA-487b (miR-487b) as well as the underlying mechanism in osteosarcoma (OS). Data downloaded from the Gene Expression Omnibus (GEO) database were used to analyze the expression and prognostic value of miR-487b/TRAK2. Cell counting kit-8, colony formation, and transwell assays were performed to investigate the biological functions of miR-487b and TRAK2. Luciferase reporter assay was applied to confirm the interactions between miR-487b and TRAK2. miR-487b was overexpressed in OS tissues and was inversely associated with the prognosis of OS patients. We discovered that miR-487b could contribute to the proliferative, clonogenic, invasive, and migratory capabilities of OS cells. Through target prediction using miRWalk and differential expression analysis based on the GEO data set, trafficking kinesin protein 2 (TRAK2) was recognized as a potential target of miR-487b, which was further verified by luciferase reporter assay. The expression of TRAK2 was decreased in OS tissues compared with normal tissues and was positively correlated with the prognosis of OS patients. A negative relevance was presented between the expression of miR-487b and TRAK2 in OS cells. Of note, further mechanistic analyses indicated that TRAK2 was implicated in the regulatory effect of miR-487b on the cell malignant behaviors in OS. To sum up, these results demonstrated that miR-487b played an oncogenic role in OS progression via directly targeting TRAK2, which could advance the development of cancer treatment.

Sodium cantharidinate suppresses human osteosarcoma MG­63 cell proliferation and induces cell cycle arrest by inhibition of PI3K/AKT activation.

The function and mechanism of sodium cantharidininate (SC) underlying its suppression of human osteosarcoma (OS) MG­63 cells were investigated for the first time in the present study. MG­63 cell proliferation was determined by WST­1 assay post SC treatment at 0, 12, 24, 48 and 72 h. The results showed that SC effectively inhibited MG­63 cell proliferation and induced cell cycle arrest at the G0/G1 phase in a dose­dependent manner. Western blotting revealed that SC induced MG­63 cell cycle arrest at the G0/G1 phase by means of inhibition of cyclin D1, CDK4 and CDK6 expression. The expression of MAPK and AKT were evaluated using western blotting and FACS experiments to determine whether such signaling pathways are involved in the antiproliferative action of SC on MG­63 cells. SC significantly inhibited the phosphorylation of AKT, but not mTOR, JNK or P38. PI3K/AKT stimulator, IGF­1, reversed the SC­induced cell cycle arrest in the MG­63 cells. Collectively, our data indicate that the phosphorylation of AKT was inhibited by SC, consequently decreasing the expression of cyclin D1, CDK4 and CDK6 and inducing MG­63 cell G0/G1 phase arrest. SC has potential as a treatment agent for human osteosarcoma.

PD­L1/PD­1 axis serves an important role in natural killer cell­induced cytotoxicity in osteosarcoma.

Osteosarcoma is a serious malignancy in pediatric patients, which comprises 2.4% of fatal cancer in children and achieves 20% of all primary bone cancers. In the present study, we employed three human osteosarcoma cell lines MG­63, HOS and U2OS for susceptibility to cytolytic activity of freshly isolated healthy donor NK cells. Cells were lysed by NK cells in a dose dependent manner. MG­63 cells exhibited less susceptibility to NK cells than HOS and U2OS cells at all cell ratios. The specific mechanism underlying the effects of NK cells on osteosarcoma cells was determined by antibody blockage experiments. The results revealed that granzyme B was the key factor in the NK cell­induced cytotoxicity of human osteosarcoma cells. To the best of our knowledge, the present study is the first to investigate the expression of PD­L1 in MG­63, HOS and U2OS cells. The relative expression of the PD­L1 gene and protein in MG­63 cell was greater than HOS and U2OS cells. The specific lysis of human osteosarcoma cells induced by NK cells was enhanced when PD­L1/PD­1 was blocked by the PD­L1 antibody. The present study proposed that the PD­L1/PD­1 axis serves an important role in NK cell­induced cytotoxicity in osteosarcoma via granzyme B secretion. Our findings may contribute to the development of precise treatments for osteosarcoma based on the expression profile of PD­L1 in patients with this disease.