Silencing tumor-intrinsic HHLA2 potentiates the anti-tumoral effect of paclitaxel on MG63 cells: Another side of immune checkpoint.

• HHLA2 silencing enhances the chemosensitivity of MG-63 cells to paclitaxel. • HHLA2 silencing increases paclitaxel antitumoral effect on MG-63 cell migration. • HHLA2 silencing increases paclitaxel antitumoral effect on MG-63 cell apoptosis. • HHLA2 silencing increases paclitaxel antitumoral effect on MG-63 cell cycle. Osteosarcoma is common type of bone cancer; however, the prognosis of patients with metastatic osteosarcoma is poor. As a new inhibitory immune checkpoint molecule, HHLA2 is upregulated in osteosarcoma. Herein, we studied the significance of tumor-intrinsic HHLA2 in MG-63 growth. Also, we examined the influence of combined therapy of HHLA2 knockdown with paclitaxel on the apoptosis, cell cycle, migration, and stemness of MG-63 cells. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay was performed to study the half-maximal inhibitory concentration (IC50) of paclitaxel and the cytotoxicity of HHLA2-small interfering RNA (siRNA) on MG-63 cells. The apoptosis and cell cycle were analyzed using flow cytometry. The wound-healing and colony formation assays were conducted to investigate the effect of paclitaxel and HHLA2 knockdown on the migration and stemness of MG-63 cells, respectively. QRT-PCR was used to determine the Bax, caspase-3, and Bcl-2 mRNA expression levels. HHLA2 silencing has enhanced the chemosensitivity of MG-63 cells to paclitaxel. Besides, HHLA2 knockdown has increased the paclitaxel-induced cytotoxic effect on MG-63 cells. In terms of stimulating apoptosis, decreasing clonogenicity, halting the cell cycle at the sub G1 phase, and inhibiting migration, tumor-intrinsic HHLA2 silencing has increased these anti-tumor effects of paclitaxel on MG-63 cells. Besides, HHLA2 knockdown has potentiated paclitaxel-mediated Bcl-2 downregulation and paclitaxel-mediated caspase-3 and Bax upregulation in MG-63 cells. Tumor-intrinsic HHLA2 knockdown increases the anti-tumoral effect of paclitaxel on MG-63 cells and enhances the chemosensitivity of MG-63 cells to paclitaxel. [ABSTRACT FROM AUTHOR]