Non‑covalent proteasome inhibitor PI‑1840 induces apoptosis and autophagy in osteosarcoma cells.

Abstract

Osteosarcoma (OS) is the predominant form of primary bone malignancy in children and adolescents. Although the combination of chemotherapy and modified surgical therapy leads to marked improvements in the survival rate, the therapeutic outcomes remain unsatisfactory. Therefore, the identification of novel drugs with higher efficacy and fewer side-effects is urgently required. Proteasome inhibitors have been approved by the Food and Drug Administration (FDA) for the treatment of certain cancers, although none of them are directed against OS. Non-covalent proteasome inhibitors, such as PI-1840, are superior to covalent ones in numerous respects in view of their chemical structure; however, to date, no studies have been published on the effects of non-covalent proteasome inhibitors on OS cells. In the present study, the antineoplastic effects of PI-1840 were systematically evaluated in the OS cell lines, MG-63 and U2-OS. Cell viability and morphological changes were assessed by Cell Counting Kit-8 (CCK-8) and live/dead assays. The cell cycle was analyzed using flow cytometry (FCM) and western blot analysis (assessing the levels of the proteins p21, p27, and the tyrosine kinase, WEE1). The extent of cell apoptosis and autophagy were assessed by FCM, western blot analysis [of the apoptosis-associated proteins, microtubule-associated protein 1 light chain 3 α (LC3) and Beclin1], and mRFP-GFP-LC3 adenovirus transfection assay. Transwell and wound healing assays, and western blot analysis of the matrix metalloproteinases (MMPs)2 and 9 were performed to preliminarily evaluate the migration and invasion capability of the cells. In the present study, our results revealed that PI-1840 inhibited the proliferation of OS cells and induced apoptosis, partly due to attenuation of the nuclear factor-κB (NF-κB) pathway. In addition, PI-1840-induced autophagy was detected, and inhibiting the autophagy of the OS cells led to an increase in the survival rate of the U2-OS cells rather than of the MG-63 cells. Furthermore, PI-1840 attenuated the migration and invasion capabilities of the OS cells. In conclusion, the present study revealed PI-1840 to be a promising drug for the treatment of OS.