S-phase kinase-associated protein 2 is involved in epithelial-mesenchymal transition in methotrexate-resistant osteosarcoma cells

Abstract

Osteosarcoma (OS), a common worldwide primary aggressive bone malignancy, arises from primitive transformed cells of mesenchymal origin and usually attacks adolescents and young adults. Methotrexate (MTX) is the anti-folate drug used as a pivotal chemotherapeutic agent in the treatment of OS. However, patients with OS often develop drug resistance, leading to poor treatment outcomes. In the present study, in order to explore the underlying mechanisms responsible for MTX resistance, we established MTX-resistant OS cells using the U2OS and MG63 cell lines and examined whether MTX-resistant OS cells underwent epithelial-mesenchymal transition (EMT) by Transwell assay, wound healing assay, MTT assay, RT-PCR and western blot analysis. We found that the viability of the MTX-resistant cells remained relatively unaltered following further treatment with MTX compared to the parental cells. The resistant cells appeared to possess a mesenchymal phenotype, with an elongated and more spindle-like shape, and acquired enhanced invasive, migratory and attachment abilities. The measurement of EMT markers also supported EMT transition in the MTX-resistant OS cells. Our result further demonstrated that the overexpression of S-phase kinase-associated protein 2 (Skp2) was closely involved in the resistance of OS cells to MTX and in the acquirement of EMT properties. Thus, the pharmacological inhibition of Skp2 may prove to be a novel therapeutic strategy with which to overcome drug resistance in OS.

Characterization of Melanoma Cell Lines Resistant to Vemurafenib and Evaluation of Their Responsiveness to EGFR- and MET-Inhibitor Treatment

“…Alterations in the morphology of cells treated with vemurafenib for 5-7 days were reported for a panel of human melanoma cells, where the flattened shape of cells was further connected to senescent phenotype [32]. An increase in stress fiber numbers was noticed in melanoma cells resistant to vemurafenib by Misek et al [33], while a spindle-like or mesenchymal-like shape and elevated adhesion were reported in a number of studies on different cancer cells exhibiting drug resistance [34][35][36]. In addition to their altered shape, we also noticed that resistant lines showed a slightly lowered proliferation rate compared to parental ones.…”

Identification of key biomarkers and functional pathways in osteosarcomas with lung metastasis: Evidence from bioinformatics analysis

“…Not only that, SKP2 has also been reported to exert an effect in DNA damage response, DNA repair, tumor angiogenesis and drug resistance [40,41] . For example, Ding et al have demonstrated that SKP2 overexpression was also closely related to the acquisition of methotrexate-resistant and EMT properties in osteosarcoma [42] . Due to the extensive carcinogenic effects of SKP2, research on the anti-tumor activity of SKP2 small molecule inhibitors is in full swing.…”

Targeting Molecular Mechanisms Underlying Treatment Efficacy and Resistance in Osteosarcoma: A Review of Current and Future Strategies

“…Targeted shRNA against SKP2 enhanced drug sensitivity in these cell lines. [394]. The novel Skp2 inhibitor, compound 25, was shown to have antitumour activities and to cooperate with chemotherapeutic agents to suppress cancer cell survival [393], making it of potential interest in osteosarcoma.…”