Effect of silencing S-phase kinase-associated protein 2 on chemosensitivity to temozolomide of human glioma cells U251

Objective: To examine the effect of silencing SKP2 on chemosensitivity of human glioma cells U251 to temozolomide (TMZ).

Methods: Adenoviruses harbouring shRNA targeting SKP2 (i.e. Ad-shSKP2) and non-targeting scrambled shRNA (i.e. Ad-shNC) were used to infect U251 cells. The transduced cells were then treated with TMZ. Cell viability after treatment was assayed using CCK8; while cell cycle and Apoptosis were examined using flow cytometry. To study the effect of silencing SKP2 on Autophagy in U251, we co-transduced the cells with Ad-mRFP-LC3 and Ad-shSKP2/Ad-shNC. The expression of Autophagy marker LC3 after TMZ treatment was studied using microscopy and Western blotting assays.

Results: The cytotoxicity of TMZ (i.e. 20-100 µM) was more significantly seen in Ad-shSKP2-transduced U251 cells than in the Ad-shNC-transduced U251 cells. The IC₅₀ values in shSKP2-U251 were significantly lower than those of the shNC-U251 (P < 0.05). Both TMZ and Ad-shSKP2 alone increased Apoptosis and promoted expression of LC3 in U251. Combined treatment of Ad-shSKP2 and TMZ further elevated Apoptosis and LC3 expression.

Conclusion: Silencing SKP2 in U251 cells increased chemosensitivity to TMZ that was accompanied with enhanced Apoptosis and Autophagy. Targeting SKP2 may be a potential approach to potentiate TMZ treatment in patients with glioma.