The proteasome is a multiprotein complex involved in the degradation of ubiquitinated proteins. Three proteasome inhibitors, calpain inhibitor I, lactacystin and MG132, induced apoptosis in several human malignant glioma cell lines. Although proteasome inhibitors induced p53 accumulation in a cell line retaining wild-type p53 activity, p53 activity was dispensable for apoptosis since transdominant-negative p53 abrogated p53-dependent p21 induction but did not modulate apoptosis. Further, p21 was induced by higher concentrations of proteasome inhibitors in a p53-independent manner both in p53 wild-type and in p53 mutant cell lines. Although there was a strong G2/M arrest in response to proteasome inhibition in glioma cells, this G2/M arrest was also observed in p21(-/-) colon carcinoma cells, suggesting that p21 is dispensable for the G2/M arrest associated with proteasome inhibition. Interestingly, the p21(-/-) cells were more resistant to protease inhibitors than parental p21(+/+) cells. In summary, our data indicate that proteasome inhibition induces a p21-independent G2/M arrest and p53-independent apoptosis in human malignant glioma cells.