Growth inhibition induced by Ro 31-8220 and calphostin C in human glioblastoma cell lines is associated with apoptosis and inhibition of CDC2 kinase.
Protein kinase C (PKC) is a central component in signal transduction and growth control and might be an appropriate target for the chemotherapy of human brain tumors. This study demonstrates that the staurosporine derivative Ro 31-8220, a potent PKC inhibitor, inhibited the growth of 7 human brain tumor cell lines with an IC50 of about 2 microM. Calphostin C, a structurally unrelated PKC inhibitor, inhibited the growth of two of these cell lines with an IC50 of about 100 to 300 nM. Drug withdrawal and clonogenicity assays indicated that the growth inhibition by both of these compounds was irreversible. Morphologic studies, DNA fragmentation studies and flow cytometric assays showed that the treated glioblastoma cells underwent apoptosis. Treatment of glioblastoma cells with Ro 31-8220 lead to a rapid decline in the level of the anti-apoptosis protein bcl-2. At least three of the glioblastoma cell lines carried mutant p53 alleles with missense mutations in the DNA binding domain of p53. Therefore, the induction of apoptosis in these cell lines occurred through a p53-independent mechanism. Furthermore treatment of these glioblastoma cell lines with Ro 31-8220 or calphostin C led to an increase of cells in the G2-M phase of the cell cycle. This correlated with a decrease in CDC2-associated histone H1 kinase activity, as well as a decrease in the level of the CDC2 protein as shown by immunoblotting. When added to subcellular assays Ro 31-8220 markedly inhibited CDC2 histone H1 kinase activity with an IC50 of 100 nM, but calphostin C directly inhibited this kinase activity only at very high concentrations (above 100 microM). Thus these compounds inhibit the growth of glioblastoma cells through novel mechanisms. Ro 31-8220, in particular, might be a useful agent for the treatment of human brain tumors.
Begemann
M
M
Herbert Irving Comprehensive Cancer Center, Columbia-Presbyterian Medical Center, New York, NY 10032, USA.
Kashimawo
S A
SA
Lunn
R M
RM
Delohery
T
T
Choi
Y J
YJ
Kim
S
S
Heitjan
D F
DF
Santella
R M
RM
Schiff
P B
PB
Bruce
J N
JN
Weinstein
I B
IB
D016588:Anticarcinogenic Agents / Q000627:therapeutic use*; D017209:Apoptosis / Q000235:genetics; D001254:Astrocytoma / Q000188:drug therapy*; D001932:Brain Neoplasms / Q000188:drug therapy*; D016203:CDC2 Protein Kinase / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects; D053938:DNA Fragmentation; D004273:DNA, Neoplasm / Q000187:drug effects; D004354:Drug Screening Assays, Antitumor; D016158:Genes, p53 / Q000187:drug effects; D005909:Glioblastoma / Q000188:drug therapy; D006801:Humans; D007211:Indoles / Q000627:therapeutic use*; D009281:Naphthalenes / Q000627:therapeutic use*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D014407:Tumor Cells, Cultured / Q000187:drug effects
D016588:Anticarcinogenic Agents; D004273:DNA, Neoplasm; D007211:Indoles; D009281:Naphthalenes; D019253:Proto-Oncogene Proteins c-bcl-2; D011493:Protein Kinase C; D016203:CDC2 Protein Kinase; C058819:calphostin C; C064758:Ro 31-8220