In vitro analysis of ovarian cancer response to cisplatin, carboplatin, and paclitaxel identifies common pathways that are also associated with overall patient survival.

BACKGROUND: Carboplatin and cisplatin, alone or in combination with paclitaxel, have similar efficacies against ovarian cancer (OVCA) yet exhibit different toxicity profiles. We characterised the common and unique cellular pathways that underlie OVCA response to these drugs and analyse whether they have a role in OVCA survival. METHODS: Ovarian cancer cell lines (n=36) were treated with carboplatin, cisplatin, paclitaxel, or carboplatin-paclitaxel (CPTX). For each cell line, IC(50) levels were quantified and pre-treatment gene expression analyses were performed. Genes demonstrating expression/IC(50) correlations (measured by Pearson; P<0.01) were subjected to biological pathway analysis. An independent OVCA clinico-genomic data set (n=142) was evaluated for clinical features associated with represented pathways. RESULTS: Cell line sensitivity to carboplatin, cisplatin, paclitaxel, and CPTX was associated with the expression of 77, 68, 64, and 25 biological pathways (P<0.01), respectively. We found three common pathways when drug combinations were compared. Expression of one pathway ('Transcription/CREB pathway') was associated with OVCA overall survival. CONCLUSION: The identification of the Transcription/CREB pathway (associated with OVCA cell line platinum sensitivity and overall survival) could improve patient stratification for treatment with current therapies and the rational selection of future OVCA therapy agents targeted to these pathways.

Human glioma PKC-iota and PKC-betaII phosphorylate cyclin-dependent kinase activating kinase during the cell cycle.

Cell cycle phase transition is regulated in part by the trimeric enzyme, cyclin-dependent kinase activating kinase (CAK) which phosphorylates and activates cyclin-dependent kinases (cdks). Protein kinase C (PKC) inhibitors prevent cell cycle phase transition, suggesting a fundamental role for PKCs in cell cycle regulation. We report that in glioma cells, CAK (cdk7) is constitutively associated with PKC-iota. In vitro phosphorylation, co-immunoprecipitation, and analysis of phosphorylated proteins by autoradiography indicate that CAK (cdk7) is a substrate for PKC-iota and PKC-betaII hyperphosphorylation. These results establish a role for PKC-iota and PKC-betaII in the activation of CAK during the glioma cell cycle.

Ultrastructural study of protein kinase C-betaII localization during the cell cycle of human glioma cells.

Transmission electron microscopy and immunogold labeling were used to determine how PKC-betaII is localized at stages in the cell cycle of the human glioma cell line U-373MG. Results show that immunogold particles in both dimethylsulfoxide (DMSO) and calphostin C (0.5 microM)-treated cells were mainly located in the cytoplasm. The concentration of gold particles in the nucleus was relatively small and constant throughout the cell cycle of both DMSO and calphostin C treated cells. Micrographs revealed changes in PKC-betaII during the cell cycle. The concentration of gold particles in the DMSO-treated cells was constant until 8 h. Subsequently, cytoplasmic PKC-betaII oscillated with an increased at 10 h, a rapid decrease at 12 h, and a rise at 14 h. The concentration of the gold particles then gradually decreased. In contrast, immunogold labeling in calphostin C-treated cells increased gradually up to 10 h. Subsequently, the pattern of PKC-betaII labeling in calphostin C-treated cells recapitulated those of control cells as seen by the rapid decline of PKC-betaII labeling at 12 h and its re-accumulation at 14 h. Additionally, there was a rapid increase at 20 h. Western blots of PKC-betaII showed constant PKC-betaII immunoreactivity throughout the cell cycle. In comparison to Western blots, in-situ immunogold labeling revealed changes in PKC-II immunoreactivity at 10 h and 14 h. This technique may represent intracellular immunoreactivity of PKC-betaII. The results from the immunogold labeling technique suggest that binding of calphostin C to the regulatory domain of PKC-betaII provokes a conformation change in PKC-betaII, preventing its activation and degradation.