Differential cyclin-dependent kinase expression and activation in human colon cancer

Background: Abnormalities of the cyclin-dependent protein kinase (CDK) machinery have been linked to cancer development. Hyperphosphorylation of the retinoblastoma (Rb) protein results in release from inhibition of progression through the G1 phase of the cell cycle. Hyperexpression of CDKI and CDK2 may enable progression through late G1, S and the G2 phases of the cell cycle. Methods/Results. To investigate tumor-associated protein kinase activities, control and tumor samples were fractionated by MonoS chromatography and tested for their ability to phosphorylate histone H1. Two major peaks of histone H1 phosphotransferase activity were resolved. The first appeared in the flow through fractions, and occasionally showed enhanced activity in the tumor samples, whilst the second was consistently increased and eluted at approximately 0.4 M NaCl. Western immunoblotting with CDKI and PSTAIRE antibodies confirmed the co-elution of CDKI and CDK2 with the second peak. Next, the phosphotransferase activities (following specific immunoprecipitation) and protein levels of CDK1, 2, 4 and 6 were determined in human colon cancer samples and their respective controls. CDK4 activity was elevated in only 3 of 7 tumor samples (range 40-160%) relative to control samples from the same patients, whereas a significant increase in CDK6 activity was observed in the same group (p<0.05). This contrasted sharply with the universal activations of CDKI (up to 18-fold, p<0.01, n=12) and CDK2 (up to 17-fold, p<0.05) in the same groups. Conclusions. CDKI especially, and to a lesser extent CDK2 and CDK6 demonstrate the most consistent biochemical activation in human colon cancer and may represent targets for pharmacological intervention. Cellular proliferation as gauged by MIBI was not directly correlated with the amplitude of activation.