1. ¹Department of Pathology and Royal Free and University College Medical School, University College London, Rockefeller Building, University Street, London, WC1E 6JJ, UK
2. ²Northern Institute for Cancer Research, University of Newcastle, Paul O'Gorman Building, Medical School, Framlington Place, Newcastle upon Tyne, NE2 4HH, UK
3. ³Department of Public Health and Primary Care, Centre for Applied Medical Statistics, Institute of Public Health, University of Cambridge, Forvie Site, Robinson Way, Cambridge, CB2 2SR, UK
4. ⁴Department of Oncology and Hutchison MRC Research Centre, University of Cambridge, Hills Road, Cambridge, CB2 2XZ, UK
5. ⁵Wolfson Institute for Biomedical Research, University College London, The Cruciform Building, Gower Street, London, WC1E 6BT, UK

Correspondence: Professor GH Williams, E-mail: gareth.williams@ucl.ac.uk

⁶These authors contributed equally to this work.

Received 27 October 2006; Revised 2 March 2007; Accepted 6 March 2007; Published online 3 April 2007.

Abstract

Activation of mitogen/extracellular-signal-regulated kinase kinase 5/extracellular signal-regulated kinase-5 (MEK5/ERK5) growth signalling is coupled to increased cell proliferation in prostate cancer (PCa). Dysregulation of the DNA replication licensing pathway, a critical step in growth control downstream of transduction signalling pathways, is associated with development of PCa. In this study we have investigated linkages between the MEK5/ERK5 pathway and DNA replication licensing during prostate carcinogenesis. The effects of increased MEK5/ERK5 signalling on the expression of replication licensing factors Mcm2 and geminin and the proliferation marker Ki67 were studied in an ecdysone-inducible system expressing a constitutively activated mutant of MEK5 in EcR293 cells and in stable ERK5 over-expressing PC3 clones. In parallel, expression of these biomarkers in PCa biopsy specimens (n=58) was studied and compared to clinicopathological parameters. In both in vitro systems induction of MEK5 expression resulted in increased levels of phosphorylated ERK5 and Mcm2, geminin and Ki67 proteins. In PCa specimens average Mcm2 expression was greater than Ki67 and geminin expression (median labelling index (LI) 36.7, 18.1, and 3.4% respectively), consistent with their differential expression according to growth status (P<0.0001). Mcm2, geminin and Ki67 expression were significantly associated with Gleason grade (P=0.0002, P=0.0003, P=0.004); however there was no link with T or M stage. There was a significant relationship between increasing ERK5 expression and increasing Mcm2 (P=0.003) and Ki67 (P=0.009) expression, with non-significant trends seen with increasing MEK5 expression. There were significant associations between Gleason grade and the number of cells traversing G1 phase (Ki67_LI-geminin_LI; (P=0.001)), with high ERK5 levels associated with both an increase in replication licensed but non-cycling cells (Mcm2_LI-Ki67_LI; (P=0.01)) and accelerated cell cycle progression (geminin_LI/Ki67_LI; (P= 0.005)), all indicative of a shift towards increasing proliferative potential. While Mcm2 and Ki67 were both prognostic factors on univariate analysis, only Mcm2 remained an independent prognostic marker on multivariate analysis. Taken together, our data show that induction of MEK5/ERK5 signalling is linked to activation of the DNA replication licensing pathway in PCa, and that the strong prognostic value of MCM proteins may result from their function as relay stations coupling growth regulatory pathways to genome duplication.