Comparison of microarray-based RNA expression with ELISA-based protein determination of HER2, uPA and PAI-1 in tumour tissue of patients with breast cancer and relation to outcome

Abstract

Prognostic and predictive markers in breast cancer are currently determined by single analysis of protein amounts. If RNA-based multi-gene analyses enter clinical practice, simultaneous determination of currently established markers like human epidermal growth factor receptor 2 (HER2), urokinase plasminogen activator (uPA) and its inhibitor (PAI-1) would represent an elegant simplification. To investigate the correlation between RNA and protein levels, we assessed HER2, uPA and PAI-1 in patients with breast cancer. In addition, we evaluated the influence of these factors on patient outcome. We collected tumour samples from 133 patients with primary breast cancer. Protein and mRNA levels were measured for HER2, uPA and PAI-1. Protein concentration was measured by ELISA, mRNA expression was analysed by Affymetrix A133U Gene Chip and validated by quantitative PCR. We were able to demonstrate a statistically significant correlation between mRNA and protein expression for HER2 (r=0.67, P<0.001) and uPA (r=0.7, P<0.001) but not for PAI-1 (r=0.27). We observed a prognostic information for PAI-1 mRNA and protein values. Patients with high PAI-1 mRNA expression had a reduced 10-year disease-free survival (DFS) rate (60 vs. 70%, P=0.071) and 10-year overall survival (OS) rate (68 vs. 79%, P=0.034). Patients with PAI-1 protein levels above 14ng/mg protein had a reduced disease-free (10-year DFS rate 54 vs. 71%, P=0.006) and overall survival rate (10-year OS-rate 63 vs. 83%, P=0.018). In the patient cohort with no chemotherapy, PAI-1 mRNA levels were the strongest prognostic factor for OS in univariate and multivariate analysis. Results of RNA-based multi-gene analyses of the prognostic and predictive markers HER2 and uPA correlate with the corresponding protein levels. This is not the case for PAI-1. However, PAI-1 mRNA expression might reveal new clinically relevant information in addition to PAI protein levels.