Her-2/neu-triggered intracellular tyrosine kinase activation: in vivo relevance of ligand-independent activation mechanisms and impact upon the efficacy of trastuzumab-based treatment

Proteolytic cleavage of the Her-2/neu extracellular domain (ECD) has been shown to initiate receptor phosphorylation representing Her-2/neu/ activation in vitro. The present investigation was performed to evaluate the clinical relevance of ECD cleavage for Her-2/ neu activation and the consequences of active intracellular Her-2/neu signalling reflected by tyrosine kinase phosphorylation in patients treated with the anti-Her-2/neu antibody trastuzumab. Sera from 62 patients receiving trastuzumab-based treatment for Her-2/neu overexpressing metastatic breast cancer were assessed for pretreatment ECD levels using an enzyme-linked immunosorbent assay. In parallel, Her-2/neu activation status of tumour specimens was assessed by immunohistochemistry using a Her-2/neu phosphorylation state specific antibody (PN2A) and correlated with the patients' ECD levels and clinical course of disease. Serum ECD levels were significantly higher in 15 (24%) patients with tumours exhibiting activated Her-2/neu as compared to those without detectable Her-2/neu phosphorylation (median 148.2 vs 28.5 ng ml^-1, P = 0.010). Whereas response rate only showed a trend to be higher in patients with Her-2/neu-phosphorylated breast cancer (47 vs 34%, P = 0.197), both uni- and multivariate analyses revealed that the median progression-free survival under trastuzumab-based treatment was significantly longer in patients with Her-2/neu-phosphorylated breast cancer- 11.7 (95% Cl 5.2-18.3) months-when compared to the progression-free survival of 4.5 (95% Cl 3.4-5.6) months observed in patients with tumours lacking phosphorylated Her-2/neu (P = 0.001). Proteolytic cleavage of the ECD represents a biologically relevant ligand-independent mechanism of Her-2/neu activation in vivo. The influence of Her-2/ neu activation status upon the outcome of trastuzumab-based therapies merits further investigation in larger prospective trials.