STAT3 activation by the ErbB2 receptor and development of peptide-based STAT3 inhibitors.

摘要

Stats (s&barbelow;ignal t&barbelow;ransducer and a&barbelow;ctivator of t&barbelow;ranscription) are latent transcription factors that translocate from the cytoplasm to nucleus. Constitutive activation of Stat3alpha by upstream oncoproteins and receptor tyrosine kinases has been found in many human tumors and tumor-derived cell lines and it is often correlated with the activation of ErbB-2. In order to explore the involvement of ErbB-2 in the activation of Stat3 and the mechanisms underlying this event, an erbB-2 point mutant was used as a model of a constitutively activated receptor. Phenylalanine mutations (Y-F) were made in the receptor's autophosphorylation sites and their ability to activate Stat3alpha was evaluated. Our results suggest that Stat3alpha and Janus tyrosine kinase 2 associates with ErbB-2 prior to tyrosine phosphorylation of the receptor and that full activation of Stat3alpha by ErbB-2 requires the participation of other non-receptor tyrosine kinases. Both Src and Jak2 kinases contribute to the activation of Stat3alpha while only Src binds to ErbB-2 only when the receptor is tyrosine phosphorylated. Our results also suggest that tyrosine 1139 may be important for Src SH2 domain association since a mutant lacking this tyrosine reduces the ability of the Src SH2 domain to bind to ErbB-2 and significantly decreases its ability to activate Stat3alpha.; In order to disrupt aberrant STAT3alpha activation which contributes to tumorigenesis, we sought small molecules which can specifically bind to the STAT3 SH2 domain, thereby abolishing its ability of being recruited into receptors, and also blocking the dimer formation required for STAT3alpha activation. A phosphopeptide derived from gp130 was found to have a high affinity to STAT3 SH2 domain, and we decided to use this peptide as the base for further modifications. A series of peptide based compounds were designed and tested using electrophoretic mobility shift assay and fluorescence polarization assay to evaluate their affinity to the STAT3 SH2 domain. Two promising compounds, DRIV-73C and BisPOM, were used for blocking STAT3alpha activity in cell culture. Either can successfully impair STAT3alpha activation induced by IL-6 stimulation in HepG2 cells. BisPOM proved to be the more effective in blocking STAT3alpha tyrosine phosphorylation in induced cells and tumor cell lines, and was the more potent in inhibiting STAT3 dependent cell growth.