Disruption of Her-2 at subdomains I and III interaction promotes an overactivation of erbB receptor family and resistance to chemotherapeutical drugs.

B51

Breast cancer growth is regulated by coordinated actions of the estrogen receptor (ER) and various growth factor receptor signaling pathways. ER, in response to E2, can interact with growth factor tyrosine kinase receptors such us HER2 and IGFR and activate their cascades. Therapies to target HER2 or ER have been developed and are currently being used successfully in many cases. However, a large population of breast cancer patients that are HER2 and ER positives are refractory to trastuzumab (used to target HER2) or to the antiestrogen tamoxifen (used to target ER positive tumors). Once the ER is blocked, cells take advantage of the Her-2 pathway and vice versa. The mechanism of how ER positive tumors that overexpress Her-2 become more aggressive and resistant to treatment is not very well understood. In this study we investigated if an extracellular domain (ECD) site in HER2 receptor, which is involved in the dimerization of the receptor, plays a role in tumor progression, resistance to antiestrogens and Her-2 therapy. We developed a deletion mutant of the sub-domain III of Her-2 ECD (Her-2 mutant). That domain contributes to most of the determinants involved in ligand binding and signal transduction. Stable BT474 cell lines overexpressing the empty vector (pLXSN) and Her-2 mutant were generated using retroviral vector. In these cell lines, first we investigated if Her-2 mutant promotes anchorage-independent growth advantage in response to E2. Her-2 mutant cells formed more colonies of a larger size than control cells, in the presence and in the absence of E2, indicating that cells that overexpress Her-2-mutant are estrogen independent and acquired a higher E2 sensitivity. However, Her-2 mutant showed to be antiestrogen sensitive as did the control cells. We did not observe differences in ERE-luciferase promoter activity between Her-2 mutant and control cells, suggesting that the growth advantage of Her-2 mutant depends on non-genomic ER activity. In addition, Her-2 mutant was resistant to Trastuzumab in anchorage-independent growth. By western blot analysis we observed that Trastuzumab treatment for 24 h causes downregulation on Her-2 levels only in the control cells. Moreover, we found that the Her-2 mutant has higher basic levels of activation of EGFR, Her-2 and Her-3, and other tyrosine kinase receptors and Akt. These results suggest that the deletion of the ECD site in the Her-2 receptor promotes elevated activation of erbB receptors, most probably by stabilization of Her-2 heterodimers with EGFR and Her-3. All together, these results suggest that there is a crosstalk between ER and erbB-family receptor that provides cell growth advantage, resistance to Her-2 therapy and estrogen independence.