Nuclear Function of ErbB-2/ErbB-3 Heterodimers in Trastuzumab-Resistant Breast Cancer Cells

ROSALIA INES CORDO RUSSO, WENDY BÉGUELIN, MARIA CELESTE DIAZ FLAQUE, VIOLETA A CHIAUZZI, LEANDRO VENTURUTTI, NATALIA M GALIGNIANA, CECILIA J PROIETTI, EDUARDO H CHARREAU, ROXANA SCHILLACI AND PATRICIA VIRGINIA ELIZALDE.

Congreso; 2014 Endocrine Society Annual Meeting; 2014

ErbB-2, a member of the ErbB family of membrane receptor tyrosine kinases, is a major player in the breast cancer (BC) scenario. ErbB-2 overexpression is associated with poor prognosis and is therapeutically targeted by trastuzumab (TZ). The dogma of ErbB-2 mechanism of action has been challenged by the demonstration that membrane ErbB-2 (MErbB-2) migrates to the nucleus (NErbB-2) of BC cells where it acts as a transcription factor or, according to our previous findings, as a transcriptional coactivator. We have recently demonstrated that NErbB-2 function is absolutely necessary for in vitro and in vivo growth of MErbB-2-positive BC cells both sensitive (BT-474) and resistant (JIMT-1) to TZ (1). Besides, we revealed that blockade of NErbB-2 localization constitutes a novel therapeutic strategy in TZ-resistant BC (1). Here, we explored the molecular mechanisms underlying NErbB-2 promotion of growth. Our findings demonstrated that heregulin (HRGβ1), a ligand of ErbB-3 and ErbB-4 which recognizes ErbB-2 as co-receptor, stimulates ErbB-2 and ErbB-3 nuclear (NErbB-3) translocation and their colocalization with signal transducer and activator of transcription 3 (Stat3) in human T47D BC cells devoid of basal NErbB-2 and NErbB-3. HRGβ1 also enhanced formation of nuclear ErbB-2/ErbB-3 heterodimers in BT-474 and JIMT-1 cells, which express basal levels of NErbB-2 and NErbB-3. To block NErbB-2 presence, we used a human ErbB-2 nuclear localization domain mutant (hErbB-2ΔNLS), unable to translocate to the nucleus, and which we have previously showed acts as a dominant negative inhibitor of endogenous NErbB-2 migration (2). Interestingly, we found that hErbB-2ΔNLS colocalized with ErbB-3 at the cytoplasm and abrogated basal and HRGβ1-induced ErbB-3 nuclear migration in BT-474 and JIMT-1 cells. Our recent findings revealed that progestin regulates cyclin D1 expression, a cancer-related gene that contains Stat3 binding sites (GAS sites) but lacks ErbB-2 response elements (HAS sites) in its proximal promoter, via the assembly of a transcriptional complex in which ErbB-2 acts as coactivator of Stat3 (2). Here, we found that HRGβ1 induces in vivobinding of Stat3, ErbB-2, and ErbB-3 to the GAS sites of the cyclin D1 promoter in T47D, BT-474, and JIMT-1 cells. Transfection of JIMT-1 cells with hErbB-2ΔNLS abolished both basal and HRGβ1-induced recruitment of ErbB-2 and ErbB-3 to the cyclin D1 promoter as well as HRGβ1-induced up-regulation of cyclin D1 protein and mRNA levels, indicating that the hErbB-2ΔNLS growth inhibitory effects on TZ-resistant cells lies in the disruption of the Stat3/ErbB-2/ErbB-3 nuclear complex driving cyclin D1 expression. These findings for the first time reveal the nuclear interaction and function of ErbB-2/ErbB-3 heterodimers and highlight the importance of targeting NErbB-2 function as a novel therapeutic strategy in TZ-resistant BC.