Ribozyme-mediated down-regulation of ErbB-4 in estrogen receptor-positive breast cancer cells inhibits proliferation both in vitro and in vivo

ErbB-4 is a recently discovered member of the class I receptor tyrosine kin ase family (ErbB), Little is known about its expression and its importance in human malignancy. To delineate the biological function of ErbB-4 recepto rs in breast cancer, we used a hammerhead ribozyme strategy to achieve down -regulation of ErbB-4 receptors in various breast cancer cell Lines. We obs erved that down-regulation of ErbB-4 in estrogen receptor-positive (ER+) hu man breast cancer cell lines (MCF-7 and T47D), which express relatively hig h levels of ErbB-4, significantly inhibited colony formation. No effects we re observed in estrogen receptor-negative (ER-) MDA-MB-453 cells, which exp ress low levels of endogenous ErbB4 and high levels of ErbB-2 and ErbB-3, T his occurred despite the fact that fluorescence-activated cell sorter analy sis of these latter cells revealed that the expression of the ErbB-4 recept or was completely abrogated by ribozyme treatment. Furthermore, down-regula tion of ErbB-4 in T47D and MCF-7 cells significantly inhibited tumor format ion in athymic nude mice (P < 0.03 and P < 0.001, respectively), In additio n, NRG-stimulated phosphorylation of ErbB-4- and NRG-induced colony formati on was significantly reduced in ribozyme-transfected T47D cells. These data provide the first evidence that elevation of ErbB-4 expression plays a rol e in the proliferation of some ER+ human breast cancer cell lines (T47D and MCF-7) that express high levels of ErbB-4, We have also investigated the expression of ErbB-4 in human primary breast carcinoma specimens, using immunohistochemical staining with an anti-ErbB-4 monoclonal antibody, ErbB-4 expression was found in 60% of the 50 primary breast tumors examined, and high intense immunoreactivity of ErbB-4 was det ected in 18% of these primary breast tumors. ErbB-4 receptor expression app eared to correlate with ER+ primary breast tumors. A similar correlation wa s also observed in the human breast cancer cell lines. These results provide a better understanding of the biological significance of ErbB-4 receptor in breast cancer. Our data suggest that elevation of th e ErbB-4 receptor plays a role in ER+ breast cancer cell proliferation. Mor eover, ribozyme technology provides a useful tool to delineate the role of a particular gene product.