Differential signaling by an anti-p185(HER2) antibody and heregulin

To understand the molecular mechanisms by which anti-p185(HER2) antibody an d the ligand heregulin inhibit tumor growth, we have investigated several s ignaling proteins and pathways, We report here that anti-p185(HER2) monoclo nal antibody ID5 induced tyrosine phosphorylation of HER2 in SKBr3 breast c ancer cells that overexpress p185(HER2). Heregulin beta 1 induced phosphory lation of both HER3 and HER2, ID5 produced a greater association of phospho lipase C (PLC)-gamma 1 with HER2 than did heregulin, Concordantly, ID5, but not heregulin, increased PLC-gamma 1 activity. However, the G(1) cell cycl e arrest and induction of p27(Kip1) produced by ID5 were not affected by th e inhibition of PLC-gamma. ID5 preferentially induced binding of the M-r 46 ,000 isoform of SHC to HER2, whereas heregulin preferentially induced bindi ng of the M-r 52,00 isoform of SHC to HER3, Heregulin, but not ID5, induced the p85 subunit of phosphatidylinositol 3'-kinase (PD-K) to interact with HER3. Heregulin induced sustained activation of PB-K signaling, whereas ID5 had only a transient effect, Heregulin, but not ID5, activated the c-Jun-N H2-terminal kinase cascade. Pretreatment of SKBr3 cells with ID5 decreased heregulin-induced association of HER2 with HER3 as well as the activation o f c-Jun-NH2-terminal kinase and PI3-K activities. Inhibition of the mitogen -activated protein kinase pathway in SKBr3 cells did not affect heregulin-i nduced G(2)-M-phase arrest, apoptosis, and differentiation. Heregulin-induc ed apoptosis could be blocked by inhibition of p70s6k, but not by inhibitio n of PI3-K, Heregulin-induced differentiation could be eliminated by inhibi tion of PI3-K, We conclude that ID5 and heregulin signal via different path ways, although both agents ran inhibit the clonogenic growth of cells that overexpress HER2.