Growth stimulation of A431 cells by epidermal growth factor: Identification of high-affinity receptors for epidermal growth factor by an anti-receptor monoclonal antibody

Epidermal growth factor (EGF) at 3 nM maximally inhibits the proliferation of A431 epidermoid carcinoma cells. We show that at lower concentrations, in the range of 3-100 pM, EGF has a mitogenic effect on A431 cells. In the presence of 100 nM anti-EGF-receptor monoclonal IgG (designated 528), which inhibits A-431 cell proliferation and blocks >95% of EGF binding, EGF becomes mitogenic for A431 cells at concentrations up to 3 nM. These results suggest that a minor population of high-affinity EGF receptors may be involved in stimulation of A431 cell proliferation. Saturation binding assays with ¹²⁵I-labeled EGF indicate that ≃0.1-0.2% of receptors for EGF are high-affinity receptors that bind EGF with an estimated K(d) of 7 x 10^-11 M. This affinity is nearly 2 orders of magnitude higher than that of the remaining EGF receptors. Although A431 cell proliferation is maximally inhibited by nonsaturating amounts of EGF (3 nM), maximal inhibition by 528 IgG (≃70% of maximal inhibition by EGF) requires saturating concentrations of antibody (≃15 nM). Unlike EGF, rapid down-regulation is not observed with 528 IgG. These results indicate different mechanisms of growth inhibition of A431 cells by EGF and 528 IgG.