The binding of bombesin to its cognate G-protein coupled receptor stimulates quiescent Swiss 3T3 cells to re-initiate DNA synthesis and cell division. Addition of a non-mitogenic concentration of insulin dramatically potentiates bombesin-induced cell proliferation. We examined whether bombesin-induced EGFR transactivation mediates synergistic cell proliferation induced by bombesin and insulin. Treatment with selective EGFR tyrosine kinase inhibitors blocked EGFR transactivation, DNA synthesis, the transition of cells from quiescence into the cell cycle, and the expression of cyclins D1 and E induced by bombesin alone. In contrast, the inhibitors prevented cell cycle progression to a much lesser degree in cells stimulated with the combination of bombesin and insulin. Our results indicate that EGFR transactivation does not mediate synergistic cell proliferation induced by bombesin and insulin, and imply that insulin compensates for the requirement for EGFR transactivation in bombesin-induced DNA synthesis. (C) 2004 Elsevier Inc. All rights reserved.