title: Inhibition of epidermal growth factor-mediated ERK1/2 activation by in situ electroporation of nonpermeant [(alkylamino)methyl]acrylophenone derivatives.
abstract: The interruption of signaling cascades in intact cells through the introduction of nonpermeant compounds inferred by in vitro studies to specifically inhibit epidermal growth factor (EGF) receptor (EGF-R) function is described. Two nonpermeant [(alkylamino)methyl]acrylophenone derivatives, [(dimethylamino)methyl] acrylo-para-[(benzoylsulfonyl)-oxy]phenone and [(dimethylamino)-methyl]acrylo-para-[(hydroxy-benzoylsulfonyl++ +)-oxy]phenone, were introduced by in situ electroporation into mouse or rat fibroblasts growing on indium-tin oxide-coated glass. Cells were subsequently stimulated with growth factors and assessed for activation of a downstream target, the extracellular signal-regulated kinase (ERK1/2), by probing with specific antibodies. Electrodes and slides were configured to provide non-electroporated control cells side by side with the electroporated ones, both growing on the same type of indium-tin oxide-coated glass surface. Using this set-up, these compounds could inhibit EGF- but not platelet-derived growth factor (PDGF)-mediated ERK1/2 activation in vivo. These results demonstrate the potential of the in situ electroporation approach for the study of tyrosine kinase action using selective but nonpermeant inhibitors that would otherwise be ineffective in intact cells.
authors.affiliation: Department of Microbiology, Queen's University, Kingston, Ontario, Canada.
authors.forename: H L
authors.initials: HL
authors.lastname: Brownell
authors.forename: N B
authors.initials: NB
authors.lastname: Lydon
authors.forename: E
authors.initials: E
authors.lastname: Schaefer
authors.forename: T M
authors.initials: TM
authors.lastname: Roberts
authors.forename: L
authors.initials: L
authors.lastname: Raptis
mesh_terms: D016475:3T3 Cells / Q000187:drug effects; D000818:Animals; D000906:Antibodies; D001557:Benzenesulfonates / Q000494:pharmacology*; D001565:Benzoates / Q000494:pharmacology*; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000276:immunology / Q000378:metabolism*; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D016895:Culture Media, Serum-Free; D018274:Electroporation / Q000379:methods*; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D004815:Epidermal Growth Factor / Q000494:pharmacology; D066246:ErbB Receptors / Q000378:metabolism; D005347:Fibroblasts / Q000187:drug effects; D051379:Mice; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D051381:Rats; D017479:Receptors, Platelet-Derived Growth Factor / Q000378:metabolism
chemical_list: C111575:((dimethylamino)methyl)acrylo-4-((benzoylsulfonyl)oxy)phenone; C111576:((dimethylamino)methyl)acrylo-4-((hydroxybenzoylsulfonyl)oxy)phenone; D000906:Antibodies; D001557:Benzenesulfonates; D001565:Benzoates; D016895:Culture Media, Serum-Free; D004791:Enzyme Inhibitors; D010982:Platelet-Derived Growth Factor; D004815:Epidermal Growth Factor; D066246:ErbB Receptors; D017479:Receptors, Platelet-Derived Growth Factor; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases