title: Quantitative analysis of protein synthesis inhibition and recovery in CRM107 immunotoxin-treated HeLa cells.
abstract: Previously a mathematical model was proposed that quantitatively related protein synthesis inhibition kinetics of antitransferrin receptor-gelonin immunotoxins to the cellular trafficking of the targeting agent. That work is here extended to describe protein synthesis inhibition kinetics of immunotoxins containing the diphtheria toxin mutant CRM107. CRM107 differs from gelonin in both translocation and ribosomal inactivation mechanisms. Targeting agents used were antitransferrin monoclonal antibodies 5E9 and OKT9, OKT9Fab, and transferrin. CRM107 conjugates inhibited protein synthesis at substantially lower concentrations than gelonin conjugates; this effect was attributed to substantially higher translocation rates for CRM107. However, under certain conditions, CRM107 immunotoxin-treated cells were able to recover completely; this behavior was never observed with gelonin immunotoxins. To quantitatively capture this phenomenon, extracellular and cytosolic degradation of the toxin as well as growth-related recovery from toxin-induced damage were incorporated into the mathematical model. Translocation and cytosolic degradation rate constants were determined for each immunotoxin. Unlike the gelonin conjugates, the translocation rate of CRM107 conjugates depended on the targeting molecule. This provided indirect evidence that CRM107 remains disulfide linked to the targeting agent for at least part of the translocation process. Although the CRM107 conjugates all had higher translocation rates and inhibited protein synthesis at lower concentrations than the gelonin conjugates, the cells' ability to recover from protein synthesis inhibition at low immunotoxin concentrations limits the utility of CRM107 conjugates for targeted cell killing.
authors: Department of Chemical Engineering, University of Wisconsin, Madison, Wisconsin 53706, USA.
authors: L A
authors: LA
authors: Wenning
authors: P T
authors: PT
authors: Yazdi
authors: R M
authors: RM
authors: Murphy
mesh_terms: D000911:Antibodies, Monoclonal / Q000378:metabolism; D001692:Biological Transport; D002455:Cell Division; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D004167:Diphtheria Toxin / Q000235:genetics / Q000494:pharmacology; D006367:HeLa Cells / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000378:metabolism; D000922:Immunotoxins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D007700:Kinetics; D008433:Mathematics; D008954:Models, Biological; D014176:Protein Biosynthesis; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology*; D011506:Proteins / Q000032:analysis* / Q000187:drug effects; D014168:Transferrin / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D004167:Diphtheria Toxin; D007140:Immunoglobulin Fab Fragments; D000922:Immunotoxins; D011500:Protein Synthesis Inhibitors; D011506:Proteins; D014168:Transferrin