Ricin binding and protein synthesis inhibition in human hematopoietic cell lines.
Previous studies showed that human blood cells exhibited varying sensitivities to ricin. To investigate the basis for these differences, ricin binding to human hematopoietic cell lines was assessed and correlated with in vitro ricin sensitivities. Resistant mutants were also isolated and characterized. Ricin binding to CEM cells was rapid, time-dependent, and blocked by unlabeled ricin, but not albumin; ricin binding approached saturation at 3 mumol/L. Scatchard analyses showed multiple classes of binding sites, with maximum and minimum Kd values estimated at 1.5 x 10(-8) mol/L and 2.5 x 10(-7) mol/L. At 4 degrees C, membrane-bound ricin dissociated slowly from the cell surface in the presence of unlabeled ricin, but greater than 95% of the surface-bound ricin was removed with 0.1 mol/L lactose. At 37 degrees C, ricin dissociated from the cell surface with biphasic kinetics. Ricin uptake at 37 degrees C increased linearly for 15 to 30 minutes and plateaued at levels representing 12% to 29% of the amount of ricin bound at 4 degrees C, depending on the cell line. Ricin binding at 4 degrees C varied two- to fivefold among hematopoietic cell lines and was reduced approximately tenfold by incubation with lactose. When compared with parent CEM cells, ricin-resistant CEM variants showed a greater than 95% reduction in ricin binding and showed no detectable binding with lactose added. However, these cells were as sensitive as parent CEM cells to an anti-T-cell ricin immunoconjugate. For all cells examined, there was a close correlation (r = +.9) between ricin bound per cell and in vitro ricin sensitivity. Human hematopoietic cells show widely varying ricin binding, indicating major differences in the carbohydrate content or structure of surface glycoproteins and glycolipids. These variations are probably the major determinant of nonspecific toxicity of ricin immunoconjugates.
Leonard
J E
JE
UC San Diego Cancer Center, La Jolla, CA 92037.
Grothaus
C D
CD
Taetle
R
R
D002460:Cell Line; D004351:Drug Resistance / Q000235:genetics; D006801:Humans; D009154:Mutation; D014411:Neoplastic Stem Cells / Q000187:drug effects* / Q000378:metabolism; D054198:Precursor Cell Lymphoblastic Leukemia-Lymphoma / Q000235:genetics / Q000378:metabolism; D011500:Protein Synthesis Inhibitors / Q000378:metabolism* / Q000633:toxicity; D011975:Receptors, Mitogen / Q000032:analysis* / Q000187:drug effects; D012276:Ricin / Q000378:metabolism* / Q000633:toxicity
D011500:Protein Synthesis Inhibitors; D011975:Receptors, Mitogen; D012276:Ricin