Different early effets of tyrphostin AG957 and geldanamycins on mitogen-activated protein kinase and p120cbl phosphorylation in anti CD-3-stimulated T-lymphoblasts.
AG957, a tyrphostin tyrosine kinase inhibitor, has been shown previously to inhibit p210(bcr-abl) phosphorylation with concurrent inhibition of p210(bcr-abl)-expressing K562 cell growth (Kaur G and Sausville EA, Anticancer Drugs 7: 815-824, 1996). To assess the specificity of the action of AG957, we have examined its effect in another tyrosine kinase-mediated system, anti CD-3-stimulated Jurkat T Acute Lymphoblastic Leukemia cells. We also compared the effects of AG957 with those of geldanamycin, which can disrupt tyrosine kinase signaling through binding to heat shock protein (hsp90), and two geldanamycin analogs, 17-amino-17-demethoxygeldanamycin (17AG) and 17-allylamino-17-demethoxygeldanamycin (17AAG). At concentrations found to produce 90% inhibition of Jurkat T-cell growth, AG957 within 4 hr of addition inhibited mitogen-activated protein (MAP) kinase activation and activity, as shown by a decreased anti CD-3-stimulated erk-2 mobility shift in lysates of treated cells and a decrease in the stimulated myelin basic protein peptide kinase activity in erk-2 immunoprecipitates, respectively. AG957 did not inhibit this activity when added directly to immunoprecipitates. Effects in cells were found to be accompanied by a decrease in the anti CD-3-stimulated phosphorylation of p120cbl. Under conditions of a similar degree of growth inhibition, geldanamycin initially did not inhibit MAP kinase activation. Geldanamycin analogs did not decrease anti CD-3-induced cbl phosphorylation, but did reduce basal p120cbl tyrosine phosphorylation. The action of AG957 occurred with an apparent shift of several tyrosine-phosphorylated proteins to apparent higher molecular weights, which also did not occur with the geldanamycins. These results suggest that growth inhibition by AG957 can alter tyrosine kinase signaling systems unrelated to p210(bcr-abl) with a prominent early effect on MAP kinase activation in T-lymphoblasts. AG957 and geldanamycin affect tyrosine kinase signaling by distinct mechanisms.
Losiewicz
M D
MD
Pharmacology and Experimental Therapeutics Section, Laboratory of Drug Discovery Research and Development, Developmental Therapeutics Program, Division of Cancer Treatment and Diagnosis, National Cancer Institute-Frederick Cancer.
Kaur
G
G
Sausville
E A
EA
D000906:Antibodies / Q000097:blood; D016227:Benzoquinones; D017252:CD3 Complex / Q000276:immunology; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D019169:Jurkat Cells; D047029:Lactams, Macrocyclic; D008213:Lymphocyte Activation; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D050721:Proto-Oncogene Proteins c-cbl; D011809:Quinones / Q000494:pharmacology*; D015398:Signal Transduction / Q000187:drug effects; D013234:Stem Cells / Q000276:immunology; D013601:T-Lymphocytes / Q000276:immunology; D020032:Tyrphostins / Q000494:pharmacology*; D044767:Ubiquitin-Protein Ligases
D000906:Antibodies; D016227:Benzoquinones; D017252:CD3 Complex; D004791:Enzyme Inhibitors; D047029:Lactams, Macrocyclic; D011518:Proto-Oncogene Proteins; D011809:Quinones; D020032:Tyrphostins; C104965:tyrphostin AG957; D050721:Proto-Oncogene Proteins c-cbl; D044767:Ubiquitin-Protein Ligases; D011505:Protein-Tyrosine Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C067419:CBL protein, human; C001277:geldanamycin