Induction of apoptosis in human leukemia cells by the tyrosine kinase inhibitor adaphostin proceeds through a RAF-1/MEK/ERK- and AKT-dependent process.
Effects of the tyrphostin tyrosine kinase inhibitor adaphostin (NSC 680410) have been examined in human leukemia cells (Jurkat, U937) in relation to mitochondrial events, apoptosis, and perturbations in signaling and cell cycle regulatory events. Exposure of cells to adaphostin concentrations > or =0.75 microM for intervals > or =6 h resulted in a pronounced release of cytochrome c and AIF, activation of caspase-9, -8, and -3, and apoptosis. These events were accompanied by the caspase-independent downregulation of Raf-1, inactivation of MEK1/2, ERK, Akt, p70S6K, dephosphorylation of GSK-3, and activation of c-Jun-N-terminal kinase (JNK) and p38 MAPK. Adaphostin also induced cleavage and dephosphorylation of pRb on CDK2- and CDK4-specific sites, as well as the caspase-dependent downregulation of cyclin D1. Inducible expression of a constitutively active MEK1 construct markedly diminished adaphostin-induced cytochrome c and AIF release, JNK activation, and apoptosis in Jurkat cells. Ectopic expression of Raf-1 or constitutively activated (myristolated) Akt also significantly attenuated adaphostin-induced apoptosis, but protection was less than that conferred by enforced activation of MEK. Lastly, antioxidants (e.g., L-N-acetylcysteine; L-NAC) opposed adaphostin-mediated mitochondrial dysfunction, Raf-1/MEK/ERK downregulation, JNK activation, and apoptosis. However, in contrast to L-NAC, enforced activation of MEK failed to block adaphostin-mediated ROS generation. Together, these findings demonstrate that the tyrphostin adaphostin induces multiple perturbations in signal transduction pathways in human leukemia cells, particularly inactivation of the cytoprotective Raf-1/MEK/ERK and Akt cascades, that culminate in mitochondrial injury, caspase activation, and apoptosis. They also suggest that adaphostin-related oxidative stress acts upstream of perturbations in these signaling pathways to trigger the cell death process.
Yu
Chunrong
C
Department of Medicine, Virginia Commonwealth University, Medical College of Virginia, Richmond, VA 23298, USA.
Rahmani
Mohamed
M
Almenara
Jorge
J
Sausville
Edward A
EA
Dent
Paul
P
Grant
Steven
S
D000218:Adamantane / Q000031:analogs & derivatives* / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D051033:Apoptosis Inducing Factor; D045304:Cytochromes c / Q000187:drug effects; D005420:Flavoproteins / Q000187:drug effects; D006801:Humans; D006873:Hydroquinones / Q000494:pharmacology*; D019169:Jurkat Cells; D007938:Leukemia / Q000188:drug therapy*; D020930:MAP Kinase Kinase Kinases / Q000378:metabolism; D008565:Membrane Proteins / Q000187:drug effects; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors*; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt; D019908:Proto-Oncogene Proteins c-raf
C494829:AIFM1 protein, human; D051033:Apoptosis Inducing Factor; D005420:Flavoproteins; D006873:Hydroquinones; D008565:Membrane Proteins; C404397:NSC 680410; D011518:Proto-Oncogene Proteins; D045304:Cytochromes c; D011505:Protein-Tyrosine Kinases; C494918:AKT1 protein, human; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D019908:Proto-Oncogene Proteins c-raf; D020928:Mitogen-Activated Protein Kinases; D020930:MAP Kinase Kinase Kinases; D000218:Adamantane