AMP-activated protein kinase protects against anti-epidermal growth factor receptor-Pseudomonas exotoxin A immunotoxin-induced MA11 breast cancer cell death.
We have shown previously that our 425.3PE immunotoxin inhibits protein synthesis and induces apoptosis in human breast cancer cells. In attempts to further elucidate the intracellular pathways implicated in its cellular effects, we found that the immunotoxin induced an initial stress response, which rapidly caused an imbalance in the cellular energy status with an increase in reactive oxygen species. The AMP-activated protein kinase (AMPK), a sensor of increased cellular AMP/ATP ratio, was activated by 425.3PE. An immunotoxin-induced activation of c-Jun NH2-terminal kinase (JNK) preceded and overlapped caspase-mediated cleavage of the alpha-subunit of AMPK in a time- and dose-dependent manner. The JNK activation occurred already at a dose level too low to induce any detectable changes in the apoptotic machinery or protein synthesis. In contrast, cycloheximide, even at a concentration causing a 90% inhibition of protein synthesis, did neither affect the ATP level nor activate JNK and AMPK. Pretreatment of the cells with the specific AMPK inhibitor compound C and JNK inhibitor SP600125 blocked activation of AMPK and JNK, respectively, and subsequently sensitized the cells to 425.3PE-induced cell death. Whereas the antioxidant N-acetyl-l-cysteine blocked the generation of reactive oxygen species and activation of JNK and AMPK, it did not block immunotoxin-induced apoptosis. Together, the results show that 425.3PE induces several parallel signaling events, observed initially as an early activation of survival pathways, protecting the cells against the toxic effects of the immunotoxin, followed by subsequent apoptosis induction and protein synthesis inhibition. Conceivably, therapeutic manipulation of the signaling intermediates AMPK and JNK might provide a means to maximize the anticancer effects of the 425.3 immunotoxin.
Andersson
Yvonne
Y
Department of Tumor Biology and Institute for Cancer Research, Norwegian Radium Hospital, 0310 Oslo, Norway. y.g.andersson@medisin.uio.no
Le
Hang
H
Juell
Siri
S
Fodstad
Øystein
Ø
D036002:ADP Ribose Transferases / Q000633:toxicity*; D055372:AMP-Activated Protein Kinases; D000255:Adenosine Triphosphate / Q000378:metabolism; D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000906:Antibodies / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D001427:Bacterial Toxins / Q000633:toxicity*; D001943:Breast Neoplasms; D020169:Caspases / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects*; D045744:Cell Line, Tumor; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D066246:ErbB Receptors / Q000276:immunology*; D005098:Exotoxins / Q000633:toxicity*; D006801:Humans; D048670:MAP Kinase Kinase 4 / Q000378:metabolism; D009097:Multienzyme Complexes / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D037521:Virulence Factors / Q000633:toxicity*; D000097668:Pseudomonas aeruginosa Exotoxin A
D036002:ADP Ribose Transferases; D055372:AMP-Activated Protein Kinases; D000255:Adenosine Triphosphate; D000590:Amino Acid Chloromethyl Ketones; D000906:Antibodies; D001427:Bacterial Toxins; D020169:Caspases; D015853:Cysteine Proteinase Inhibitors; D066246:ErbB Receptors; D005098:Exotoxins; D048670:MAP Kinase Kinase 4; D009097:Multienzyme Complexes; D017346:Protein Serine-Threonine Kinases; D037521:Virulence Factors; D000097668:Pseudomonas aeruginosa Exotoxin A; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone