17-Allylamino-17-demethoxygeldanamycin enhances the lethality of deoxycholic acid in primary rodent hepatocytes and established cell lines.
Ansamycin antibiotics that target heat shock protein 90 function are being developed as anticancer agents but are also known to be dose limiting in patients due to hepatotoxicity. Herein, to better understand how the normal tissue toxicity of geldanamycins could be ameliorated to improve the therapeutic index of these agents, we examined the interactions of 17-allylamino-17-demethoxygeldanamycin (17AAG) and the secondary bile acid deoxycholic acid (DCA) in hepatocytes and fibroblasts. DCA and 17AAG interacted in a greater than additive fashion to cause hepatocyte cell death within 2 to 6 h of coadministration. As single agents DCA, but not 17AAG, enhanced the activity of extracellular signal-regulated kinase 1/2, AKT, c-Jun NH(2)-terminal kinase 1/2 (JNK1/2), and p38 mitogen-activated protein kinase (MAPK). Combined exposure of cells to DCA and 17AAG further enhanced JNK1/2 and p38 MAPK activity. Inhibition of JNK1/2 or p38 MAPK, but not activator protein-1, suppressed the lethality of 17AAG and of 17AAG and DCA. Constitutive activation of AKT, but not MAPK/extracellular signal-regulated kinase kinase 1/2, suppressed 17AAG- and DCA-induced cell killing and reduced activation of JNK1/2. DCA and 17AAG exposure promoted association of BAX with mitochondria, and functional inhibition of BAX or caspase-9, but not of BID and caspase-8, suppressed 17AAG and DCA lethality. DCA and 17AAG interacted in a greater than additive fashion to promote and prolong the generation of reactive oxygen species (ROS). ROS-quenching agents, inhibition of mitochondrial function, expression of dominant-negative thioredoxin reductase, or expression of dominant-negative apoptosis signaling kinase 1 suppressed JNK1/2 and p38 MAPK activation and reduced cell killing after 17AAG and DCA exposure. The potentiation of DCA-induced ROS production by 17AAG was abolished by Ca(2+) chelation and ROS generation, and cell killing following 17AAG and DCA treatment was abolished in cells lacking expression of PKR-like endoplasmic reticulum kinase. Thus, DCA and 17AAG interact to stimulate Ca(2+)-dependent and PKR-like endoplasmic reticulum kinase-dependent ROS production; high levels of ROS promote intense activation of the p38 MAPK and JNK1/2 pathways that signal to activate the intrinsic apoptosis pathway.
Mitchell
Clint
C
Department of Biochemistry, Massey Cancer Center, Box 980035, Virginia Commonwealth University, Richmond, VA 23298-0035, USA.
Park
Margaret A
MA
Zhang
Guo
G
Han
Song Iy
SI
Harada
Hisashi
H
Franklin
Richard A
RA
Yacoub
Adly
A
Li
Pin-Lan
PL
Hylemon
Philip B
PB
Grant
Steven
S
Dent
Paul
P
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D016227:Benzoquinones / Q000494:pharmacology*; D015153:Blotting, Western; D002118:Calcium; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy / Q000378:metabolism; D002518:Ceramides / Q000378:metabolism; D003840:Deoxycholic Acid / Q000494:pharmacology*; D003902:Detergents / Q000494:pharmacology*; D004359:Drug Therapy, Combination; D022781:Hepatocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D047029:Lactams, Macrocyclic / Q000494:pharmacology*; D008113:Liver Neoplasms / Q000188:drug therapy / Q000378:metabolism; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors; D051381:Rats; D017207:Rats, Sprague-Dawley; D026361:Reactive Nitrogen Species / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism
D016227:Benzoquinones; D002518:Ceramides; D003902:Detergents; D047029:Lactams, Macrocyclic; D026361:Reactive Nitrogen Species; D017382:Reactive Oxygen Species; D003840:Deoxycholic Acid; C112765:tanespimycin; D017346:Protein Serine-Threonine Kinases; D020928:Mitogen-Activated Protein Kinases; D002118:Calcium