17-Allylamino-17-demethoxygeldanamycin enhances the lethality of deoxycholic acid in primary rodent hepatocytes and established cell lines

Abstract

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 PKRlike 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. [Mol Cancer Ther 2007;6(2):618 -32]

Activation of focal adhesion kinase and JNK contributes to the extracellular matrix and cAMP-GEF mediated survival from bile acid induced apoptosis in rat hepatocytes

“…Since bile acid apoptotic signaling involves activation of the stress activated kinases, JNK and p38 MAPK (17,19,20), we examined whether adherence to different ECM influenced bile acid phosphorylation of these kinases. Hepatocytes plated on CI had a 4-5 fold increase in JNK phosphorylation in response to GCDC, while hepatocytes adhered to LM or PL had a 15 to 30 fold increase in GCDC induced JNK phosphorylation (Fig 8 A, B).…”

“…GCDC phosphorylated p38 MAPK equally on CI, PL and LM (Fig 9A,B). Since p38 MAPK signaling has been implicated in both pro- (19,20) and anti-apoptotic bile acid signaling (29,36), we wished to clarify the role of p38 MAPK in GCDC induced apoptosis in our system. Inhibition of p38 MAPK with SB203580, had no effect on GCDC induced apoptosis in hepatocytes plated on C1, PL or LM (Fig 9 C).…”

“…Bile acid mediated JNK activation is pro-apoptotic in hepatocytes (17,19,20). In the current study, hepatocytes plated on LM and PL had profound increases in JNK phosphorylation in response to GCDC compared to hepatocytes plated on Type 1 collagen, and chemical inhibition of JNK activation prevented bile acid induced apoptosis on all the matrices.…”

“…Bile acid induced apoptosis involves activation of the pro-apoptotic stress activated kinase, c-Jun N terminal kinase (17) and in some instances activation of p38 mitogen activated kinase kinase (p38 MAPK) (19,20). Emerging evidence suggests that bile acids may also modulate FAK.…”

Low-Dose BBR3610 Toxicity in Colon Cancer Cells Is p53-Independent and Enhanced by Inhibition of Epidermal Growth Factor Receptor (ERBB1)-Phosphatidyl Inositol 3 Kinase Signaling

“…The Bcl-xL and activated MEK1 EE adenoviruses were kindly provided by Dr. J. Moltken (University of Cincinnati, Cincinnati, OH). Other reagents were of the highest quality commercially available (Yacoub et al, 2006Mitchell et al, 2007).…”