Abstract
We observed that glioma cells are differentially sensitive to N-{4-[4-(4′-chloro-biphenyl-2-ylmethyl)-piperazin-1-yl]-benzoyl}-4-(3-dimethylamino-1-phenylsulfanylmethyl-propylamino)-3-nitro-benzenesulfonamide (ABT-737) and administration of ABT-737 at clinically achievable doses failed to induce apoptosis. Although elevated Bcl-2 levels directly correlated with sensitivity to ABT-737, overexpression of Bcl-2 did not influence sensitivity to ABT-737. To understand the molecular basis for variable and relatively modest sensitivity to the Bcl-2 homology domain 3 mimetic drug ABT-737, the abundance of Bcl-2 family members was assayed in a panel of glioma cell lines. Bcl-2 family member proteins, Bcl-xL, Bcl-w, Mcl-1, Bax, Bak, Bid, and Noxa, were found to be expressed ubiquitously at similar levels in all cell lines tested. We then examined the contribution of other apoptosis-resistance pathways to ABT-737 resistance. Bortezomib, an inhibitor of nuclear factor-kappaB (NF-κB), was found to enhance sensitivity of ABT-737 in phosphatase and tensin homolog on chromosome 10 (PTEN)-wild type, but not PTEN-mutated glioma cell lines. We therefore investigated the association between phosphatidylinositol 3-kinase (PI3K)/Akt activation and resistance to the combination of ABT-737 and bortezomib in PTEN-deficient glioma cells. Genetic and pharmacological inhibition of PI3K inhibition sensitized PTEN-deficient glioma cells to bortezomib- and ABT-737-induced apoptosis by increasing cleavage of Bid protein, activation and oligomerization of Bax, and loss of mitochondrial membrane potential. Our data further suggested that PI3K/Akt-dependent protection may occur upstream of the mitochondria. This study demonstrates that interference with multiple apoptosis-resistance signaling nodes, including NF-κB, Akt, and Bcl-2, may be required to induce apoptosis in highly resistant glioma cells, and therapeutic strategies that target the PI3K/Akt pathway may have a selective role for cancers lacking PTEN function.
Footnotes
This work was supported by the National Institutes of Health National Institute of Neurological Disorders and Stroke [Grant P01NS40923] (to I.F.P.); and The Walter L. Copeland Fund of The Pittsburgh Foundation (D.R.P.).
Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
ABBREVIATIONS:
- NF-κB
- nuclear factor-kappaB
- Ad
- adenovirus
- BH3
- Bcl-2 homology domain 3
- CHAPS
- 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acid
- CMV
- cytomegalovirus
- DiOC6
- 3′,3′-dihexyloxacarbo-cyanine iodide
- DMSO
- dimethyl sulfoxide
- DN
- dominant negative
- DSP
- dithiobis(succinimidylpropionate)
- E64d
- (2S,3S)-trans-epoxysuccinyl-l-leucylamido-3-methylbutane ethyl ester
- MTS
- 3-[4,5-dimethylthiazol- 2yl]-5-[3-carboxymethoxyphenyl]-2-[4-sulfophenyl]-2H, tetrazolium
- FACS
- fluorescence-activated cell sorting
- FITC
- fluorescein isothiocyanate
- HDACI
- histone deacetylase inhibitor
- MOI
- multiplicity of infection
- NAC
- N-acetyl-l-cysteine
- PBS
- phosphate-buffered saline
- PAGE
- polyacrylamide gel electrophoresis
- PARP
- poly(ADP)-ribose polymerase
- PI
- propidium iodide
- PI3K
- phosphatidylinositol 3-kinase
- PTEN
- phosphatase and tensin homolog on chromosome 10
- ROS
- reactive oxygen species
- Smac/DIABLO
- second mitochondria-derived activator of caspases/direct inhibitor of apoptosis binding protein with low PI
- t-Bid
- truncated Bid
- TRAIL
- tumor necrosis factor-related apoptosis-inducing ligand
- ABT-737
- N-{4-[4-(4′-chloro-biphenyl-2-ylmethyl)-piperazin-1-yl]-benzoyl}-4-(3-dimethylamino-1-phenylsulfanylmethyl-propylamino)-3-nitro-benzenesulfonamide
- ABT-263
- (R)-4-(4-((4′-chloro-4,4-dimethyl-3,4,5,6-tetrahydro-[1,1′-biphenyl]-2-yl)methyl)piperazin-1-yl)-N-((4-((4-morpholino-1-(phenylthio)butan-2-yl)amino)-3-((trifluoromethyl)sulfonyl)phenyl)sulfonyl)benzamide
- PI-103
- 3-[4-(4-morpholinylpyrido[3′,2′:4,5]furo[3,2-d]pyrimidin-2-yl]phenol
- LY294002
- 2-morpholin-4-yl-8-phenylchromen-4-one
- Δψm
- mitochondrial membrane potential.
- Received January 6, 2012.
- Accepted March 2, 2012.
- Copyright © 2012 by The American Society for Pharmacology and Experimental Therapeutics
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