Abstract
Melanoma differentiation associated gene-7/interleukin 24 (mda-7/IL-24) is a unique interleukin (IL)-10 family cytokine displaying selective apoptosis-inducing activity in transformed cells without harming normal cells. The present studies focused on defining the mechanism(s) by which recombinant adenoviral delivery of MDA-7/IL-24 inhibits cell survival of human ovarian carcinoma cells. Expression of MDA-7/IL-24 induced phosphorylation of protein kinase R-like endoplasmic reticulum kinase (PERK) and eukaryotic initiation factor2α (eIF2α). In a PERK-dependent fashion, MDA-7/IL-24 reduced ERK1/2 and AKT phosphorylation and activated c-Jun NH2-terminal kinase (JNK) 1/2 and p38 mitogen-activated protein kinase (MAPK). MDA-7/IL-24 reduced MCL-1 and BCL-XL and increased BAX levels via PERK signaling; cell-killing was mediated via the intrinsic pathway, and cell killing was primarily necrotic as judged using Annexin V/propidium iodide staining. Inhibition of p38 MAPK and JNK1/2 abolished MDA-7/IL-24 toxicity and blocked BAX and BAK activation, whereas activation of mitogen-activated extracellular-regulated kinase (MEK) 1/2 or AKT suppressed enhanced killing and JNK1/2 activation. MEK1/2 signaling increased expression of the MDA-7/IL-24 and PERK chaperone BiP/78-kDa glucose regulated protein (GRP78), and overexpression of BiP/GRP78 suppressed MDA-7/IL-24 toxicity. MDA-7/IL-24-induced LC3-green fluorescent protein vesicularization and processing of LC3; and knockdown of ATG5 suppressed MDA-7/IL-24-mediated toxicity. MDA-7/IL-24 and cisplatin interacted in a greater than additive fashion to kill tumor cells that was dependent on a further elevation of JNK1/2 activity and recruitment of the extrinsic CD95 pathway. MDA-7/IL-24 toxicity was enhanced in a weak additive fashion by paclitaxel; paclitaxel enhanced MDA-7/IL-24 + cisplatin lethality in a greater than additive fashion via BAX. Collectively, our data demonstrate that MDA-7/IL-24 induces an endoplasmic reticulum stress response that activates multiple proapoptotic pathways, culminating in decreased ovarian tumor cell survival.
Footnotes
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↵ The online version of this article (available at http://molpharm.aspetjournals.org) contains supplemental material.
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This work was supported by the National Institutes of Health National Cancer Institute [Grants P01-CA104177, R01-CA108325, R01-CA63753, R01-CA77141, R01-CA097318, R01-CA12764101]; the National Institutes of Health National Institute of Diabetes and Digestive and Kidney Diseases [Grant R01-DK52825]; the National Institutes of Health National Institute of Neurological Disorders and Stroke [Grant P01-NS031492]; the V Foundation; and The Samuel Waxman Cancer Research Foundation.
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Article, publication date, and citation information can be found at http://molpharm.aspetjournals.org.
doi:10.1124/mol.109.061820
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ABBREVIATIONS:
- MDA-7/IL-24
- melanoma differentiation associated gene-7/interleukin-24
- ERK
- extracellular signal-regulated kinase
- MEK
- mitogen-activated extracellular regulated kinase
- JNK
- c-Jun NH2-terminal kinase
- PERK
- protein kinase R-like endoplasmic reticulum kinase
- MAPK
- mitogen-activated protein kinase
- IL
- interleukin
- OCC
- ovarian cancer cell
- GFP
- green fluorescent protein
- ER
- endoplasmic reticulum
- GST
- glutathione transferase
- PAGE
- polyacrylamide gel electrophoresis
- m.o.i.
- multiplicity of infection
- CDDP
- cisplatin [cis-diamminedichloroplatinum(II)]
- FBS
- fetal bovine serum
- GBM
- glioblastoma
- GRP78
- 78-kDa glucose regulated protein
- PARP1
- poly(ADP-ribose) polymerase 1
- siRNA
- small interfering RNA
- IP
- inhibitory peptide
- GAPDH
- glyceraldehyde-3-phosphate dehydrogenase
- MCL-1
- myeloid cell leukemia sequence 1
- IRE1
- inositol requirement 1
- BBR3464
- (SP-4–1)-diamminebis((SP-4–2)-diamminechloroplatinum(π) (μ-hexane-1,6-diamine))platinum tetranitrate
- LY294002
- 2-(4-morpholinyl)-8-phenyl-1(4H)-benzopyran-4-one hydrochloride
- eIF2α
- eukaryotic initiation factor 2α.
- Received October 20, 2009.
- Accepted November 11, 2009.
- Copyright © 2010 The American Society for Pharmacology and Experimental Therapeutics
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