Adenoviral transfer of mda-7 leads to BAX up-regulation and apoptosis in mesothelioma cells, and is abrogated by over-expression of BCL-XL.
BACKGROUND
Malignant pleural mesothelioma (MPM) is unresponsive to conventional therapies. Forced expression of the novel tumor suppressor mda-7 gene in other cell types has resulted in decreased growth and apoptosis. We evaluated cell growth, apoptosis and tumor suppressor characteristics following forced expression of this gene in mesothelioma cell lines.


METHODS
MDA-7 expression in human MPM cells at baseline, following pharmacologic differentiation and viral mda-7 transduction (Ad-mda7) were evaluated with Western blot. Cell viability was evaluated with a colorimetric (XTT) assay, and apoptosis with subG1 FACS and Hoescht. Caspase-3 expression was evaluated by functional assay. These parameters were also evaluated in a stable bcl-xl hyper-expressing MPM cell line. Bax mRNA levels were evaluated with real-time PCR.


RESULTS
No baseline or differentiated MPM MDA7 expression was found, but was noted following Ad-mda7 exposure. More than 50% of MPM cells were killed at 5 days following Ad-mda7 exposure (p < 0.001). Apoptosis was accompanied by caspase-3 cleavage and increased BAX expression at both the protein (translational) and mRNA (transcriptional) level. These findings were reduced in a bcl-xl hyper-expressing cell line (P < 0.01).


CONCLUSIONS
Although mda-7 does not appear to be a MPM suppressor gene, adenoviral-mediated expression in cell lines induces apoptotic cellular death related to BAX upregulation and caspase cleavage. This is supported by abrogation of effect in a bcl-xl hyper-expressing cell line.
Cao
Xiaobo X
XX
Department of Thoracic and Cardiovascular Surgery, Section of Thoracic Molecular Oncology, The University of Texas M.D. Anderson Cancer Center, Houston, USA.
Mohuiddin
Imran
I
Chada
Sunil
S
Mhashilkar
Abner M
AM
Ozvaran
Mustafa K
MK
McConkey
David J
DJ
Miller
Steven D
SD
Daniel
Jonathon C
JC
Smythe
W Roy
WR
D000256:Adenoviridae; D017209:Apoptosis / Q000502:physiology*; D020169:Caspases / Q000378:metabolism; D002454:Cell Differentiation / Q000502:physiology; D002460:Cell Line; D016147:Genes, Tumor Suppressor; D005822:Genetic Vectors; D006801:Humans; D007378:Interleukins / Q000235:genetics / Q000378:metabolism*; D008654:Mesothelioma / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D015854:Up-Regulation / Q000502:physiology*; D051028:bcl-2-Associated X Protein; D051020:bcl-X Protein; D000099180:Interleukin-24
D020169:Caspases; D007378:Interleukins; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D051020:bcl-X Protein; D000099180:Interleukin-24; C494826:BAX protein, human; C494811:BCL2L1 protein, human