Adenoviral-mediated gene transfer of a constitutively active retinoblastoma gene inhibits human pancreatic tumor cell proliferation.
BACKGROUND
The development of pancreatic cancer involves an accumulation of genetic changes, including oncogene activation and mutations in tumor suppressor genes important in cell cycle regulation. As a step to developing gene therapies to restore cell cycle control, we hypothesized that adenoviral-mediated gene transfer of a constitutively active, nonphosphorylatable form of the retinoblastoma gene (AdRb) would inhibit human pancreatic tumor cell proliferation.


METHODS
Transfection efficiency was assessed by beta-gal staining with an adenovirus expressing the beta-galactosidase gene (AdLacZ). The effect of AdRb on DNA synthesis in pancreatic cancer cell lines was determined by tritiated thymidine incorporation. Western blotting with an antihemagglutinin antibody directed to the hemagglutinin-tagged AdRb construct was performed to confirm transfection of pancreatic cancer cells. Apoptosis was evaluated with a TUNEL assay.


RESULTS
Efficient transfection of human pancreatic cancer cell lines was achieved with AdLacZ. AdRb inhibited tritiated thymidine uptake in the cancer cell lines BxPC-3, MIA PaCa-2, and PANC-1. Western blotting confirmed transfection of cancer cells with AdRb. AdRb did not inhibit growth by apoptosis.


CONCLUSIONS
Adenoviral-mediated gene delivery of constitutively active Rb produces significant growth inhibition in human pancreatic cancer cell lines and is not a result of apoptosis. Further studies examining the role of Rb in pancreatic cancer are warranted.
Simeone
D M
DM
Department of Surgery, University of Michigan Medical School, Ann Arbor, USA.
Cascarelli
A
A
Logsdon
C D
CD
D000256:Adenoviridae; D002455:Cell Division; D004273:DNA, Neoplasm / Q000096:biosynthesis; D016161:Genes, Retinoblastoma; D005822:Genetic Vectors; D006801:Humans; D007700:Kinetics; D010190:Pancreatic Neoplasms / Q000473:pathology*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D016160:Retinoblastoma Protein / Q000096:biosynthesis*; D013936:Thymidine / Q000378:metabolism; D013997:Time Factors; D014162:Transfection; D014407:Tumor Cells, Cultured; D001616:beta-Galactosidase / Q000096:biosynthesis
D004273:DNA, Neoplasm; D011993:Recombinant Fusion Proteins; D016160:Retinoblastoma Protein; D001616:beta-Galactosidase; D013936:Thymidine