Effects of adenoviral wild-type p53 gene transfer in p53-mutated lymphoma cells.
The present study assessed the role of adenoviral vector-mediated wild-type p53 gene transfer in B lymphoma cells. Deficiency of p53-mediated cell death is common in human cancer contributing to both tumorigenesis and chemoresistance. Lymphoma cells are being considered as suitable targets for gene therapy protocols. Recently, we reported an adenoviral protocol leading to highly efficient gene transfer to B lymphoma cells. All lymphoma cell lines (n=5) tested here showed mutations in the p53 gene locus. The aim of this work was to transduce lymphoma cells with the wild-type p53 gene. Using this protocol, 88% of Raji, 75% of Daudi, and 45% of OCI-Ly8-LAM53 cells were transfected with the reporter gene green fluorescent protein at a multiplicity of infection of 200. The expression of green fluorescent protein in CA46 and BL41 cells was 27% and 42%, respectively. At this multiplicity of infection, growth characteristics of lymphoma cell lines were not changed significantly. In contrast, cells transduced with wild-type p53 gene showed an inhibition of proliferation as well as an increase in apoptosis. Cell loss by apoptosis after p53 gene transfer was up to 40% as compared to transduction with an irrelevant vector. In addition, we determined the effects of DNA damage produced by the DNA topoisomerase II inhibitor etoposide on wild-type p53 transfected lymphoma cells. In Ad-p53-transfected Raji cells, treatment with the drug resulted in a marked increase of cell loss in comparison to Ad-beta-Gal-transfected cells (45% vs. 77%). Interestingly, performing cytotoxicity studies, we could show an increased sensitivity of Raji and Daudi cells against immunological effector cells. In conclusion, transduction of wild-type p53 into lymphoma cells expressing mutated p53 was efficient and led to inhibition of proliferation and increase in apoptotic rate in some cell lines dependent on p53 mutation. This protocol should have an impact on the use of lymphoma cells in cancer gene therapy protocols.
Buttgereit
P
P
Medizinische Klinik und Poliklinik I, Rheinische Friedrich-Wilhelms Universität, Bonn, Germany.
Schakowski
F
F
Märten
A
A
Brand
K
K
Renoth
S
S
Ziske
C
C
Schöttker
B
B
Ebert
O
O
Schroers
R
R
Schmidt-Wolf
I G
IG
D000256:Adenoviridae / Q000235:genetics*; D017209:Apoptosis; D002455:Cell Division; D002460:Cell Line; D005047:Etoposide / Q000494:pharmacology; D005434:Flow Cytometry; D018014:Gene Transfer Techniques; D016158:Genes, p53 / Q000235:genetics*; D049452:Green Fluorescent Proteins; D006801:Humans; D015151:Immunoblotting; D007694:Killer Cells, Natural / Q000378:metabolism; D008164:Luminescent Proteins / Q000378:metabolism; D008223:Lymphoma / Q000235:genetics* / Q000378:metabolism; D008856:Microscopy, Fluorescence; D009154:Mutation; D009336:Necrosis; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D011993:Recombinant Fusion Proteins; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D014161:Transduction, Genetic; D014162:Transfection
D008164:Luminescent Proteins; D011500:Protein Synthesis Inhibitors; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D005047:Etoposide