Abstract
Nutlin-3a is a preclinical drug that stabilizes p53 by blocking the interaction between p53 and MDM2. In our previous study, Nutlin-3a promoted a tetraploid G(1) arrest in two p53 wild-type cell lines (HCT116 and U2OS), and both cell lines underwent endoreduplication after Nutlin-3a removal. Endoreduplication gave rise to stable tetraploid clones resistant to therapy-induced apoptosis. Prior knowledge of whether cells are susceptible to Nutlin-induced endoreduplication and therapy resistance could help direct Nutlin-3a-based therapies. In the present study, Nutlin-3a promoted a tetraploid G(1) arrest in multiple p53 wild-type cell lines. However, some cell lines underwent endoreduplication to relatively high extents after Nutlin-3a removal whereas other cell lines did not. The resistance to endoreduplication observed in some cell lines was associated with a prolonged 4N arrest after Nutlin-3a removal. Knockdown of either p53 or p21 immediately after Nutlin-3a removal could drive endoreduplication in otherwise resistant 4N cells. Finally, 4N-arrested cells retained persistent p21 expression; expressed senescence-associated beta-galactosidase; displayed an enlarged, flattened phenotype; and underwent a proliferation block that lasted at least 2 weeks after Nutlin-3a removal. These findings demonstrate that transient Nutlin-3a treatment can promote an apparently permanent proliferative block in 4N cells of certain cell lines associated with persistent p21 expression and resistance to endoreduplication.
Highlights
JULY 23, 2010 VOLUME 285 NUMBER 30 results in the stabilization and activation of p53 and subsequent p53-dependent growth arrest and/or apoptosis [4, 5]
These findings demonstrate that transient Nutlin-3a treatment can promote an apparently permanent proliferative block in 4N cells of certain cell lines associated with persistent p21 expression and resistance to endoreduplication
We reported that two p53 (DO-1; Santa Cruz Biotechnology) and p21 (F-5; Santa p53 wild-type cancer cell lines (U2OS and HCT116) treated
Summary
Dept. of Anatomy and Cell Biology, Rush University Medical Center, 1750 W. Exposure to 10 M Nutlin-3a for up to 6 days induced a senescent-like arrest that included expression of senescence-associated -galactosidase in multiple p53 wild-type cancer cell lines. Despite this arrest, nearly 100% of Nutlin-3a-treated cells resumed cycling after Nutlin removal, as determined by BrdUrd incorporation. 4N-arrested cells retained persistent p21 expression and displayed an enlarged, flattened phenotype and proliferation block that lasted at least 2 weeks after Nutlin-3a removal These findings demonstrate that transient Nutlin-3a treatment can promote an apparently permanent proliferative block in 4N cells of certain cell lines associated with persistent p21 expression and resistance to endoreduplication
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