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An Orthotopic Bladder Cancer Model for Gene Delivery Studies
Published on: December 2, 2013
The growth inhibitory effect of p21 adenovirus on human bladder cancer cells
1Department of Urology, University of Texas Southwestern Medical Center, Dallas 75235-9110, USA.
Purpose:
To evaluate whether p21 (WAF-1/CIP1) should be considered a potential candidate for human bladder cancer gene therapy, we determined: (1) the basal level of p21 expression in bladder cancer cell lines, (2) the response of bladder cancer cells to increased p21 expression following p21 adenovirus infection, and (3) the mechanism of growth inhibition produced by p21 overexpression.
Materials And Methods:
Five established human bladder cancer cell lines and one primary culture derived from an invasive transitional cell carcinoma were used in this study. To examine the effect of p21 protein on the growth of human bladder cancer cells, a recombinant adenovirus vector system containing p21 cDNA, under the control of cytomegalovirus promoter, was constructed. A control virus containing p21 in an antisense orientation was used to eliminate potential artifacts caused by viral toxicity.
Results:
Human bladder cancer cell lines exhibit variable endogenous p21 levels which correlate with the in vitro growth status. Significant, but highly variable increases in the steady-state level of p21 were detected in p21 adenovirus infected cells. Human bladder cancer cell lines responded heterogeneously to p21 adenovirus infection. Growth of the WH cell line was substantially inhibited in a dose and time-course dependent fashion. The mechanism of p21 growth inhibition was found to be due to G0/G1 arrest and not the induction of apoptosis. In contrast, p21 adenovirus failed to inhibit the growth of T24 bladder cancer cells because T24 cells were resistant to viral infection. The 253J bladder cancer cells exhibited marked sensitivity to adenovirus; substantial growth inhibition was seen with both sense and antisense p21 very early in the time course of infection.
Conclusions:
We found significant variation in the basal level of p21 protein expression in several human bladder cancer cell lines. Increased p21 expression as a result of adenoviral infection may be a potent growth suppressor in some human bladder cancer because it elicits cell cycle arrest in G0/G1 stage, but not the induction of apoptosis. Bladder cancer cells exhibit a wide spectrum of sensitivity to adenoviral infection that may be caused by the presence of viral receptor heterogeneity. This wide spectrum of sensitivity has significant basic scientific and clinical implications and warrants further study.
Insights
Gene therapy using p21 (WAF-1/CIP1) shows potential for bladder cancer by inducing cell cycle arrest. However, variable cell sensitivity to adenovirus vectors requires further investigation for clinical application.
Area of Science:
- Oncology
- Molecular Biology
- Gene Therapy
Background:
- The protein p21 (WAF-1/CIP1) plays a crucial role in cell cycle regulation.
- Understanding p21's role in bladder cancer is essential for developing targeted therapies.
- Gene therapy approaches are being explored for various cancers, including bladder cancer.
Purpose of the Study:
- To assess p21 (WAF-1/CIP1) as a candidate for human bladder cancer gene therapy.
- To determine basal p21 expression levels in bladder cancer cell lines.
- To investigate the response of bladder cancer cells to p21 overexpression via adenovirus infection and its growth inhibition mechanisms.
Main Methods:
- Utilized five human bladder cancer cell lines and one primary culture.
- Constructed a recombinant adenovirus vector carrying p21 cDNA under a CMV promoter.
- Employed a control virus with antisense p21 to mitigate viral toxicity artifacts.
Main Results:
- Bladder cancer cell lines displayed variable endogenous p21 levels correlating with growth status.
- Adenovirus-mediated p21 overexpression led to significant but heterogeneous increases in p21 levels.
- Growth inhibition occurred via G0/G1 cell cycle arrest, not apoptosis, in sensitive cell lines (e.g., WH).
- T24 cells showed resistance to p21 adenovirus due to poor viral infection, while 253J cells were highly sensitive.
Conclusions:
- Basal p21 protein expression varies significantly across human bladder cancer cell lines.
- Adenoviral p21 overexpression can inhibit bladder cancer growth by inducing G0/G1 cell cycle arrest.
- Heterogeneity in bladder cancer cell sensitivity to adenovirus infection, potentially due to viral receptor differences, has significant implications for gene therapy development.
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