p21 Expression by human urothelial carcinoma cells modulates the phenotypic response to BCG.
OBJECTIVE
The direct phenotypic effects of BCG on human urothelial carcinoma (UC) cells include cell cycle arrest, apoptotic resistance, and caspase-independent cell death. These effects are associated with increased expression of the cyclin dependant kinase inhibitor (CDKI) p21. This study assessed the role of p21 expression in mediating the phenotypic effects observed in response to BCG.


METHODS
Inducible systems for the autocrine expression of p21, or the blockade of p21 expression in response to BCG, were established in the human UC line T24. The effect of increasing or inhibiting p21 expression on tumor phenotype was assessed using assays for cell cycle compartmentalization (flow cytometry), apoptotic sensitivity (caspase 3 activation), and cytotoxicity (vital dye exclusion).


RESULTS
p21 Overexpression resulted in cell cycle arrest with an increase in the percentage of the cell in G0/G1 phase when compared with the untreated group. p21 Expression decreased basal caspase-3 expression compared with the untreated group, and reversed camptothecin-induced caspase-3 activity compared with camptothecin alone group. p21 Overexpression increased BCG's direct cytotoxicity. shRNA-mediated inhibition of p21 expression in response to BCG failed to reverse BCG-induced changes in cell cycle compartmentalization. p21 Inhibition partially reversed the antiapoptotic effect of BCG. The expression of p21 was required for the direct cytotoxic effect of BCG.


CONCLUSIONS
p21 Expression is sufficient but not necessary for BCG-induced cell cycle arrest. It is both sufficient and necessary for the full antiapoptotic effect of BCG. p21 Expression alone is not sufficient for caspase-independent cytotoxicity but is necessary for BCG's direct cytotoxic effect.
See
William A
WA
Department of Urology, Medical College of Wisconsin, Milwaukee, WI 53226, USA. cziebell@mcw.edu
Zhang
Guangjian
G
Chen
Fanghong
F
Cao
Yanli
Y
D001500:BCG Vaccine / Q000627:therapeutic use*; D002166:Camptothecin / Q000494:pharmacology; D053148:Caspase 3 / Q000378:metabolism; D002453:Cell Cycle; D045744:Cell Line, Tumor; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000235:genetics / Q000502:physiology*; D004441:Ecdysterone / Q000031:analogs & derivatives / Q000494:pharmacology; D006801:Humans; D010641:Phenotype; D001749:Urinary Bladder Neoplasms / Q000378:metabolism / Q000473:pathology / Q000628:therapy*
D001500:BCG Vaccine; C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D004441:Ecdysterone; C008999:ponasterone A; D053148:Caspase 3; D002166:Camptothecin