p16^INK4A silencing augments DNA damage-induced apoptosis in cervical cancer cells Free

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Cancer of the cervix is the second most common cancer among women worldwide, with a higher prevalence in the developing countries. The p16^INK4A tumor suppressor gene has been reported to be frequently inactivated in human cancer. However, for cervical cancer cells, the accumulation of p16 was observed, possibly resulting from the inactivation of Rb by E7. Consequently, p16 has been suggested to be a potential early diagnostic bio-marker for cervical cancer. Despite this, the functional impact of p16 over-expression in cervical cancer has not been thoroughly investigated. In this study, we have demonstrated the consistent and significant up-regulation of p16 in cervical cancer tissues when compared to both matched non-tumourous tissues of the same patient and normal cervical tissues from non-cancerous patients. To gain further insights into the over-expression of p16 in cervical carcinogenesis, we employed p16-specific siRNA to down-regulate the expression of p16. Although the silencing of p16 was accompanied by the up-regulation of p53, p21 and Rb protein expression, no significant effect on cell cycle progression was observed. However, when the p16 siRNA-silenced cells were subjected to DNA damage stress including UV-irradiation and cisplatin treatments, a significantly higher percentage of apoptotic cells could be detected in the p16-siRNA silenced cells compared to control siRNA-treated cells. The induction of apoptosis was associated with the activation of p53 through phosphorylation, and this process, when analyzed by gene profiling experiments, demonstrated the involvement of both the intrinsic mitochondrial and extrinsic death-receptor apoptotic pathways. Of particular interest, it was noted that target genes related to p53-mediated DNA damage, including p21, PCNA, and GADD45A were up-regulated, while the anti-apoptotic genes BAG1, TIMP3, AKT1, MCL1 and FAIM were down-regulated. Similarly, apoptotic genes belonging to both the mitochondrial and death receptor pathways, such as caspase 8, Bcl family-related genes, APAF1, FAS, FADD and TNFR-family genes were also up-regulated in UV-irradiated, p16-silenced cervical cancer cells. In summary, the observation that p16-silencing could augment DNA damage-induced apoptosis in cervical cancer cells offers alternative strategies for anti-cancer therapies for human cervical cancer.