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Perforin-1 (PRF), a cytotoxic lymphocyte pore-forming protein, plays an important role in the action of cytotoxic T cells and natural killer cells in that it causes the lysis of abnormal body cells and the elimination of virus-infected cells and tumors. Upon degranulation, PRF inserts itself into the target cell's plasma membrane, forming a pore. The subsequent translocation of pro-apoptotic granzymes (including granzyme B, A, M et al.) into the cytoplasm provides the proteases with access to numerous protein substrates that promote apoptosis after cleavage. These proteases are believed to be the main executioners of target cell apoptosis. Although the PRF and granzyme components are both critical to this process and in some way involved in inducing cell death in target cells, the inhibition of tumor growth could still be efficient in granzymedeficient mice. It is unclear whether PRF alone can suppress tumors. In this study, we discovered that forced ectopic expression of PRF alone, in the absence of granzymes, could mediate cell death in cancer cells. Notably, transient expression of both full-length and truncated active-form PRF in human Hep G2, SK-BR-3, and HeLa cells was found to induce apparent cell growth inhibition and cell death, as evidenced by chromosome condensation and DNA fragmentation, increased caspase-3 activity, and the release of apoptosis inducing factor (AIF) and cytochrome c from the mitochondria. This PRFinduced cell death could be abrogated by pan-caspase inhibitor (Z-VAD) and mitochondria protector (TAT-BH4). The implication of these results is that ectopically expressed PRF has apoptosis-inducing abilities, and PRF alone is sufficient to induce apoptotic cell death in cells with ectopic expression. Taking this into consideration, our results suggest the possibility of using PRF as a pro-apoptotic gene for tumor therapeutics. [ABSTRACT FROM AUTHOR]

Background: Chemo-resistance to cisplatin-centered cancer therapy is a major obstacle to the effective treatment of human ovarian cancer. Previous reports indicated that arsenic trioxide (ATO) induces cell apoptosis in both drug-sensitive and - resistant ovarian cancer cells. Principal Findings: In this study, we determined the molecular mechanism of ATO-induced apoptosis in ovarian cancer cells. Our data demonstrated that ATO induced cell apoptosis by decreasing levels of phosphorylated AKT (p-AKT) and activating caspase-3 and caspase-9. Importantly, BIM played a critical role in ATO-induced apoptosis. The inhibition of BIM expression prevented AKT dephosphorylation and inhibited caspase-3 activation during cell apoptosis. However, surprisingly, gene silencing of AKT or FOXO3A had little effect on BIM expression and phosphorylation. Moreover, the activation of caspase-3 by ATO treatment improved AKT dephosphorylation, not only by cleaving the regulatory A subunit of protein phosphatase 2A (PP2A), but also by increasing its activation. Furthermore, our data indicated that the c-Jun Nterminal kinases (JNK) pathway is involved in the regulation of BIM expression. Conclusions: We demonstrated the roles of BIM in ATO-induced apoptosis and the molecular mechanisms of BIM expression regulated by ATO during ovarian cancer cell apoptosis. Our findings suggest that BIM plays an important role in regulating p-AKT by activating caspase-3 and that BIM mediates the level of AKT phosphorylation to determine the threshold for overcoming cisplatin resistance in ovarian cancer cells. [ABSTRACT FROM AUTHOR]

Background: ABCC1 and ABCG2 are ubiquitous ATP-binding cassette transmembrane proteins that play an important role in multidrug resistance (MDR). In this study, we evaluated the possible interaction of vandetanib, an orally administered drug inhibiting multiple receptor tyrosine kinases, with ABCC1 and ABCG2 in vitro. Methodology and Principal Findings: MDR cancer cells overexpressing ABCC1 or ABCG2 and their sensitive parental cell lines were used. MTT assay showed that vandetanib had moderate and almost equal-potent anti-proliferative activity in both sensitive parental and MDR cancer cells. Concomitant treatment of MDR cells with vandetanib and specific inhibitors of ABCC1 or ABCG2 did not alter their sensitivity to the former drug. On the other hand, clinically attainable but non-toxic doses of vandetanib were found to significantly enhance the sensitivity of MDR cancer cells to ABCC1 or ABCG2 substrate antitumor drugs. Flow cytometric analysis showed that vandetanib treatment significantly increase the intracellular accumulation of doxorubicin and rhodamine 123, substrates of ABCC1 and ABCG2 respectively, in a dose-dependent manner (P,0.05). However, no significant effect was shown in sensitive parental cell lines. Reverse transcription-PCR and Western blot analysis showed that vandetanib did not change the expression of ABCC1 and ABCG2 at both mRNA and protein levels. Furthermore, total and phosphorylated forms of AKT and ERK1/2 remained unchanged after vandetanib treatment in both sensitive and MDR cancer cells. Conclusions: Vandetanib is unlikely to be a substrate of ABCC1 or ABCG2. It overcomes ABCC1- and ABCG2-mediated drug resistance by inhibiting the transporter activity, independent of the blockade of AKT and ERK1/2 signal transduction pathways. [ABSTRACT FROM AUTHOR]