MSeC is being used as a chemotherapeutic modulator of different anticancer agents in different cancers (27
). The reported increase in the therapeutic index of irinotecan in nude mice against multiple human tumor xenografts by Cao et al. led to the initiation of the concept of testing MSeC in combination with docetaxel against prostate cancer. It has been reported that organic selenium compounds (e.g., MSeC) and inorganic selenium compounds (e.g., sodium selenite) affect cell growth in multiple human cancer cell lines through different pathways (28
). Whereas most of the focus is on inorganic selenium compounds, this is the first report, to our knowledge, to investigate the combination treatment of organic selenium compound (MSeC) with docetaxel against prostate cancer. The goal of this study is to evaluate MSeC as a modulator of docetaxel efficacy and to understand the mechanism of the synergy between MSeC and docetaxel against prostate cancer. Understanding the mechanism of action of this combination regimen is important and could translate into improved clinical outcomes, thereby improving quality of life of men with prostate cancer.
In this study, we showed that pretreatment with MSeC is essential for the in vitro enhancement of docetaxel antitumor activity. At equitoxic concentration, pretreatment with MSeC decreased the concurrent combination dose that achieved 50% inhibition of cell growth (IC50 of concurrent combination) by 80%. In addition, at equimolar concentration, pretreatment with MSeC increased the inhibitory effect of the concurrent combination treatment by 30% (). Thus, MSeC enhancement of docetaxel antitumor activity is dependent on MSeC schedule.
This docetaxel enhanced inhibition of cell growth by the sequential combination treatment (MSeC/docetaxel) observed at the level of the population (sulforhodamine B assays) was determined to be due to an increase in apoptotic cell death and not due to cytostatic or growth-slowing effects. Trypan blue assay and time-lapse video showed that the vast majority of cells were trypan blue positive and were subject to an apoptotic type of death (shrinking, displaying total cessation of all membrane movement, and eventually breaking into smaller parts and forming apoptotic bodies; Figs. and ).
The enhanced docetaxel antitumor activity was further investigated to determine whether the combination effect is additive or synergistic. The result showed that the enhanced combination effect is synergistic when docetaxel and MSeC are given at different percentages of their individual concentrations that achieve IC90 or IC50 effects (). Specifically, administration of 20% of MSeC alone concentration that achieved 90% inhibition of cell growth (20% of IC90) and 15% of docetaxel alone concentration that achieved 90% inhibition of cell growth (15% of IC90) resulted in 90% combination inhibition of cell growth (IC90 of the combination; ). In other words, 90% inhibition of cell growth with the combination treatment required 0.2 of MSeC and 0.15 of docetaxel of their individual IC90 doses. If the effect of the combination treatment of MSeC and docetaxel was additive, only 35% inhibition of cell growth would have been achieved. On the other hand, when docetaxel and MSeC were administered at different percentages of their individual IC10 concentrations, the combination effect was additive ().
Several reports documented the finding that MSeC induced apoptosis through different mechanisms in various cancer lines including leukemia, ovarian, and mammary cells (17
). In this study, to investigate the effect of the combination treatment (MSeC/docetaxel) on apoptosis and determine the mechanism of action behind the synergy, caspase-3 activity assay and detection of apoptotic cell death were done. MSeC/docetaxel treatment significantly increased the caspase-3 activity and apoptotic cell death when compared with each drug alone and untreated control (). The induced apoptotic cell death by MSeC/docetaxel treatment was significantly reduced when cells were pretreated with a pan-caspase inhibitor (). Conversion of synergy between MSeC and docetaxel to antagonism after pretreatment with a pancaspase inhibitor further confirms that caspase activation plays a critical role in the observed synergy (). In other words, blocking caspase-dependent apoptosis eliminates the synergy between MSeC and docetaxel.
The role of survivin in control and drug resistance in various in vitro
and in vivo
models, including prostate cancer, was documented by many reports. In summary, the higher the survivin level, the less apoptotic response achieved after drug treatment (35
). Furthermore, the relationship between survivin and the response to docetaxel was also documented in several published reports; briefly, the higher the survivin level, the more resistance to docetaxel treatment (38
). In this study, survivin level was down-regulated by MSeC/docetaxel treatment when MSeC was given at IC25
and docetaxel was given at IC90
, which corresponded with the maximum synergy. However, MSeC/docetaxel treatment with MSeC at IC25
and docetaxel at IC10
, which corresponded with additive effect, did not change the survivin level (). These results associate survivin expression level after MSeC/docetaxel treatment with the observed MSeC/docetaxel synergy. The lower the survivin level in C2G cells after the MSeC/docetaxel treatment, the more synergy is observed.
In summary, pretreatment with MSeC was essential to markedly sensitize cells to docetaxel. The synergy between MSeC and docetaxel in C2G prostate cancer cells is associated with increased level of caspase-dependent apoptosis and decreased level of survivin. Future studies will determine whether the mechanism of action of MSeC/docetaxel treatment down-regulating survivin is transcriptional or translational.