Mifepristone abrogates repopulation of ovarian cancer cells in between courses of cisplatin treatment

Abstract

Abstract. Repopulation of cancer cells escaping lethal chemotherapy is a critical factor hindering treatment success. One strategy to inhibit tumor cell repopulation is the use of cytostatic compounds between courses of lethal chemotherapy.In this study, we tested the hypothesis that mifepristone (MF), a steroid compound with demonstrated growth inhibition activity in ovarian cancer, should be efficacious in inducing cytostasis and preventing repopulation of ovarian cancer cells if given among rounds of cisplatin (CDDP) treatment. We established an in vitro approach wherein ovarian cancer cells with high (OV2008) or low (SK-OV-3) sensitivity to CDDP were exposed to 3 (OV2008) or 2 (SK-OV-3) rounds of lethal doses of CDDP for 1 h, 12 (OV2008) or 24 (SK-OV-3) days apart. Every 4 or 8 days cell number, cell viability, cell cycle traverse, and colony-forming capacity of viable cells was analyzed. Although CDDP killed the vast majority of cells, there were remnant cells escaping CDDP lethality and repopulating the culture, as evidenced by increased cell number, improved clonogenic capacity of viable cells, and normalization of DNA synthesis. Conversely, when cells were exposed to CDDP for 1 h, and 5, 10 or 20 μM MF was present in the culture medium after CDDP removal, the number, clonogenic capacity, and DNA synthesis ability of the cells were reduced in a dose-dependent manner. The blockage by MF of post-CDDP repopulation was accompanied by a remarkable increase in the percentage of cells expressing the cell death marker cleaved poly(ADP-ribose) polymerase and the mitotic marker phospho-histone H3, suggesting that MF also potentiated CDDP lethality and that the cells likely die due to mitotic failure. In summary, this is the first study reporting that presence of cytostatic concentrations of MF after courses of lethal doses of CDDP prevents repopulation of remnant ovarian cancer cells surviving CDDP treatment.

Antiprogestin mifepristone inhibits the growth of cancer cells of reproductive and non-reproductive origin regardless of progesterone receptor expression

“…Following a period of 24 h allotted for cell adherence, the cells were cultured in the continuous presence of MF or DMSO for 72 h. Triplicate cultures were trypsinized, pelleted by centrifugation at 500 g for 5 min, and resuspended in the appropriate growth medium. An aliquot of each cell suspension was combined with ViaCount reagent (Guava Technologies, Hayward, CA) resulting in a 1:10 (v/v) dilution and then studied using the Guava ViaCount application in the Guava EasyCyte Mini microcapillary cytometer (Guava Technologies) as we previously described [ 15 ]. The data were acquired and analyzed using the CytoSoft 4.1 software (Guava Technologies).…”

“…In the few existing reports that investigated the effects of MF in epithelial ovarian cancer cells, our laboratory [ 13 - 15 ] and others [ 16 , 17 ], demonstrated the efficacy of MF as a growth inhibitory agent. We have shown that MF inhibits the growth of ovarian cancer cells of different genetic backgrounds in a dose-dependent manner in vitro and in vivo [ 13 ].…”

Resistance to cisplatin and paclitaxel does not affect the sensitivity of human ovarian cancer cells to antiprogestin-induced cytotoxicity

“…Due to the low toxicity of these drugs, their potential use as maintenance therapy or antirepopulation therapy following standard of care is anticipated. In this regard we have demonstrated that mifepristone was capable of abrogating the regrowth of cancer cells that escaped CDDP [ 9 ] or the combination CDDP/PTX therapy [ 10 ]. The results presented herein highlight the fact that other antiprogestins in addition to mifepristone can be efficacious against platinum/taxane double-resistant ovarian cancer cells.…”

“…Following the indicated treatments, triplicate cultures were trypsinized, pelleted by centrifugation at 500 g for 5 min, and washed with PBS. The cells were resuspended in ViaCount reagent (Guava Technologies, Hayward, CA) and studied using the Guava ViaCount application in the Guava EasyCyte Mini microcapillary cytometer (Guava Technologies) as we previously reported [ 9 ]. When indicated, the concentration of drugs that caused inhibition of 50% in growth (IC50) were determined using software designed to study drug interaction, which calculates the median effective dose or Dm that is similar to the IC50 (Calcusyn, Biosoft, Cambridge, UK).…”

“…One such possibility for maintenance therapy is the use of antiprogestin compounds that can be chronically given with minimal toxicity [ 7 ]. Our laboratory studied the effect of antiprogestin mifepristone, which successfully blocked the growth of ovarian cancer cells in vitro and in vivo [ 8 ], and prevented the repopulation of ovarian cancer cells that escaped cisplatin (CDDP) [ 9 ] or CDDP-paclitaxel (PTX) [ 10 ] therapies. We have also found that antiprogestins mifepristone, ORG-31710, and ulipristal (CDB-2914), when used at pharmacologic concentrations, cause cytostasis by blocking the activity of cyclin-dependent kinase 2 (Cdk2), thus preventing the cells from moving towards the G1/S transition, and, consequently, synthesizing DNA; moreover, these compounds increased the accumulation of cyclin-dependent kinase inhibitors p21 cip1 and p27 kip1 , as well as promoted their association to Cdk2, leading to its reduced activity [ 8 , 11 ].…”