Clear cell carcinoma (CCC) of the ovary is known to show reduced sensitivity to platinum-based frontline chemotherapy and is associated with a worse prognosis than serous adenocarcinoma.1–4
The lack of an effective chemotherapy for recurrent CCC after frontline treatment is an important problem in the clinical management of CCC. To improve the prognosis of patients with CCC, the development of novel treatment strategies for both first-line treatment and the salvage treatment of recurrent disease is needed.
Trabectedin, which was formerly known as ecteinascidin-743 (ET-743), is an antineoplastic agent that was originally derived from the Caribbean marine tunicate Ecteinascidia turbinata. It covalently binds to the minor groove of DNA, bending it toward the major groove and disrupting transcription, leading to G2-M cell cycle arrest and ultimately apoptosis.5 Although the exact mechanism of trabectedin’s cytotoxicity is unclear, it is assumed to be based on the inhibition of transcription-dependent nucleotide excision repair via the trapping of the proteins responsible for nucleotide excision repair, which causes the affected cells to undergo apoptosis.6 In a preclinical study, trabectedin displayed significant in vitro and in vivo antitumor activity against a range of solid tumors, including soft tissue sarcoma (STS); ovarian, breast, prostate, and renal cancer; melanoma; and non–small cell lung cancer.7–10 On the basis of the promising results obtained in preclinical and clinical studies,9,11,12 trabectedin has been approved in Europe for the treatment of STS after anthracycline and ifosfamide failure as well as for treating patients who cannot receive these agents.13
In ovarian cancer, trabectedin has been studied in several phase 1 and 2 clinical trials and exhibited a favorable toxicity profile and promising activity in recurrent ovarian cancer patients.14,15 After the encouraging results of these studies, a phase 3 trial investigating the activity of trabectedin plus pegylated liposomal doxorubicin (PLD) versus PLD alone in patients with recurrent ovarian cancer has recently been conducted. As this study demonstrated that trabectedin-based combination chemotherapy conferred a survival benefit, trabectedin has become a focus of attention for researchers investigating the treatment of epithelial ovarian cancer.16,17 Currently, a phase 3 study is being conducted to compare the activity of trabectedin plus PLD versus carboplatin plus PLD in patients with recurrent ovarian cancer (the INOVATYON study). However, because of the small number of CCC patients included in the abovementioned studies,14–17 the clinical activity of trabectedin against CCC remains unclear.
We have recently reported the results of a preclinical study that showed that trabectedin is more effective against CCC than the currently available anticancer agents including cisplatin, paclitaxel, and SN-38 (an active metabolite of irinotecan) and proposed that trabectedin is the most effective single agent for treating CCC.18 However, no previous preclinical studies (including ours) have investigated the optimal cytotoxic agents for combining with trabectedin during the treatment of ovarian cancer. In the current study, to develop effective combination chemotherapies for CCC, we investigated the best anticancer agents for combining with trabectedin using chemonaive and chemorefractory CCC cells. Moreover, as the inhibition of mammalian target of rapamycin complex 1 (mTORC1) significantly enhanced the therapeutic efficacy of trabectedin and prevented CCC cells from acquiring resistance to trabectedin in a previous study,18 we also investigated the benefit of adding the mTORC1 inhibitor everolimus to trabectedin-based combination chemotherapy against CCC.
MATERIALS AND METHODS
Reagents and Antibiotics
Trabectedin was obtained from PharmaMar (Madrid, Spain). Everolimus was obtained from Novartis Pharma AG (Basel, Switzerland). Cisplatin, paclitaxel, 7-ethyl-10-hydroxycamptothecin (SN-38), and doxorubicin were purchased from Sigma (St Louis, MO). Topotecan was supplied by Nippon Kayaku (Tokyo, Japan). Enhanced chemiluminescence Western blotting detection reagents were purchased from Perkin Elmer (Boston, MA). Antibodies against poly(ADP-ribose) polymerase (PARP) and β-actin were obtained from Cell Signaling Technology (Beverly, MA). Antibodies against P-glycoprotein (P-gp) were purchased from Thermo Scientific (Waltham, MA). The CellTiter 96-well proliferation assay kit was obtained from Promega (Madison, WI).
Trabectedin was prepared as a 10-µmol/L stock solution in ethanol. Cisplatin and doxorubicin were dissolved in sterilized double-distilled water to a final concentration of 10 mmol/L. Paclitaxel, SN-38, and topotecan were dissolved in dimethyl sulfoxide to final concentrations of 10 mmol/L.
The human ovarian CCC cell lines RMG1, RMG2, KOC7C, and HAC2 were kindly provided by Dr Itamochi (Tottori University, Tottori, Japan). These cells were maintained as monolayer cultures in Dulbecco’s modified Eagle’s medium (DMEM Ham’s F-12; Gibco Ltd) containing 10% fetal bovine serum.
Cell Proliferation Assay
The MTS assay was used to analyze the effects of each chemical. Cells were plated in 96-well plates and exposed to the drugs at different concentrations. After 48 hours of incubation, the number of surviving cells was assessed by determining the A490nm of the dissolved formazan product after the addition of MTS for 1 hour, as described by the manufacturer (Promega). Cell viability is expressed as follows: Aexp group/Acontrol × 100.
The Isobologram Method and the Combination Index
The isobologram method relies on the measurement of the combined concentrations of D1 (trabectedin) and D2 (another drug) that cause a given effect (in our study, a fractional kill value of 50% or 70%). For each experimental concentration of trabectedin, the concentration of D2 that caused the desired effect when used in combination with D1 was found by nonlinear fitting of the concentration-effect relationship of D2 to the given trabectedin concentration. Conversely, for each experimental concentration of D2, the trabectedin concentration that caused the desired effect when used in combination with D2 was found by nonlinear fitting of the concentration-effect relationship of trabectedin to the particular D2 concentration. In this manner, multiple pairs of drug concentrations that achieved the isoeffect were found. For each pair of drug concentrations (Dtrabectedin, DD2) that produced effect X when used in combination, the combination index (CI) was calculated as follows: CI = Dtrabectedin/ICx, trabectedin + DD2/ICx, D2, where ICx, trabectedin and ICx, D2 are the concentrations of each drug that would produce effect X when administered alone. CI less than 1, CI equal to 1, and CI greater than 1 indicate synergism, additive effect, and antagonism, respectively. The significance of the differences in the mean values from CI equal to 1 was evaluated using a 2-tailed t test.
Western Blot Analysis
The cells were treated with 1 of the drug combinations for 24 hours and then lysed in lysis buffer. The protein concentrations of the cell lysates were determined using the Bio-Rad protein assay reagent. Equal amounts of protein were applied to 10% polyacrylamide gels, and then the electrophoresed proteins were transblotted onto nitrocellulose membranes. After the membranes had been blocked, they were incubated with anti-PARP, anti–β-actin antibody, or anti–P-gp antibodies. The immunoblots were visualized with appropriate horseradish peroxidase-coupled immunoglobulins using the enhanced chemiluminescence Western blotting system.
Establishment of Chemoresistant Cell Lines
Cisplatin-resistant and trabectedin-resistant sublines derived from RMG1 and RMG2 were developed in our laboratory by continuously exposing the cells to cisplatin or trabectedin, as described previously.18,19 Briefly, cells of both lines were exposed to stepwise increases in the concentration of cisplatin or trabectedin. The initial exposures involved concentrations of 10 nmol/L and 0.1 nmol/L for cisplatin and trabectedin, respectively. After the cells had regained their exponential growth rate, the cisplatin or trabectedin concentration was doubled, and the procedure was repeated until cisplatin and trabectedin doses of 80 µmol/L and 10 nmol/L, respectively, had been reached. The resulting cisplatin-resistant (RMG1-CR and RMG2-CR) and trabectedin-resistant sublines (RMG1-YR and RMG2-YR) were subcultured weekly and treated monthly with 80 µmol/L cisplatin or 10 nmol/L trabectedin to maintain a high level of chemoresistance.
Paclitaxel-resistant sublines (RMG1-PR and RMG2-PR) were also established from RMG1 and RMG2 cells by increasing the paclitaxel concentration from 0.1 nmol/L to 120 nmol/L in a stepwise manner.
Cell proliferation was analyzed using the Student t test. P values of less than 0.05 were considered significant.
Determination of IC50 Values
The in vitro activities of cisplatin, SN-38, topotecan, doxorubicin, paclitaxel, and trabectedin were evaluated in 4 human CCC cell lines after 48 hours of treatment. The IC50 values obtained in each experiment are summarized in Table 1. As shown, trabectedin demonstrated significantly greater antitumor activity against the CCC cell lines than the other anticancer agents.
Effects of Combination Treatment With Trabectedin and Other Antineoplastic Agents
Figure 1 shows the CI values for each combination treatment (trabectedin plus cisplatin, SN-38, topotecan, or doxorubicin) obtained in the 4 CCC cell lines. Representative isobolograms are also shown in Figure 1, Supplemental Digital Content, available at http://links.lww.com/IGC/A218. Although we tried to examine the effects of combination treatment with paclitaxel and trabectedin, CI values could not be obtained for this combination using the isobologram method because of the low sensitivity of some CCC cell lines to paclitaxel, even at high concentrations. As shown in Figure 1, among the 4 combination treatments tested, SN-38 combined with trabectedin demonstrated the lowest CI value, which indicated that it displayed the strongest synergism, in each cell line. Combination treatment with topotecan and trabectedin displayed the second lowest CI value in each cell line. Combination treatment with trabectedin and cisplatin or doxorubicin demonstrated CI values of roughly 0.9 to 1.0, indicating mild synergy or an additive effect.
The Mechanisms Responsible for the Synergism Between Trabectedin and Irinotecan or Topotecan
To investigate the mechanisms underlying the synergism between trabectedin and irinotecan or topotecan, we first examined the effects of SN-38 and topotecan on trabectedin-induced apoptosis in CCC cells. As shown in Figure 2A, the addition of SN-38 or topotecan to trabectedin enhanced the trabectedin-induced cleavage of PARP in RMG1 cells. A similar result was obtained in the RMG2 cells (data not shown). These results indicate that both SN-38 and topotecan enhanced the antitumor activity of trabectedin by enhancing apoptosis.
We next investigated whether P-gp, a member of the ATP-binding cassette transporter superfamily, was involved in observed synergistic effects. For this purpose, we established trabectedin-resistant sublines (RMG1-YR) derived from RMG1 cells. As shown in Figure 2B, significantly greater P-gp expression was observed in trabectedin-resistant cell line than in their representative parental cell line, indicating the involvement of P-gp in trabectedin resistance. Treatment with SN-38 or topotecan as single agents significantly reduced the expression of P-gp in a dose-dependent manner (Fig. 2C); however, such effects were not observed in cells treated with cisplatin or doxorubicin (data not shown). These results suggest that the reduced P-gp expression induced by SN-38 or topotecan is, at least in part, involved in the synergistic effects of trabectedin plus irinotecan or topotecan.
Effects of Combination Treatment With Trabectedin and SN-38 or Topotecan on Chemoresistant CCC
To examine the effectiveness of combination treatment with trabectedin and SN-38 or topotecan against cisplatin-resistant or paclitaxel-resistant CCC, we established cisplatin-resistant or paclitaxel-resistant sublines (RMG1-CR, RMG2-CR, RMG1-PR, and RMG2-PR) from RMG1 and RMG2 cells, as described in Materials and Methods. The acquired resistance of these cell lines to cisplatin or paclitaxel was confirmed by MTS assays. As shown, the chemoresistant sublines displayed markedly reduced paclitaxel or cisplatin sensitivity compared with their respective parental cell lines (Fig. 2A–D, Supplemental Digital Content, http://links.lww.com/IGC/A218). Figures 3A and B show the CI values of each combination treatment at the IC50 levels observed in these resistant cell lines. Representative isobolograms are also shown in Figure 3, Supplemental Digital Content, available at http://links.lww.com/IGC/A218. As shown in Table 2, although all 4 chemoresistant cell lines displayed relatively higher IC50 values for trabectedin, SN-38, and topotecan than their respective parental cell lines, the combination treatment with trabectedin and SN-38 or topotecan exhibited synergistic effects in these resistant cell lines.
According to previous studies, the peak plasma concentrations of SN-38, topotecan, and trabectedin are 166.0 nmol/L, 385.0 nmol/L, and 10.3 nmol/L, respectively.20–22 As shown in Figures 3C and D, combination treatment with trabectedin and SN-38 or topotecan at clinically achievable concentrations (below the peak plasma concentrations) was highly effective against both the parental and chemoresistant CCC cell lines, indicating that the trabectedin-containing chemotherapies proposed in the current study are reasonable and clinically achievable.
mTOR Inhibition by Everolimus Enhances the Antitumor Activity of Trabectedin Plus SN-38 or Topotecan
We have previously reported that mTOR inhibition by everolimus significantly enhanced the efficacy of trabectedin and prevented CCC cells from acquiring resistance to trabectedin.18 To examine whether the addition of everolimus would further enhance the antitumor activity of trabectedin plus SN-38 or topotecan, we conducted MTS assays (Fig. 4). As shown, treatment with everolimus significantly enhanced the antitumor effect of trabectedin plus SN-38 or topotecan in both the parental and chemoresistant CCC cell lines.
Several preclinical investigations have suggested that trabectedin acts synergistically in combination with other cytotoxic agents. For example, trabectedin has displayed synergistic effects against STS when combined with cisplatin,23 doxorubicin,24,25 paclitaxel,25 or irinotecan.26 However, only limited information is available about the efficacy of such combinations against ovarian cancer.23 In the current study, we attempted to find agents that produce strong synergistic effects against ovarian CCC, when combined with trabectedin. As a result, we found that combination treatment with trabectedin and SN-38 produced the strongest synergistic effect against CCC in vitro. Combination treatment with trabectedin and topotecan also resulted in mild synergistic effects against CCC. According to previous studies, the peak plasma concentrations of SN-38, topotecan, and trabectedin are 166.0 nmol/L, 385.0 nmol/L, and 10.3 nmol/L, respectively,20–22 indicating that the trabectedin-containing chemotherapies proposed in the current study are reasonable and clinically achievable.
The mechanisms underlying the synergistic effects of combination treatment with trabectedin and SN-38 or topotecan remain unknown. However, molecular pharmacology studies have indicated that irinotecan and topotecan interact with DNA via different mechanisms than trabectedin. Both irinotecan and topotecan are inhibitors of the type I topoisomerase enzyme, which binds to topoisomerase I DNA complex, resulting in the formation of irreversible double-strand breaks and, hence, cell death.27 In contrast, trabectedin binds to the N2 position of guanine in the minor groove of DNA, forms intrastrand and interstrand cross-links, and disrupts transcription, resulting in cell death.5 Thus, in the current study, combining trabectedin with SN-38 or topotecan might have caused increased genotoxicity in CCC cells through the induction of multiple DNA damage mechanisms.
To investigate the mechanisms responsible for the synergistic interactions observed in the current study, we examined the effects of trabectedin, irinotecan, and topotecan, both as single and combined agents, on the major survival signals that are reported to be involved in the mechanisms of chemoresistance, including the AKT and MAPK pathways.28 However, we did not detect any changes in the levels of these signals after treatment with trabectedin, irinotecan, and/or topotecan (data not shown). Thus, we next investigated whether P-gp is involved in the observed synergistic effects. P-glycoprotein is encoded by the multidrug resistance 1 gene and is known to be involved in multidrug resistance, including resistance to irinotecan29 and topotecan.30 As shown in Figure 2B, RMG1-YR, trabectedin-resistant cells, displayed significantly greater P-gp expression than their representative parental cell lines, which strongly indicated that P-gp was involved in their trabectedin resistance. Importantly, when these cell lines were treated with SN-38 or topotecan, their P-gp expression was attenuated in a dose-dependent manner. These results suggest that the reductions in P-gp expression induced by SN-38 or topotecan are, at least in part, involved in the observed synergistic effects of trabectedin and SN-38 or topotecan on CCC. Further investigations are needed to obtain a deeper understanding of the mechanisms underlying the synergistic effects of trabectedin-based combination chemotherapy.
On the basis of previous preclinical and clinical studies, irinotecan is considered to be an effective clinical treatment of CCC of the ovary.31–36 Currently, the Japanese Gynecologic Oncology Group (JGOG) is conducting a randomized phase 3 trial comparing paclitaxel plus carboplatin (TC) with irinotecan plus cisplatin as first-line chemotherapies.37 Thus, the clinical activity of combination treatment with trabectedin plus irinotecan or topotecan against CCC should be evaluated in the front-line therapy setting.
Another important finding of our study was the significant antitumor activity of trabectedin plus irinotecan against cisplatin-resistant and paclitaxel-resistant CCC because the lack of an effective chemotherapy for recurrent CCC after frontline platinum-based combination chemotherapy is a major clinical problem in the management of CCC. In the current study, although the antitumor effects of trabectedin and irinotecan as single agents against the cisplatin-resistant and paclitaxel-resistant CCC cells were slightly milder than their effects against the parental cells, concurrent treatment with trabectedin and SN-38 resulted in synergistic interactions in these cells. As the RMG1-CR, RMG1-PR, RMG2-CR, and RMG2-PR cells used in this study mimic the clinical resistance seen in cisplatin-treated or paclitaxel-treated patients, our results suggest that cisplatin and paclitaxel refractory CCC are also candidates for combination treatment involving trabectedin plus irinotecan or topotecan.
In this study, we also investigated the role of the mTORC1 in the sensitivity of CCC to trabectedin plus irinotecan or topotecan. Importantly, as shown in Figure 4, treatment with everolimus, an mTORC1 inhibitor, enhanced the antitumor efficacy of combination treatment with trabectedin plus irinotecan or topotecan in our experimental model. This finding is consistent with our previous report showing that AKT-mTORC1 signaling is involved in trabectedin resistance.18 Moreover, this finding is also consistent with a recent case report showing the potential clinical activity of trabectedin in combination with mTORC1 inhibitor in patients with recurrent CCC.38 Previous studies have indicated that mTORC1 is frequently activated in CCC of the ovary and can be targeted therapeutically.19 ,39 On the basis of promising results from preclinical investigations,19 ,39 the Gynecologic Oncology Group is currently conducting a phase 2 trial (protocol GOG0268) evaluating the efficacy of combination treatment with temsirolimus and carboplatin plus paclitaxel as a first-line chemotherapy for patients with stage III to IV CCC of the ovary. In addition, the Japanese Gynecologic Oncology Group will soon initiate a phase 2 trial of everolimus monotherapy for patients with recurrent CCC of the ovary (protocol JGOG3012). Considering the results of our preclinical investigations, the clinical activity of trabectedin plus irinotecan or topotecan, in combination with an mTORC1 inhibitor, should be investigated in future trials in patients with CCC of the ovary.
We have to recognize the potential weakness of our study, that is, its in vitro experimental design. To evaluate the antitumor activity and adverse effects of combination treatment with trabectedin and irinotecan or topotecan, further studies using animal models and clinical trials should be performed in the future.
In conclusion, our findings indicate that the combination treatment with trabectedin plus irinotecan is a promising regimen for treating CCC of the ovary, both as a frontline treatment and as a salvage treatment of recurrence after platinum-based or paclitaxel-based chemotherapy. We consider that our preclinical data provide significant rationale for future clinical trials of trabectedin plus irinotecan or topotecan in combination with an mTORC1 inhibitor in this patient population.
The authors thank Yuko Morishita for her secretarial support.
1. Winter WE, Maxwell GL, Tian C, et al. Prognostic factors for stage III epithelial ovarian cancer: a Gynecologic Oncology Group Study. J Clin Oncol. 2007; 25: 3621–3627.
2. Sugiyama T, Kamura T, Kigawa J, et al. Clinical characteristics of clear cell carcinoma of the ovary: a distinct histologic type with poor prognosis and resistance to platinum-based chemotherapy. Cancer. 2000; 88: 2584–2589.
3. Bamias A, Psaltopoulou T, Sotiropoulou M, et al. Mucinous but not clear cell histology is associated with inferior survival in patients with advanced stage ovarian carcinoma treated with platinum-paclitaxel chemotherapy. Cancer. 2010; 116: 1462–1468.
4. Crotzer DR, Sun CC, Coleman RL, et al. Lack of effective systemic therapy for recurrent clear cell carcinoma of the ovary. Gynecol Oncol. 2007; 105: 404–408.
5. Carter NJ, Keam SJ. Trabectedin
: a review of its use in soft tissue sarcoma and ovarian cancer. Drugs. 2010; 70: 355–376.
6. Herrero AB, Martín-Castellanos C, Marco E, et al. Cross-talk between nucleotide excision and homologous recombination DNA repair pathways in the mechanism of action of antitumor trabectedin
. Cancer Res. 2006; 66: 8155–8162.
7. Valoti G, Nicoletti MI, Pellegrino A, et al. Ecteinascidin-743, a new marine natural product with potent antitumor activity on human ovarian carcinoma xenografts. Clin Cancer Res. 1998; 4: 1977–1983.
8. Hendriks HR, Fiebig HH, Giavazzi R, et al. High antitumour activity of ET743 against human tumour xenografts from melanoma, non–small-cell lung and ovarian cancer. Ann Oncol. 1999; 10: 1233–1240.
9. Morioka H, Weissbach L, Vogel T, et al. Antiangiogenesis treatment combined with chemotherapy produces chondrosarcoma necrosis. Clin Cancer Res. 2003; 9: 1211–1217.
10. Donald S, Verschoyle RD, Greaves P, et al. Complete protection by high-dose dexamethasone against the hepatotoxicity of the novel antitumor drug yondelis (ET-743) in the rat. Cancer Res. 2003; 63: 5902–5908.
11. Delaloge S, Yovine A, Taamma A, et al. Ecteinascidin-743: a marine-derived compound in advanced, pretreated sarcoma patients—preliminary evidence of activity. J Clin Oncol. 2001; 19: 1248–1255.
12. Yovine A, Riofrio M, Blay JY, et al. Phase II study of ecteinascidin-743 in advanced pretreated soft tissue sarcoma patients. J Clin Oncol. 2004; 22: 890–899.
13. Ganjoo KN, Patel SR. Trabectedin
: an anticancer drug from the sea. Expert Opin Pharmacother. 2009; 10: 2735–2743.
14. Krasner CN, McMeekin DS, Chan S, et al. A phase II study of trabectedin
single agent in patients with recurrent ovarian cancer previously treated with platinum-based regimens. Br J Cancer. 2007; 97: 1618–1624.
15. Sessa C, De Braud F, Perotti A, et al. Trabectedin
for women with ovarian carcinoma after treatment with platinum and taxanes fails. J Clin Oncol. 2005; 23: 1867–1874.
16. Monk BJ, Herzog TJ, Kaye SB, et al. Trabectedin
plus pegylated liposomal doxorubicin in recurrent ovarian cancer. J Clin Oncol. 2010; 28: 3107–3114.
17. Monk BJ, Herzog TJ, Kaye SB, et al. Trabectedin
plus pegylated liposomal doxorubicin (PLD) versus PLD in recurrent ovarian cancer: overall survival analysis. Eur J Cancer. 2012; 48: 2361–2368.
18. Mabuchi S, Hisamatsu T, Kawase C, et al. The activity of trabectedin
as a single agent or in combination with everolimus for clear cell carcinoma of the ovary. Clin Cancer Res. 2011; 17: 4462–4473.
19. Mabuchi S, Kawase C, Altomare DA, et al. mTOR is a promising therapeutic target both in cisplatin-sensitive and cisplatin-resistant clear cell carcinoma of the ovary. Clin Cancer Res. 2009; 15: 5404–5413.
20. Pitot HC, Goldberg RM, Reid JM, et al. Phase I dose-finding and pharmacokinetic trial of irinotecan
hydrochloride (CPT-11) using a once-every-three-week dosing schedule for patients with advanced solid tumor malignancy. Clin Cancer Res. 2000; 6: 2236–2244.
21. Molina JR, Kaufmann SH, Reid JM, et al. Evaluation of lapatinib and topotecan
combination therapy: tissue culture, murine xenograft, and phase I clinical trial data. Clin Cancer Res. 2008; 14: 7900–7908.
22. von Mehren M, Schilder RJ, Cheng JD, et al. A phase I study of the safety and pharmacokinetics of trabectedin
in combination with pegylated liposomal doxorubicin in patients with advanced malignancies. Ann Oncol. 2008; 19: 1802–1809.
23. D’Incalci M, Colombo T, Ubezio P, et al. The combination of yondelis and cisplatin is synergistic against human tumor xenografts. Eur J Cancer. 2003; 39: 1920–1926.
24. Meco D, Colombo T, Ubezio P, et al. Effective combination of ET-743 and doxorubicin in sarcoma: preclinical studies. Cancer Chemother Pharmacol. 2003; 52: 131–138.
25. Takahashi N, Li WW, Banerjee D, et al. Sequence-dependent enhancement of cytotoxicity produced by ecteinascidin 743 (ET-743) with doxorubicin or paclitaxel in soft tissue sarcoma cells. Clin Cancer Res. 2001; 7: 3251–3257.
26. Riccardi A, Meco D, Ubezio P, et al. Combination of trabectedin
is highly effective in a human rhabdomyosarcoma xenograft. Anticancer Drugs. 2005; 16: 811–815.
27. Mathijssen RH, Loos WJ, Verweij J, et al. Pharmacology of topoisomerase I inhibitors irinotecan
(CPT-11) and topotecan
. Curr Cancer Drug Targets. 2002; 2: 103–123.
28. Mabuchi S, Ohmichi M, Kimura A, et al. Inhibition of phosphorylation of BAD and Raf-1 by Akt sensitizes human ovarian cancer cells to paclitaxel. J Biol Chem. 2002; 277: 33490–33500.
29. Kroetz DL. Role for drug transporters beyond tumor resistance: hepatic functional imaging and genotyping of multidrug resistance transporters for the prediction of irinotecan
toxicity. J Clin Oncol. 2006; 24: 4225–4227.
30. Vanhoefer U, Müller MR, Hilger RA, et al. Reversal of MDR1-associated resistance to topotecan
by PAK-200S, a new dihydropyridine analogue, in human cancer cell lines. Br J Cancer. 1999; 81: 1304–1310.
Ovarian clear cell carcinoma; Trabectedin; Irinotecan; Topotecan; Isobologram
Supplemental Digital Content
© 2014 by the International Gynecologic Cancer Society and the European Society of Gynaecological Oncology.