J Gynecol Oncol.  2013 Jul;24(3):273-279. 10.3802/jgo.2013.24.3.273.

Selective cyclooxygenase inhibitors increase paclitaxel sensitivity in taxane-resistant ovarian cancer by suppressing P-glycoprotein expression

• Lee JP
• Hahn HS
• Hwang SJ
• Choi JY
• Park JS
• Lee IH
• Kim TJ

• KMID: 2288535
• DOI: http://doi.org/10.3802/jgo.2013.24.3.273

Abstract

OBJECTIVE
The purpose of this study was to investigate whether selective cyclooxygenase (COX) inhibitors promote paclitaxel-induced apoptosis in taxane-resistant ovarian cancer cells by suppressing MDR1/P-glycoprotein (P-gp) expression.
METHODS
Taxane-resistant ovarian cancer cells were cultured with paclitaxel alone or combined with a selective COX inhibitors. The expression patterns of MDR1/P-gp and the ability of COX inhibitors to inhibit growth of taxane-resistant ovarian cancer cells were measured. The efficacy of prostaglandin E2 (PGE2) supplementation was measured to evaluate the mechanisms involved in suppressing MDR1 gene expression.
RESULTS
P-gp was upregulated in taxane-resistant ovarian cancer cells compared to paired paclitaxel-sensitive ovarian cancer cells. An 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay showed that selective COX inhibitors significantly enhanced the cytotoxic effects of paclitaxel in taxane-resistant ovarian cancer cells via a prostaglandin-independent mechanism. These increased apoptotic effects were further verified by measuring an increased percentage of cells in sub-G1 stage using flow cytometry. Selective COX inhibitors suppressed MDR1 and P-gp expression. Moreover, combined treatment with paclitaxel and selective COX inhibitors increased poly (ADP-ribose) polymerase (PARP) cleavage in taxane-resistant ovarian cancer cells.
CONCLUSION
Selective COX inhibitors significantly promote paclitaxel-induced cell death in taxane-resistant ovarian cancer cells in a prostaglandin-independent manner. COX inhibitors could be potent therapeutic tools to promote paclitaxel sensitization of taxane-resistant ovarian cancers by suppressing MDR1/P-gp, which is responsible for the efflux of chemotherapeutic agents.

Keyword

Chemosensitizer; Cyclooxygenase inhibitor; Ovarian cancer; Paclitaxel; P-glycoprotein

MeSH Terms

Apoptosis
Cell Death
Cyclooxygenase Inhibitors
Dinoprostone
Flow Cytometry
Ovarian Neoplasms
P-Glycoprotein
Paclitaxel
Prostaglandin-Endoperoxide Synthases
Tetrazolium Salts
Thiazoles
Cyclooxygenase Inhibitors
Dinoprostone
P-Glycoprotein
Paclitaxel
Prostaglandin-Endoperoxide Synthases
Tetrazolium Salts
Thiazoles

Figure

• 

  Fig. 1 P-glycoprotein and cyclooxygenase (COX) expression in ovarian cancer cell lines. HMDR, STR, H, and SK represent HeyA8-MDR, SKOV3ip2-TR, HeyA8 and SKOV3ip1, respectively.

• 

  Fig. 2 Treatment of taxane-resistant ovarian cancer cells with selective cyclooxygenase (COX) inhibitors combined with paclitaxel. Cell viability was determined by using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The viability of the vehicle treated cells was considered as 100%. Statistical significance is indicated by asterisks. *p<0.05. (A) HeyA8-MDR, (B) SKOV3ip2-TR cell line.

• 

  Fig. 3 Selective cyclooxygenase (COX) inhibitors increase apoptosis in taxane-resistant ovarian cancer cells treated with paclitaxel. (A, B) Fluorescence activated cell sorting (FACS) analyses were performed to examine distribution of cells in various cell cycle stages when treated with paclitaxel (100 nM) alone or combined with SC560 (20 µM) or NS398 (100 µM). (C) Increased apoptosis in taxane-resistant ovarian cancer cells treated with paclitaxel combined with a selective COX inhibitor. Taxane-resistant cells were treated with paclitaxel (100 nM) alone or combined with a COX inhibitor (SC560, 20 µM or NS398, 100 µM) and Western blotting for cleaved poly (ADP-ribose) polymerase (PARP) was performed to examine apoptosis.

• 

  Fig. 4 Adding prostaglandin E2 (PGE2) to ovarian cancer cells treated with paclitaxel combined with SC560 or NS398. PGE2 (10 µM) was added to cultures with paclitaxel (100 nM) and SC560 or NS398 for 72 hours. Cell viability was determined using the MTT assay. The viability of the vehicle treated cells was considered as 100%. Statistical significance is indicated by asterisks. *p<0.05. (A) HeyA8-MDR, (B) SKOV3ip2-TR cell line.

• 

  Fig. 5 Reverse transcription polymerase chain reaction (RT-PCR) for MDR1 mRNA and Western blotting for P-glycoprotein (P-gp) protein in HeyA8-MDR and SKOV3ip2-TR cells. HeyA8-MDR and SKOV3ip2-TR cells were treated with paclitaxel, SC560, or NS398, or a combination for 24 hours and cell lysates were harvested for RT-PCR or Western blotting. RT-PCR for 18S rRNA and Western blotting for actin were performed as internal controls. (A) RT-PCR, (B) Western blotting.

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