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Ascites Increases Expression/Function of Multidrug Resistance Proteins in Ovarian Cancer Cells.

Mo L, Pospichalova V, Huang Z, Murphy SK, Payne S, Wang F, Kennedy M, Cianciolo GJ, Bryja V, Pizzo SV, Bachelder RE - PLoS ONE (2015)

Bottom Line: One mechanism behind chemo-resistance involves the upregulation of multidrug resistance (MDR) genes (ABC transporters) that effectively transport (efflux) drugs out of the tumor cells.As a common symptom in stage III/IV ovarian cancer patients, ascites is associated with cancer progression.Functional studies show ascites-driven efflux is suppressible by specific inhibitors of either of two ABC transporters [Multidrug Related Protein (MRP1); Breast Cancer Related Protein (BCRP)].

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Duke University Medical Center, Durham, North Carolina, 27710, United States of America.

ABSTRACT
Chemotherapy resistance is the major reason for the failure of ovarian cancer treatment. One mechanism behind chemo-resistance involves the upregulation of multidrug resistance (MDR) genes (ABC transporters) that effectively transport (efflux) drugs out of the tumor cells. As a common symptom in stage III/IV ovarian cancer patients, ascites is associated with cancer progression. However, whether ascites drives multidrug resistance in ovarian cancer cells awaits elucidation. Here, we demonstrate that when cultured with ascites derived from ovarian cancer-bearing mice, a murine ovarian cancer cell line became less sensitive to paclitaxel, a first line chemotherapeutic agent for ovarian cancer patients. Moreover, incubation of murine ovarian cancer cells in vitro with ascites drives efflux function in these cells. Functional studies show ascites-driven efflux is suppressible by specific inhibitors of either of two ABC transporters [Multidrug Related Protein (MRP1); Breast Cancer Related Protein (BCRP)]. To demonstrate relevance of our findings to ovarian cancer patients, we studied relative efflux in human ovarian cancer cells obtained from either patient ascites or from primary tumor. Immortalized cell lines developed from human ascites show increased susceptibility to efflux inhibitors (MRP1, BCRP) compared to a cell line derived from a primary ovarian cancer, suggesting an association between ascites and efflux function in human ovarian cancer. Efflux in ascites-derived human ovarian cancer cells is associated with increased expression of ABC transporters compared to that in primary tumor-derived human ovarian cancer cells. Collectively, our findings identify a novel activity for ascites in promoting ovarian cancer multidrug resistance.

No MeSH data available.


Related in: MedlinePlus

Ascites derived from ovarian cancer-bearing mice increases chemo-resistance of ID8 cells.Murine ovarian cancer cells (ID8) from three different sources were studied: 1) ID8 cells from normal culture medium (Medium), 2) ID8 cells freshly isolated from ascites fluid in a syngeneic model (In vivo cells), and 3) ID8 cells treated for 7 days in vitro with ascites supernatant (Ascites treated 7 days). Each of these cell populations was exposed to paclitaxel(A) or docetaxel(B) at the indicated concentration for 24 h and [3H]-thymidine incorporation was determined. Three independent experiments were performed and a representative result is shown. Error bar represents SD of quadruplicates in each condition. * indicates p<0.05, *** p<0.001, Two way ANOVA. A. ID8 cells pre-treated with acellular ascites for 7 days (Ascites treated 7 days) or isolated from ascites (in vivo cells) have increased resistance to Paclitaxel compared to ID8 cells from normal medium. B. ID8 cells pre-treated with acellular ascites for 7 days (Ascites treated 7 days) have increased resistance to docetaxel at 2 and 4 nM compared to ID8 cells from normal culture. ID8 cells isolated from ascites (in vivo cells) have increased resistance to docetaxel compared to ID8 cells from normal medium (Medium). C-D. 7-AAD uptake was measured by flow cytometric analysis. The percent 7-AAD positive cells from three independent experiments is shown for paclitaxel (C) and docetaxel (D)-treated cells. Error bars represent SD. * indicates p<0.05, ** p<0.01, *** p<0.001, Two way ANOVA. ID8 cells pre-treated with acellular ascites for 7 days (Ascites treated 7 days) are more resistant to chemotherapy-induced cell death (i.e.,exhibit fewer 7-AAD+ cells) than ID8 cells from normal culture.
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pone.0131579.g001: Ascites derived from ovarian cancer-bearing mice increases chemo-resistance of ID8 cells.Murine ovarian cancer cells (ID8) from three different sources were studied: 1) ID8 cells from normal culture medium (Medium), 2) ID8 cells freshly isolated from ascites fluid in a syngeneic model (In vivo cells), and 3) ID8 cells treated for 7 days in vitro with ascites supernatant (Ascites treated 7 days). Each of these cell populations was exposed to paclitaxel(A) or docetaxel(B) at the indicated concentration for 24 h and [3H]-thymidine incorporation was determined. Three independent experiments were performed and a representative result is shown. Error bar represents SD of quadruplicates in each condition. * indicates p<0.05, *** p<0.001, Two way ANOVA. A. ID8 cells pre-treated with acellular ascites for 7 days (Ascites treated 7 days) or isolated from ascites (in vivo cells) have increased resistance to Paclitaxel compared to ID8 cells from normal medium. B. ID8 cells pre-treated with acellular ascites for 7 days (Ascites treated 7 days) have increased resistance to docetaxel at 2 and 4 nM compared to ID8 cells from normal culture. ID8 cells isolated from ascites (in vivo cells) have increased resistance to docetaxel compared to ID8 cells from normal medium (Medium). C-D. 7-AAD uptake was measured by flow cytometric analysis. The percent 7-AAD positive cells from three independent experiments is shown for paclitaxel (C) and docetaxel (D)-treated cells. Error bars represent SD. * indicates p<0.05, ** p<0.01, *** p<0.001, Two way ANOVA. ID8 cells pre-treated with acellular ascites for 7 days (Ascites treated 7 days) are more resistant to chemotherapy-induced cell death (i.e.,exhibit fewer 7-AAD+ cells) than ID8 cells from normal culture.

Mentions: We tested the effect of ascites on ovarian cancer cell resistance to paclitaxel, a first line chemotherapeutic agent. Comparison was performed between 1) ID8 cells grown in normal culture medium, 2) ID8 cells freshly isolated from ascites fluid in a syngeneic model, and 3) ID8 cells treated for 7 days in vitro with ascites supernatant derived from ID8-bearing mice. Chemotherapy dosing was optimized for a range to achieve 0 to 95% cell growth inhibition, as measured by [3H]-thymidine incorporation. Both the ID8 cells treated for 7 days with ascites supernatant and ID8 cells freshly isolated from ascites fluid were more resistant to paclitaxel than ID8 cells cultured in normal medium, as measured by [3H]-thymidine incorporation assay (Fig 1A). To validate the hypothesis that ascites drives ovarian cancer cell chemo-resistance, we next studied the response of ovarian cancer cells (+/- ascites treatment) to another chemotherapeutic reagent in the taxane family (docetaxel). Notably, ID8 cells treated for 7 days with ascites supernatant, as well as ID8 cells freshly isolated from ascites fluid were significantly more resistant to docetaxel than ID8 cells from normal culture (Fig 1B).


Ascites Increases Expression/Function of Multidrug Resistance Proteins in Ovarian Cancer Cells.

Mo L, Pospichalova V, Huang Z, Murphy SK, Payne S, Wang F, Kennedy M, Cianciolo GJ, Bryja V, Pizzo SV, Bachelder RE - PLoS ONE (2015)

Ascites derived from ovarian cancer-bearing mice increases chemo-resistance of ID8 cells.Murine ovarian cancer cells (ID8) from three different sources were studied: 1) ID8 cells from normal culture medium (Medium), 2) ID8 cells freshly isolated from ascites fluid in a syngeneic model (In vivo cells), and 3) ID8 cells treated for 7 days in vitro with ascites supernatant (Ascites treated 7 days). Each of these cell populations was exposed to paclitaxel(A) or docetaxel(B) at the indicated concentration for 24 h and [3H]-thymidine incorporation was determined. Three independent experiments were performed and a representative result is shown. Error bar represents SD of quadruplicates in each condition. * indicates p<0.05, *** p<0.001, Two way ANOVA. A. ID8 cells pre-treated with acellular ascites for 7 days (Ascites treated 7 days) or isolated from ascites (in vivo cells) have increased resistance to Paclitaxel compared to ID8 cells from normal medium. B. ID8 cells pre-treated with acellular ascites for 7 days (Ascites treated 7 days) have increased resistance to docetaxel at 2 and 4 nM compared to ID8 cells from normal culture. ID8 cells isolated from ascites (in vivo cells) have increased resistance to docetaxel compared to ID8 cells from normal medium (Medium). C-D. 7-AAD uptake was measured by flow cytometric analysis. The percent 7-AAD positive cells from three independent experiments is shown for paclitaxel (C) and docetaxel (D)-treated cells. Error bars represent SD. * indicates p<0.05, ** p<0.01, *** p<0.001, Two way ANOVA. ID8 cells pre-treated with acellular ascites for 7 days (Ascites treated 7 days) are more resistant to chemotherapy-induced cell death (i.e.,exhibit fewer 7-AAD+ cells) than ID8 cells from normal culture.
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pone.0131579.g001: Ascites derived from ovarian cancer-bearing mice increases chemo-resistance of ID8 cells.Murine ovarian cancer cells (ID8) from three different sources were studied: 1) ID8 cells from normal culture medium (Medium), 2) ID8 cells freshly isolated from ascites fluid in a syngeneic model (In vivo cells), and 3) ID8 cells treated for 7 days in vitro with ascites supernatant (Ascites treated 7 days). Each of these cell populations was exposed to paclitaxel(A) or docetaxel(B) at the indicated concentration for 24 h and [3H]-thymidine incorporation was determined. Three independent experiments were performed and a representative result is shown. Error bar represents SD of quadruplicates in each condition. * indicates p<0.05, *** p<0.001, Two way ANOVA. A. ID8 cells pre-treated with acellular ascites for 7 days (Ascites treated 7 days) or isolated from ascites (in vivo cells) have increased resistance to Paclitaxel compared to ID8 cells from normal medium. B. ID8 cells pre-treated with acellular ascites for 7 days (Ascites treated 7 days) have increased resistance to docetaxel at 2 and 4 nM compared to ID8 cells from normal culture. ID8 cells isolated from ascites (in vivo cells) have increased resistance to docetaxel compared to ID8 cells from normal medium (Medium). C-D. 7-AAD uptake was measured by flow cytometric analysis. The percent 7-AAD positive cells from three independent experiments is shown for paclitaxel (C) and docetaxel (D)-treated cells. Error bars represent SD. * indicates p<0.05, ** p<0.01, *** p<0.001, Two way ANOVA. ID8 cells pre-treated with acellular ascites for 7 days (Ascites treated 7 days) are more resistant to chemotherapy-induced cell death (i.e.,exhibit fewer 7-AAD+ cells) than ID8 cells from normal culture.
Mentions: We tested the effect of ascites on ovarian cancer cell resistance to paclitaxel, a first line chemotherapeutic agent. Comparison was performed between 1) ID8 cells grown in normal culture medium, 2) ID8 cells freshly isolated from ascites fluid in a syngeneic model, and 3) ID8 cells treated for 7 days in vitro with ascites supernatant derived from ID8-bearing mice. Chemotherapy dosing was optimized for a range to achieve 0 to 95% cell growth inhibition, as measured by [3H]-thymidine incorporation. Both the ID8 cells treated for 7 days with ascites supernatant and ID8 cells freshly isolated from ascites fluid were more resistant to paclitaxel than ID8 cells cultured in normal medium, as measured by [3H]-thymidine incorporation assay (Fig 1A). To validate the hypothesis that ascites drives ovarian cancer cell chemo-resistance, we next studied the response of ovarian cancer cells (+/- ascites treatment) to another chemotherapeutic reagent in the taxane family (docetaxel). Notably, ID8 cells treated for 7 days with ascites supernatant, as well as ID8 cells freshly isolated from ascites fluid were significantly more resistant to docetaxel than ID8 cells from normal culture (Fig 1B).

Bottom Line: One mechanism behind chemo-resistance involves the upregulation of multidrug resistance (MDR) genes (ABC transporters) that effectively transport (efflux) drugs out of the tumor cells.As a common symptom in stage III/IV ovarian cancer patients, ascites is associated with cancer progression.Functional studies show ascites-driven efflux is suppressible by specific inhibitors of either of two ABC transporters [Multidrug Related Protein (MRP1); Breast Cancer Related Protein (BCRP)].

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Duke University Medical Center, Durham, North Carolina, 27710, United States of America.

ABSTRACT
Chemotherapy resistance is the major reason for the failure of ovarian cancer treatment. One mechanism behind chemo-resistance involves the upregulation of multidrug resistance (MDR) genes (ABC transporters) that effectively transport (efflux) drugs out of the tumor cells. As a common symptom in stage III/IV ovarian cancer patients, ascites is associated with cancer progression. However, whether ascites drives multidrug resistance in ovarian cancer cells awaits elucidation. Here, we demonstrate that when cultured with ascites derived from ovarian cancer-bearing mice, a murine ovarian cancer cell line became less sensitive to paclitaxel, a first line chemotherapeutic agent for ovarian cancer patients. Moreover, incubation of murine ovarian cancer cells in vitro with ascites drives efflux function in these cells. Functional studies show ascites-driven efflux is suppressible by specific inhibitors of either of two ABC transporters [Multidrug Related Protein (MRP1); Breast Cancer Related Protein (BCRP)]. To demonstrate relevance of our findings to ovarian cancer patients, we studied relative efflux in human ovarian cancer cells obtained from either patient ascites or from primary tumor. Immortalized cell lines developed from human ascites show increased susceptibility to efflux inhibitors (MRP1, BCRP) compared to a cell line derived from a primary ovarian cancer, suggesting an association between ascites and efflux function in human ovarian cancer. Efflux in ascites-derived human ovarian cancer cells is associated with increased expression of ABC transporters compared to that in primary tumor-derived human ovarian cancer cells. Collectively, our findings identify a novel activity for ascites in promoting ovarian cancer multidrug resistance.

No MeSH data available.


Related in: MedlinePlus