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Prostasin may contribute to chemoresistance, repress cancer cells in ovarian cancer, and is involved in the signaling pathways of CASP/PAK2-p34/actin.

Yan BX, Ma JX, Zhang J, Guo Y, Mueller MD, Remick SC, Yu JJ - Cell Death Dis (2014)

Bottom Line: In this study, we show that prostasin is decreased in an ovarian cancer drug-resistant cell line and in ovarian cancer patients with high levels of excision repair cross-complementing 1, a marker for chemoresistance.In addition, the surviving cells grow at a much lower rate compared with non-overexpressed cells.Our investigation of the molecular mechanisms suggests that prostasin may repress cancer cells and/or contribute to chemoresistance by modulating the CASP/P21-activated protein kinase (PAK2)-p34 pathway, and thereafter PAK2-p34/JNK/c-jun and PAK2-p34/mlck/actin signaling pathways.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Biochemistry, School of Medicine, Department of Basic Pharmaceutical Sciences, School of Pharmacy, and Mary Babb Randolph Cancer Center, Robert C. Byrd Health Sciences Center, West Virginia University, Morgantown, WV 26506, USA [2] IcesnowYanyan Bioscience Association, Beijing 00094, China.

ABSTRACT
Ovarian cancer is the deadliest of gynecologic cancers, largely due to the development of drug resistance in chemotherapy. Prostasin may have an essential role in the oncogenesis. In this study, we show that prostasin is decreased in an ovarian cancer drug-resistant cell line and in ovarian cancer patients with high levels of excision repair cross-complementing 1, a marker for chemoresistance. Our cell cultural model investigation demonstrates prostasin has important roles in the development of drug resistance and cancer cell survival. Forced overexpression of prostasin in ovarian cancer cells greatly induces cell death (resulting in 99% cell death in a drug-resistant cell line and 100% cell death in other tested cell lines). In addition, the surviving cells grow at a much lower rate compared with non-overexpressed cells. In vivo studies indicate that forced overexpression of prostasin in drug-resistant cells greatly inhibits the growth of tumors and may partially reverse drug resistance. Our investigation of the molecular mechanisms suggests that prostasin may repress cancer cells and/or contribute to chemoresistance by modulating the CASP/P21-activated protein kinase (PAK2)-p34 pathway, and thereafter PAK2-p34/JNK/c-jun and PAK2-p34/mlck/actin signaling pathways. Thus, we introduce prostain as a potential target for treating/repressing some ovarian tumors and have begun to identify their relevant molecular targets in specific signaling pathways.

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Related in: MedlinePlus

Prostasin has important roles in chemoresistance and cell death in cell culture model. (a) Prostasin decreased in paclitaxel-resistance cancer cell line. O432: ovarian cancer cell line Ovca432 (sensitive to paclitaxel); O432-RP: paclitaxel resistance cell line generated from Ovca432. The prostasin protein levels in O432 and O432-RP cells are shown in immunoblots with specific antibodies. (b) Prostasin siRNAs transfection reduced prostasin. O432-pro-D cells (transfected with prostasin siRNA) express lower prostasin, compared with control cells of O432 (transfected with reagent only) and O432-Cs (transfected with no-targeting siRNA). The prostasin protein levels are shown in immunoblots with specific antibodies. (c) Downregulation of prostasin in O432 cells resulted in increase of chemoresistant activity. Cells were treated with paclitaxel at different concentrations for 24 h (starting 48 h after siRNAs transfection) and cultured with normal medium for an additional 7 to 10 days before cell survival was assayed. Relative cell survival rates of each cell line are shown. (d) Overexpression of prostasin greatly induces cell death in ovarian cancer cells. The cell survival rates are shown after forced overexpression of prostasin in several cell lines from day-0 to day-8, respectively. (e) Prostasin cDNA transfection resulted in overexpression of prostasin in chemoresistant O432-RP cells. O432-RP-pro-O cells (transfected with prostasin cDNA) express higher prostasin compared with control cells O432-RP and O432-RP-C (transfected with control vector). The prostasin protein levels are shown in immunoblots with specific antibodies. (f) Forced overexpression of prostasin represses growth of chemoresistant cells. Relative cell growth rates are shown for O432-RP-pro-O and control cells O432-RP and O432-RP-C. (g) Forced overexpression of prostasin in O432-RP cells re-sensitizes chemoresistant cells. Cells were plated at about 10–20% confluence and treated with paclitaxel at different concentrations for 24 h, cultured with normal medium for additional 7 to 10 days, then assayed for cell survival. Relative survival rates of cell lines are shown
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fig2: Prostasin has important roles in chemoresistance and cell death in cell culture model. (a) Prostasin decreased in paclitaxel-resistance cancer cell line. O432: ovarian cancer cell line Ovca432 (sensitive to paclitaxel); O432-RP: paclitaxel resistance cell line generated from Ovca432. The prostasin protein levels in O432 and O432-RP cells are shown in immunoblots with specific antibodies. (b) Prostasin siRNAs transfection reduced prostasin. O432-pro-D cells (transfected with prostasin siRNA) express lower prostasin, compared with control cells of O432 (transfected with reagent only) and O432-Cs (transfected with no-targeting siRNA). The prostasin protein levels are shown in immunoblots with specific antibodies. (c) Downregulation of prostasin in O432 cells resulted in increase of chemoresistant activity. Cells were treated with paclitaxel at different concentrations for 24 h (starting 48 h after siRNAs transfection) and cultured with normal medium for an additional 7 to 10 days before cell survival was assayed. Relative cell survival rates of each cell line are shown. (d) Overexpression of prostasin greatly induces cell death in ovarian cancer cells. The cell survival rates are shown after forced overexpression of prostasin in several cell lines from day-0 to day-8, respectively. (e) Prostasin cDNA transfection resulted in overexpression of prostasin in chemoresistant O432-RP cells. O432-RP-pro-O cells (transfected with prostasin cDNA) express higher prostasin compared with control cells O432-RP and O432-RP-C (transfected with control vector). The prostasin protein levels are shown in immunoblots with specific antibodies. (f) Forced overexpression of prostasin represses growth of chemoresistant cells. Relative cell growth rates are shown for O432-RP-pro-O and control cells O432-RP and O432-RP-C. (g) Forced overexpression of prostasin in O432-RP cells re-sensitizes chemoresistant cells. Cells were plated at about 10–20% confluence and treated with paclitaxel at different concentrations for 24 h, cultured with normal medium for additional 7 to 10 days, then assayed for cell survival. Relative survival rates of cell lines are shown

Mentions: To study the function of prostasin in chemoresistance, we examined prostasin expression in ovarian cancer paclitaxel-resistant cell line O432-RP (it also cross-resistant to cisplatin), a cell line we generated previously from ovarian cancer cell Ovca432 (O432),32 and performed the functional investigations. Western blot analysis showed that prostasin is deceased in drug-resistant O432-RP cell line, compared with the parental-sensitive O432 cell line (Figure 2a), suggesting prostasin may be involved in acquired chemoresistance in ovarian cancer cells. We then examined chemoresistant phenotype after knockdown prostasin in sensitive O432 cell and overexpression of prostasin in resistant cell O432-RP. siRNA knockdown prostasin in O432 cells obtained greatly reduced prostasin cells (O432-pro-D: Figure 2b) compared with the control cells. These cells were treated with paclitaxel at several doses and drug sensitivity was assessed by cell survival assays. We observed that O432-pro-D cells have a higher survival rate compared with O432 cells, which transfected with no-targeting siRNA or reagent (Figure 2c). The data demonstrated that O432-pro-D cells are more resistant to paclitaxel treatment compared with the control cells, suggesting that downregulation of prostasin alone may result in increasing chemoresistance. In contrast, we overexpressed prostasin by transfecting prostasin cDNA containing vector into O432-RP cells. To our surprise, we observed that forced overexpression of prostasin resulted in inducing cell death dramatically, with more than 99.99% cells dead after successful transfection (Figure 2d). Less than 0.01% of the surviving cells (named O432-RP-pro-O), which expressed higher prostasin (Figure 2e), were found to have a very low growth rate compared with the control cells O432-RP and O432-RP-C (transfected with empty vector; Figure 2f). These three cell sub-lines were then treated with paclitaxel at several doses and were assessed for the drug sensitivity by cell survival assays. The survival rate of O432-RP-pro-O was dramatically reduced compared with the control cells (Figure 2g), suggesting that chemoresistance may be reversed. We also tried to overexpress prostasin in other ovarian cancer cell lines, such as Uci101, OVCAR-3, and Caov-3. As seen in the O432-RP cell line, cell death was greatly induced upon prostasin overexpression. However, we did not detect any surviving cells in these cell lines after prostasin cDNA was successfully transfected, which was assessed by geneticin selection and control transfections (Figure 2e). These results indicate that forced overexpression of prostasin in ovarian cancer cells dramatically induces cell death – 100% in some cell lines and more than 99.99% in the other cell line – and represses the cell growth of surviving cells, and may partially reverse chemoresistance. These functional studies indicate prostasin may have critical roles in cell survival and chemoresistance in the studied cell models.


Prostasin may contribute to chemoresistance, repress cancer cells in ovarian cancer, and is involved in the signaling pathways of CASP/PAK2-p34/actin.

Yan BX, Ma JX, Zhang J, Guo Y, Mueller MD, Remick SC, Yu JJ - Cell Death Dis (2014)

Prostasin has important roles in chemoresistance and cell death in cell culture model. (a) Prostasin decreased in paclitaxel-resistance cancer cell line. O432: ovarian cancer cell line Ovca432 (sensitive to paclitaxel); O432-RP: paclitaxel resistance cell line generated from Ovca432. The prostasin protein levels in O432 and O432-RP cells are shown in immunoblots with specific antibodies. (b) Prostasin siRNAs transfection reduced prostasin. O432-pro-D cells (transfected with prostasin siRNA) express lower prostasin, compared with control cells of O432 (transfected with reagent only) and O432-Cs (transfected with no-targeting siRNA). The prostasin protein levels are shown in immunoblots with specific antibodies. (c) Downregulation of prostasin in O432 cells resulted in increase of chemoresistant activity. Cells were treated with paclitaxel at different concentrations for 24 h (starting 48 h after siRNAs transfection) and cultured with normal medium for an additional 7 to 10 days before cell survival was assayed. Relative cell survival rates of each cell line are shown. (d) Overexpression of prostasin greatly induces cell death in ovarian cancer cells. The cell survival rates are shown after forced overexpression of prostasin in several cell lines from day-0 to day-8, respectively. (e) Prostasin cDNA transfection resulted in overexpression of prostasin in chemoresistant O432-RP cells. O432-RP-pro-O cells (transfected with prostasin cDNA) express higher prostasin compared with control cells O432-RP and O432-RP-C (transfected with control vector). The prostasin protein levels are shown in immunoblots with specific antibodies. (f) Forced overexpression of prostasin represses growth of chemoresistant cells. Relative cell growth rates are shown for O432-RP-pro-O and control cells O432-RP and O432-RP-C. (g) Forced overexpression of prostasin in O432-RP cells re-sensitizes chemoresistant cells. Cells were plated at about 10–20% confluence and treated with paclitaxel at different concentrations for 24 h, cultured with normal medium for additional 7 to 10 days, then assayed for cell survival. Relative survival rates of cell lines are shown
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Related In: Results  -  Collection

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Show All Figures
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fig2: Prostasin has important roles in chemoresistance and cell death in cell culture model. (a) Prostasin decreased in paclitaxel-resistance cancer cell line. O432: ovarian cancer cell line Ovca432 (sensitive to paclitaxel); O432-RP: paclitaxel resistance cell line generated from Ovca432. The prostasin protein levels in O432 and O432-RP cells are shown in immunoblots with specific antibodies. (b) Prostasin siRNAs transfection reduced prostasin. O432-pro-D cells (transfected with prostasin siRNA) express lower prostasin, compared with control cells of O432 (transfected with reagent only) and O432-Cs (transfected with no-targeting siRNA). The prostasin protein levels are shown in immunoblots with specific antibodies. (c) Downregulation of prostasin in O432 cells resulted in increase of chemoresistant activity. Cells were treated with paclitaxel at different concentrations for 24 h (starting 48 h after siRNAs transfection) and cultured with normal medium for an additional 7 to 10 days before cell survival was assayed. Relative cell survival rates of each cell line are shown. (d) Overexpression of prostasin greatly induces cell death in ovarian cancer cells. The cell survival rates are shown after forced overexpression of prostasin in several cell lines from day-0 to day-8, respectively. (e) Prostasin cDNA transfection resulted in overexpression of prostasin in chemoresistant O432-RP cells. O432-RP-pro-O cells (transfected with prostasin cDNA) express higher prostasin compared with control cells O432-RP and O432-RP-C (transfected with control vector). The prostasin protein levels are shown in immunoblots with specific antibodies. (f) Forced overexpression of prostasin represses growth of chemoresistant cells. Relative cell growth rates are shown for O432-RP-pro-O and control cells O432-RP and O432-RP-C. (g) Forced overexpression of prostasin in O432-RP cells re-sensitizes chemoresistant cells. Cells were plated at about 10–20% confluence and treated with paclitaxel at different concentrations for 24 h, cultured with normal medium for additional 7 to 10 days, then assayed for cell survival. Relative survival rates of cell lines are shown
Mentions: To study the function of prostasin in chemoresistance, we examined prostasin expression in ovarian cancer paclitaxel-resistant cell line O432-RP (it also cross-resistant to cisplatin), a cell line we generated previously from ovarian cancer cell Ovca432 (O432),32 and performed the functional investigations. Western blot analysis showed that prostasin is deceased in drug-resistant O432-RP cell line, compared with the parental-sensitive O432 cell line (Figure 2a), suggesting prostasin may be involved in acquired chemoresistance in ovarian cancer cells. We then examined chemoresistant phenotype after knockdown prostasin in sensitive O432 cell and overexpression of prostasin in resistant cell O432-RP. siRNA knockdown prostasin in O432 cells obtained greatly reduced prostasin cells (O432-pro-D: Figure 2b) compared with the control cells. These cells were treated with paclitaxel at several doses and drug sensitivity was assessed by cell survival assays. We observed that O432-pro-D cells have a higher survival rate compared with O432 cells, which transfected with no-targeting siRNA or reagent (Figure 2c). The data demonstrated that O432-pro-D cells are more resistant to paclitaxel treatment compared with the control cells, suggesting that downregulation of prostasin alone may result in increasing chemoresistance. In contrast, we overexpressed prostasin by transfecting prostasin cDNA containing vector into O432-RP cells. To our surprise, we observed that forced overexpression of prostasin resulted in inducing cell death dramatically, with more than 99.99% cells dead after successful transfection (Figure 2d). Less than 0.01% of the surviving cells (named O432-RP-pro-O), which expressed higher prostasin (Figure 2e), were found to have a very low growth rate compared with the control cells O432-RP and O432-RP-C (transfected with empty vector; Figure 2f). These three cell sub-lines were then treated with paclitaxel at several doses and were assessed for the drug sensitivity by cell survival assays. The survival rate of O432-RP-pro-O was dramatically reduced compared with the control cells (Figure 2g), suggesting that chemoresistance may be reversed. We also tried to overexpress prostasin in other ovarian cancer cell lines, such as Uci101, OVCAR-3, and Caov-3. As seen in the O432-RP cell line, cell death was greatly induced upon prostasin overexpression. However, we did not detect any surviving cells in these cell lines after prostasin cDNA was successfully transfected, which was assessed by geneticin selection and control transfections (Figure 2e). These results indicate that forced overexpression of prostasin in ovarian cancer cells dramatically induces cell death – 100% in some cell lines and more than 99.99% in the other cell line – and represses the cell growth of surviving cells, and may partially reverse chemoresistance. These functional studies indicate prostasin may have critical roles in cell survival and chemoresistance in the studied cell models.

Bottom Line: In this study, we show that prostasin is decreased in an ovarian cancer drug-resistant cell line and in ovarian cancer patients with high levels of excision repair cross-complementing 1, a marker for chemoresistance.In addition, the surviving cells grow at a much lower rate compared with non-overexpressed cells.Our investigation of the molecular mechanisms suggests that prostasin may repress cancer cells and/or contribute to chemoresistance by modulating the CASP/P21-activated protein kinase (PAK2)-p34 pathway, and thereafter PAK2-p34/JNK/c-jun and PAK2-p34/mlck/actin signaling pathways.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Biochemistry, School of Medicine, Department of Basic Pharmaceutical Sciences, School of Pharmacy, and Mary Babb Randolph Cancer Center, Robert C. Byrd Health Sciences Center, West Virginia University, Morgantown, WV 26506, USA [2] IcesnowYanyan Bioscience Association, Beijing 00094, China.

ABSTRACT
Ovarian cancer is the deadliest of gynecologic cancers, largely due to the development of drug resistance in chemotherapy. Prostasin may have an essential role in the oncogenesis. In this study, we show that prostasin is decreased in an ovarian cancer drug-resistant cell line and in ovarian cancer patients with high levels of excision repair cross-complementing 1, a marker for chemoresistance. Our cell cultural model investigation demonstrates prostasin has important roles in the development of drug resistance and cancer cell survival. Forced overexpression of prostasin in ovarian cancer cells greatly induces cell death (resulting in 99% cell death in a drug-resistant cell line and 100% cell death in other tested cell lines). In addition, the surviving cells grow at a much lower rate compared with non-overexpressed cells. In vivo studies indicate that forced overexpression of prostasin in drug-resistant cells greatly inhibits the growth of tumors and may partially reverse drug resistance. Our investigation of the molecular mechanisms suggests that prostasin may repress cancer cells and/or contribute to chemoresistance by modulating the CASP/P21-activated protein kinase (PAK2)-p34 pathway, and thereafter PAK2-p34/JNK/c-jun and PAK2-p34/mlck/actin signaling pathways. Thus, we introduce prostain as a potential target for treating/repressing some ovarian tumors and have begun to identify their relevant molecular targets in specific signaling pathways.

Show MeSH
Related in: MedlinePlus