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Knock-down of PRAME increases retinoic acid signaling and cytotoxic drug sensitivity of Hodgkin lymphoma cells.

Kewitz S, Staege MS - PLoS ONE (2013)

Bottom Line: DNA microarray analysis of HL cells after PRAME knock-down indicated regulation of several genes including down-regulation of known anti-apoptotic factors.Increased retinoic acid signaling in these cells was revealed by increased expression of the retinoic acid metabolizing cytochrome P450 (CYP26B1), a transcriptional target of retinoic acid signaling.Our data suggest that PRAME inhibits retinoic acid signaling in HL cells and that the knock-down of PRAME might be an interesting option for the development of new therapy strategies for patients with chemo-resistant HL.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, Martin-Luther-University Halle-Wittenberg, Halle, Germany.

ABSTRACT
The prognosis for patients with Hodgkin lymphoma (HL) has improved in recent decades. On the other hand, not all patients can be cured with the currently established therapy regimes and this therapy is associated with several adverse late effects. Therefore it is necessary to develop new therapy strategies. After treatment of L-540 HL cells with 5'-azacytidine (5AC), we observed increased expression of the preferentially expressed antigen in melanoma (PRAME). In addition, we detected an increased resistance of 5AC-treated cells against cytotoxic drugs. We analyzed the influence of PRAME on cell survival of HL cells by knocking down PRAME in the chemotherapy resistant cell line L-428, a cell line that express PRAME at a high level. After knock-down of PRAME using vector based RNA interference we observed increased sensitivity for cisplatin, etoposide and retinoic acid. DNA microarray analysis of HL cells after PRAME knock-down indicated regulation of several genes including down-regulation of known anti-apoptotic factors. Increased retinoic acid signaling in these cells was revealed by increased expression of the retinoic acid metabolizing cytochrome P450 (CYP26B1), a transcriptional target of retinoic acid signaling. Our data suggest that PRAME inhibits retinoic acid signaling in HL cells and that the knock-down of PRAME might be an interesting option for the development of new therapy strategies for patients with chemo-resistant HL.

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Knock-down of PRAME increased sensitivity for cisplatin and ATRA.Cells of the HL cell line L-428 after PRAME knock-down or transfection with control vector were treated for 24 hours with 25 µg/mL cisplatin. Cells were pre-incubated for four days with ATRA or DMSO and then treated for 24 hours with cisplatin. The viability was assessed by propidium iodide staining. In the experiments without pre-incubation, the number of living cells in the samples without cisplatin was set as 100%. In the experiments with pre-incubation, the number of living cells in the control cells with DMSO without cisplatin was set as 100%. Presented are means and standard errors from quadruplicate determination. Asterisks indicate significance (p<0.05; Students t test).
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pone-0055897-g005: Knock-down of PRAME increased sensitivity for cisplatin and ATRA.Cells of the HL cell line L-428 after PRAME knock-down or transfection with control vector were treated for 24 hours with 25 µg/mL cisplatin. Cells were pre-incubated for four days with ATRA or DMSO and then treated for 24 hours with cisplatin. The viability was assessed by propidium iodide staining. In the experiments without pre-incubation, the number of living cells in the samples without cisplatin was set as 100%. In the experiments with pre-incubation, the number of living cells in the control cells with DMSO without cisplatin was set as 100%. Presented are means and standard errors from quadruplicate determination. Asterisks indicate significance (p<0.05; Students t test).

Mentions: We asked whether PRAME knock-down influences the sensitivity for other drugs. We incubated cells with PRAME knock-down and control cells for 24 hours with 25 µg/mL cisplatin. As shown in Figure 5, the knock-down of PRAME led to an increased sensitivity for cisplatin. Cells with the control vector showed a viability of 87.8% whereas cells with PRAME knock-down show a viability of only 69%. Similar results were obtained when we treated the transfected cells with etoposide (Figure S6). Pre-incubation with RA further increases the sensitivity of cells with PRAME knock-down. Cells pre-incubated with DMSO (control) showed the same viability as cells treated with cisplatin alone. Cells pre-incubated with all-trans RA and than treated with cisplatin showed a decreased viability (Figure 5).


Knock-down of PRAME increases retinoic acid signaling and cytotoxic drug sensitivity of Hodgkin lymphoma cells.

Kewitz S, Staege MS - PLoS ONE (2013)

Knock-down of PRAME increased sensitivity for cisplatin and ATRA.Cells of the HL cell line L-428 after PRAME knock-down or transfection with control vector were treated for 24 hours with 25 µg/mL cisplatin. Cells were pre-incubated for four days with ATRA or DMSO and then treated for 24 hours with cisplatin. The viability was assessed by propidium iodide staining. In the experiments without pre-incubation, the number of living cells in the samples without cisplatin was set as 100%. In the experiments with pre-incubation, the number of living cells in the control cells with DMSO without cisplatin was set as 100%. Presented are means and standard errors from quadruplicate determination. Asterisks indicate significance (p<0.05; Students t test).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3569423&req=5

pone-0055897-g005: Knock-down of PRAME increased sensitivity for cisplatin and ATRA.Cells of the HL cell line L-428 after PRAME knock-down or transfection with control vector were treated for 24 hours with 25 µg/mL cisplatin. Cells were pre-incubated for four days with ATRA or DMSO and then treated for 24 hours with cisplatin. The viability was assessed by propidium iodide staining. In the experiments without pre-incubation, the number of living cells in the samples without cisplatin was set as 100%. In the experiments with pre-incubation, the number of living cells in the control cells with DMSO without cisplatin was set as 100%. Presented are means and standard errors from quadruplicate determination. Asterisks indicate significance (p<0.05; Students t test).
Mentions: We asked whether PRAME knock-down influences the sensitivity for other drugs. We incubated cells with PRAME knock-down and control cells for 24 hours with 25 µg/mL cisplatin. As shown in Figure 5, the knock-down of PRAME led to an increased sensitivity for cisplatin. Cells with the control vector showed a viability of 87.8% whereas cells with PRAME knock-down show a viability of only 69%. Similar results were obtained when we treated the transfected cells with etoposide (Figure S6). Pre-incubation with RA further increases the sensitivity of cells with PRAME knock-down. Cells pre-incubated with DMSO (control) showed the same viability as cells treated with cisplatin alone. Cells pre-incubated with all-trans RA and than treated with cisplatin showed a decreased viability (Figure 5).

Bottom Line: DNA microarray analysis of HL cells after PRAME knock-down indicated regulation of several genes including down-regulation of known anti-apoptotic factors.Increased retinoic acid signaling in these cells was revealed by increased expression of the retinoic acid metabolizing cytochrome P450 (CYP26B1), a transcriptional target of retinoic acid signaling.Our data suggest that PRAME inhibits retinoic acid signaling in HL cells and that the knock-down of PRAME might be an interesting option for the development of new therapy strategies for patients with chemo-resistant HL.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, Martin-Luther-University Halle-Wittenberg, Halle, Germany.

ABSTRACT
The prognosis for patients with Hodgkin lymphoma (HL) has improved in recent decades. On the other hand, not all patients can be cured with the currently established therapy regimes and this therapy is associated with several adverse late effects. Therefore it is necessary to develop new therapy strategies. After treatment of L-540 HL cells with 5'-azacytidine (5AC), we observed increased expression of the preferentially expressed antigen in melanoma (PRAME). In addition, we detected an increased resistance of 5AC-treated cells against cytotoxic drugs. We analyzed the influence of PRAME on cell survival of HL cells by knocking down PRAME in the chemotherapy resistant cell line L-428, a cell line that express PRAME at a high level. After knock-down of PRAME using vector based RNA interference we observed increased sensitivity for cisplatin, etoposide and retinoic acid. DNA microarray analysis of HL cells after PRAME knock-down indicated regulation of several genes including down-regulation of known anti-apoptotic factors. Increased retinoic acid signaling in these cells was revealed by increased expression of the retinoic acid metabolizing cytochrome P450 (CYP26B1), a transcriptional target of retinoic acid signaling. Our data suggest that PRAME inhibits retinoic acid signaling in HL cells and that the knock-down of PRAME might be an interesting option for the development of new therapy strategies for patients with chemo-resistant HL.

Show MeSH
Related in: MedlinePlus