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Ulipristal Acetate Inhibits Progesterone Receptor Isoform A-Mediated Human Breast Cancer Proliferation and BCl2-L1 Expression.

Esber N, Le Billan F, Resche-Rigon M, Loosfelt H, Lombès M, Chabbert-Buffet N - PLoS ONE (2015)

Bottom Line: P4 significantly stimulated MDA-iPRA expressing cells proliferation.UPA decreased cell proliferation and repressed BCL2-L1 expression in the presence of PRA, correlating with PRA and SRC1 but not RNA Pol II recruitment.Further confirmation in the clinical setting is required.

View Article: PubMed Central - PubMed

Affiliation: Institut National de la Santé et de la Recherche Médicale, Unité Mixte de Recherche-Scientifique 1185, Faculté de Médecine Paris Sud, Le Kremlin-Bicêtre, France; Université Paris-Sud, Faculté de Médecine Paris Sud, Unité Mixte de Recherche-Scientifique 1185, Le Kremlin-Bicêtre, France; HRA-Pharma, Paris, France.

ABSTRACT
The progesterone receptor (PR) with its isoforms and ligands are involved in breast tumorigenesis and prognosis. We aimed at analyzing the respective contribution of PR isoforms, PRA and PRB, in breast cancer cell proliferation in a new estrogen-independent cell based-model, allowing independent PR isoforms analysis. We used the bi-inducible human breast cancer cell system MDA-iPRAB. We studied the effects and molecular mechanisms of action of progesterone (P4) and ulipristal acetate (UPA), a new selective progesterone receptor modulator, alone or in combination. P4 significantly stimulated MDA-iPRA expressing cells proliferation. This was associated with P4-stimulated expression of the anti-apoptotic factor BCL2-L1 and enhanced recruitment of PRA, SRC-1 and RNA Pol II onto the +58 kb PR binding motif of the BCL2-L1 gene. UPA decreased cell proliferation and repressed BCL2-L1 expression in the presence of PRA, correlating with PRA and SRC1 but not RNA Pol II recruitment. These results bring new information on the mechanism of action of PR ligands in controlling breast cancer cell proliferation through PRA in an estrogen independent model. Evaluation of PR isoforms ratio, as well as molecular signature studies based on PRA target genes could be proposed to facilitate personalized breast cancer therapy. In this context, UPA could be of interest in endocrine therapy. Further confirmation in the clinical setting is required.

No MeSH data available.


Related in: MedlinePlus

Hormonal regulation of P4-PRA-upregulated genes.(a) BCL2-L1, (b) DUSP6, (c) WNT5A, (d) LGR4 mRNA expression levels were determined in MDA-iPRA cells. Cells were treated for 6 h with vehicle, P4 (1 nM) and/or UPA (1 μM) in steroid-free medium, following 24 h induction of PRA expression using RSL1 (0.5 μM). RT-qPCR analyses were performed as described in Materials and Methods. Data are expressed as fold induction compared to vehicle condition arbitrarily set at 1, and are means ± SEM from three independent cell cultures measured in duplicate. *, **, *** symbols indicate p< 0.05, 0.01 and 0.001 respectively compared to the vehicle-treated MDA-iPR- cells while x, xx, xxx symbols indicate p<0.05, 0.01 and 0.001 respectively compared to the V or P4-treated MDA-iPRA cells (non-parametric Mann Whitney t-tests).
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pone.0140795.g003: Hormonal regulation of P4-PRA-upregulated genes.(a) BCL2-L1, (b) DUSP6, (c) WNT5A, (d) LGR4 mRNA expression levels were determined in MDA-iPRA cells. Cells were treated for 6 h with vehicle, P4 (1 nM) and/or UPA (1 μM) in steroid-free medium, following 24 h induction of PRA expression using RSL1 (0.5 μM). RT-qPCR analyses were performed as described in Materials and Methods. Data are expressed as fold induction compared to vehicle condition arbitrarily set at 1, and are means ± SEM from three independent cell cultures measured in duplicate. *, **, *** symbols indicate p< 0.05, 0.01 and 0.001 respectively compared to the vehicle-treated MDA-iPR- cells while x, xx, xxx symbols indicate p<0.05, 0.01 and 0.001 respectively compared to the V or P4-treated MDA-iPRA cells (non-parametric Mann Whitney t-tests).

Mentions: We have previously identified by microarray analysis on the same MDA-iPRAB cell model, a subset of potential target gene candidates regulated in a P4-dependent and/or PR isoform specific manner [6]. Given the importance of PRA isoform, its impact on P4 signaling in mammary tumorigenesis processes such as cell growth, death and migration, and taking into account its role in mediating cell proliferation, we chose to focus our attention on some PRA-dependent upregulated (Table 1) or downregulated genes (Table 2) in the presence of P4. It would be thus possible to correlate P4-stimulated MDA-iPRA cell proliferation to the corresponding gene expression and to further evaluate the activity of the antiprogestin UPA in these processes. Among PRA-targeted genes transcriptionally stimulated (Fig 3) or inhibited by P4 (Fig 4), we examined the expression of few genes (BCL2-L1, DUSP6, WNT5A, LGR4, TGFβ2, EREG, F3, F2RL1), owing to their role in apoptosis, cell proliferation, metastasis, invasiveness, angiogenesis (Tables 1 and 2). We first validated the regulated expression of these genes by RT-qPCR analysis, after a 6 hr treatment by P4 following the same experimental duration as previously described [6]. We found that P4 significantly enhanced expression of BCL2-L1 by 25% (Fig 3A), that of DUSP6 by 50% (Fig 3B), that of WNT5A by 30% (Fig 3C) while LGR4 expression was increased by 60% (Fig 3D). Interestingly, we demonstrated that the expression of these P4-upregulated genes was repressed by UPA alone or in combination with P4, confirming the antagonistic activity of UPA (Fig 3).


Ulipristal Acetate Inhibits Progesterone Receptor Isoform A-Mediated Human Breast Cancer Proliferation and BCl2-L1 Expression.

Esber N, Le Billan F, Resche-Rigon M, Loosfelt H, Lombès M, Chabbert-Buffet N - PLoS ONE (2015)

Hormonal regulation of P4-PRA-upregulated genes.(a) BCL2-L1, (b) DUSP6, (c) WNT5A, (d) LGR4 mRNA expression levels were determined in MDA-iPRA cells. Cells were treated for 6 h with vehicle, P4 (1 nM) and/or UPA (1 μM) in steroid-free medium, following 24 h induction of PRA expression using RSL1 (0.5 μM). RT-qPCR analyses were performed as described in Materials and Methods. Data are expressed as fold induction compared to vehicle condition arbitrarily set at 1, and are means ± SEM from three independent cell cultures measured in duplicate. *, **, *** symbols indicate p< 0.05, 0.01 and 0.001 respectively compared to the vehicle-treated MDA-iPR- cells while x, xx, xxx symbols indicate p<0.05, 0.01 and 0.001 respectively compared to the V or P4-treated MDA-iPRA cells (non-parametric Mann Whitney t-tests).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4608808&req=5

pone.0140795.g003: Hormonal regulation of P4-PRA-upregulated genes.(a) BCL2-L1, (b) DUSP6, (c) WNT5A, (d) LGR4 mRNA expression levels were determined in MDA-iPRA cells. Cells were treated for 6 h with vehicle, P4 (1 nM) and/or UPA (1 μM) in steroid-free medium, following 24 h induction of PRA expression using RSL1 (0.5 μM). RT-qPCR analyses were performed as described in Materials and Methods. Data are expressed as fold induction compared to vehicle condition arbitrarily set at 1, and are means ± SEM from three independent cell cultures measured in duplicate. *, **, *** symbols indicate p< 0.05, 0.01 and 0.001 respectively compared to the vehicle-treated MDA-iPR- cells while x, xx, xxx symbols indicate p<0.05, 0.01 and 0.001 respectively compared to the V or P4-treated MDA-iPRA cells (non-parametric Mann Whitney t-tests).
Mentions: We have previously identified by microarray analysis on the same MDA-iPRAB cell model, a subset of potential target gene candidates regulated in a P4-dependent and/or PR isoform specific manner [6]. Given the importance of PRA isoform, its impact on P4 signaling in mammary tumorigenesis processes such as cell growth, death and migration, and taking into account its role in mediating cell proliferation, we chose to focus our attention on some PRA-dependent upregulated (Table 1) or downregulated genes (Table 2) in the presence of P4. It would be thus possible to correlate P4-stimulated MDA-iPRA cell proliferation to the corresponding gene expression and to further evaluate the activity of the antiprogestin UPA in these processes. Among PRA-targeted genes transcriptionally stimulated (Fig 3) or inhibited by P4 (Fig 4), we examined the expression of few genes (BCL2-L1, DUSP6, WNT5A, LGR4, TGFβ2, EREG, F3, F2RL1), owing to their role in apoptosis, cell proliferation, metastasis, invasiveness, angiogenesis (Tables 1 and 2). We first validated the regulated expression of these genes by RT-qPCR analysis, after a 6 hr treatment by P4 following the same experimental duration as previously described [6]. We found that P4 significantly enhanced expression of BCL2-L1 by 25% (Fig 3A), that of DUSP6 by 50% (Fig 3B), that of WNT5A by 30% (Fig 3C) while LGR4 expression was increased by 60% (Fig 3D). Interestingly, we demonstrated that the expression of these P4-upregulated genes was repressed by UPA alone or in combination with P4, confirming the antagonistic activity of UPA (Fig 3).

Bottom Line: P4 significantly stimulated MDA-iPRA expressing cells proliferation.UPA decreased cell proliferation and repressed BCL2-L1 expression in the presence of PRA, correlating with PRA and SRC1 but not RNA Pol II recruitment.Further confirmation in the clinical setting is required.

View Article: PubMed Central - PubMed

Affiliation: Institut National de la Santé et de la Recherche Médicale, Unité Mixte de Recherche-Scientifique 1185, Faculté de Médecine Paris Sud, Le Kremlin-Bicêtre, France; Université Paris-Sud, Faculté de Médecine Paris Sud, Unité Mixte de Recherche-Scientifique 1185, Le Kremlin-Bicêtre, France; HRA-Pharma, Paris, France.

ABSTRACT
The progesterone receptor (PR) with its isoforms and ligands are involved in breast tumorigenesis and prognosis. We aimed at analyzing the respective contribution of PR isoforms, PRA and PRB, in breast cancer cell proliferation in a new estrogen-independent cell based-model, allowing independent PR isoforms analysis. We used the bi-inducible human breast cancer cell system MDA-iPRAB. We studied the effects and molecular mechanisms of action of progesterone (P4) and ulipristal acetate (UPA), a new selective progesterone receptor modulator, alone or in combination. P4 significantly stimulated MDA-iPRA expressing cells proliferation. This was associated with P4-stimulated expression of the anti-apoptotic factor BCL2-L1 and enhanced recruitment of PRA, SRC-1 and RNA Pol II onto the +58 kb PR binding motif of the BCL2-L1 gene. UPA decreased cell proliferation and repressed BCL2-L1 expression in the presence of PRA, correlating with PRA and SRC1 but not RNA Pol II recruitment. These results bring new information on the mechanism of action of PR ligands in controlling breast cancer cell proliferation through PRA in an estrogen independent model. Evaluation of PR isoforms ratio, as well as molecular signature studies based on PRA target genes could be proposed to facilitate personalized breast cancer therapy. In this context, UPA could be of interest in endocrine therapy. Further confirmation in the clinical setting is required.

No MeSH data available.


Related in: MedlinePlus