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. 2014 Aug 21;9(8):e104302.
doi: 10.1371/journal.pone.0104302. eCollection 2014.

Poly(ADP-ribose) polymerase 1 (PARP1) overexpression in human breast cancer stem cells and resistance to olaparib

Marine Gilabert  1 , Simon Launay  2 , Christophe Ginestier  3 , François Bertucci  4 , Stéphane Audebert  5 , Mathieu Pophillat  5 , Yves Toiron  5 , Emilie Baudelet  5 , Pascal Finetti  3 , Tetsuro Noguchi  6 , Hagay Sobol  7 , Daniel Birnbaum  3 , Jean-Paul Borg  8 , Emmanuelle Charafe-Jauffret  9 , Anthony Gonçalves  1
Affiliations

Poly(ADP-ribose) polymerase 1 (PARP1) overexpression in human breast cancer stem cells and resistance to olaparib

Marine Gilabert et al. PLoS One. .

Abstract

Background: Breast cancer stem cells (BCSCs) have been recognized as playing a major role in various aspects of breast cancer biology. To identify specific biomarkers of BCSCs, we have performed comparative proteomics of BCSC-enriched and mature cancer cell populations from the human breast cancer cell line (BCL), BrCA-MZ-01 VSports手机版. .

Methods: ALDEFLUOR assay was used to sort BCSC-enriched (ALDH+) and mature cancer (ALDH-) cell populations. Total proteins were extracted from both fractions and subjected to 2-Dimensional Difference In-Gel Electrophoresis (2-D DIGE) V体育安卓版. Differentially-expressed spots were excised and proteins were gel-extracted, digested and identified using MALDI-TOF MS. .

Results: 2-D DIGE identified poly(ADP-ribose) polymerase 1 (PARP1) as overexpressed in ALDH+ cells from BrCA-MZ-01. This observation was confirmed by western blot and extended to four additional human BCLs. ALDH+ cells from BRCA1-mutated HCC1937, which had the highest level of PARP1 overexpression, displayed resistance to olaparib, a specific PARP1 inhibitor V体育ios版. .

Conclusion: An unbiased proteomic approach identified PARP1 as upregulated in ALDH+, BCSC-enriched cells from various human BCLs, which may contribute to clinical resistance to PARP inhibitors. VSports最新版本.

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Conflict of interest statement

Competing Interests: The authors have read the journal’s policy and have the following conflicts: This work was supported by a grant from Sanofi-Aventis. The authors declare that they have no other conflict of interest V体育平台登录. The authors confirm that this does not alter their adherence to PLOS ONE policies on sharing data and materials.

Figures

Figure 1
Figure 1. 2D-DIGE analysis of ALDH+ versus ALDH− BrCA-MZ-01 cells. Upper panel,
Representatives three-(left) and two-(right) color merged images of 2D-DIGE gel: red spots are from the Cy5-labeled ALDH+ cells, green spots are from Cy3-labeled ALDH− cells and blue spots are from standardized samples. Lower panel, Three-dimensional simulation of the relative abundance of PARP1 protein in ALDH+ (a), standardized (b) and ALDH− samples (c).
Figure 2
Figure 2. Up-regulated expression of PARP1 in ALDH+ relative to ALDH− cells from human breast cancer cell lines.
A. Western blotting image. Presented blots are representative of at least 2 independent experiments. B. The quantitative comparison of PARP1 expression between ALDH+ and ALDH− cells, expressed as ratio. PARP1 protein expression was first normalized to Tubulin expression. * PARP1 protein expression was compared between ALDH+ and ALDH− cells using Wilcoxon signed rank test.
Figure 3
Figure 3. Effect of olaparib treatment on BCSCs from BCLs according to PARP1 expression.
Human BCLs were seeded in culture flask and incubated 72 hours in olaparib- or DMSO-containing medium. At each time and for each condition, percentage of ALDH+ and ALDH− cells were determined using ALDEFLUOR assay and absolute number of cells was counted using trypan blue. Results are presented after normalization by numbers of cells in DMSO-treated conditions and Log-2 transformation. * Mean values of ALDH+ and ALDH− cells were compared using T-test.
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References

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