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By promoting cell differentiation, miR-100 sensitizes basal-like breast cancer stem cells to hormonal therapy.

Petrelli A, Carollo R, Cargnelutti M, Iovino F, Callari M, Cimino D, Todaro M, Mangiapane LR, Giammona A, Cordova A, Montemurro F, Taverna D, Daidone MG, Stassi G, Giordano S - Oncotarget (2015)

Bottom Line: Moreover, miR-100 favors BrCSC differentiation, converting a basal like phenotype into luminal.It induces the expression of a functional estrogen receptor (ER) and renders basal-like BrCSCs responsive to hormonal therapy.Our findings indicate a new possible therapeutic strategy, which could make aggressive breast cancers responsive to standard treatments.

View Article: PubMed Central - PubMed

Affiliation: University of Torino School of Medicine, Candiolo Cancer Institute-FPO, IRCCS, Str. Provinciale, Candiolo, Torino, Italy.

ABSTRACT
Basal-like breast cancer is an aggressive tumor subtype with a poor response to conventional therapies. Tumor formation and relapse are sustained by a cell subset of Breast Cancer Stem Cells (BrCSCs). Here we show that miR-100 inhibits maintenance and expansion of BrCSCs in basal-like cancer through Polo-like kinase1 (Plk1) down-regulation. Moreover, miR-100 favors BrCSC differentiation, converting a basal like phenotype into luminal. It induces the expression of a functional estrogen receptor (ER) and renders basal-like BrCSCs responsive to hormonal therapy. The key role played by miR-100 in breast cancer free-survival is confirmed by the analysis of a cohort of patients' tumors, which shows that low expression of miR-100 is a negative prognostic factor and is associated with gene signatures of high grade undifferentiated tumors. Our findings indicate a new possible therapeutic strategy, which could make aggressive breast cancers responsive to standard treatments.

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MiR-100 inhibition induces a stem-like phenotype in breast cancer cellsA, phase contrast images of MCF7 cells transiently transfected with a control (ctr) or a miR-100 specific antagomir (antag). Following miR-100 antagomir transfection, obtained mammospheres retained the ability to differentiate when cultured in DMEM 10% Foetal Bovine Serum (antag 10%FBS 24h; antag 10%FBS 7 days). Magnification 4x. B, stem cell transcription factors expression in control and antagomir transfected cells, analyzed by quantitative RT-PCR. Data are average ± SD of biological replicates. MCF7 cells and mammospheres obtained from MCF7 cells upon growth in stem cell conditions (MCFS) were used as controls. * P< 0.05. C, stemness and pluripotency gene expression profiling of the cells described in (B) performed using TaqMan gene expression arrays. Gene expression is reported as −ΔCT (CT gene – CT GAPDH) median-centered. A, B, C and D indicate biological replicates.
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Figure 1: MiR-100 inhibition induces a stem-like phenotype in breast cancer cellsA, phase contrast images of MCF7 cells transiently transfected with a control (ctr) or a miR-100 specific antagomir (antag). Following miR-100 antagomir transfection, obtained mammospheres retained the ability to differentiate when cultured in DMEM 10% Foetal Bovine Serum (antag 10%FBS 24h; antag 10%FBS 7 days). Magnification 4x. B, stem cell transcription factors expression in control and antagomir transfected cells, analyzed by quantitative RT-PCR. Data are average ± SD of biological replicates. MCF7 cells and mammospheres obtained from MCF7 cells upon growth in stem cell conditions (MCFS) were used as controls. * P< 0.05. C, stemness and pluripotency gene expression profiling of the cells described in (B) performed using TaqMan gene expression arrays. Gene expression is reported as −ΔCT (CT gene – CT GAPDH) median-centered. A, B, C and D indicate biological replicates.

Mentions: Expression profiling studies showed that miR-100 is deregulated in various types of cancers [25-27]. Here, attention was focused on human breast cancer, where the biological role of miR-100 in tumor onset and progression remains elusive. The aim was to modulate miR-100 expression in vitro in breast cancer cells and study the biological consequences. The breast cancer cell line MCF7 was transiently transfected in the absence of serum, either with a miR-100 specific antagomir or a control antagomir. MiR-100 antagomir transfected cells acquired a mammosphere-like phenotype. These mammospheres retained the ability to differentiate when cultured in the presence of serum, acquiring an adherent shape (Fig. 1A). In order to ensure that antagomir-induced mammospheres showed stem cell characteristics, we analyzed the expression of the stem cell transcription factors Nanog, Oct4 and Sox2. As shown in Fig. 1B, miR-100 depleted cells expressed higher levels of the three transcription factors, compared to cells transfected with the control antagomir and to mammospheres obtained from MCF7 cells cultured in standard stem cell conditions. A wider gene expression analysis revealed that miR-100 knockdown led to a global gene reprogramming that could be responsible for the acquisition of the stem-like phenotype (Fig. 1C). Also employed was a complementary approach, evaluating miR-100 expression in mammospheres generated from breast cancer cell lines cultured in standard stem cell conditions. Consistently, the expression of the miRNA was lower in mammospheres than in the original adherent cells (Supplementary Fig. 1A,B).


By promoting cell differentiation, miR-100 sensitizes basal-like breast cancer stem cells to hormonal therapy.

Petrelli A, Carollo R, Cargnelutti M, Iovino F, Callari M, Cimino D, Todaro M, Mangiapane LR, Giammona A, Cordova A, Montemurro F, Taverna D, Daidone MG, Stassi G, Giordano S - Oncotarget (2015)

MiR-100 inhibition induces a stem-like phenotype in breast cancer cellsA, phase contrast images of MCF7 cells transiently transfected with a control (ctr) or a miR-100 specific antagomir (antag). Following miR-100 antagomir transfection, obtained mammospheres retained the ability to differentiate when cultured in DMEM 10% Foetal Bovine Serum (antag 10%FBS 24h; antag 10%FBS 7 days). Magnification 4x. B, stem cell transcription factors expression in control and antagomir transfected cells, analyzed by quantitative RT-PCR. Data are average ± SD of biological replicates. MCF7 cells and mammospheres obtained from MCF7 cells upon growth in stem cell conditions (MCFS) were used as controls. * P< 0.05. C, stemness and pluripotency gene expression profiling of the cells described in (B) performed using TaqMan gene expression arrays. Gene expression is reported as −ΔCT (CT gene – CT GAPDH) median-centered. A, B, C and D indicate biological replicates.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4385854&req=5

Figure 1: MiR-100 inhibition induces a stem-like phenotype in breast cancer cellsA, phase contrast images of MCF7 cells transiently transfected with a control (ctr) or a miR-100 specific antagomir (antag). Following miR-100 antagomir transfection, obtained mammospheres retained the ability to differentiate when cultured in DMEM 10% Foetal Bovine Serum (antag 10%FBS 24h; antag 10%FBS 7 days). Magnification 4x. B, stem cell transcription factors expression in control and antagomir transfected cells, analyzed by quantitative RT-PCR. Data are average ± SD of biological replicates. MCF7 cells and mammospheres obtained from MCF7 cells upon growth in stem cell conditions (MCFS) were used as controls. * P< 0.05. C, stemness and pluripotency gene expression profiling of the cells described in (B) performed using TaqMan gene expression arrays. Gene expression is reported as −ΔCT (CT gene – CT GAPDH) median-centered. A, B, C and D indicate biological replicates.
Mentions: Expression profiling studies showed that miR-100 is deregulated in various types of cancers [25-27]. Here, attention was focused on human breast cancer, where the biological role of miR-100 in tumor onset and progression remains elusive. The aim was to modulate miR-100 expression in vitro in breast cancer cells and study the biological consequences. The breast cancer cell line MCF7 was transiently transfected in the absence of serum, either with a miR-100 specific antagomir or a control antagomir. MiR-100 antagomir transfected cells acquired a mammosphere-like phenotype. These mammospheres retained the ability to differentiate when cultured in the presence of serum, acquiring an adherent shape (Fig. 1A). In order to ensure that antagomir-induced mammospheres showed stem cell characteristics, we analyzed the expression of the stem cell transcription factors Nanog, Oct4 and Sox2. As shown in Fig. 1B, miR-100 depleted cells expressed higher levels of the three transcription factors, compared to cells transfected with the control antagomir and to mammospheres obtained from MCF7 cells cultured in standard stem cell conditions. A wider gene expression analysis revealed that miR-100 knockdown led to a global gene reprogramming that could be responsible for the acquisition of the stem-like phenotype (Fig. 1C). Also employed was a complementary approach, evaluating miR-100 expression in mammospheres generated from breast cancer cell lines cultured in standard stem cell conditions. Consistently, the expression of the miRNA was lower in mammospheres than in the original adherent cells (Supplementary Fig. 1A,B).

Bottom Line: Moreover, miR-100 favors BrCSC differentiation, converting a basal like phenotype into luminal.It induces the expression of a functional estrogen receptor (ER) and renders basal-like BrCSCs responsive to hormonal therapy.Our findings indicate a new possible therapeutic strategy, which could make aggressive breast cancers responsive to standard treatments.

View Article: PubMed Central - PubMed

Affiliation: University of Torino School of Medicine, Candiolo Cancer Institute-FPO, IRCCS, Str. Provinciale, Candiolo, Torino, Italy.

ABSTRACT
Basal-like breast cancer is an aggressive tumor subtype with a poor response to conventional therapies. Tumor formation and relapse are sustained by a cell subset of Breast Cancer Stem Cells (BrCSCs). Here we show that miR-100 inhibits maintenance and expansion of BrCSCs in basal-like cancer through Polo-like kinase1 (Plk1) down-regulation. Moreover, miR-100 favors BrCSC differentiation, converting a basal like phenotype into luminal. It induces the expression of a functional estrogen receptor (ER) and renders basal-like BrCSCs responsive to hormonal therapy. The key role played by miR-100 in breast cancer free-survival is confirmed by the analysis of a cohort of patients' tumors, which shows that low expression of miR-100 is a negative prognostic factor and is associated with gene signatures of high grade undifferentiated tumors. Our findings indicate a new possible therapeutic strategy, which could make aggressive breast cancers responsive to standard treatments.

Show MeSH
Related in: MedlinePlus