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Cripto-1 as a novel therapeutic target for triple negative breast cancer.

Castro NP, Fedorova-Abrams ND, Merchant AS, Rangel MC, Nagaoka T, Karasawa H, Klauzinska M, Hewitt SM, Biswas K, Sharan SK, Salomon DS - Oncotarget (2015)

Bottom Line: The use of laser-capture microdissection combined with Nanostring mRNA and microRNA analysis revealed overexpression of either epithelial and miRNA-200 family or mesenchymal markers in adenocarcinoma and mesenchymal regions, respectively.Cripto-1 knockout by the CRISPR-Cas9 system inhibited tumor growth and pulmonary metastasis.Our findings show characterization of a novel mouse model that mimics the TNBC and reveal Cripto-1 as a TNBC target hence may offer alternative treatment strategies for TNBC.

View Article: PubMed Central - PubMed

Affiliation: Tumor Growth Factor Section, Mouse Cancer Genetics Program, National Cancer Institute, Frederick, MD, USA.

ABSTRACT
Triple-negative breast cancer (TNBC) presents the poorest prognosis among the breast cancer subtypes and no current standard therapy. Here, we performed an in-depth molecular analysis of a mouse model that establishes spontaneous lung metastasis from JygMC(A) cells. These primary tumors resembled the triple-negative breast cancer (TNBC) both phenotypically and molecularly. Morphologically, primary tumors presented both epithelial and spindle-like cells but displayed only adenocarcinoma-like features in lung parenchyma. The use of laser-capture microdissection combined with Nanostring mRNA and microRNA analysis revealed overexpression of either epithelial and miRNA-200 family or mesenchymal markers in adenocarcinoma and mesenchymal regions, respectively. Cripto-1, an embryonic stem cell marker, was present in spindle-like areas and its promoter showed activity in primary tumors. Cripto-1 knockout by the CRISPR-Cas9 system inhibited tumor growth and pulmonary metastasis. Our findings show characterization of a novel mouse model that mimics the TNBC and reveal Cripto-1 as a TNBC target hence may offer alternative treatment strategies for TNBC.

No MeSH data available.


Related in: MedlinePlus

Orthotopic metastasis of JygMC(A) cells and epithelial mesenchymal characterization in vitroLung metastasis by orthotopic cell injection in Balb/C nude mice. B. Representation of bioluminescent imaging of animals. Animals imaged at day 15, 22, 29, 42 and 52 post JygMC(A)-GFP/Luc cell injection. Ex-vivo imaging of lymph nodes and lung metastases. C. Immunofluorescence of luminal epithelial and basal mesenchymal markers in JygMC(A) parental cells. Epithelial marker: CDH1 (Alexa 488), luminal marker: CK18 (Alexa 488), basal marker: K5 and K14 (Alexa 488) and mesenchymal marker: Vimentin (Alexa 594). Nuclear staining in blue (DAPI). Scale bars: 200 μm.
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Figure 1: Orthotopic metastasis of JygMC(A) cells and epithelial mesenchymal characterization in vitroLung metastasis by orthotopic cell injection in Balb/C nude mice. B. Representation of bioluminescent imaging of animals. Animals imaged at day 15, 22, 29, 42 and 52 post JygMC(A)-GFP/Luc cell injection. Ex-vivo imaging of lymph nodes and lung metastases. C. Immunofluorescence of luminal epithelial and basal mesenchymal markers in JygMC(A) parental cells. Epithelial marker: CDH1 (Alexa 488), luminal marker: CK18 (Alexa 488), basal marker: K5 and K14 (Alexa 488) and mesenchymal marker: Vimentin (Alexa 594). Nuclear staining in blue (DAPI). Scale bars: 200 μm.

Mentions: In order to assess the metastatic progression of JygMC(A) mouse mammary tumor cells, we generated JygMC(A) cells stably expressing the firefly luciferase and eGFP reporter genes (JygMC(A)-GFP/Luc) to facilitate the in vivo tracking of primary and metastatic tumor cells. Cells were injected into the fourth mammary fat pad on day 1, after 30 days primary tumors were removed. Lung metastases were accessed on day 40-50 (see schematic chart of orthotopic metastasis on Figure 1A). Animals were imaged in different time-points using in vivo bioluminescent imaging (Figure 1B). To determine whether the tagged cells would have similar metastatic and tumor-initiating capacity as the parental cells, four groups of 3 animals were injected as follows: Group A = 500,000 cells, Group B = 50,000 cells, Group C = 5,000 cells, Group D = 500 cells. At day 30 after injection, the primary tumors were removed, except for Group D where tumors were removed at day 52. The parental and tagged cells exhibited similar tumor volumes (Figure S1A), primary tumor incidence, tumor-initiating capacity, metastases frequencies (Table S1). JygMC(A) cells exhibited high propensity to metastasize to the lungs and liver, and, to a small extent, the spleen and kidney when injected into the fourth mammary fat pad, as shown in Table S1.


Cripto-1 as a novel therapeutic target for triple negative breast cancer.

Castro NP, Fedorova-Abrams ND, Merchant AS, Rangel MC, Nagaoka T, Karasawa H, Klauzinska M, Hewitt SM, Biswas K, Sharan SK, Salomon DS - Oncotarget (2015)

Orthotopic metastasis of JygMC(A) cells and epithelial mesenchymal characterization in vitroLung metastasis by orthotopic cell injection in Balb/C nude mice. B. Representation of bioluminescent imaging of animals. Animals imaged at day 15, 22, 29, 42 and 52 post JygMC(A)-GFP/Luc cell injection. Ex-vivo imaging of lymph nodes and lung metastases. C. Immunofluorescence of luminal epithelial and basal mesenchymal markers in JygMC(A) parental cells. Epithelial marker: CDH1 (Alexa 488), luminal marker: CK18 (Alexa 488), basal marker: K5 and K14 (Alexa 488) and mesenchymal marker: Vimentin (Alexa 594). Nuclear staining in blue (DAPI). Scale bars: 200 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4494913&req=5

Figure 1: Orthotopic metastasis of JygMC(A) cells and epithelial mesenchymal characterization in vitroLung metastasis by orthotopic cell injection in Balb/C nude mice. B. Representation of bioluminescent imaging of animals. Animals imaged at day 15, 22, 29, 42 and 52 post JygMC(A)-GFP/Luc cell injection. Ex-vivo imaging of lymph nodes and lung metastases. C. Immunofluorescence of luminal epithelial and basal mesenchymal markers in JygMC(A) parental cells. Epithelial marker: CDH1 (Alexa 488), luminal marker: CK18 (Alexa 488), basal marker: K5 and K14 (Alexa 488) and mesenchymal marker: Vimentin (Alexa 594). Nuclear staining in blue (DAPI). Scale bars: 200 μm.
Mentions: In order to assess the metastatic progression of JygMC(A) mouse mammary tumor cells, we generated JygMC(A) cells stably expressing the firefly luciferase and eGFP reporter genes (JygMC(A)-GFP/Luc) to facilitate the in vivo tracking of primary and metastatic tumor cells. Cells were injected into the fourth mammary fat pad on day 1, after 30 days primary tumors were removed. Lung metastases were accessed on day 40-50 (see schematic chart of orthotopic metastasis on Figure 1A). Animals were imaged in different time-points using in vivo bioluminescent imaging (Figure 1B). To determine whether the tagged cells would have similar metastatic and tumor-initiating capacity as the parental cells, four groups of 3 animals were injected as follows: Group A = 500,000 cells, Group B = 50,000 cells, Group C = 5,000 cells, Group D = 500 cells. At day 30 after injection, the primary tumors were removed, except for Group D where tumors were removed at day 52. The parental and tagged cells exhibited similar tumor volumes (Figure S1A), primary tumor incidence, tumor-initiating capacity, metastases frequencies (Table S1). JygMC(A) cells exhibited high propensity to metastasize to the lungs and liver, and, to a small extent, the spleen and kidney when injected into the fourth mammary fat pad, as shown in Table S1.

Bottom Line: The use of laser-capture microdissection combined with Nanostring mRNA and microRNA analysis revealed overexpression of either epithelial and miRNA-200 family or mesenchymal markers in adenocarcinoma and mesenchymal regions, respectively.Cripto-1 knockout by the CRISPR-Cas9 system inhibited tumor growth and pulmonary metastasis.Our findings show characterization of a novel mouse model that mimics the TNBC and reveal Cripto-1 as a TNBC target hence may offer alternative treatment strategies for TNBC.

View Article: PubMed Central - PubMed

Affiliation: Tumor Growth Factor Section, Mouse Cancer Genetics Program, National Cancer Institute, Frederick, MD, USA.

ABSTRACT
Triple-negative breast cancer (TNBC) presents the poorest prognosis among the breast cancer subtypes and no current standard therapy. Here, we performed an in-depth molecular analysis of a mouse model that establishes spontaneous lung metastasis from JygMC(A) cells. These primary tumors resembled the triple-negative breast cancer (TNBC) both phenotypically and molecularly. Morphologically, primary tumors presented both epithelial and spindle-like cells but displayed only adenocarcinoma-like features in lung parenchyma. The use of laser-capture microdissection combined with Nanostring mRNA and microRNA analysis revealed overexpression of either epithelial and miRNA-200 family or mesenchymal markers in adenocarcinoma and mesenchymal regions, respectively. Cripto-1, an embryonic stem cell marker, was present in spindle-like areas and its promoter showed activity in primary tumors. Cripto-1 knockout by the CRISPR-Cas9 system inhibited tumor growth and pulmonary metastasis. Our findings show characterization of a novel mouse model that mimics the TNBC and reveal Cripto-1 as a TNBC target hence may offer alternative treatment strategies for TNBC.

No MeSH data available.


Related in: MedlinePlus