Elucidation of altered pathways in tumor-initiating cells of triple-negative breast cancer: A useful cell model system for drug screening

Anne G Christensen, Sidse Ehmsen, Mikkel G Terp, Richa Batra, Nicolas Alcaraz, Jan Baumbach, Julie Boertmann Noer, José M A Moreira, Rikke Leth-Larsen, Martin R Larsen, Henrik J Ditzel

Publikation: Bidrag til tidsskriftTidsskriftartikelForskningpeer review

Resumé

A limited number of cancer cells within a tumor are thought to have self-renewing and tumor-initiating capabilities that produce the remaining cancer cells in a heterogeneous tumor mass. Elucidation of central pathways preferentially used by tumor-initiating cells/cancer stem cells (CSCs) may allow their exploitation as potential cancer therapy targets. We used single cell cloning to isolate and characterize four isogenic cell clones from a triple-negative breast cancer cell line; two exhibited mesenchymal-like and two epithelial-like characteristics. Within these pairs, one, but not the other, resulted in tumors in immunodeficient NOD/Shi-scid/IL-2 Rγ null mice and efficiently formed mammospheres. Quantitative proteomics and phosphoproteomics were used to map signaling pathways associated with the tumor-initiating ability. Signaling associated with apoptosis was suppressed in tumor-initiating versus nontumorigenic counterparts with pro-apoptotic proteins, such as Bcl2-associated agonist of cell death (BAD), FAS-associated death domain protein (FADD), and myeloid differentiation primary response protein (MYD88), downregulated in tumor-initiating epithelial-like cells. Functional studies confirmed significantly lower apoptosis in tumor-initiating versus nontumorigenic cells. Moreover, central pathways, including β-catenin and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB)-related signaling, exhibited increased activation in the tumor-initiating cells. To evaluate the CSC model as a tool for drug screening, we assessed the effect of separately blocking NF-κB and Wnt/β-catenin signaling and found markedly reduced mammosphere formation, particularly for tumor-initiating cells. Similar reduction was also observed using patient-derived primary cancer cells. Furthermore, blocking NF-κB signaling in mice transplanted with tumor-initiating cells significantly reduced tumor outgrowth. Our study demonstrates that suppressed apoptosis, activation of pathways associated with cell viability, and CSCs are the major differences between tumor-initiating and nontumorigenic cells independent of their epithelial-like/mesenchymal-like phenotype. These altered pathways may provide targets for future drug development to eliminate CSCs, and the cell model may be a useful tool in such drug screenings. Stem Cells 2017;35:1898–1912.

OriginalsprogEngelsk
TidsskriftStem Cells
Vol/bind35
Udgave nummer8
Sider (fra-til)1898–1912
ISSN1066-5099
DOI
StatusUdgivet - aug. 2017

Fingeraftryk

Neoplasms
Apoptosis
Apoptosis Regulatory Proteins
Interleukin-2
Organism Cloning
Cell Survival
Proteins
Cell Death
Down-Regulation
Clone Cells
Epithelial Cells
Cell Line
Pharmaceutical Preparations

Citer dette

@article{7a3e311c20c34c7a950922dd09c05a69,
title = "Elucidation of altered pathways in tumor-initiating cells of triple-negative breast cancer: A useful cell model system for drug screening",
abstract = "A limited number of cancer cells within a tumor are thought to have self-renewing and tumor-initiating capabilities that produce the remaining cancer cells in a heterogeneous tumor mass. Elucidation of central pathways preferentially used by tumor-initiating cells/cancer stem cells (CSCs) may allow their exploitation as potential cancer therapy targets. We used single cell cloning to isolate and characterize four isogenic cell clones from a triple-negative breast cancer cell line; two exhibited mesenchymal-like and two epithelial-like characteristics. Within these pairs, one, but not the other, resulted in tumors in immunodeficient NOD/Shi-scid/IL-2 Rγ null mice and efficiently formed mammospheres. Quantitative proteomics and phosphoproteomics were used to map signaling pathways associated with the tumor-initiating ability. Signaling associated with apoptosis was suppressed in tumor-initiating versus nontumorigenic counterparts with pro-apoptotic proteins, such as Bcl2-associated agonist of cell death (BAD), FAS-associated death domain protein (FADD), and myeloid differentiation primary response protein (MYD88), downregulated in tumor-initiating epithelial-like cells. Functional studies confirmed significantly lower apoptosis in tumor-initiating versus nontumorigenic cells. Moreover, central pathways, including β-catenin and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB)-related signaling, exhibited increased activation in the tumor-initiating cells. To evaluate the CSC model as a tool for drug screening, we assessed the effect of separately blocking NF-κB and Wnt/β-catenin signaling and found markedly reduced mammosphere formation, particularly for tumor-initiating cells. Similar reduction was also observed using patient-derived primary cancer cells. Furthermore, blocking NF-κB signaling in mice transplanted with tumor-initiating cells significantly reduced tumor outgrowth. Our study demonstrates that suppressed apoptosis, activation of pathways associated with cell viability, and CSCs are the major differences between tumor-initiating and nontumorigenic cells independent of their epithelial-like/mesenchymal-like phenotype. These altered pathways may provide targets for future drug development to eliminate CSCs, and the cell model may be a useful tool in such drug screenings. Stem Cells 2017;35:1898–1912.",
keywords = "Journal Article, NF-κB, Targeted treatment, Cancer stem cells, Mammospheres, Apoptosis, Triple-negative breast cancer, Cell Proliferation, Antigens, CD/metabolism, Humans, Neoplastic Stem Cells/pathology, Protein Interaction Maps, Triple Negative Breast Neoplasms/drug therapy, Cell Shape, Mass Spectrometry, Epithelial-Mesenchymal Transition, Female, Drug Evaluation, Preclinical, Wnt Signaling Pathway, Spheroids, Cellular/pathology, Reproducibility of Results, Signal Transduction, Cell Survival, Carcinogenesis/metabolism, Animals, Models, Biological, Proteomics, Cell Line, Tumor, Mice, Biomarkers, Tumor/metabolism",
author = "Christensen, {Anne G} and Sidse Ehmsen and Terp, {Mikkel G} and Richa Batra and Nicolas Alcaraz and Jan Baumbach and Noer, {Julie Boertmann} and Moreira, {Jos{\'e} M A} and Rikke Leth-Larsen and Larsen, {Martin R} and Ditzel, {Henrik J}",
note = "{\circledC} 2017 AlphaMed Press.",
year = "2017",
month = "8",
doi = "10.1002/stem.2654",
language = "English",
volume = "35",
pages = "1898–1912",
journal = "Stem Cells",
issn = "1066-5099",
publisher = "AlphaMed Press, Inc.",
number = "8",

}

Elucidation of altered pathways in tumor-initiating cells of triple-negative breast cancer : A useful cell model system for drug screening. / Christensen, Anne G; Ehmsen, Sidse; Terp, Mikkel G; Batra, Richa; Alcaraz, Nicolas; Baumbach, Jan; Noer, Julie Boertmann; Moreira, José M A; Leth-Larsen, Rikke; Larsen, Martin R; Ditzel, Henrik J.

I: Stem Cells, Bind 35, Nr. 8, 08.2017, s. 1898–1912.

Publikation: Bidrag til tidsskriftTidsskriftartikelForskningpeer review

TY - JOUR

T1 - Elucidation of altered pathways in tumor-initiating cells of triple-negative breast cancer

T2 - A useful cell model system for drug screening

AU - Christensen, Anne G

AU - Ehmsen, Sidse

AU - Terp, Mikkel G

AU - Batra, Richa

AU - Alcaraz, Nicolas

AU - Baumbach, Jan

AU - Noer, Julie Boertmann

AU - Moreira, José M A

AU - Leth-Larsen, Rikke

AU - Larsen, Martin R

AU - Ditzel, Henrik J

N1 - © 2017 AlphaMed Press.

PY - 2017/8

Y1 - 2017/8

N2 - A limited number of cancer cells within a tumor are thought to have self-renewing and tumor-initiating capabilities that produce the remaining cancer cells in a heterogeneous tumor mass. Elucidation of central pathways preferentially used by tumor-initiating cells/cancer stem cells (CSCs) may allow their exploitation as potential cancer therapy targets. We used single cell cloning to isolate and characterize four isogenic cell clones from a triple-negative breast cancer cell line; two exhibited mesenchymal-like and two epithelial-like characteristics. Within these pairs, one, but not the other, resulted in tumors in immunodeficient NOD/Shi-scid/IL-2 Rγ null mice and efficiently formed mammospheres. Quantitative proteomics and phosphoproteomics were used to map signaling pathways associated with the tumor-initiating ability. Signaling associated with apoptosis was suppressed in tumor-initiating versus nontumorigenic counterparts with pro-apoptotic proteins, such as Bcl2-associated agonist of cell death (BAD), FAS-associated death domain protein (FADD), and myeloid differentiation primary response protein (MYD88), downregulated in tumor-initiating epithelial-like cells. Functional studies confirmed significantly lower apoptosis in tumor-initiating versus nontumorigenic cells. Moreover, central pathways, including β-catenin and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB)-related signaling, exhibited increased activation in the tumor-initiating cells. To evaluate the CSC model as a tool for drug screening, we assessed the effect of separately blocking NF-κB and Wnt/β-catenin signaling and found markedly reduced mammosphere formation, particularly for tumor-initiating cells. Similar reduction was also observed using patient-derived primary cancer cells. Furthermore, blocking NF-κB signaling in mice transplanted with tumor-initiating cells significantly reduced tumor outgrowth. Our study demonstrates that suppressed apoptosis, activation of pathways associated with cell viability, and CSCs are the major differences between tumor-initiating and nontumorigenic cells independent of their epithelial-like/mesenchymal-like phenotype. These altered pathways may provide targets for future drug development to eliminate CSCs, and the cell model may be a useful tool in such drug screenings. Stem Cells 2017;35:1898–1912.

AB - A limited number of cancer cells within a tumor are thought to have self-renewing and tumor-initiating capabilities that produce the remaining cancer cells in a heterogeneous tumor mass. Elucidation of central pathways preferentially used by tumor-initiating cells/cancer stem cells (CSCs) may allow their exploitation as potential cancer therapy targets. We used single cell cloning to isolate and characterize four isogenic cell clones from a triple-negative breast cancer cell line; two exhibited mesenchymal-like and two epithelial-like characteristics. Within these pairs, one, but not the other, resulted in tumors in immunodeficient NOD/Shi-scid/IL-2 Rγ null mice and efficiently formed mammospheres. Quantitative proteomics and phosphoproteomics were used to map signaling pathways associated with the tumor-initiating ability. Signaling associated with apoptosis was suppressed in tumor-initiating versus nontumorigenic counterparts with pro-apoptotic proteins, such as Bcl2-associated agonist of cell death (BAD), FAS-associated death domain protein (FADD), and myeloid differentiation primary response protein (MYD88), downregulated in tumor-initiating epithelial-like cells. Functional studies confirmed significantly lower apoptosis in tumor-initiating versus nontumorigenic cells. Moreover, central pathways, including β-catenin and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB)-related signaling, exhibited increased activation in the tumor-initiating cells. To evaluate the CSC model as a tool for drug screening, we assessed the effect of separately blocking NF-κB and Wnt/β-catenin signaling and found markedly reduced mammosphere formation, particularly for tumor-initiating cells. Similar reduction was also observed using patient-derived primary cancer cells. Furthermore, blocking NF-κB signaling in mice transplanted with tumor-initiating cells significantly reduced tumor outgrowth. Our study demonstrates that suppressed apoptosis, activation of pathways associated with cell viability, and CSCs are the major differences between tumor-initiating and nontumorigenic cells independent of their epithelial-like/mesenchymal-like phenotype. These altered pathways may provide targets for future drug development to eliminate CSCs, and the cell model may be a useful tool in such drug screenings. Stem Cells 2017;35:1898–1912.

KW - Journal Article

KW - NF-κB

KW - Targeted treatment

KW - Cancer stem cells

KW - Mammospheres

KW - Apoptosis

KW - Triple-negative breast cancer

KW - Cell Proliferation

KW - Antigens, CD/metabolism

KW - Humans

KW - Neoplastic Stem Cells/pathology

KW - Protein Interaction Maps

KW - Triple Negative Breast Neoplasms/drug therapy

KW - Cell Shape

KW - Mass Spectrometry

KW - Epithelial-Mesenchymal Transition

KW - Female

KW - Drug Evaluation, Preclinical

KW - Wnt Signaling Pathway

KW - Spheroids, Cellular/pathology

KW - Reproducibility of Results

KW - Signal Transduction

KW - Cell Survival

KW - Carcinogenesis/metabolism

KW - Animals

KW - Models, Biological

KW - Proteomics

KW - Cell Line, Tumor

KW - Mice

KW - Biomarkers, Tumor/metabolism

U2 - 10.1002/stem.2654

DO - 10.1002/stem.2654

M3 - Journal article

C2 - 28600813

VL - 35

SP - 1898

EP - 1912

JO - Stem Cells

JF - Stem Cells

SN - 1066-5099

IS - 8

ER -