ECM1 secreted by HER2-overexpressing breast cancer cells promotes formation of a vascular niche accelerating cancer cell migration and invasion

Sophie Sarah Steinhaeuser; Erika Morera; Zuzana Budkova; Alexander Schepsky; Qiong Wang; Óttar Rolfsson; Angela Riedel; Aileen Krueger; Bylgja Hilmarsdóttir; Gunhild Mari Maelandsmo; Bryndis Valdimarsdottir; Anna Karen Sigurðardóttir; Bjarni Agnar Agnarsson; Jón Gunnlaugur Jónasson; Sævar Ingþórsson; Gunnhildur Ásta Traustadóttir; Thordur Oskarsson; Thorarinn Gudjonsson

doi:10.1038/s41374-020-0415-6

ECM1 secreted by HER2-overexpressing breast cancer cells promotes formation of a vascular niche accelerating cancer cell migration and invasion

Sophie Sarah Steinhaeuser
, Erika Morera
, Zuzana Budkova
, Alexander Schepsky
, Qiong Wang
, Óttar Rolfsson
, Angela Riedel
, Aileen Krueger
, Bylgja Hilmarsdóttir
, Gunhild Mari Maelandsmo
, Bryndis Valdimarsdottir
, Anna Karen Sigurðardóttir
, Bjarni Agnar Agnarsson
, Jón Gunnlaugur Jónasson
, Sævar Ingþórsson
, Gunnhildur Ásta Traustadóttir
, Thordur Oskarsson
, Thorarinn Gudjonsson

University of Iceland, Landspitali

Research output: Contribution to journal › Article › peer-review

Abstract

The tumor microenvironment is increasingly recognized as key player in cancer progression. Investigating heterotypic interactions between cancer cells and their microenvironment is important for understanding how specific cell types support cancer. Forming the vasculature, endothelial cells (ECs) are a prominent cell type in the microenvironment of both normal and neoplastic breast gland. Here, we sought out to analyze epithelial–endothelial cross talk in the breast using isogenic non-tumorigenic vs. tumorigenic breast epithelial cell lines and primary ECs. The cellular model used here consists of D492, a breast epithelial cell line with stem cell properties, and two isogenic D492-derived EMT cell lines, D492M and D492HER2. D492M was generated by endothelial-induced EMT and is non-tumorigenic while D492HER2 is tumorigenic, expressing the ErbB2/HER2 oncogene. To investigate cellular cross talk, we used both conditioned medium (CM) and 2D/3D co-culture systems. Secretome analysis of D492 cell lines was performed using mass spectrometry and candidate knockdown (KD), and overexpression (OE) was done using siRNA and CRISPRi/CRISPRa technology. D492HER2 directly enhances endothelial network formation and activates a molecular axis in ECs promoting D492HER2 migration and invasion, suggesting an endothelial feedback response. Secretome analysis identified extracellular matrix protein 1 (ECM1) as potential angiogenic inducer in D492HER2. Confirming its involvement, KD of ECM1 reduced the ability of D492HER2-CM to increase endothelial network formation and induce the endothelial feedback, while recombinant ECM1 (rECM1) increased both. Interestingly, NOTCH1 and NOTCH3 expression was upregulated in ECs upon treatment with D492HER2-CM or rECM1 but not by CM from D492HER2 with ECM1 KD. Blocking endothelial NOTCH signaling inhibited the increase in network formation and the ability of ECs to promote D492HER2 migration and invasion. In summary, our data demonstrate that cancer-secreted ECM1 induces a NOTCH-mediated endothelial feedback promoting cancer progression by enhancing migration and invasion. Targeting this interaction may provide a novel possibility to improve cancer treatment.

Original language	English
Pages (from-to)	928-944
Number of pages	17
Journal	Laboratory Investigation
Volume	100
Issue number	7
DOIs	https://doi.org/10.1038/s41374-020-0415-6
Publication status	Published - 1 Jul 2020

Bibliographical note

Funding Information: Acknowledgements We thank the FingerPrints Proteomics Facility at the University of Dundee for performing the mass spectrometry and the Center for Systems Biology at the University of Iceland for the technical support during mass spectrometry data analysis. Further, we want to thank the HI-STEM laboratory and the sequencing core facility at the German Cancer Research Center, DKFZ Heidelberg, Germany, for technical support and performing the RNA microarray. Finally, we thank Prof. Dr Haraldur Haraldsson, University of Iceland for providing primary human umbilical vascular ECs. Human umbilical vascular ECs were obtained from umbilical chords at the child-birth department, University Hospital Iceland (ethical application nr. 35/2013). This work was supported by Grants from Landspitali University Hospital Science Fund, University of Iceland Research Fund (Grant of Excellence: #152144051 Doctoral Grant: #185042051), and Icelandic Science and Technology Policy. Publisher Copyright: © 2020, The Author(s), under exclusive licence to United States and Canadian Academy of Pathology.

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

SDG 3 Good Health and Well-being

Other keywords

Brjóstakrabbamein

Access to Document

10.1038/s41374-020-0415-6

Cite this

Steinhaeuser, S. S., Morera, E., Budkova, Z., Schepsky, A., Wang, Q., Rolfsson, Ó., Riedel, A., Krueger, A., Hilmarsdóttir, B., Maelandsmo, G. M., Valdimarsdottir, B., Sigurðardóttir, A. K., Agnarsson, B. A., Jónasson, J. G., Ingþórsson, S., Traustadóttir, G. Á., Oskarsson, T., & Gudjonsson, T. (2020). ECM1 secreted by HER2-overexpressing breast cancer cells promotes formation of a vascular niche accelerating cancer cell migration and invasion. Laboratory Investigation, 100(7), 928-944. https://doi.org/10.1038/s41374-020-0415-6

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title = "ECM1 secreted by HER2-overexpressing breast cancer cells promotes formation of a vascular niche accelerating cancer cell migration and invasion",

abstract = "The tumor microenvironment is increasingly recognized as key player in cancer progression. Investigating heterotypic interactions between cancer cells and their microenvironment is important for understanding how specific cell types support cancer. Forming the vasculature, endothelial cells (ECs) are a prominent cell type in the microenvironment of both normal and neoplastic breast gland. Here, we sought out to analyze epithelial–endothelial cross talk in the breast using isogenic non-tumorigenic vs. tumorigenic breast epithelial cell lines and primary ECs. The cellular model used here consists of D492, a breast epithelial cell line with stem cell properties, and two isogenic D492-derived EMT cell lines, D492M and D492HER2. D492M was generated by endothelial-induced EMT and is non-tumorigenic while D492HER2 is tumorigenic, expressing the ErbB2/HER2 oncogene. To investigate cellular cross talk, we used both conditioned medium (CM) and 2D/3D co-culture systems. Secretome analysis of D492 cell lines was performed using mass spectrometry and candidate knockdown (KD), and overexpression (OE) was done using siRNA and CRISPRi/CRISPRa technology. D492HER2 directly enhances endothelial network formation and activates a molecular axis in ECs promoting D492HER2 migration and invasion, suggesting an endothelial feedback response. Secretome analysis identified extracellular matrix protein 1 (ECM1) as potential angiogenic inducer in D492HER2. Confirming its involvement, KD of ECM1 reduced the ability of D492HER2-CM to increase endothelial network formation and induce the endothelial feedback, while recombinant ECM1 (rECM1) increased both. Interestingly, NOTCH1 and NOTCH3 expression was upregulated in ECs upon treatment with D492HER2-CM or rECM1 but not by CM from D492HER2 with ECM1 KD. Blocking endothelial NOTCH signaling inhibited the increase in network formation and the ability of ECs to promote D492HER2 migration and invasion. In summary, our data demonstrate that cancer-secreted ECM1 induces a NOTCH-mediated endothelial feedback promoting cancer progression by enhancing migration and invasion. Targeting this interaction may provide a novel possibility to improve cancer treatment.",

keywords = "Brj{\'o}stakrabbamein",

author = "Steinhaeuser, \{Sophie Sarah\} and Erika Morera and Zuzana Budkova and Alexander Schepsky and Qiong Wang and {\'O}ttar Rolfsson and Angela Riedel and Aileen Krueger and Bylgja Hilmarsd{\'o}ttir and Maelandsmo, \{Gunhild Mari\} and Bryndis Valdimarsdottir and Sigur{\dh}ard{\'o}ttir, \{Anna Karen\} and Agnarsson, \{Bjarni Agnar\} and J{\'o}nasson, \{J{\'o}n Gunnlaugur\} and S{\ae}var Ing{\th}{\'o}rsson and Traustad{\'o}ttir, \{Gunnhildur {\'A}sta\} and Thordur Oskarsson and Thorarinn Gudjonsson",

note = "Funding Information: Acknowledgements We thank the FingerPrints Proteomics Facility at the University of Dundee for performing the mass spectrometry and the Center for Systems Biology at the University of Iceland for the technical support during mass spectrometry data analysis. Further, we want to thank the HI-STEM laboratory and the sequencing core facility at the German Cancer Research Center, DKFZ Heidelberg, Germany, for technical support and performing the RNA microarray. Finally, we thank Prof. Dr Haraldur Haraldsson, University of Iceland for providing primary human umbilical vascular ECs. Human umbilical vascular ECs were obtained from umbilical chords at the child-birth department, University Hospital Iceland (ethical application nr. 35/2013). This work was supported by Grants from Landspitali University Hospital Science Fund, University of Iceland Research Fund (Grant of Excellence: \#152144051 Doctoral Grant: \#185042051), and Icelandic Science and Technology Policy. Publisher Copyright: {\textcopyright} 2020, The Author(s), under exclusive licence to United States and Canadian Academy of Pathology.",

year = "2020",

month = jul,

day = "1",

doi = "10.1038/s41374-020-0415-6",

language = "English",

volume = "100",

pages = "928--944",

journal = "Laboratory Investigation",

issn = "0023-6837",

publisher = "Elsevier B.V.",

number = "7",

}

Steinhaeuser, SS, Morera, E, Budkova, Z, Schepsky, A, Wang, Q , Rolfsson, Ó, Riedel, A, Krueger, A, Hilmarsdóttir, B, Maelandsmo, GM, Valdimarsdottir, B, Sigurðardóttir, AK, Agnarsson, BA, Jónasson, JG , Ingþórsson, S , Traustadóttir, GÁ, Oskarsson, T & Gudjonsson, T 2020, 'ECM1 secreted by HER2-overexpressing breast cancer cells promotes formation of a vascular niche accelerating cancer cell migration and invasion', Laboratory Investigation, vol. 100, no. 7, pp. 928-944. https://doi.org/10.1038/s41374-020-0415-6

TY - JOUR

T1 - ECM1 secreted by HER2-overexpressing breast cancer cells promotes formation of a vascular niche accelerating cancer cell migration and invasion

AU - Steinhaeuser, Sophie Sarah

AU - Morera, Erika

AU - Budkova, Zuzana

AU - Schepsky, Alexander

AU - Wang, Qiong

AU - Rolfsson, Óttar

AU - Riedel, Angela

AU - Krueger, Aileen

AU - Hilmarsdóttir, Bylgja

AU - Maelandsmo, Gunhild Mari

AU - Valdimarsdottir, Bryndis

AU - Sigurðardóttir, Anna Karen

AU - Agnarsson, Bjarni Agnar

AU - Jónasson, Jón Gunnlaugur

AU - Ingþórsson, Sævar

AU - Traustadóttir, Gunnhildur Ásta

AU - Oskarsson, Thordur

AU - Gudjonsson, Thorarinn

N1 - Funding Information: Acknowledgements We thank the FingerPrints Proteomics Facility at the University of Dundee for performing the mass spectrometry and the Center for Systems Biology at the University of Iceland for the technical support during mass spectrometry data analysis. Further, we want to thank the HI-STEM laboratory and the sequencing core facility at the German Cancer Research Center, DKFZ Heidelberg, Germany, for technical support and performing the RNA microarray. Finally, we thank Prof. Dr Haraldur Haraldsson, University of Iceland for providing primary human umbilical vascular ECs. Human umbilical vascular ECs were obtained from umbilical chords at the child-birth department, University Hospital Iceland (ethical application nr. 35/2013). This work was supported by Grants from Landspitali University Hospital Science Fund, University of Iceland Research Fund (Grant of Excellence: #152144051 Doctoral Grant: #185042051), and Icelandic Science and Technology Policy. Publisher Copyright: © 2020, The Author(s), under exclusive licence to United States and Canadian Academy of Pathology.

PY - 2020/7/1

Y1 - 2020/7/1

N2 - The tumor microenvironment is increasingly recognized as key player in cancer progression. Investigating heterotypic interactions between cancer cells and their microenvironment is important for understanding how specific cell types support cancer. Forming the vasculature, endothelial cells (ECs) are a prominent cell type in the microenvironment of both normal and neoplastic breast gland. Here, we sought out to analyze epithelial–endothelial cross talk in the breast using isogenic non-tumorigenic vs. tumorigenic breast epithelial cell lines and primary ECs. The cellular model used here consists of D492, a breast epithelial cell line with stem cell properties, and two isogenic D492-derived EMT cell lines, D492M and D492HER2. D492M was generated by endothelial-induced EMT and is non-tumorigenic while D492HER2 is tumorigenic, expressing the ErbB2/HER2 oncogene. To investigate cellular cross talk, we used both conditioned medium (CM) and 2D/3D co-culture systems. Secretome analysis of D492 cell lines was performed using mass spectrometry and candidate knockdown (KD), and overexpression (OE) was done using siRNA and CRISPRi/CRISPRa technology. D492HER2 directly enhances endothelial network formation and activates a molecular axis in ECs promoting D492HER2 migration and invasion, suggesting an endothelial feedback response. Secretome analysis identified extracellular matrix protein 1 (ECM1) as potential angiogenic inducer in D492HER2. Confirming its involvement, KD of ECM1 reduced the ability of D492HER2-CM to increase endothelial network formation and induce the endothelial feedback, while recombinant ECM1 (rECM1) increased both. Interestingly, NOTCH1 and NOTCH3 expression was upregulated in ECs upon treatment with D492HER2-CM or rECM1 but not by CM from D492HER2 with ECM1 KD. Blocking endothelial NOTCH signaling inhibited the increase in network formation and the ability of ECs to promote D492HER2 migration and invasion. In summary, our data demonstrate that cancer-secreted ECM1 induces a NOTCH-mediated endothelial feedback promoting cancer progression by enhancing migration and invasion. Targeting this interaction may provide a novel possibility to improve cancer treatment.

AB - The tumor microenvironment is increasingly recognized as key player in cancer progression. Investigating heterotypic interactions between cancer cells and their microenvironment is important for understanding how specific cell types support cancer. Forming the vasculature, endothelial cells (ECs) are a prominent cell type in the microenvironment of both normal and neoplastic breast gland. Here, we sought out to analyze epithelial–endothelial cross talk in the breast using isogenic non-tumorigenic vs. tumorigenic breast epithelial cell lines and primary ECs. The cellular model used here consists of D492, a breast epithelial cell line with stem cell properties, and two isogenic D492-derived EMT cell lines, D492M and D492HER2. D492M was generated by endothelial-induced EMT and is non-tumorigenic while D492HER2 is tumorigenic, expressing the ErbB2/HER2 oncogene. To investigate cellular cross talk, we used both conditioned medium (CM) and 2D/3D co-culture systems. Secretome analysis of D492 cell lines was performed using mass spectrometry and candidate knockdown (KD), and overexpression (OE) was done using siRNA and CRISPRi/CRISPRa technology. D492HER2 directly enhances endothelial network formation and activates a molecular axis in ECs promoting D492HER2 migration and invasion, suggesting an endothelial feedback response. Secretome analysis identified extracellular matrix protein 1 (ECM1) as potential angiogenic inducer in D492HER2. Confirming its involvement, KD of ECM1 reduced the ability of D492HER2-CM to increase endothelial network formation and induce the endothelial feedback, while recombinant ECM1 (rECM1) increased both. Interestingly, NOTCH1 and NOTCH3 expression was upregulated in ECs upon treatment with D492HER2-CM or rECM1 but not by CM from D492HER2 with ECM1 KD. Blocking endothelial NOTCH signaling inhibited the increase in network formation and the ability of ECs to promote D492HER2 migration and invasion. In summary, our data demonstrate that cancer-secreted ECM1 induces a NOTCH-mediated endothelial feedback promoting cancer progression by enhancing migration and invasion. Targeting this interaction may provide a novel possibility to improve cancer treatment.

KW - Brjóstakrabbamein

UR - https://www.scopus.com/pages/publications/85082834537

U2 - 10.1038/s41374-020-0415-6

DO - 10.1038/s41374-020-0415-6

M3 - Article

C2 - 32203150

SN - 0023-6837

VL - 100

SP - 928

EP - 944

JO - Laboratory Investigation

JF - Laboratory Investigation

IS - 7

ER -

ECM1 secreted by HER2-overexpressing breast cancer cells promotes formation of a vascular niche accelerating cancer cell migration and invasion

Abstract

Bibliographical note

UN SDGs

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