Characterizing the heterogeneity of triple-negative breast cancers using microdissected normal ductal epithelium and RNA-sequencing

Milan Radovich; Susan E. Clare; Rutuja Atale; Ivanesa Pardo; Bradley A. Hancock; Jeffrey P. Solzak; Nawal Kassem; Theresa Mathieson; Anna Maria V Storniolo; Connie Rufenbarger; Heather A. Lillemoe; Rachel J. Blosser; Mi Ran Choi; Candice A. Sauder; Diane Doxey; Jill E. Henry; Eric E. Hilligoss; Onur Sakarya; Fiona C. Hyland; Matthew Hickenbotham; Jin Zhu; Jarret Glasscock; Sunil Badve; Mircea Ivan; Yunlong Liu; George W. Sledge; Bryan P. Schneider

doi:10.1007/s10549-013-2780-y

Characterizing the heterogeneity of triple-negative breast cancers using microdissected normal ductal epithelium and RNA-sequencing

Milan Radovich^*, Susan E. Clare, Rutuja Atale, Ivanesa Pardo, Bradley A. Hancock, Jeffrey P. Solzak, Nawal Kassem, Theresa Mathieson, Anna Maria V Storniolo, Connie Rufenbarger, Heather A. Lillemoe, Rachel J. Blosser, Mi Ran Choi, Candice A. Sauder, Diane Doxey, Jill E. Henry, Eric E. Hilligoss, Onur Sakarya, Fiona C. Hyland, Matthew HickenbothamJin Zhu, Jarret Glasscock, Sunil Badve, Mircea Ivan, Yunlong Liu, George W. Sledge, Bryan P. Schneider

^*Corresponding author for this work

Surgery, Breast Surgery Division

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

27 Scopus citations

Abstract

Triple-negative breast cancers (TNBCs) are a heterogeneous set of tumors defined by an absence of actionable therapeutic targets (ER, PR, and HER-2). Microdissected normal ductal epithelium from healthy volunteers represents a novel comparator to reveal insights into TNBC heterogeneity and to inform drug development. Using RNA-sequencing data from our institution and The Cancer Genome Atlas (TCGA) we compared the transcriptomes of 94 TNBCs, 20 microdissected normal breast tissues from healthy volunteers from the Susan G. Komen for the Cure Tissue Bank, and 10 histologically normal tissues adjacent to tumor. Pathway analysis comparing TNBCs to optimized normal controls of microdissected normal epithelium versus classic controls composed of adjacent normal tissue revealed distinct molecular signatures. Differential gene expression of TNBC compared with normal comparators demonstrated important findings for TNBC-specific clinical trials testing targeted agents; lack of over-expression for negative studies and over-expression in studies with drug activity. Next, by comparing each individual TNBC to the set of microdissected normals, we demonstrate that TNBC heterogeneity is attributable to transcriptional chaos, is associated with non-silent DNA mutational load, and explains transcriptional heterogeneity in addition to known molecular subtypes. Finally, chaos analysis identified 146 core genes dysregulated in >90 % of TNBCs revealing an over-expressed central network. In conclusion, use of microdissected normal ductal epithelium from healthy volunteers enables an optimized approach for studying TNBC and uncovers biological heterogeneity mediated by transcriptional chaos.

Original language	English (US)
Pages (from-to)	57-68
Number of pages	12
Journal	Breast Cancer Research and Treatment
Volume	143
Issue number	1
DOIs	https://doi.org/10.1007/s10549-013-2780-y
State	Published - Jan 2014

Funding

Acknowledgments We would like to thank Mark Mooney, James Elliott, Darryl Leon, and Ryan Richt for discussions of next-generation sequencing and analysis. We also like to thank Benjamin Haibe-Kains for assistance with PAM50 analysis and to thank Carla Bullitt, Stuart Tugendreich, Gordon Janaway, and Bryant Macy for assistance with Ingenuity Pathway Analysis. We also like to thank the IUSCC Tissue Procurement and Distribution Core for providing tissues for IHC and qPCR validations. Finally, we would like to thank and acknowledge the TCGA for the sample procurement, production of the TNBC RNA-seq data, and the clinical annotation of TCGA samples used in this publication. This work was supported by the Susan G. Komen for the Cure® (S.E.C., G.W.S., A.V.S., S.B., B.P.S.), Breast Cancer Research Foundation (S.E.C., A.V.S.) and the Catherine Peachey Fund (M.R., S.E.C., G.W.S., A.V.S., C.R., B.P.S.).. M.R. was supported by pre-doctoral fellowships from the National Institutes of Health, NRSA 1T32CA111198 Cancer Biology Training Program and 5TL1RR025759 Indiana Clinical and Translational Sciences Institute Career Development Award.

Keywords

Adjacent normal
Ductal epithelium
Normal breast
RNA-seq
TCGA
Triple-negative breast cancer

ASJC Scopus subject areas

Oncology
Cancer Research

UN SDGs

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

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10.1007/s10549-013-2780-y

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Radovich, M., Clare, S. E., Atale, R., Pardo, I., Hancock, B. A., Solzak, J. P., Kassem, N., Mathieson, T., Storniolo, A. M. V., Rufenbarger, C., Lillemoe, H. A., Blosser, R. J., Choi, M. R., Sauder, C. A., Doxey, D., Henry, J. E., Hilligoss, E. E., Sakarya, O., Hyland, F. C., ... Schneider, B. P. (2014). Characterizing the heterogeneity of triple-negative breast cancers using microdissected normal ductal epithelium and RNA-sequencing. Breast Cancer Research and Treatment, 143(1), 57-68. https://doi.org/10.1007/s10549-013-2780-y

@article{d0bb223d2daa440e8e22a893eda0448f,

title = "Characterizing the heterogeneity of triple-negative breast cancers using microdissected normal ductal epithelium and RNA-sequencing",

abstract = "Triple-negative breast cancers (TNBCs) are a heterogeneous set of tumors defined by an absence of actionable therapeutic targets (ER, PR, and HER-2). Microdissected normal ductal epithelium from healthy volunteers represents a novel comparator to reveal insights into TNBC heterogeneity and to inform drug development. Using RNA-sequencing data from our institution and The Cancer Genome Atlas (TCGA) we compared the transcriptomes of 94 TNBCs, 20 microdissected normal breast tissues from healthy volunteers from the Susan G. Komen for the Cure Tissue Bank, and 10 histologically normal tissues adjacent to tumor. Pathway analysis comparing TNBCs to optimized normal controls of microdissected normal epithelium versus classic controls composed of adjacent normal tissue revealed distinct molecular signatures. Differential gene expression of TNBC compared with normal comparators demonstrated important findings for TNBC-specific clinical trials testing targeted agents; lack of over-expression for negative studies and over-expression in studies with drug activity. Next, by comparing each individual TNBC to the set of microdissected normals, we demonstrate that TNBC heterogeneity is attributable to transcriptional chaos, is associated with non-silent DNA mutational load, and explains transcriptional heterogeneity in addition to known molecular subtypes. Finally, chaos analysis identified 146 core genes dysregulated in >90 % of TNBCs revealing an over-expressed central network. In conclusion, use of microdissected normal ductal epithelium from healthy volunteers enables an optimized approach for studying TNBC and uncovers biological heterogeneity mediated by transcriptional chaos.",

keywords = "Adjacent normal, Ductal epithelium, Normal breast, RNA-seq, TCGA, Triple-negative breast cancer",

author = "Milan Radovich and Clare, {Susan E.} and Rutuja Atale and Ivanesa Pardo and Hancock, {Bradley A.} and Solzak, {Jeffrey P.} and Nawal Kassem and Theresa Mathieson and Storniolo, {Anna Maria V} and Connie Rufenbarger and Lillemoe, {Heather A.} and Blosser, {Rachel J.} and Choi, {Mi Ran} and Sauder, {Candice A.} and Diane Doxey and Henry, {Jill E.} and Hilligoss, {Eric E.} and Onur Sakarya and Hyland, {Fiona C.} and Matthew Hickenbotham and Jin Zhu and Jarret Glasscock and Sunil Badve and Mircea Ivan and Yunlong Liu and Sledge, {George W.} and Schneider, {Bryan P.}",

note = "Funding Information: Acknowledgments We would like to thank Mark Mooney, James Elliott, Darryl Leon, and Ryan Richt for discussions of next-generation sequencing and analysis. We also like to thank Benjamin Haibe-Kains for assistance with PAM50 analysis and to thank Carla Bullitt, Stuart Tugendreich, Gordon Janaway, and Bryant Macy for assistance with Ingenuity Pathway Analysis. We also like to thank the IUSCC Tissue Procurement and Distribution Core for providing tissues for IHC and qPCR validations. Finally, we would like to thank and acknowledge the TCGA for the sample procurement, production of the TNBC RNA-seq data, and the clinical annotation of TCGA samples used in this publication. This work was supported by the Susan G. Komen for the Cure{\textregistered} (S.E.C., G.W.S., A.V.S., S.B., B.P.S.), Breast Cancer Research Foundation (S.E.C., A.V.S.) and the Catherine Peachey Fund (M.R., S.E.C., G.W.S., A.V.S., C.R., B.P.S.).. M.R. was supported by pre-doctoral fellowships from the National Institutes of Health, NRSA 1T32CA111198 Cancer Biology Training Program and 5TL1RR025759 Indiana Clinical and Translational Sciences Institute Career Development Award.",

year = "2014",

month = jan,

doi = "10.1007/s10549-013-2780-y",

language = "English (US)",

volume = "143",

pages = "57--68",

journal = "Breast Cancer Research and Treatment",

issn = "0167-6806",

publisher = "Springer",

number = "1",

}

Radovich, M, Clare, SE, Atale, R, Pardo, I, Hancock, BA, Solzak, JP, Kassem, N, Mathieson, T, Storniolo, AMV, Rufenbarger, C, Lillemoe, HA, Blosser, RJ, Choi, MR, Sauder, CA, Doxey, D, Henry, JE, Hilligoss, EE, Sakarya, O, Hyland, FC, Hickenbotham, M, Zhu, J, Glasscock, J, Badve, S, Ivan, M, Liu, Y, Sledge, GW & Schneider, BP 2014, 'Characterizing the heterogeneity of triple-negative breast cancers using microdissected normal ductal epithelium and RNA-sequencing', Breast Cancer Research and Treatment, vol. 143, no. 1, pp. 57-68. https://doi.org/10.1007/s10549-013-2780-y

TY - JOUR

T1 - Characterizing the heterogeneity of triple-negative breast cancers using microdissected normal ductal epithelium and RNA-sequencing

AU - Radovich, Milan

AU - Clare, Susan E.

AU - Atale, Rutuja

AU - Pardo, Ivanesa

AU - Hancock, Bradley A.

AU - Solzak, Jeffrey P.

AU - Kassem, Nawal

AU - Mathieson, Theresa

AU - Storniolo, Anna Maria V

AU - Rufenbarger, Connie

AU - Lillemoe, Heather A.

AU - Blosser, Rachel J.

AU - Choi, Mi Ran

AU - Sauder, Candice A.

AU - Doxey, Diane

AU - Henry, Jill E.

AU - Hilligoss, Eric E.

AU - Sakarya, Onur

AU - Hyland, Fiona C.

AU - Hickenbotham, Matthew

AU - Zhu, Jin

AU - Glasscock, Jarret

AU - Badve, Sunil

AU - Ivan, Mircea

AU - Liu, Yunlong

AU - Sledge, George W.

AU - Schneider, Bryan P.

N1 - Funding Information: Acknowledgments We would like to thank Mark Mooney, James Elliott, Darryl Leon, and Ryan Richt for discussions of next-generation sequencing and analysis. We also like to thank Benjamin Haibe-Kains for assistance with PAM50 analysis and to thank Carla Bullitt, Stuart Tugendreich, Gordon Janaway, and Bryant Macy for assistance with Ingenuity Pathway Analysis. We also like to thank the IUSCC Tissue Procurement and Distribution Core for providing tissues for IHC and qPCR validations. Finally, we would like to thank and acknowledge the TCGA for the sample procurement, production of the TNBC RNA-seq data, and the clinical annotation of TCGA samples used in this publication. This work was supported by the Susan G. Komen for the Cure® (S.E.C., G.W.S., A.V.S., S.B., B.P.S.), Breast Cancer Research Foundation (S.E.C., A.V.S.) and the Catherine Peachey Fund (M.R., S.E.C., G.W.S., A.V.S., C.R., B.P.S.).. M.R. was supported by pre-doctoral fellowships from the National Institutes of Health, NRSA 1T32CA111198 Cancer Biology Training Program and 5TL1RR025759 Indiana Clinical and Translational Sciences Institute Career Development Award.

PY - 2014/1

Y1 - 2014/1

N2 - Triple-negative breast cancers (TNBCs) are a heterogeneous set of tumors defined by an absence of actionable therapeutic targets (ER, PR, and HER-2). Microdissected normal ductal epithelium from healthy volunteers represents a novel comparator to reveal insights into TNBC heterogeneity and to inform drug development. Using RNA-sequencing data from our institution and The Cancer Genome Atlas (TCGA) we compared the transcriptomes of 94 TNBCs, 20 microdissected normal breast tissues from healthy volunteers from the Susan G. Komen for the Cure Tissue Bank, and 10 histologically normal tissues adjacent to tumor. Pathway analysis comparing TNBCs to optimized normal controls of microdissected normal epithelium versus classic controls composed of adjacent normal tissue revealed distinct molecular signatures. Differential gene expression of TNBC compared with normal comparators demonstrated important findings for TNBC-specific clinical trials testing targeted agents; lack of over-expression for negative studies and over-expression in studies with drug activity. Next, by comparing each individual TNBC to the set of microdissected normals, we demonstrate that TNBC heterogeneity is attributable to transcriptional chaos, is associated with non-silent DNA mutational load, and explains transcriptional heterogeneity in addition to known molecular subtypes. Finally, chaos analysis identified 146 core genes dysregulated in >90 % of TNBCs revealing an over-expressed central network. In conclusion, use of microdissected normal ductal epithelium from healthy volunteers enables an optimized approach for studying TNBC and uncovers biological heterogeneity mediated by transcriptional chaos.

AB - Triple-negative breast cancers (TNBCs) are a heterogeneous set of tumors defined by an absence of actionable therapeutic targets (ER, PR, and HER-2). Microdissected normal ductal epithelium from healthy volunteers represents a novel comparator to reveal insights into TNBC heterogeneity and to inform drug development. Using RNA-sequencing data from our institution and The Cancer Genome Atlas (TCGA) we compared the transcriptomes of 94 TNBCs, 20 microdissected normal breast tissues from healthy volunteers from the Susan G. Komen for the Cure Tissue Bank, and 10 histologically normal tissues adjacent to tumor. Pathway analysis comparing TNBCs to optimized normal controls of microdissected normal epithelium versus classic controls composed of adjacent normal tissue revealed distinct molecular signatures. Differential gene expression of TNBC compared with normal comparators demonstrated important findings for TNBC-specific clinical trials testing targeted agents; lack of over-expression for negative studies and over-expression in studies with drug activity. Next, by comparing each individual TNBC to the set of microdissected normals, we demonstrate that TNBC heterogeneity is attributable to transcriptional chaos, is associated with non-silent DNA mutational load, and explains transcriptional heterogeneity in addition to known molecular subtypes. Finally, chaos analysis identified 146 core genes dysregulated in >90 % of TNBCs revealing an over-expressed central network. In conclusion, use of microdissected normal ductal epithelium from healthy volunteers enables an optimized approach for studying TNBC and uncovers biological heterogeneity mediated by transcriptional chaos.

KW - Adjacent normal

KW - Ductal epithelium

KW - Normal breast

KW - RNA-seq

KW - TCGA

KW - Triple-negative breast cancer

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DO - 10.1007/s10549-013-2780-y

M3 - Article

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AN - SCOPUS:84892799530

SN - 0167-6806

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SP - 57

EP - 68

JO - Breast Cancer Research and Treatment

JF - Breast Cancer Research and Treatment

IS - 1

ER -

Characterizing the heterogeneity of triple-negative breast cancers using microdissected normal ductal epithelium and RNA-sequencing

Abstract

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Keywords

ASJC Scopus subject areas

UN SDGs

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