Breast Cancer Research and Treatment

, Volume 132, Issue 2, pp 487–498 | Cite as

Pilot and feasibility study: prospective proteomic profiling of mammary epithelial cells from high-risk women provides evidence of activation of pro-survival pathways

  • Catherine Ibarra-DrendallEmail author
  • Michelle M. Troch
  • William T. Barry
  • Gloria Broadwater
  • Emanuel F. PetricoinIII
  • Julia Wulfkuhle
  • Lance A. Liotta
  • Siya Lem
  • Joseph C. BakerJr.
  • Anne C. Ford
  • Lee G. Wilke
  • Carola Zalles
  • Nicole M. Kuderer
  • Abigail W. Hoffman
  • Melanie Shivraj
  • Priya Mehta
  • Jamila Williams
  • Nora Tolbert
  • Laurie W. Lee
  • Patrick G. Pilie
  • Dihua Yu
  • Victoria L. Seewaldt
Preclinical study


Normal mammary gland homeostasis requires the coordinated regulation of protein signaling networks. However, we have little prospective information on whether activation of protein signaling occurs in premalignant mammary epithelial cells, as represented by cells with cytological atypia from women who are at high risk for breast cancer. This information is critical for understanding the role of deregulated signaling pathways in the initiation of breast cancer and for developing targeted prevention and/or treatment strategies for breast cancer in the future. In this pilot and feasibility study, we examined the expression of 52 phosphorylated, total, and cleaved proteins in 31 microdissected Random Periareolar Fine Needle Aspiration (RPFNA) samples by high-throughput Reverse Phase Protein Microarray. Unsupervised hierarchical clustering analysis indicated the presence of four clusters of proteins that represent the following signaling pathways: (1) receptor tyrosine kinase/Akt/mammalian target of rapamycin (RTK/Akt/mTOR), (2) RTK/Akt/extracellular signal-regulated kinase (RTK/Akt/ERK), (3) mitochondrial apoptosis, and (4) indeterminate. Clusters 1 through 3 comprised moderately to highly expressed proteins, while Cluster 4 comprised proteins that are lowly expressed in a majority of RPFNA samples. Our exploratory study showed that the interlinked components of mitochondrial apoptosis pathway are highly expressed in all mammary epithelial cells obtained from high-risk women. In particular, the expression levels of anti-apoptotic Bcl-xL and pro-apoptotic Bad are positively correlated in both non-atypical and atypical samples (unadjusted P < 0.0001), suggesting a delicate balance between the pro-apoptotic and anti-apoptotic regulation of cell proliferation during the early steps of mammary carcinogenesis. Our feasibility study suggests that the activation of key proteins along the RTK/Akt pathway may tip this balance to cell survival. Taken together, our results demonstrate the feasibility of mapping proteomic signaling networks in limited RPFNA samples obtained from high-risk women and the promise of developing rational drug targets or preventative strategies for breast cancer in future proteomic studies with a larger cohort of high-risk women.


Breast cancer risk Atypical mammary cytology Protein microarray Biomarker development Cell survival 



Apoptotic protease activating factor-1


Bcl2-antagonist of cell death


Bcl2 associated X-protein


B-cell lymphoma-2


Bcl2 interacting protein


Breast cancer-associated gene 1/2


Casein kinase II or 2


Ductal carcinoma in situ


Epidermal growth factor receptor


Estrogen receptor


Erythroblastic leukemia viral oncogene homolog


Extracellular signal-regulated kinase


Fas-associated via death domain




Insulin-like growth factor receptor


Lobular carcinoma in situ


Laser capture microdissection


Mammalian target of rapamycin


Operating room


Phosphatidylinositol 3-kinase


Phosphatase and tensin homolog


Random Periareolar Fine Needle Aspiration


Reverse phase protein microarray


Receptor tyrosine kinase



The authors thank Susan G. Komen for the Cure foundation for financial support to Drs. Yu and Seewaldt (KG091020) and Dr. Ibarra-Drendall (KG090730). Dr. Seewaldt is also supported by the National Institute of Health/National Cancer Institute (R01CA88799 and R01CA114068). Additional financial support was provided by NCI/AVON Partners in Progress Grant 3 P30 CA014236-32S1 to Drs. Seewaldt and Yu.


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Copyright information

© Springer Science+Business Media, LLC. 2011

Authors and Affiliations

  • Catherine Ibarra-Drendall
    • 1
    Email author
  • Michelle M. Troch
    • 1
  • William T. Barry
    • 2
  • Gloria Broadwater
    • 2
  • Emanuel F. PetricoinIII
    • 3
  • Julia Wulfkuhle
    • 3
  • Lance A. Liotta
    • 3
  • Siya Lem
    • 1
  • Joseph C. BakerJr.
    • 1
  • Anne C. Ford
    • 4
  • Lee G. Wilke
    • 5
  • Carola Zalles
    • 6
  • Nicole M. Kuderer
    • 1
  • Abigail W. Hoffman
    • 1
  • Melanie Shivraj
    • 1
  • Priya Mehta
    • 1
  • Jamila Williams
    • 1
  • Nora Tolbert
    • 1
  • Laurie W. Lee
    • 1
  • Patrick G. Pilie
    • 1
  • Dihua Yu
    • 7
  • Victoria L. Seewaldt
    • 1
  1. 1.Division of Medical OncologyDuke University Medical CenterDurhamUSA
  2. 2.Department of Biostatistics & BioinformaticsDuke University Medical CenterDurhamUSA
  3. 3.Center for Applied Proteomics and Molecular MedicineGeorge Mason UniversityManassasUSA
  4. 4.Department of Obstetrics and GynecologyDuke University Medical CenterDurhamUSA
  5. 5.Division of General SurgeryUniversity of WisconsinMadisonUSA
  6. 6.Texas A&M Health Sciences CenterCollege of MedicineRound RockUSA
  7. 7.Department of Molecular and Cellular OncologyMD Anderson Cancer CenterHoustonUSA

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