An increase in the mammographic density of breast tissue is correlated to a four to six-fold increased risk of developing breast carcinoma, making it the single biggest risk factor for breast carcinoma. Despite this, the molecular mechanism by which breast density links to carcinoma formation is unknown. We have recently shown that in a mouse model of increased collagen deposition, mammary tumor incidence, invasion, and metastasis increase three-fold, suggesting that an increase in collagen per se is part of the mechanism by which increased breast density is correlated with increased risk in humans. The purpose of this proposal is to understand how physical properties of the ECM regulate breast cell behavior. Our hypothesis is that locally dense ECM enhances the formation of matrix adhesions that result in activation of signaling pathways linked to FAK, and that FAK is a central mediator by which matrix density regulates gene expression to promote proliferation and invasion. This hypothesis will be tested in the following Specific Aims:
Aim 1 : Define changes in the expression of proliferation and metastasis-associated genes regulated by collagen density and alignment. Gene expression of the proliferation and the metastasis gene signatures, will be determined in human breast carcinoma samples, using tissue arrays linked to patient outcome, and with DCIS and microinvasive DCIS. Metastasis genes will be screened by siRNA and shRNA knock-down for their role in mediating invasion into 3D matrices in vitro and in vivo.
Aim 2 : Test the hypothesis that FAK regulates proliferation and invasion in response to collagen density and alignment Tumors that are FAK-/- do not invade or metastasize in vivo, even in the context of a FAK+/+ stroma. We will use in vitro 3D invasion assays and our in vivo FAK-/- mice to investigate the role of FAK in cell proliferation, matrix alignment and 3D invasion. Conditional FAK knockout mice will be used to determine the role of FAK in enhanced tumor progression in the collagenase-resistant transgenic mouse model (Col1a1tm1Jae) having a dense collagen matrix. Phosphorylation of FAK at pY397 will be determined in mammary tissues from Col1a1 mice, mouse tumor, and human pathologic samples to determine if pY397FAK correlates to dense breast tissue in vivo.
Aim 3 : Test the link between collagen density, FAK and downstream signaling events, ERK, Src, and PI3K, in regulating cell proliferation and invasion. FAK-/- cells lose activation of ERK and PI3K pathways. Using pharmacologic inhibitors and siRNA/shRNA approaches, we will inhibit each of these pathways and determine their role in mediating the proliferation and invasion of human breast cells into dense and aligned collagen matrices in vitro. Xenografts of human carcinoma, and mice bearing tumors in wt and dense collagen stroma (Col1a1 mouse model) will be treated in vivo with pharmacologic inhibitors to test the role of these molecules in tumor progression in vivo.

Public Health Relevance

Understanding the role the physical properties of the extracellular matrix plays in cancer progression is of great health relevance as breast density accounts for a 4-6 fold increase in carcinoma risk. Moreover, we have found that collagen alignment carries a 5 fold risk of disease relapse, suggesting that understanding how collagen alignment occurs, and how it contributes to progression will help us understand breast carcinoma. These experiments are designed to understand the underlying molecular mechanisms by which the dense extracellular matrix regulates breast cell behavior and invasion and progression, and could suggest future targets for therapy.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
1R01CA114462-01A2
Application #
7782617
Study Section
Tumor Progression and Metastasis Study Section (TPM)
Program Officer
Woodhouse, Elizabeth
Project Start
2010-01-01
Project End
2014-12-31
Budget Start
2010-01-01
Budget End
2010-12-31
Support Year
1
Fiscal Year
2010
Total Cost
$304,917
Indirect Cost
Name
University of Wisconsin Madison
Department
Biochemistry
Type
Other Domestic Higher Education
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715
Le, Lily Thao-Nhi; Cazares, Oscar; Mouw, Janna K et al. (2016) Loss of miR-203 regulates cell shape and matrix adhesion through ROBO1/Rac/FAK in response to stiffness. J Cell Biol 212:707-19
García-Mendoza, María G; Inman, David R; Ponik, Suzanne M et al. (2016) Neutrophils drive accelerated tumor progression in the collagen-dense mammary tumor microenvironment. Breast Cancer Res 18:49
Oudin, Madeleine J; Jonas, Oliver; Kosciuk, Tatsiana et al. (2016) Tumor Cell-Driven Extracellular Matrix Remodeling Drives Haptotaxis during Metastatic Progression. Cancer Discov 6:516-31
Burkel, Brian; Morris, Brett A; Ponik, Suzanne M et al. (2016) Preparation of 3D Collagen Gels and Microchannels for the Study of 3D Interactions In Vivo. J Vis Exp :
Miller, Cassandra L; Muthupalani, Sureshkumar; Shen, Zeli et al. (2016) Lamellipodin-Deficient Mice: A Model of Rectal Carcinoma. PLoS One 11:e0152940
Szulczewski, Joseph M; Inman, David R; Entenberg, David et al. (2016) In Vivo Visualization of Stromal Macrophages via label-free FLIM-based metabolite imaging. Sci Rep 6:25086
Tung, Jason C; Barnes, J Matthew; Desai, Shraddha R et al. (2015) Tumor mechanics and metabolic dysfunction. Free Radic Biol Med 79:269-80
Barcus, Craig E; Holt, Elizabeth C; Keely, Patricia J et al. (2015) Dense collagen-I matrices enhance pro-tumorigenic estrogen-prolactin crosstalk in MCF-7 and T47D breast cancer cells. PLoS One 10:e0116891
Curran, Colleen S; Carrillo, Esteban R; Ponik, Suzanne M et al. (2015) Collagen density regulates xenobiotic and hypoxic response of mammary epithelial cells. Environ Toxicol Pharmacol 39:114-24
Riching, Kristin M; Keely, Patricia J (2015) Rho family GTPases: making it to the third dimension. Int J Biochem Cell Biol 59:111-5

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