Breast cancer is a heterogeneous disease comprised of at least 5 major tumor subtypes that coalesce as the second leading cause of cancer death in women in the United States. Amongst individual breast cancer subtypes, those classified as being triple-negative breast cancers (TNBCs) are clinically unique via their presentation of aggressive and metastatic phenotypes, and their high propensity to recur rapidly following conventional chemotherapy treatment. TNBCs are also noteworthy by their failure to express hormone receptors (estrogen and progesterone) and ErbB2/HER2, a phenotype that renders targeted chemotherapies (e.g., hormonal or HER2-directed) ineffective and contributes to the poor prognosis of TNBC patients. Although our understanding of the molecular features and clinical manifestations of TNBCs has increased in recent years, science and medicine still lack sufficient knowledge of TNBC development and progression to permit the synthesis of novel pharmaceuticals capable of specifically targeting and alleviating this lethal breast cancer subtype. Transforming growth factor-? (TGF-?) is a powerful suppressor of mammary tumorigenesis. Interestingly, late-stage TNBCs respond to TGF-? as if this cytokine were a tumor promoter, leading to the acquisition of metastatic and stem cell phenotypes. Although the molecular mechanisms underlying the conversion of TGF-? function from that of a tumor suppressor to a tumor promoter in TNBCs remains incompletely understood, our laboratory recently defined a novel integrin-based signaling module that facilitates oncogenic TGF-? signaling in TNBCs. Along these lines, we find the expression and activity of lysyl oxidase (LOX) to contribute to oncogenic TGF-? signaling in part due to alterations in mechanotransduction. Based on these and other preliminary findings, we hypothesize that integrin switching underlies metastatic progression of TNBCs driven TGF-?. A corollary states that developing novel chemotherapeutics to prevent integrin switching and oncogenic TGF-? signaling will significantly improve the overall survival rates of TNBC patients. These hypotheses will be addressed by four Specific Aims.
Aim 1 will determine the role of integrins and focal adhesion complexes during initiation of the "TGF-B Paradox" by mechanotransduction. We will manipulate, both positively and negatively, the expression of B1 and B3 integrin and their effectors to gauge their function in coupling mechanotransduction to the oncogenic activities of TGF-? both in vitro and in vivo. Likewise, the ability of mechanotransduction and Smad2/3 signaling to epigenetically silence E-cadherin expression during TNBC metastasis will be assessed.
Aim 2 will determine the role of LOX family members in mediating oncogenic TGF-? signaling in TNBCs. Additionally, genetic and pharmacological inactivation of LOX family members will be undertaken to assess their function in driving TNBC metastasis stimulated by TGF-B.
Aim 3 will map the transcriptome and epigenetic events coupled to TNBC development and metastatic progression. The presence of identified epigenetic marks in patient breast cells isolated by random periareolar fine needle aspiration will be determined to assess their utility as predictive TNBC biomarkers. Lastly, Aim 4 will visualize TGF-? signaling during metastatic progression of TNBCs through the use of dual bioluminescent imaging. Simultaneous CTL1-based MRI approaches will be employed to monitor corresponding changes in tumor reactive stroma, as well as determine how conventional chemotherapies impact these metastatic events. Collectively, these studies will provide valuable information on how TGF-?, integrins, and mechanotransduction cooperate in promoting the EMT, invasion, and metastasis of TNBCs, and more importantly, on how to control these deadly processes by inactivating the oncogenic activities of TGF-?. Moreover, translating our epigenetic biomarkers to clinical diagnostic applications will enable science and medicine to significantly improve the overall survival of patients with TNBCs.

Public Health Relevance

Triple-negative breast cancers (TNBCs) are an unusually aggressive and metastatic subtype of breast cancer that also exhibits a rapid and ardent recurrence rate and, consequently, possesses a dismal prognosis for women bearing TNBC tumors. Oncogenic TGF-? signaling figures prominently in promoting the development and metastatic progression of TNBCs. In studies supported by the parent grant, we delineated two oncogenic TGF-? signaling pathways operant in promoting the acquisition of EMT and metastatic phenotypes by TNBCs. Key effector molecules that mediate these events are integrins and LOX, both of which couple mechanotransduction to the conversion of TGF-? function from that of a tumor suppressor to a tumor promoter. Along these lines, we recently created a novel TNBC model whose initiation and evolution is absolutely dependent upon TGF-? and its stimulation of EMT. Unfortunately, how these individual events coalesce to promote TNBC development and metastatic progression remains to be determined definitively. Moreover, these knowledge gaps have prevented science and medicine from inventing treatments effective in alleviating the oncogenic activities of TGF-? in TNBCs. Our proposed studies are highly innovative and medically relevant because they will (i) provide valuable information on how TNBCs develop and progress, as well as on how TGF-? cooperates with integrins and LOX family members to promote these processes;(ii) identify the epigenetic features that manifest TNBC development and progression, and validate these marks in patient breast tissue samples that range from normal to atypia to invasive carcinoma;and (iv) visualize the spatiotemporal events coupled to TGF-? and microenvironmental signaling in TNBCs, as well as how these events are impacted by administration of conventional chemotherapies. Collectively, this study will provide novel insights into how TNBCs develop and metastasize, and in doing so, will lead to improved clinical outcomes of patients bearing this deadly disease.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
2R01CA129359-06
Application #
8320506
Study Section
Tumor Progression and Metastasis Study Section (TPM)
Program Officer
Ault, Grace S
Project Start
2007-07-11
Project End
2017-06-30
Budget Start
2012-08-17
Budget End
2013-06-30
Support Year
6
Fiscal Year
2012
Total Cost
$289,351
Indirect Cost
$105,051
Name
Case Western Reserve University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
077758407
City
Cleveland
State
OH
Country
United States
Zip Code
44106
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Parvani, Jenny G; Galliher-Beckley, Amy J; Schiemann, Barbara J et al. (2013) Targeted inactivation of *1 integrin induces *3 integrin switching, which drives breast cancer metastasis by TGF-*. Mol Biol Cell 24:3449-59
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Taylor, Molly A; Davuluri, Gangarao; Parvani, Jenny G et al. (2013) Upregulated WAVE3 expression is essential for TGF-*-mediated EMT and metastasis of triple-negative breast cancer cells. Breast Cancer Res Treat 142:341-53
Wendt, M K; Schiemann, B J; Parvani, J G et al. (2013) TGF-ýý stimulates Pyk2 expression as part of an epithelial-mesenchymal transition program required for metastatic outgrowth of breast cancer. Oncogene 32:2005-15
Taylor, Molly A; Sossey-Alaoui, Khalid; Thompson, Cheryl L et al. (2013) TGF-* upregulates miR-181a expression to promote breast cancer metastasis. J Clin Invest 123:150-63
Wendt, Michael K; Tian, Maozhen; Schiemann, William P (2012) Deconstructing the mechanisms and consequences of TGF-ýý-induced EMT during cancer progression. Cell Tissue Res 347:85-101
Wendt, Michael K; Molter, Joseph; Flask, Christopher A et al. (2011) In vivo dual substrate bioluminescent imaging. J Vis Exp :

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