Novel approaches for chemoprevention of breast cancer are desirable because many risk factors associated with this disease are not easily modifiable. Moreover, some of the currently available chemopreventive options targeted against breast cancer are sub-optimal. Needless to emphasize that breast cancer continues to be a leading cause of cancer-related death in women worldwide despite tremendous advances towards targeted therapies. Ongoing objective of this research project is to develop a non-endocrine strategy for chemoprevention of breast cancer using cruciferous vegetable constituent Benzyl Isothiocyanate (BITC). Research objectives of the current funded grant were nearly fully achieved with notable publications. For example, we demonstrated that BITC administration in the diet confers significant protection against mammary cancer in a transgenic mouse model (MMTV-neu) without any signs of overt toxicity. We are extremely excited with our more recent published as well as unpublished observations underpinning novel actions of BITC potentially contributing to its chemopreventive effect, including inhibition of epithelial-mesenchymal transition (EMT) in vitro and in vivo (published) and suppression of self-renewal of breast cancer stem cells (bCSC) in vitro (unpublished observations). We also found that BITC treatment activates Notch signaling, which is a positive regulator of EMT as well as CSC self-renewal. Thus it is only logical to experimentally test whether Notch activation by BITC has negative impact on its chemopreventive response. At the same time, expression of urokinase-type plasminogen activator (uPA) and its receptor (uPAR) is significantly downregulated by BITC treatment. Because uPAR overexpression is sufficient to drive both EMT and stemness in breast cancer cells, further investigation of the role of uPA/uPAR in BITC-mediated inhibition of EMT and bCSC self-renewal is equally meritorious. Hypothesis: Research design in the present renewal application logically builds upon these exciting and novel observations to test a stimulating hypothesis that mammary cancer chemoprevention by BITC is mediated by inhibition of the uPA/uPAR system leading to suppression of EMT and self-renewal of bCSC, which may be amenable to augmentation by pharmacological suppression of Notch using a ?-secretase inhibitor.
Specific Aims : Proposed research utilizes relevant cellular and in vivo animal models of breast cancer to determine: (1) the impact of Notch activation by BITC on its effects contributing to mammary cancer chemoprevention;(2) the contribution of uPA/uPAR suppression in BITC-mediated inhibition of EMT;and (3) obtain in vivo evidence for efficacy of BITC against bCSC self-renewal and to study the role of uPA/uPAR system in this response. Translational Impact of the Proposed Research: Studies conducted thus far provide compelling preclinical evidence for chemopreventive efficacy of BITC against breast cancer, but efficient translation of these findings into a clinical setting is dependent on a full understanding of the molecular pharmacology of BITC driving its chemopreventive responses. Clinical trial design without a full appreciation of the molecular pharmacology of BITC may be sub-optimal. Defining innovation of the current application includes translational merit of the proposed research as well as novel research directions. For example, intrinsic value of the studies proposed in Aim 1 resides in potential rational design of a combination regimen involving BITC and a ?-secretase inhibitor to achieve even greater chemopreventive efficacy.
Aim 2 may identify biomarkers of BITC response (e.g., uPA and uPAR) potentially useful in future clinical investigations. Discovery of biomarker(s) predictive of BITC response is an equally meritorious objective because cancer incidence is too rigorous of an end point for malignancies with long latency such as breast cancer. Likewise, experimental validation of the in vivo efficacy of BITC against bCSC may (in future) lead to novel BITC- based regimens for clinical management of breast cancer because the existing mechanistic model stipulates critical role for bCSC not only in cancer development and progression but also in resistance to therapy.

Public Health Relevance

Breast cancer is a leading cause of cancer-related death in women worldwide. Novel approaches for chemoprevention of breast cancer are clinically attractive. Long-term objective of this research project is to develop a safe and inexpensive but effective regimen for chemoprevention of breast cancer using cruciferous vegetable constituent benzyl isothiocyanate (BITC). Research conducted in the previous funding period provided preclinical evidence for efficacy of BITC against breast cancer, but efficient translation of these observations into a clinical setting depends on full appreciation of the molecular pharmacology of BITC. Clinical trial design without a full appreciation of the molecular pharmacology of BITC may be sub-optimal. Defining innovation of the current application includes translational merit of the proposed research as well as novel research directions.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
2R01CA129347-06
Application #
8368331
Study Section
Chemo/Dietary Prevention Study Section (CDP)
Program Officer
Emenaker, Nancy J
Project Start
2007-09-07
Project End
2017-05-31
Budget Start
2012-08-01
Budget End
2013-05-31
Support Year
6
Fiscal Year
2012
Total Cost
$279,768
Indirect Cost
$95,468
Name
University of Pittsburgh
Department
Pharmacology
Type
Schools of Medicine
DUNS #
004514360
City
Pittsburgh
State
PA
Country
United States
Zip Code
15213
Antony, Marie L; Lee, Joomin; Hahm, Eun-Ryeong et al. (2014) Growth arrest by the antitumor steroidal lactone withaferin A in human breast cancer cells is associated with down-regulation and covalent binding at cysteine 303 of ?-tubulin. J Biol Chem 289:1852-65
Sehrawat, Anuradha; Sakao, Kozue; Singh, Shivendra V (2014) Notch2 activation is protective against anticancer effects of zerumbone in human breast cancer cells. Breast Cancer Res Treat 146:543-55
Sehrawat, Anuradha; Kim, Su-Hyeong; Vogt, Andreas et al. (2013) Suppression of FOXQ1 in benzyl isothiocyanate-mediated inhibition of epithelial-mesenchymal transition in human breast cancer cells. Carcinogenesis 34:864-73
Kim, Su-Hyeong; Sehrawat, Anuradha; Singh, Shivendra V (2013) Dietary chemopreventative benzyl isothiocyanate inhibits breast cancer stem cells in vitro and in vivo. Cancer Prev Res (Phila) 6:782-90
Antony, Marie Lue; Kim, Su-Hyeong; Singh, Shivendra V (2012) Critical role of p53 upregulated modulator of apoptosis in benzyl isothiocyanate-induced apoptotic cell death. PLoS One 7:e32267
Xiao, Dong; Bommareddy, Ajay; Kim, Su-Hyeong et al. (2012) Benzyl isothiocyanate causes FoxO1-mediated autophagic death in human breast cancer cells. PLoS One 7:e32597
Hahm, Eun-Ryeong; Singh, Shivendra V (2012) Bim contributes to phenethyl isothiocyanate-induced apoptosis in breast cancer cells. Mol Carcinog 51:465-74
Kim, Su-Hyeong; Nagalingam, Arumugam; Saxena, Neeraj K et al. (2011) Benzyl isothiocyanate inhibits oncogenic actions of leptin in human breast cancer cells by suppressing activation of signal transducer and activator of transcription 3. Carcinogenesis 32:359-67
Sehrawat, Anuradha; Singh, Shivendra V (2011) Benzyl isothiocyanate inhibits epithelial-mesenchymal transition in cultured and xenografted human breast cancer cells. Cancer Prev Res (Phila) 4:1107-17
Warin, Renaud; Xiao, Dong; Arlotti, Julie A et al. (2010) Inhibition of human breast cancer xenograft growth by cruciferous vegetable constituent benzyl isothiocyanate. Mol Carcinog 49:500-7

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