HEALTH BENEFITS OF DIALLYL TRISULFIDE, A BIOACTIVE COMPOUND FROM ALLIUM VEGETABLES

• Sponsoring Institution: National Institute of Food and Agriculture
• Project Status: COMPLETE
• Funding Source: HATCH
• Reporting Frequency: Annual
• Accession No.: 1018789
• Project Start Date: May 1, 2019
• Project End Date: Jun 30, 2021
• Program Code: [(N/A)]- (N/A)

Recipient Organization
UNIVERSITY OF NEVADA
(N/A)
RENO,NV 89557

Performing Department
Nutrition

Non Technical Summary
Breast cancer is the most frequently diagnosed type of cancer in women in the United States. Ductal carcinoma in situ is an early stage, localized form of breast cancer that can progress to an invasive breast cancer phenotype and increase the lifelong risk of developing a metastatic and lethal breast cancer. The survival of patients with ductal carcinoma in situ depends on blocking the progression of ductal carcinoma in situ lesions to an invasive form of breast cancer. Currently there are no effective chemopreventive strategies to prevent the development of breast ductal carcinoma in situ. Epidemiological observations have suggested an inverse association between consumption of Allium vegetables (e.g., garlic and onions) and a lower risk of developing cancer, including breast cancer. Diallyl trisulfide is a bioactive compound found naturally in Allium vegetables, that has been shown to have multiple anticancer effects. In this project we propose to develop a novel strategy to target the growth and progression of ductal carcinoma in situ by using diallyl trisulfide. This project will provide a proof-of-principle for the effect of diallyl trisulfide against breast ductal carcinoma in situ. The long term goal of this project is to develop a novel chemopreventive strategy for breast cancer that uses diet-derived bioactive compounds.

Animal Health Component

100%

Research Effort Categories

Basic

(N/A)

Applied

100%

Developmental

(N/A)

Classification

Knowledge Area (KA)	Subject of Investigation (SOI)	Field of Science (FOS)	Percent
702	1451	1010	100%

Knowledge Area
702 - Requirements and Function of Nutrients and Other Food Components;

Subject Of Investigation
1451 - Onion, garlic, leek, shallot;

Field Of Science
1010 - Nutrition and metabolism;

Keywords

allium vegetables

diallyl trisulfide

breast ductal carcinoma in situ

chemoprevention

Goals / Objectives
The primary goal of this project is to provide a proof-of-principle for the effect of dially trisulfide, a bioactive compound found in Allium vegetables, against breast ductal carcinoma in situ cell growth. The objectives of this project are to determine the effect of diallyl trisulfide on 1) cell growth and colony formation ability; 2) induction of cell cylcle arrest; and 3) induction of apoptosis in breast ductal carcinoma in situ cells.

Project Methods
We will use well-established cell-based models to test the hypothesis that diallyl trisulfide inhibits breast ductal carcinoma in situ tumor growth through induction of cell cycle arrest and apoptosis in ductal carcinoma in situ cells. As a cellular model we will use two breast ductal carcinoma in situ cell lines (MCF10DCIS and SUM102PT) and a normal-like breast epithelial cell line (MCF10A) as a negative control. MCF10DCIS cells (also known as MCF10DCIS.com cells) were isolated from a xenograft originating from premalignant MCF10AT cells and were shown to form ductal carcinoma in situ-like comedo lesions that spontaneously progress to invasive ductal carcinoma when xenografted in immunodeficient mice (21). SUM102PT cells were isolated from a 56-years-old female patient with minimally invasive breast adenocarcinoma and with extensive ductal carcinoma in situ and have been well described (21-23). MCF10A cells were isolated from a 36-years-old female patient with fibrocystic disease and represent a non-tumorigenic breast epithelial cell line that will be used as a negative control. The specific aims of this project are:Aim 1. Determine the effect of diallyl trisulfide on cell growth and cell cycle progression in breast ductal carcinoma in situ cells. First we will determine the effect of diallyl trisulfide on cell cycle progression, and then we will employ gene overexpression or gene knockdown experiments, as appropriate, to determine the contribution of specific cell cycle pathway components on the overall anti-cancer effect of diallyl trisulfide in ductal carcinoma in situ cells.1.1 Determine the effect of diallyl trisulfide on cell growth and colony formation ability of ductal carcinoma in situ cells. Cell survival and clonogenic assays will be performed to determine the effect of diallyl trisulfide on ductal carcinoma in situ cell growth and colony formation ability using methodology described by us previously (22,24). The inhibitory concentration (IC50) for each cell line will be determined.1.2 Determine the effect of diallyl trisulfide on cell cycle progression in ductal carcinoma in situ cells. We will characterize the effect of diallyl trisulfide on cell cycle arrest by using MCF10DCIS and SUM102PT cell lines as a model. Cells will be treated with diallyl trisulfide in a dose- and time-dependent manner and then processed for cell cycle analysis by flow cytometry using methodology described by us previously (24,25). We will then determine the effect of diallyl trisulfide treatment on cell cycle molecular markers. Cells will be treated with diallyl trisulfide in a dose- and time-dependent manner and then analyzed by Western immunoblotting for the effect of diallyl trisulfide on cell cycle markers such as cyclin B1, phospho histone H3, Chk2, cdc25A and cdc25C.Aim 2. Determine the effect of diallyl trisulfide on apoptosis induction in ductal carcinoma in situ cells. Cells will be treated with diallyl trisulfide in a dose- and time-dependent manner and then processed for analysis of apoptosis markers by Western immunoblotting and by an ELISA assay for quantifying cytoplasmic histone-associated DNA fragmentation using methodology described by us previously (24). The effect of diallyl trisulfide on cell apoptosis markers such as cleaved PARP, cleaved caspase 3, Bcl2, BclXL, Bak, Bax and Bim will be determined. Once the effect of diallyl trisulfide on apoptosis pathway components is determined, gene overexpression or gene knockdown experiments, as appropriate, will be conducted to understand the contribution of diallyl trisulfide regulation of specific pathway components on induction of apoptosis in ductal carcinoma in situ cells.In summary, the proposed studies will provide a proof-of-principle for the effect and mechanism of action of diallyl trisulfide against ductal carcinoma in situ cell growth. This contribution is significant because it may lead to a novel strategy to target the progression of ductal carcinoma in situ tumors by using diallyl trisulfide, a diet-derived promising chemopreventive agent. The results of this study will provide a foundation for future pre-clinical and clinical studies of diallyl trisulfide as a chemopreventive agent for breast ductal carcinoma in situ.

Progress 05/01/19 to 06/30/21

Outputs
Target Audience: Nothing Reported Changes/Problems:The COVID-19 pandemic caused some delays and disruptions during the duration of this project (2020-2021). What opportunities for training and professional development has the project provided?Trained two graduate students (Biotechnology and Nutrition Science) and one undergraduate student (Biotechnology). How have the results been disseminated to communities of interest?1) Published peer-reviewed journal articles: 1.1. Stan S.D., Abtahi M. Diallyl trisulfide induces apoptosis in breast ductal carcinoma in situ derived and minimally invasive breast cancer cells. Nutrients, 2022, 14(7): 1455. 1.2. Kiesel, V.A., Stan S.D. Modulation of Notch signaling pathway by bioactive dietary agents. International Journal of Molecular Sciences, 2022, 23(7): 3532. 2) Presented research at a national conference; Stan S.D. and Abtahi M. "Mechanisms of action of diallyl trisulfide in breast ductal carcinoma in situ cells" at NUTRITION 2020 LIVE ONLINE, June 1- June 4, 2020, American Society for Nutrition Annual Meeting. 3) Presented at a local community outreach event; Abtahi M. and Stan S.D. "Health benefits of Allium vegetables-derived bioactive compounds" Poster presentation at Nevada Field Day, October 19, 2019. This community outreach event was attended by over 500 attendees. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? Performed research activities to provide a proof-of-principle for the effect of diallyl trisulfide, a bioactive compound found in Allium vegetables, against breast ductal carcinoma in situ cell growth, colony formation ability, induction of cell cycle arrest, and induction of apoptosis in breast ductal carcinoma in situ cells.

Publications

• Type: Journal Articles Status: Published Year Published: 2022 Citation: Stan S.D., Abtahi M. Diallyl trisulfide induces apoptosis in breast ductal carcinoma in situ derived and minimally invasive breast cancer cells. Nutrients, 2022, 14(7): 1455.
• Type: Journal Articles Status: Published Year Published: 2022 Citation: Kiesel, V.A., Stan S.D. Modulation of Notch signaling pathway by bioactive dietary agents. International Journal of Molecular Sciences, 2022, 23(7): 3532.
• Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Stan S.D., Abtahi M. Mechanisms of action of diallyl trisulfide in breast ductal carcinoma in situ cells. American Society for Nutrition Annual Meeting, June 2020, Current Developments in Nutrition, vol. 4, supplement 2, page 353.

Progress 10/01/19 to 09/30/20

Outputs
Target Audience: Nothing Reported Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Trained one graduate student and one undergraduate student on the project. How have the results been disseminated to communities of interest?Presented research at a national conference and at a local community outreach event. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? Performed research activities to provide a proof-of-principle for the effect of diallyl trisulfide, a bioactive compound found in Allium vegetables, against breast ductal carcinoma in situ cell growth, colony formation ability and induction of apoptosis in breast ductal carcinoma in situ cells.

Publications

• Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Stan S.D. and Abtahi M. Mechanisms of action of diallyl trisulfide in breast ductal carcinoma in situ cells. Current Developments in Nutrition, Volume 4, Issue Supplement 2, June 2020, Page 353.

Progress 05/01/19 to 09/30/19

Outputs
Target Audience: Nothing Reported Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Trained one graduate student and one undergraduate student on the project. How have the results been disseminated to communities of interest? Nothing Reported What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? Performed research activities to provide a proof-of-principle for the effect of diallyl trisulfide, a bioactive compound found in Allium vegetables, against breast ductal carcinoma in situ cell growth, colony formation ability and induction of apoptosis in breast ductal carcinoma in situ cells.

Publications