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Original Article

Withania somnifera extract reduces the invasiveness of MDA-MB-231
breast cancer and inhibits cytokines associated with metastasis

Kamel F. Khazal , Donald L. Hill 2
1
1 Department of Biomedical Sciences, School of Veterinary Medicine, Tuskegee University, Tuskegee, Alabama 36088, USA.
2 Division of Preventive Medicine, University of Alabama at Birmingham, Birmingham, Alabama 35294, USA.
Correspondence to: Dr. Kamel F. Khazal, Department of Biomedical Sciences, School of Veterinary Medicine, Tuskegee University, Tuskegee,
Alabama 36088, USA. E-mail: kamel@mytu.tuskegee.edu

ABSTRACT
Aim: The aim was to examine the anti-proliferative effect of a Withania somnifera (WS) root extract in cell cultures and
nude mouse xenografts of breast cancer cell line MDA-MB-231. Methods: WS root extract was used to treat tumor cells at
concentrations up to 100 μg and for nude mouse experiments, the mice received daily WS at 300 mg/kg by oral gavage for
8 weeks. Results: The WS extract reduced viability of MDA-MB-231 cells by 75% and 88% after exposure of the cells to 50 and
100 μg/mL, respectively, compared to vehicle-treated controls. WS extract caused a dose-dependent increase in the percentage
of cells in the sub-G1 phase compared to untreated controls by 6% and 10% after exposure to 25 and 50 μg/mL WS extract,
respectively. WS extract also inhibited proliferation of xenografted MDA-MB-231 cells. The WS extract caused reductions in
xenograft size by 60% compared to the untreated control after 8 weeks of treatment. Six of ten mice in the control group showed
tumor metastasis to the lung, whereas there was none in the mice treated with the WS extract. At the gene level, WS caused a 75%
reduction in chemokine CCL2 expression (P < 0.05) in the xenografted tumors of the treated mice. Conclusion: WS root extract
inhibited proliferation of breast cancer cells in vitro and in vivo and signifi cantly reduced expression of the cytokine, CCL2. These
results warrant further studies to assess the underlying molecular mechanism of the anti-tumor activity of the WS extract in breast
cancer.

Key words: Withania somnifera extract, MDA-MB-231, breast cancer, metastasis, animal model

Introduction to spread and recur. TNBCs are different from other
[2]
kinds of breast cancer in that they are highly metastatic
Invasive breast cancer is considered one of the great and resistant to conventional therapies, such as anticancer
challenges for clinicians to control and improve survival drugs and radiation. [2]
of patients. In 2013, an estimated 232,340 new cases of
invasive breast cancer were diagnosed in women in the In a search for an agent that inhibits proliferation and
USA, along with other 64,640 cases of non-invasive invasion of TNBCs, we evaluated an extract derived from
[1]
breast cancer. For women under 45, deadly forms an Indian herb, Withania somnifera (WS), which is a
of this type of breast cancer are more common in nightshade medicinal plant that contains active components
African-American women than white women, and for the treatment of a variety of ailments, including
African-American women are more likely to die of breast cancer. [7-10] The use of WS root extract is practical since
[2]
cancer. Despite three decades of advances in treatment it contains the active compounds present in the plant. In
of breast cancer using hormone receptor modulators, TNBC cells, sub-cytotoxic concentrations of withaferin A,
[11]
aromatase inhibitors, and surgery, [3-5] mortality remains derived from WS, reduce various effectors of metastasis.
high due to tumor metastasis to the lymph nodes, liver, In the present study, we assessed the effect of the WS
and lung. Triple-negative breast cancer (TNBC) extract on proliferation and metastasis of MDA-MB-231
[6]
accounts for 10-20% of diagnosed breast cancers and cells, derived from a TNBC, in cell cultures, and in mice.
is more likely to affect younger African Americans,
Hispanics, and/or those with BRCA1 mutations. TNBCs Methods
are more aggressive, diffi cult to treat, and more likely Preparation of WS extract

Roots of WS were ground to a paste, and then extracted
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with 5 volumes of 70% ethanol by stirring for 2 days.
Quick Response Code: The alcoholic extract was fi ltered, and the solvent was
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www.jcmtjournal.com evaporated under a vacuum. The extract was then dried
to a powder and kept in a closed container until use. To
[12]
avoid variations in the activity of different preparations,
DOI:
10.4103/2394-4722.157601 the suffi cient extract was obtained in one batch for use
throughout the experiments.

94 Journal of Cancer Metastasis and Treatment ¦ Volume 1 ¦ Issue 2 ¦ July 15, 2015 ¦