Abstract P5-03-09: Phenotypic modulation of mammary stem cells by CAPE (caffeic acid phenethyl ester)
Authors: Omene C , Martinez H , Dave H , Vijayakumar S , Shah N , Li XL , Iizuka D , Barcellos-Hoff MH 
Published: August 2014

Background: Over 30% of cancer patients and up to 60% of breast cancer patients routinely use alternative formulations, commonly in combination with allopathic treatment, which may be linked with fewer adverse events. The naturopathic formulation propolis, and its active component Caffeic Acid Phenethyl Ester (CAPE), is a widely available honeybee product with an extended safety history credited with anti-inflammatory, antioxidant, and antitumor properties. We have previously shown that CAPE, inhibits growth of MCF-7 (ER+/PR+) and MDA-231 (TNBC) cells by cell cycle arrest and apoptosis; inhibits mdr-1 gene expression, NF-kB, and VEGF (1). In addition, CAPE inhibits the growth and tumorigenic potential of breast cancer stem cells derived from MDA-231 cells (2). We have demonstrated that CAPE, which is structurally similar to the hydroxamic acid class of HDAC inhibitors, mediates its effects on breast cancer through epigenetic modifications via histone deacetylase inhibition (unpublished data). We hypothesized that CAPE would inhibit mammary stem cells, which was tested in primary mouse mammary epithelial cells and a human breast epithelial cell line.

Materials and Methods: Single mammary epithelial cells were isolated from Balb/c mice and cultured for 8 days in Ultra Low Attachment 96 well plates using mammosphere media containing 5%FBS and 5% Matrigel (3). The first generation of mammospheres were dissociated into single cells and recultured in the presence of CAPE for 8 days in secondary and tertiary passages. Alternatively, a Let7c-miRNA reporter expressed in MCF10a non-malignant human mammary epithelial cells were sorted to isolate Let7c-negative cells and placed in mammosphere culture and similarly treated with CAPE for 7 to 10 days. The mammospheres were stained with luminal marker K18 and basal marker K14 and examined using immunofluorescent microscopy.

Results: CAPE treatment results in a dose dependent decrease in mammosphere forming efficiency (MFE) of mammospheres obtained from a primary culture of mammary gland derived epithelial cells as well as those derived from the MCF10A cell line. Furthermore, MFE of tertiary cultures is reduced even after CAPE is removed. Interestingly, mammospheres shifted from predominantly K14 to K18 positive in those mammospheres that formed at the lowest CAPE treatment concentration.

Conclusion: CAPE inhibits mammosphere formation from normal mouse mammary gland and from a non-malignant human mammary cell line. The mechanism of CAPE effects may lie in its established role in inducing cell cycle arrest, apoptosis or via epigenetic mechanisms, among others. Interestingly, CAPE causes a shift from a basal to a luminal cell phenotype in mammospheres, suggesting an effect on lineage commitment.

References

1) *Wu J, *Omene C, Karkaszka J, et al. CANCER LETTERS. 2011 Sep 1;308(1):43-53. Epub 2011 May 13. (* Share 1st authorship)

2) Omene C, Wu J, Frenkel K. INVESTIGATIONAL NEW DRUGS. 2012 Aug;30(4):1279-88. Epub 2011 May 3.

3) Guo W, Keckesova Z, Donaher JL, et al. CELL. 2012 Mar 2;148(5):1015-28.

Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P5-03-09.