Molecular analysis of the pro-apoptotic effect of the herbal extracts BN107 and BN108 on breast cancer cells

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While advances in early detection and adjuvant therapy for breast cancer have had a favorable impact on survival, patients who develop metastatic breast cancer generally succumb to death. Commonly used hormonal and chemotherapeutic agents can lead to transient regression of tumors as well as palliate symptoms related to cancer. However, these treatments are often accompanied by toxicities and intolerable side effects. Novel therapies with minimal toxicities are urgently needed for this patient population.
 Botanical medicine is one of the most popular complementary and alternative medical approaches, and Chinese herbal therapies are frequently sought and used by breast cancer patients. However, the molecular mechanisms through which certain herbal extracts exert growth inhibitory activity on breast cancer cells remain largely unknown. Clinical data on the efficacy and safety of the herbal medicines are even scarcer.
 We present preclinical data on the potential mechanisms of the growth-inhibitory effect of two herbs Anemarrhena asphodeloides (BN108) and Gleditsia sinensis (BN107) on breast cancer cells.
 Numerous studies have shown that the extract from the fruits of Gleditsia sinensis has cytotoxic activity against various tumors in vitro. We have investigated the potential mechanisms underlying the growth-inhibitory activity of BN107, an aqueous extract prepared from the thorns/modified leaves of Gleditsia sinensis, on breast cancer cells. A panel of breast cancer cell lines was examined for responses to BN107. Significant growth inhibition occurred in most of the cell lines tested. Apoptosis appears to be the major cellular pathways mediating the growth inhibitory effect of BN107 as evident from Annexin V binding, dissipation of mitochondrial potential, activation of caspases, and DNA fragmentation. Among all cell lines tested, absence of functional estrogen receptor (ER) appears to correlate positively with the sensitivity to BN107. Transcriptome analyses comparing sensitive versus resistant cell lines revealed distinct patterns of gene expression in response to BN107.
 Treatment of breast cancer cell lines with the aqueous extract Anemarrhena asphodeloides Bunge (BN108) induced cell death, in particular in lines overexpressing HER2 receptor. Normal mammary epithelial cells and fibroblasts were resistant to the cytotoxic effects of BN108. Breast cancer cells sensitive to BN108 underwent apoptotic death confirmed by DNA fragmentation, caspase activation, cleavage of PARP and Annexin V staining. In addition to caspase 3, we observed activation of caspases 4 and 9 which are linked to apoptosis induced by endoplasmic reticulum stress. BN108 induced rapid inactivation of AKT and mTOR kinases in breast cancer but not non-transformed cells. Expression of several genes that have well known pro-apoptotic and anti-proliferative characteristics was induced by BN108.