A Novel Herbal Medicine for the Treatment of Breast Cancer

Lay Abstract Breast cancer is the most common cancer affecting women and the second leading cause of death. Commonly used treatments cause serious side effects and may result in drug resistance. Our goal is to identi fy promising new dmg leads derived from plant-based products with anti-carcinogenic activity. An herbal formulation, SCinecarpus Lehyam (SL), used in Asian countries to treat breast cancer, was the focus of preliminary investigations in our laboratory. Studies were conducted on a type of breast cancer cell that is sensitive to the effects of estrogen and a type that is not sensitive to estrogen. These ceJl types were treated with five different extracts of SL (chloroform, ethyl acetate, n-butanol, n-hexane and \vatcr). We found a significant reduction in cell viability and an increase in cell death, particularly in cells that are not sensitive to estrogen, when they were treated with the n-hexane extract of SL, which we designated hSL. These results were further confirmed in animal models. Moreover, hSL did not induce toxIcity in normal breast cells. Recently, we identified a single major component in hSL, which we refer to as hSL-compound I (hSL-CI), which appears to be more potent than SL. Based on our preliminary data, we hypothesize that hSL-CI imparts its chcmotherapeutic and/or chcmopreventive effects by inhibiting progression of the ceIl cycle and inducing apoptosis. To test this hypothesis, three specific aims are proposed: I) to determine the cell-kiJling effect of hSL-CI in breast cancer using cell culture and animal model approaches; 2) to characterize hSL-CI by nuclear magnetic resonance (NMR) spectroscopy and mass spectrometry (MS) and elucidate its structure; 3) to analyze the mode of action of hSL-CI in breast cancer cells. In aim I, we wi II extend our studies to five additional types of breast cancer cells and normal breast cells and will analyze the effect ofhSL-CI through various bioassays aimed at helping us advance our understanding of the anti-cancer http://www .komen.org/ResearchGrants/Suhmit/printSection 1.aspx ?secid=section2 8/3/2005 ApplIcatIOn - Pnnt Prevlcw Page 2 at 20 properties of this potent extract of SL. Thcsc results will bc confirmed in animal tumor studics. [n aim 2, wc will use analytical tools to dctcrmine thc molecular structurc of hSL-CI. In aim 3, we will study the effect of hSL-C[ on genes that are responsible for cell death and survival. The outcomes of this study may lead to significant advances in our understanding of the anticancer effects of hSL-CI and may lead to drug discovery efforts for the treatment of breast cancer. Scientific Abstract Breast cancer (BCa) is the most commonly occurring cancer in women, comprising almost one third of all malignancies. Our goal is to identify promising new drug leads derived from phytochemicals with anti-carcinogenic activity. Recently we reported the anti-cancer effect of an herbal formulation, Scmecarpus Lchyam (SL), for the trcatment of SCa. Estrogen receptor (ER)-positive (MCF-7) and ER-negative (MDA-231) BCa cell lines were treated with different fractions of SL (chloroform, ethyl acetate, n-butanol, n-hexane and water). The n-hexane fraction of SL (hSL) significantly reduced cell viability and induced apoptosis with little toxicity to normal breast epithelial cells; this effect appears to be more potent in ER- than FR' BCa cells. Further, hSL inhibited the growth of xenografts in nude mice derived from MDA-231 cells. Subsequently, we isolated a major potent component from hSL, designated hSL-compound I (hSL-CI), which appears to have more potent anti-cancer effects than hSL when tested in MCF-7 and MDA-231 BCa cells. Based on our preliminary data, we hypothesize that hSL-CI imparts its chemotherapeutic and/or chemoprcventive effects by inhibiting progression of the cell cyele and inducing apoptosis. To test this hypothesis, three specific aims are proposed: 1) to determine the apoptotic effect of hSL-CI in BCa using in vitro and in vivo approaches; 2) to characterize hSL-CI by nuclear magnetic resonance (NMR) spectroscopy and mass spectrometry (MS) and elucidate its structure; 3) to analyze the molecular mechanisms of the apoptotic signal transduction pathways of hSL-CI in breast cancer cells. In aim I, we will analyze the effect of hSL-CI on additional ER+ and ER- Bea cell lines and normal prostate epithelial cells by MTl, Annexin V-FITC, propidium iodide (PI) staining and TUNEL assays. These in vitro findings will be confirmed by in vivo tumor regression studies (xenografts). In aim 2, hSL-CI will be characterized by NMR and MS to elucidate its structure. In aim 3, we will analyze MAP kinase signaling in ER+ BCa and NFkB signaling in EW BCa by Western blot analysis, promoter assays and electro-mobility shift analysis to detemline the molecular function of hSL-CI when exposed to BCa cells. The outcomes of this study may lead to significant advances in our understanding of the effects ofhSL-CI and may lead to drug discovery efforts for the treatment of breast cancer. Project Description