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Chemical Composition, Cytotoxicity and Molecular Profiling of Cordia Africana Lam

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Chemical Composition, Cytotoxicity and Molecular Profiling of Cordia Africana Lam Natural Product Research Formerly Natural Product Letters ISSN: 1478-6419 (Print) 1478-6427 (Online) Journal homepage: https://www.tandfonline.com/loi/gnpl20 Chemical composition, cytotoxicity and molecular profiling of Cordia africana Lam. on human breast cancer cell line Abeer M. Ashmawy, Iriny M. Ayoub & Omayma A. Eldahshan To cite this article: Abeer M. Ashmawy, Iriny M. Ayoub & Omayma A. Eldahshan (2020): Chemical composition, cytotoxicity and molecular profiling of Cordiaafricana Lam. on human breast cancer cell line, Natural Product Research, DOI: 10.1080/14786419.2020.1736064 To link to this article: https://doi.org/10.1080/14786419.2020.1736064 View supplementary material Published online: 05 Mar 2020. Submit your article to this journal View related articles View Crossmark data Full Terms & Conditions of access and use can be found at https://www.tandfonline.com/action/journalInformation?journalCode=gnpl20 NATURAL PRODUCT RESEARCH https://doi.org/10.1080/14786419.2020.1736064 SHORT COMMUNICATION Chemical composition, cytotoxicity and molecular profiling of Cordia africana Lam. on human breast cancer cell line Abeer M. Ashmawya, Iriny M. Ayoubb and Omayma A. Eldahshanb aCancer Biology Department, Biochemistry Unit, National Cancer Institute, Cairo University, Cairo, Egypt; bFaculty of Pharmacy, Pharmacognosy Department, Ain Shams University, Cairo, Egypt ABSTRACT ARTICLE HISTORY Metabolic profiling of the hexane extract and essential oil Received 4 January 2020 of Cordia africana Lam., an economically important agroforestry Accepted 13 February 2020 species, was performed using GC-FID and GC-MS. Cytotoxicity was assessed in MCF-7 cancer cells using sulphorhodamine-B assay. KEYWORDS Cordia africana Caspase-8, BcL-2 and Ki-67 levels were evaluated using ; cytotoxicity; MCF-7; BcL-2; Ki-67; enzyme-linked immunosorbent assay. Forty-one compounds were caspase-8; apoptosis identified in the hexane extract, meanwhile, eighty compounds were annotated in the essential oil. n-Decane (9.52%), b-sitosterol (8.13%), undecane (6.32%) and palmitaldehyde (5.50 %) were the major compounds identified in the hexane extract, whereas, b-caryophyllene (32.0%), germacrene D (10.12%) and d-cadinene (7.20%) predominated in the essential oil. The half maximal inhibi- tory concentrations (IC50) of the extract and oil were 4.55 mg/mL and 12.90 mg/mL, respectively. Caspase-8 activity increased signifi- cantly in hexane extract treated cells. Furthermore, BcL-2 and Ki- 67 protein expression levels were significantly decreased. Cordia africana Lam exhibit potent cytotoxicity against breast cancer cell line, most likely through apoptosis regulation. CONTACT Omayma A. Eldahshan [email protected] Faculty of Pharmacy, Pharmacognosy Department, Ain Shams University, African Union Organization Street, Cairo 11566, Egypt. Supplemental data for this article can be accessed at https://doi.org/10.1080/14786419.2020.1736064. ß 2020 Informa UK Limited, trading as Taylor & Francis Group 2 A. M. ASHMAWY ET AL. 1. Introduction Globally, cancer is considered to be the second leading cause of death (Fitzmaurice et al. 2017). Breast cancer is the most common cancer among women. According to the World Health Organization, 2.1 million women are diagnosed with breast cancer every year. Moreover, breast cancer is responsible for the high- est rates of cancer deaths among women. In 2018, it was estimated that about 627,000 women died from breast cancer representing 15% of all cancer-related deaths among women (WHO 2018). Apoptosis is considered a promising target for anticancer therapy (Pfeffer and Singh 2018). Caspases are cysteine proteases that act as key mediators of apoptosis (Thornberry 1998). In cases of apoptotic cell death, caspases are activated resulting in breakdown of critical cellular substrates triggering dramatic morphological changes normally associated with apoptosis (Bratton et al. 2000). Caspase-dependent apoptosis follows intrinsic and extrinsic pathways. The intrinsic pathway arises as a consequence of DNA damage or cellular stress i.e., a signal originating within the cell. In contrast, the extrinsic pathway is elicited by stimulation of cell surface receptors, known as “death receptors” (Kominami et al. 2012). In cancer, the apoptotic pathway is inhibited through the overexpression of antiapoptotic proteins and the under-expression of proapoptotic proteins (Pfeffer and Singh 2018). Nature combats cancer by its medicinal plants. Naturally occurring secondary metabolites from plants offer promising new anticancer therapies that exhibit anticancer activity by activating the apoptotic pathway (Pfeffer and Singh 2018). Researchers have identified a myriad of plant extracts, essential oils and phytochemicals possessing promising anticancer properties (Eldahshan 2013; Ayoub et al. 2014; Al-Sayed et al. 2016; Taha and Eldahshan 2017;Elkadyand Ayoub 2018;Ashmawyetal.2019). Cordia africana Lam. (syn.: C. holstii Gurke)€ is an evergreen tree belonging to the borage family (Boraginaceae) (Gebreegziabher 2016). It is an important agroforestry species native to Eastern and Southern Africa (Schmidt and Mwaura 2010; Derero et al. 2011). C. africana is more valued for timber than medicinal purposes, represent- ing an economically important species (Balemie and Kebebew 2006; Regassa 2013). In folk medicine, C. africana is used for the treatment of migraine, broken bones, wounds, gastritis and constipation (Zenebe et al. 2012). Reported pharmacological activities include antibacterial (Geyid et al. 2005), anti-inflammatory and cytotoxic activities (Isa et al. 2014). Despite its applications in traditional medicine, little is known on the phytochemical profile and biological activities of C. africana. Scientific evidence addressing the cellular and molecular mechanisms is lacking. Only few reports were traced for the pharmacological activities of C. africana including antibac- terial (Geyid et al. 2005), antifungal (Moshi et al. 2007), anti-inflammatory and cytotoxic activities (Isa et al. 2014). In this study, GC-MS based metabolic profiling and cytotox- icity of C. africana leaf essential oil and hexane extract were performed. Effects on the expression levels of the molecular biomarkers caspase-8, Bcl-2 and Ki-67 were assessed to unveil the possible underlying mechanisms. To the best of our knowledge, this is the first report on the chemical profile and biological activity of the essential oil and hexane fraction of C. africana leaves. NATURAL PRODUCT RESEARCH 3 2. Results and discussion 2.1. Chemical composition of the hexane extract and essential oil Hydrodistillation of the leaves yielded 0.016 % (w/w), colorless essential oil having a characteristic odor. Forty-one compounds were identified in the hexane extract, accounting for 70.03% of total composition. Meanwhile, 80 compounds were identified in the leaf essential oil, representing 89.45% of total oil composition. Hydrocarbons and phytosterols predominated in the hexane extract, with n-decane (9.52%), b-sitosterol (8.13%), undecane (6.32%), palmitaldehyde (5.5%), nonane (5.24%) and phytol (5.20%) representing the major constituents identified. However, sesquiterpene hydrocarbons predominated in the leaf essential oil including b-caryophyllene (32.0%), germacrene D (10.12%), d-cadinene (7.20%) in addition to phytol (5.27%). (Figures S1 and S2, Tables S1 and S2, Supplementary material). 2.2. Cytotoxicity and apoptosis 2.2.1. Determination of half maximal inhibitory concentration (IC50) Both the hexane extract and essential oil showed promising anti-proliferative activity exhibiting IC50 values of 4.55 and 12.90 mg/mL, respectively. Cytotoxicity of CA-HE was comparable to that of doxorubicin (IC50 3.83 mg/mL). Consequently, the possible underlying mechanism for the observed cytotoxic activity of the more potent CA-HE was investigated. 2.2.2. Apoptosis and proliferation Caspase-8 enzymatic activity was significantly increased in MCF-7 cells treated with CA-HE at a dose equivalent to the IC50 value (4.55 mg/mL) (Figure S3, Supplementary material). Meanwhile, BcL-2 protein expression was significantly decreased (Figure S4, Supplementary material). Moreover, expression of the proliferative marker (Ki-67) protein expression level in MCF-7 cells was significantly reduced compared to the untreated group, P < 0.05 (Figure S5, Supplementary material). Hydrocarbons and phytosterols dominated in C. africana hexane extract (CA-HE). Phytol, an acyclic monounsaturated diterpene alcohol, induced significant anticancer activity against breast cancer at IC50 125 lg/mL (Sheeja et al. 2016). Moreover, it induced concentration-dependent cytotoxic response in many cancerous cell lines, being the most effective against the breast adenocarcinoma MCF-7 with IC50 of 8.79 mM (Pejin et al. 2014). Phytosterols possess anti-cancer effects against estrogen-dependent human breast cancer cell lines (Ju et al. 2004). Phytosterols inhibit the production of carcinogens, cancer-cell growth, invasion, metastasis, and promotes apoptosis of cancerous cells (Meric et al. 2006). b-Sitosterol treatment of breast cancer cells at 16 mM has been observed to reduce the cell growth by 50% (Awad et al. 2007). It increased activity of caspase-3 of human endothelial cells in vitro by more than 60% (Rubis et al. 2008). This could be explained through down regulation of Bcl-2. Major components of CA-EO identified include a and b-caryophyllene, germacrene D, d-cadinene and phytol. Caryophyllene significantly reduced tumors developed from orthotopically
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