Vol 13, Issue 10, 2020 Online - 2455-3891 Print - 0974-2441 IN VITRO AND IN SILICO STUDIES ON EVALUATING EREMANTHIN AS POTENTIAL THERAPEUTIC DRUG AGAINST BREAST CANCER ANITA ROY A, ANGEL MARY, INDU SABAPATHY, MANIKKAM RAJALAKSHMI* Bioinformatics Centre (BIF), Department of Biotechnology and Bioinformatics, Holy Cross College (Autonomous), Tiruchirappalli, Tamil Nadu, India. Email: mdraji@gmail.com Received: 10 January 2020, Revised and Accepted: 11 August 2020 ABSTRACT Objective: The present study was aimed to evaluate the anticancer property of eremanthin isolated from Costus speciosus against breast cancer using in vitro and in silico approaches and thereby to develop eremanthin as a typical phytotherapeutic drug against cancer. Methods: The presence of specific biologically active extract was confirmed under GC–MS/MS (gas chromatography–mass spectrometry) analysis. The cell proliferation inhibitory effect of the eremanthin was confirmed by MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide) and LDH (lactate dehydrogenase) assay. In silico studies were performed to predict the targeted interaction of eremanthin with cancer proteins. Results: The GC–MS/MS analysis confirmed the presence of eremanthin with peak value of RA: 20.03. The MTT and LDH assays revealed the antiproliferative activity of eremanthin on MCF-7 and MDA-MB-231 breast cancer cell lines. The results provide stable interaction between eremanthin and cancer target proteins. Conclusion: Thus, the compound can be used as an effective herbal therapeutic molecule to treat cancer with further explorations. Keywords: Costus speciosus, Eremanthin, Cancer, Molecular docking. INTRODUCTION Cancer is a complex genetic disease that is caused by gene mutations, dysregulation of cellular pathways [1] or through environmental factors, the carcinogens [2]. Surgery combined with chemotherapy and radiotherapy has become the local therapy for cancer, which has been often found to result in significant side effects and related toxicities [3]. Alternative and traditional medicines, mostly herbal, are now regarded as important source of therapy but underutilized tools against disease which have better compatibility and lesser side effects [4]. Natural product-based cancer therapy is flourishing with substantial utilization of anticancer drugs in clinics being either natural or derived from natural products [5]. The therapeutic potential of Costus speciosus [6] is referred in Ayurveda and can be emphasized for identifying an array of biologically active compounds in different parts of the plant [7,8]. Thus, the present study aspires to estimate the anticancer property of a biologically active constituent isolated from C. speciosus, eremanthin against breast cancer. Eremanthin is a common secondary metabolite present in medicinally valuable plants and a bioactive compound with various pharmacological properties [9]. In recent times, search for new cancer drugs has moved toward a more mechanistic approach on identifying molecular targets for cell transformation rather than developing drugs that kill tumors cells [10]. We evaluated the potentials of eremanthin isolated from n-hexane extract of C. speciosus rhizome on breast cancer cells and cancer targeted protein. METHODS Chemicals All chemicals were purchased from Sigma-Aldrich Chemicals Pvt. Ltd. (USA). Solvents were obtained from Fisher Scientific Ltd., India. All the chemicals used were extra pure and were of culture grade. Plant collection The rhizome of C. speciosus was collected from the local market of Tiruchirappalli District, Tamil Nadu, India. The species was authenticated by the Department of Botany, Holy Cross College (Autonomous), Tiruchirappalli, India. The voucher specimen is preserved in the herbarium of the department. The rhizomes were dried under shade and mechanically reduced to moderate coarse powder and sieved. Preparation of plant extract The rhizome powder was collected and soaked in organic solvents such as hexane, ethyl acetate, and methanol and used for the preparation of extract. Five hundred grams powder was extracted with 1.5l of hexane (1:3 w/v) for 72 h with frequent mixing and filtered. The filtrate was evaporated to dry under reduced pressure using rotary evaporator at 40°C. The remaining plant material are used for further extraction with ethyl acetate and methanol sequentially in a similar manner and obtained for the present study. Phytochemical screening Chemical tests were carried out on the rhizome extracts of C. speciosus using standard procedures to identify the constituents as described by Harborne (1973) for alkaloids, flavonoids, terpenoids, quinines, phenols, volatile oil, glycosides, tannins, and saponins [11]. Isolation and identification of the active compound One kilogram powder was soaked in 3l of hexane for 72 h with intermittent shaking and after filtering through Buchner funnel, it was concentrated using vacuum rotary evaporator at 40°C. Twenty-five gram of active crude hexane extract was chromatographed on a silica gel column (Merck 10–200 mesh, 750 g 3.5 i.d.×60 cm) and successively eluted with stepwise gradient of petroleum ether and hexane solvent system (5%, 10%, 20%, 30%,50%, 70%, and 100%). A total of 116 fractions were collected and each fraction was spotted on a pre-coated silica gel (Merck-60 F254, 0.25 mm thick) plate and eluted in hexane:ethyl acetate (3:1). An oil substance was obtained in subfraction. The oil substance was checked on TLC (thin- layer chromatography). It showed single spot on TLC. It was subjected to GC–MS/MS analysis. © 2020 The Authors. Published by Innovare Academic Sciences Pvt Ltd. This is an open access article under the CC BY license (http://creativecommons. org/licenses/by/4. 0/) DOI: http://dx.doi.org/10.22159/ajpcr.2020.v13i10.36818 Research Article