Original article Synthesis of 2-alkoxy and 2-benzyloxy analogues of estradiol as anti- breast cancer agents through microtubule stabilization B. Sathish Kumar a , Amit Kumar b , Jyotsna Singh b , Mohammad Hasanain b , Arjun Singh a , Kaneez Fatima a , Dharmendra K. Yadav a , Vinay Shukla b , Suaib Luqman a , Feroz Khan a , Debabrata Chanda a , Jayanta Sarkar b , Rituraj Konwar b , Anila Dwivedi b , Arvind S. Negi a, * a CSIR-Central Institute of Medicinal and Aromatic Plants (CSIR-CIMAP), Kukrail Picnic Spot Road, P.O. CIMAP, Lucknow 226015, India b CSIR-Central Drug Research Institute (CSIR-CDRI), B.S.10/1, Sector 10, Jankipuram Extension, Sitapur Road, Lucknow 226031, India article info Article history: Received 4 June 2014 Received in revised form 25 August 2014 Accepted 9 September 2014 Available online 10 September 2014 Keywords: Breast cancer Microtubule stabilization Cell cycle Estrogenicity Antiestrogenicity Acute oral toxicity abstract 2-Methoxyestradiol (2ME2) is an investigational anticancer drug. In the present study, 2-alkoxyesters/ acid and 2-benzyloxy analogues of estradiol have been synthesized as analogues of 2ME2. Three of the derivatives exhibited significant anticancer activity against human breast cancer cell lines. The best analogue of the series i.e. 24 showed stabilization of tubulin polymerisation process. It was substantiated by confocal microscopy and molecular docking studies where 24 occupied ‘paclitaxel binding pocket’ to stabilize the polymerisation process. Compound 24 significantly inhibited MDA-MB-231 cells (IC 50 : 7 mM) and induced arrest of cell cycle and apoptosis in MDA-MB-231 cells. In acute oral toxicity, 24 was found to be non-toxic and well tolerated in Swiss albino mice up to 1000 mg/kg dose. © 2014 Elsevier Masson SAS. All rights reserved. 1. Introduction Breast cancer is a leading cause of women mortality worldwide. It is one of the most common malignant cancers for women causing 1.7 million patients and about 0.52 million deaths in 2012 [1]. Many factors are believed to contribute to this burden of breast cancer including genetic, environmental, life style and biological etc. The mechanism involved in cell proliferation, invasion and metastasis of breast cancer are not fully understood. Breast cancer is a het- erogeneous disease consisting of multiple molecular subtypes [2aec]. Based on responsiveness, it is mainly subdivided in to hormone responsive (ER/PR/Her-2) and hormone non-responsive. Various approaches have been developed to tackle these. For ER positive breast cancers selective estrogen receptor modulators (SERMs) and aromatase inhibitors have been developed while cytotoxic drugs like paclitaxel, doxorubicin, cyclophosphamide etc. are used to combat ER-negative cancers. However, there is a general consensus that breast cancer treatment needs multimodality approach to eradicate residual cancer cells and prevent recurrence of the disease. Several anti-breast cancer drugs have been devel- oped, but the morbidity and mortality of the disease is so high that it is still a challenge to scientific fraternity. Microtubules have been considered as an ideal target for anti- cancer drug development due to their essential role in mitosis forming the dynamic spindle apparatus. Microtubules play crucial role in the maintenance of cell structure, chromosomal segregation, protein trafficking and mitosis [3]. Both, microtubule stabilisers like paclitaxel, epothilones, discodermolide etc. and microtubule in- hibitors like colchicine, podophyllotoxin, combretastatins, vinca alkaloids etc. induce cell cycle arrest and apoptosis and thus act as anticancer agents. In the recent past, 2-methoxyestradiol (2ME2, 1) a metabolite of endogenous estrogen 17b-estradiol has emerged as a potent anti- cancer molecule. It is an investigational drug. 2ME2 is an anti- angiogenic drug which induces G2/M arrest and apoptosis [4]. It selectively targets endothelial cell adhesion and migration. It also inhibits microtubule assembly after binding to tubulin near the colchicine binding site [5e7]. Two of its water soluble prodrugs 3- mono sodium phosphate and 3,17-diphosphate salts have also been prepared for better bioavailability [8]. Being an important investi- gational drug, several routes of its total synthesis have also been * Corresponding author. E-mail addresses: arvindcimap@rediffmail.com, arvindcimap@gmail.com (A.S. Negi). Contents lists available at ScienceDirect European Journal of Medicinal Chemistry journal homepage: http://www.elsevier.com/locate/ejmech http://dx.doi.org/10.1016/j.ejmech.2014.09.033 0223-5234/© 2014 Elsevier Masson SAS. All rights reserved. European Journal of Medicinal Chemistry 86 (2014) 740e751