Original article Design, synthesis and in vitro anti-proliferative activity of 4,6-quinazolinediamines as potent EGFR-TK inhibitors Samar Mowafy a , Nahla A. Farag a , Khaled A.M. Abouzid b, * a Pharmaceutical Chemistry Department, Faculty of Pharmacy, Misr International University, Abbassia,11566 Cairo, Egypt b Pharmaceutical Chemistry Department, Faculty of Pharmacy, Ain Shams University, Abbassia,11566 Cairo, Egypt article info Article history: Received 2 February 2012 Received in revised form 24 September 2012 Accepted 11 October 2012 Available online 27 October 2012 Keywords: 4-Anilinoquinazoline EGFR-TK inhibitors Docking study Anti-proliferative activity abstract 4-Anilino-6-substituted-quinazolines were designed, synthesized and evaluated for EGFR-TK and tumor growth inhibitory activities. The target compounds were designed with enamine ester or urea moieties appended at the C-6 of quinazoline as additional hydrogen bond acceptor functions. Most of the synthesized compounds displayed potent EGFR-TK inhibitory activity at 10 mM and the 6-ureido-anili- noquinazoline derivative 7a showed IC 50 value of 0.061 mM. Moreover, six compounds were tested by National Cancer Institute (NCI), USA for their anti-proliferative activity at 10 mM in full NCI 60 cell panel. Compound 7a was further assayed for five dose molar ranges in full NCI 60 cell panel and exhibited remarkable growth inhibitory activity pattern against Non-Small Cell Lung Cancer EKVX (GI 50 ¼ 0.37 mM), NCI-H322M (GI 50 ¼ 0.36 mM), Renal Cancer A498 (GI 50 ¼ 0.46 mM), TK-10 (GI 50 ¼ 0.99 mM) and Breast Cancer MDA-MB-468 (GI 50 ¼ 1.096 mM) which are of high EGFR expres- sion. Docking study was performed for the active compounds into ATP binding site of EGFR-TK which showed similar binding mode to gefitinib and additional binding with Cys-773 at the gatekeeper of EGFR-TK enzyme. Ó 2012 Elsevier Masson SAS. All rights reserved. 1. Introduction The epidermal growth factor receptor (EGFR) protein belongs to the ErbB family of receptor tyrosine kinases (RTKs) which plays an important role in the regulation of cell growth, differentiation, and survival [1]. EGFR is a glycoprotein that contains an extracellular ligand-binding domain, a transmembrane region, and an intracel- lular domain with kinase activity [2]. EGFR is over expressed in several human tumors including Non-Small Cell Lung Cancer (NSCLC), prostate, breast and ovarian cancers [3e10] and correlates with a poor prognosis in many cancer patients [11]. Thus, EGFR is an attractive target for the design and development of compounds that can specifically bind to the receptor and inhibit its tyrosine kinase (TK) activity and its signal transduction pathway in cancer cells. Inhibition of EGFR has been achieved through two main approaches: by blocking ligand binding to the extracellular domain with monoclonal antibodies and by using small-molecule inhibitors that interact at the ATP-binding site [12]. A well-studied class of these latter inhibitors is represented by 4-anilinoquinazolines, exempli- fied by the lead compound gefitinib, erlotinib which are approved by the FDA for the treatment of advanced Non-Small Cell Lung Cancer [13], Fig. 1 . These agents belong to the 4-anilinoquinazoline class of inhibitors and the key features between the receptor and this template have been revealed as follows [14,15]. (1) The quinazoline core fits into the ATP binding pocket of the kinase domain, where the N-1 nitrogen of the quinazoline nucleus interacts with the backbone NH of Met-769 via a hydrogen bond, and water mediated hydrogen bonding is observed between the N-3 of the quinazoline and the Thr-766 side chain. (2) The aniline ring fills an adjacent lipophilic pocket. (3) The solubilizing side chains at C-6 and/or C-7 of the quinazoline core act to improve physical properties and confer a more favorable pharmacokinetic profile. In the past decade, the SAR study of quinazoline as EGFR inhibitors was directed toward the modification of the C-4 aryla- mino group [16], by varying the aniline substitution pattern, fine tuning of the kinase selectivity profile can be achieved [17]. Also, the substitution at C-6 position of quinazoline has received increasing attention in the development of more potent & selective inhibition [9,17]. Almost all substituents in this position were designed to bind with Cys-773 in EGFR through hydrogen bonding or covalent bonding [16,18,19]. Since 6-position of quinazoline points toward the outside of the protein & theoretical studies have indicated that bulky substituents are tolerated at the 6-position. Moreover the 6-position substitution allows optimum distance for * Corresponding author. Tel.: þ20 1222165624; fax: þ20 225080728. E-mail address: Khaled.abouzid@pharm.asu.edu.eg (K.A.M. Abouzid). Contents lists available at SciVerse ScienceDirect European Journal of Medicinal Chemistry journal homepage: http://www.elsevier.com/locate/ejmech 0223-5234/$ e see front matter Ó 2012 Elsevier Masson SAS. All rights reserved. http://dx.doi.org/10.1016/j.ejmech.2012.10.017 European Journal of Medicinal Chemistry 61 (2013) 132e145