Original article Nonclassical antifolates, part 5. Benzodiazepine analogs as a new class of DHFR inhibitors: Synthesis, antitumor testing and molecular modeling study q Hussein I. El-Subbagh a, * , Ghada S. Hassan b , Shahenda M. El-Messery c , Sarah T. Al-Rashood d , Fatmah A.M. Al-Omary d , Yasmin S. Abulfadl e , Marwa I. Shabayek e a Department of Pharmaceutical Chemistry, Faculty of Pharmaceutical Sciences & Pharmaceutical Industries, Future University,12311 Cairo, Egypt b Department of Medicinal Chemistry, Faculty of Pharmacy, Mansoura University, P.O. Box 35516, Mansoura, Egypt c Department of Pharmaceutical Organic Chemistry, Faculty of Pharmacy, Mansoura University, P.O. Box 35516, Mansoura, Egypt d Department of Pharmaceutical Chemistry, College of Pharmacy, King Saud University, P.O. Box 2457, Riyadh 11451, Saudi Arabia e Department of Pharmacology (Biochemistry Section), Faculty of Pharmaceutical Sciences & Pharmaceutical Industries, Future University,12311 Cairo, Egypt article info Article history: Received 15 November 2013 Received in revised form 3 January 2014 Accepted 6 January 2014 Available online 13 January 2014 Keywords: Synthesis Tetrahydro-quinazolines Dibenzo[b,e][1,4]diazepines DHFR inhibition Molecular modeling study abstract A new series of tetrahydro-quinazoline and tetrahydro-1H-dibenzo[b,e][1,4]diazepine analogs were synthesized and tested for their DHFR inhibition and in vitro antitumor activity. Compound 35 showed a remarkable DHFR inhibitory potency (IC 50 , 0.004 mM) which is twenty fold more active than metho- trexate (MTX). Compounds 17 and 23 proved to be fifteen fold more active than the known antitumor 5- FU, with MG-MID GI 50 , TGI, and LC 50 values of 1.5, 46.8, 93.3 and 1.4,17.4, 93.3 mM, respectively. Com- puter modeling studies allowed the identification that methoxy and methyl substituents, the p-system of the chalcone core, the nitrogen atoms, on the dibenzodiazepine ring as pharmacophoric features essential for activity. These mark points could be used as template model for further future optimization. Ó 2014 Elsevier Masson SAS. All rights reserved. 1. Introduction Dihydrofolate reductase (DHFR) is the key enzyme in folate metabolism; it plays a major role in the biosynthesis of nucleic acids through the catalysis of the NADPH reduction of 7,8- dihydrofolate to 5,6,7,8-tetrahydrofolate and intimately couples with thymidylate synthase for purine and pyrimidine production [1,2]. As a result, DHFR becomes an important target for anticancer and antimicrobial agents [3]. Pre-clinical and clinical studies identified a plethora of mechanisms of drug resistance that are the primary hindrance for the allocation of curative cancer chemo- therapy. Drug discovery of novel antifolates with improved prop- erties and superior activities remains an attractive strategy both in academic research and in the pharmaceutical industry [3,4]. Many quinazoline analogs are known to possess anticancer activity through DHFR inhibition as non-classical antifolates such as tri- metrexate (TMQ) and piritrexim (PTX), Chart 1 [5e11]. On the other hand, benzodiazepine derivatives have been reported to possess antiviral, antimicrobial, and antitumor activities [12]. Meanwhile, the a,b-unsaturated ketone function condensed with variety of cyclizing agents such as hydrazine hydrate, thiourea, malononitrile, ethylcyanoacetate, and 2-aminothiazoles produced compounds possessing broad spectrum antineoplastic activity against variety of tumor cell lines [13e18]. Recently, a new series of substituted-quinazolin-4-ones was designed, synthesized, and evaluated for their in vitro DHFR inhi- bition in our laboratories. This study allowed the allocation of active DHFR inhibitors with IC 50 values around 0.4 mM, bearing aromatic p-systems. Molecular modeling study of this class of compounds revealed the importance of the main pharmacophoric groups (the 4-carbonyl fragment, the basic nitrogen atom at N-1, and the hy- drophobic p-system regions) as well as of their relative spatial distances in regard to the quinazoline nucleus. The substitution pattern and spatial considerations of the p-systems proved to be critical for DHFR inhibition [19e21]. q For parts 1e4 see Refs. [19e22]. * Corresponding author. Tel./fax: þ20 2 26186111. E-mail address: subbagh@yahoo.com (H.I. El-Subbagh). Contents lists available at ScienceDirect European Journal of Medicinal Chemistry journal homepage: http://www.elsevier.com/locate/ejmech 0223-5234/$ e see front matter Ó 2014 Elsevier Masson SAS. All rights reserved. http://dx.doi.org/10.1016/j.ejmech.2014.01.004 European Journal of Medicinal Chemistry 74 (2014) 234e245