Synthesis of sulfonamides incorporating piperazinyl-ureido moieties and their carbonic anhydrase I, II, IX and XII inhibitory activity Cenzo Congiu a,⇑ , Valentina Onnis a , Alessandro Deplano a , Gianfranco Balboni a , Nurcan Dedeoglu b , Claudiu T. Supuran b,c,⇑ a Department of Life and Environmental Sciences, Unit of Pharmaceutical, Pharmacological and Nutraceutical Sciences, University of Cagliari, Via Ospedale 72, Cagliari I-09124, Italy b Università degli Studi di Firenze, Polo Scientifico, Laboratorio di Chimica Bioinorganica, Rm. 188, Via della Lastruccia 3, 50019 Sesto Fiorentino (Florence), Italy c Università degli Studi di Firenze, NEUROFARBA Department, Section of Pharmaceutical and Nutraceutical Chemistry, Via Ugo Schiff 6, 50019 Sesto Fiorentino (Florence), Italy article info Article history: Received 19 June 2015 Revised 17 July 2015 Accepted 18 July 2015 Available online 26 July 2015 Keywords: Carbonic anhydrase Sulfonamide Inhibitor Piperazine Tumor-associated isoform abstract By using SLC-0111 (4-fluorophenylureido-benzenesulfonamide), a sulfonamide carbonic anhydrase (CA, EC 4.2.1.1) inhibitor in Phase I clinical trials as an antitumor agent as lead molecule, a series of benzenesulfonamide derivatives incorporating ureido moieties was synthesized. The new compounds contain a 4-N-substituted piperazine fragment in which the ureido linker has been included, and were tested as inhibitors of the cytosolic human (h) hCA I and II isoforms, as well as the transmembrane, tumor-associated enzymes hCA IX and XII. Depending on the substitution pattern at the piperazine ring, low nanomolar inhibitors were detected against all four isoforms, making the new class of sulfonamides of interest for various pharmacologic applications. Ó 2015 Elsevier Ltd. All rights reserved. The sulfonamides are the most investigated class of inhibitors of the metallo-enzyme carbonic anhydrase (CA, EC 4.2.1.1), 1–4 with many representatives in clinical use as diuretics, antiglaucoma agents, anticonvulsants, antiobesity and more recently anti-tumor drugs. This is due to the fact that in humans (h) (and in other ver- tebrates) there are at least 15 different a-CA isoforms (hCA I–hCA XIV, with two mitochondrial enzymes hCA VA and VB), with a diverse physiologic role, catalytic activity, subcellular localization, affinity for various classes of modulators of their activity (inhibi- tors and activators), and pH optimum for the catalysis of CO 2 hydration/bicarbonate dehydration. 5–8 Antiglaucoma CA inhibitor (CAI) drugs mainly target isoforms CA II, IV and XII (involved in aqueous humor secretion within the eye); 1,2,8 the diuretics CA II, IV, XII and XIV (highly abundant in various segments of the kidneys); 1,3,6 the antiepileptics CA VII and XIV. 8 The selective inhi- bition of the CA IX and XII isoforms leads to antitumor and antime- tastatic effects, since these enzymes are highly over-expressed in hypoxic tumors, being involved in the survival of the cancer cells in acidic and hypoxic environments. 1,5 However the main drawback associated to the use of CAIs is represented by the lack of selectivity in inhibiting the various isoforms, thus resulting in a plethora of side effects. 1–4 In this contest many efforts have been made for the development of iso- form-selective CAIs, and some remarkable results have been achieved in the last 15 years since the introduction of the tail approach. 9,10 Currently a sulfonamide CA IX inhibitor (SLC-0111) entered in Phase I clinical trials for the treatment of patients with advanced solid, metastatic tumors over-expressing CA IX/XII. 11 Unlike the prototypical sulfonamide CAI acetazolamide (AAZ), SLC-0111 is a selective inhibitor for the transmembrane isoforms hCA IX/XII (over the cytosolic widespread isoforms hCA I and II). 12 N H F N H O SO 2 NH 2 S N N AcNH SO 2 NH 2 SLC-0111 AAZ The salient feature of SLC-0111 and the series of compounds to which it belongs, 12a is the presence of the ureido functionality as linker between the benzene sulfonamide fragment (which coordinates in deprotonated form at the sulfamoyl group to the zinc ion from the CA active site) and the tail of the inhibitor. By means of X-ray crystallography for adducts of SLC-0111 and other four of its congeners bound to hCA II, we demonstrated that the reasons for the isoform selectivity of these sulfonamides is the ureido linker, which allows a great flexibility to the tails http://dx.doi.org/10.1016/j.bmcl.2015.07.060 0960-894X/Ó 2015 Elsevier Ltd. All rights reserved. ⇑ Corresponding authors. Tel.: +39 070 675 8630; fax: +39 070 675 8612 (C.C.); tel.: +39 055 4573005; fax: +39 055 4573385 (C.T.S.). E-mail addresses: ccongiu@unica.it (C. Congiu), claudiu.supuran@unifi.it (C.T. Supuran). Bioorganic & Medicinal Chemistry Letters 25 (2015) 3850–3853 Contents lists available at ScienceDirect Bioorganic & Medicinal Chemistry Letters journal homepage: www.elsevier.com/locate/bmcl