Carbonic anhydrases (CAs, EC 4.2.1.1) are a family of enzymes widespread in all life kingdoms. In mammals, isozyme CA IX is highly overexpressed in many cancer types being present in few normal tissues. Its expression is strongly induced by hypoxia present in many tumors, being regulated by the HIF transcription factor and correlated with a poor response to classical chemo- and radiotherapies. CA IX was recently shown to contribute to acidification of the tumor environment, by efficiently catalyzing the hydration of carbon dioxide to bicarbonate and protons with its extracellularly situated active site, leading both to the acquisition of metastasic phenotypes and to chemoresistance with weakly basic anticancer drugs. Inhibition of this enzymatic activity by specific and potent inhibitors was shown to revert these acidification processes, establishing a clear-cut role of CA IX in tumorigenesis. The development of a wide range of potent and selective CA IX inhibitors belonging to diverse chemical classes, such as membrane-impermeant, fluorescent or metal-containing such agents, could thus provide useful tools for highlighting the exact role of CA IX in hypoxic cancers, to control the pH (im)balance of tumor cells, and to develop novel diagnostic or therapeutic applications for the management of tumors. Indeed, both fluorescent inhibitors or positively charged, membrane impermeant sulfonamides have been recently developed as potent CA IX inhibitors and used as proof-of-concept tools for demonstrating that CA IX constitutes a novel and interesting target for the anticancer drug development.

The carbonic anhydrases (CAs, EC 4.2.1.1) are ubiquitous metalloenzymes with five independently evolved (R, ,γ,δ,ς) classes reported up to date. [1][2][3] These enzymes catalyze the reversible hydration of carbon dioxide to bicarbonate and protons by means of a metalhydroxide (Lig 3 M 2+ (OH) -) mechanism. 2 In addition to the established role of the carbonic anhydrase inhibitors (CAIs) as diuretics and antiglaucoma drugs, it has recently emerged that CAIs could have potential as novel antiobesity, anticancer, and anti-infective drugs. 1,2 A critical problem in the design of CAIs is related to the high number of isoforms in mammals (15), their diffuse localization in many tissues/organs, and the lack of isozyme selectivity of the presently available inhibitors. [1][2][3][4] Recently, nanoparticles received great attention for their biomedical applications both for the site-specific delivery of drugs 7 or for imaging purposes. 7,8 Several recent such examples include HIV inhibition with a CCR5 antagonist attached to multivalent gold nanoparticles (Au NPs), tumor targeting with NPs loaded with hydroxycamptothecin, paclitaxel, or fumagillin, and magnetic resonance imaging (MRI) techniques of tumors based on integrins targeting with Fe 3 O 4 NPs, computer tomography (CT) or MRI imaging with gadolinium chelate Au NPs, etc. 7,8 Human CA IX (hCA IX) is an extracellular, transmembrane isoform which was recently shown to constitute a novel and interesting target for the anticancer therapy due to its overexpression in many cancer tissues and not in their normal counterparts. 1b,9 Its expression is strongly induced by hypoxia present in many tumor tissues and correlated with a bad response to classical chemo-and radiotherapies. 1b,9 CA IX was shown to acidify the extratumoral medium leading both to the acquisition of metastasic phenotypes and to chemoresistance with many anticancer drugs, these processes being reverted by inhibition of the enzyme catalytic activity with sulfonamide inhibitors. 1b,9 The development of selective CA IX inhibitors might provide useful tools for highlighting the exact role of CA IX in hypoxic cancers, to control the pH imbalance of tumor cells and lead to novel diagnostic or therapeutic applications for the management of such tumors. 1b,9 Although many sulfonamide/sulfamate/sulfamide potent CA IX inhibitors were reported, few of them show an acceptable level of selectivity for inhibiting the transmembrane-tumor associated target isoform IX over the cytosolic, ubiquitous isozymes hCA I and II. 1b,9 Considering the extracellular localization of the target CA isoform and the fact that nanomaterials generally show membrane impermeability, 10 we report here the synthesis of CAI coated Au NPs which show excellent CA IX inhibitory properties and selectivity for the inhibition of the tumor-associated isoform over hCA I and II.The key intermediate 3a was synthesized by coupling lipoic acid 1 with 4-aminoethylbenzene sulfonamide 2a in the presence of EDCI/ DMAP, as outlined in Scheme 1. The CAI coated...

A series of nitroimidazoles incorporating sulfonamide/sulfamide/sulfamate moieties were designed and synthesized as radio/chemosensitizing agent targeting the tumor-associated carbonic anhydrase (CA) isoforms IX and XII. Most of the new compounds were nanomolar inhibitors of these isoforms. Crystallographic studies on the complex of hCA II with the lead sulfamide derivative of this series clarified the binding mode of this type of inhibitors in the enzyme active site cavity. Some of the best nitroimidazole CA IX inhibitors showed significant activity in vitro by reducing hypoxia-induced extracellular acidosis in HT-29 and HeLa cell lines. In vivo testing of the lead molecule in the sulfamide series, in cotreatment with doxorubicin, demonstrated a chemosensitization of CA IX containing tumors. Such CA inhibitors, specifically targeting the tumor-associated isoforms, are candidates for novel treatment strategies against hypoxic tumors overexpressing extracellular CA isozymes.