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Fibroblast activation protein (FAP) is a very reliable biomarker for tissue remodeling. FAP has so far mainly been studied in oncology, but there is growing interest in the enzyme in other diseases like fibrosis. Recently, FAP-targeting diagnostics and therapeutics have emerged, of which the so-called FAPIs are among the most promising representatives. FAPIs typically have a relatively high molecular weight and contain very polar, multicharged chelator moieties. While this is not limiting the application of FAPIs in oncology, more druglike FAPIs could be required to optimally study diseases characterized by denser, less permeable tissue. In response, we designed the first druglike 18 F-labeled FAPIs. We report target potencies, biodistribution, and pharmacokinetics and demonstrate FAP-dependent uptake in murine tumor xenografts. Finally, this paper puts forward compound 10 as a highly promising, druglike FAPI for 18 F-PET imaging. This molecule is fit for additional studies in fibrosis and its preclinical profile warrants clinical investigation.

The ability to image early treatment response to radiotherapy in head and neck squamous cell carcinoma (HNSCC) will enable the identification of radioresistant tumor volumes suitable for treatment intensification. Here, we propose the system x c - radiotracer (4 S )-4-(3-[ 18 F]fluoropropyl)-L-glutamate ([ 18 F]FSPG) as a non-invasive method to monitor radiation response in HNSCC. We assessed temporal changes in cell death, antioxidant status, and [ 18 F]FSPG retention following a single dose of 10 Gy irradiation in FaDU HNSCC cells. Next, using a fractionated course of radiotherapy, we assessed tumor volume changes and performed [ 18 F]FSPG-PET imaging in FaDU-bearing mouse xenografts, followed by ex vivo response assessment. In cells, 10 Gy irradiation reduced [ 18 F]FSPG retention, coinciding with the induction of apoptosis and the production of reactive oxygen species. In vivo, [ 18 F]FSPG tumor retention was halved seven days after the start of treatment, which preceded radiotherapy-induced tumor shrinkage, thereby confirming [ 18 F]FSPG-PET as an early and sensitive marker of radiation response., Competing Interests: Competing interestsT.H.W. is the Editor-in-Chief of npj Imaging. He was blinded to this manuscript's editorial and peer review process. All other authors declare no competing financial or non-financial interests., (© The Author(s) 2024.)

Mouse models are invaluable tools for radiotracer development and validation. They are, however, expensive, low throughput, and are constrained by animal welfare considerations. Here, we assessed the chicken chorioallantoic membrane (CAM) as an alternative to mice for preclinical cancer imaging studies. NCI-H460 FLuc cells grown in Matrigel on the CAM formed vascularized tumors of reproducible size without compromising embryo viability. By designing a simple method for vessel cannulation it was possible to perform dynamic PET imaging in ovo, producing high tumor-to-background signal for both 18 F-2-fluoro-2-deoxy-D-glucose ( 18 F-FDG) and (4S)-4-(3- 18 F-fluoropropyl)-L-glutamate ( 18 F-FSPG). The pattern of 18 F-FDG tumor uptake were similar in ovo and in vivo, although tumor-associated radioactivity was higher in the CAM-grown tumors over the 60 min imaging time course. Additionally, 18 F-FSPG provided an early marker of both treatment response to external beam radiotherapy and target inhibition in ovo. Overall, the CAM provided a low-cost alternative to tumor xenograft mouse models which may broaden access to PET and SPECT imaging and have utility across multiple applications., Competing Interests: Competing Interests The authors declare no competing interests.

Resistance against anti-cancer therapy is one of the major challenges during treatment of multiple cancers. Gemcitabine is a standard first-line chemotherapeutic drug, yet autophagy is highly activated in the hypoxic microenvironment of solid tumors and enhances the survival of tumor cells against gemcitabine chemotherapy. Recently, we showed the add-on effect of autophagy inhibitor UAMC-2526 to prevent HT-29 colorectal tumor growth in CD1 -/- Foxn1nu mice treated with oxaliplatin. In this study, we aimed to investigate the potential beneficial effects of UAMC-2526 in a syngeneic Panc02 mouse model of pancreatic ductal adenocarcinoma (PDAC). Our data showed that UAMC-2526 combined with gemcitabine significantly reduced tumor growth as compared to the individual treatments. However, in contrast to in vitro experiments with Panc02 cells in culture, we were unable to detect autophagy inhibition by UAMC-2526 in Panc02 tumor tissue, neither via western blot analysis of autophagy markers LC3 and p62, nor by transmission electron microscopy. In vitro experiments revealed that UAMC-2526 enhances the potential of gemcitabine to inhibit Panc02 cell proliferation without obvious induction of cell death. Altogether, we conclude that although the combination treatment of UAMC-2526 with gemcitabine did not inhibit autophagy in the Panc02 mouse model, it has a beneficial effect on tumor growth inhibition., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Takhsha, Vangestel, Tanc, De Bruycker, Berg, Pintelon, Stroobants, De Meyer, Van Der Veken and Martinet.)

Fibroblast activation protein (FAP) is a proline-selective protease that belongs to the S9 family of serine proteases. It is typically highly expressed in the tumor microenvironment (TME) and especially in cancer-associated fibroblasts, the main cell components of the tumor stroma. The exact role of its enzymatic activity in the TME remains largely unknown. Hence, tools that enable selective, activity-based visualization of FAP within the TME can help to unravel FAP's function. We describe the synthesis, biochemical characterization, and application of three different activity-based probes (biotin-, Cy3-, and Cy5-labeled) based on the FAP-inhibitor UAMC1110, an in-house developed molecule considered to be the most potent and selective FAP inhibitor available. We demonstrate that the three probes have subnanomolar FAP affinity and pronounced selectivity with respect to the related S9 family members. Furthermore, we report that the fluorescent Cy3- and Cy5-labeled probes are capable of selectively detecting FAP in a cellular context, making these chemical probes highly suitable for further biological studies. Moreover, proof of concept is provided for in situ FAP activity staining in patient-derived cryosections of urothelial tumors., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Van Rymenant, Tanc, Van Elzen, Bracke, De Wever, Augustyns, Lambeir, Kockx, De Meester and Van Der Veken.)

Autophagy is an intracellular degradation/recycling pathway that provides nutrients and building blocks to cellular metabolism and keeps the cytoplasm clear of obsolete proteins and organelles. During recent years, dysregulated autophagy activity has been reported to be a characteristic of many different disease types, including cancer and neurodegenerative disorders. This has created a strong case for development of autophagy modulating compounds as potential treatments for these diseases. Inhibitors of autophagy have been proposed as a therapeutic intervention in, e.g., advanced cancer, and inhibiting the cysteine protease Atg4B has been put forward as a main strategy to block autophagy. We recently identified and demonstrated -both in vitro and in vivo - that compounds with a benzotropolone basic structure targeting Atg4B, can significantly slow down tumor growth and potentiate the effect of classical chemotherapy. In this study we report the synthesis and inhibition profile of new benzotropolone derivatives with additional structural modifications at 6 different positions. To obtain a solid inhibition profile, all compounds were evaluated on three levels, including two cell-based assays to confirm autophagy and intracellular Atg4B inhibition and an SDS-PAGE-based experiment to assess in vitro Atg4B affinity. Several molecules with a promising profile were identified., (Copyright © 2019 Elsevier Inc. All rights reserved.)

The reaction mechanism of the carbonic anhydrase-mediated hydrolysis of sulfocoumarins to sulfonic acids has been investigated on an enzyme cluster model using the B3LYP hybrid density functional theory (DFT) and the QST procedure for the Transition State (TS) search. A multistep process was highlighted, with the rate-determining step identified in the initial dual nucleophilic/acidic attack of the zinc-bound hydroxide ion to the sulfocoumarin sulfur atom and to the C3=C4 double bond. The reported multi-step process, combined to SAR analysis on a new set of derivatives, highlighted unprecedented mechanistic aspects of the CA-mediated prodrug activation, which in turn possess relevant consequences to the isoforms-selective inhibition profiles reported by such a class of compounds., (© 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

A series of ureido and bis-ureido derivatives were prepared by reacting histamine with alkyl/aryl-isocyanates or di-isocyanates. The obtained derivatives were assayed as activators of the enzyme carbonic anhydrase (CA, EC 4.2.1.1), due to the fact that histamine itself has this biological activity. Although inhibition of CAs has pharmacological applications in the field of antiglaucoma, anticonvulsant, anticancer, and anti-infective agents, activation of these enzymes is not yet properly exploited pharmacologically for cognitive enhancement or Alzheimer's disease treatment, conditions in which a diminished CA activity was reported. The ureido/bis-ureido histamine derivatives investigated here showed activating effects only against the cytosolic human (h) isoform hCA I, having no effect on the widespread, physiologically dominant isoform hCA II. This is the first report in which CA I-selective activators were identified. Such compounds may constitute interesting tools for better understanding the physiological/pharmacological effects connected to activation of this widespread CA isoform, whose physiological function is not fully understood.

In this paper, we describe the discovery of the 3-hydroxyquinazoline-2,4-dione as a useful scaffold to obtain potent inhibitors of the tumor-associated human carbonic anhydrases (hCAs) IX and XII. A set of derivatives (1-29), bearing different substituents on the fused benzo ring (Cl, NO 2 , NH 2 , CF 3 , ureido, amido, heterocycles), were synthesized, and several of them showed nanomolar activity in inhibiting the hCA IX and XII isoforms, while they were ineffective against the cytosolic enzymes hCAs I and II. Some selected compounds were tested for their antiproliferative activity against HT-29 colon cancer cell lines. After 48 h of treatment with the lower dose (30 μM), derivatives 12, 14, 15, and 19 were significantly active, inducing a mortality by about 50% in both normoxia and hypoxia. This finding led us to hypothesize for these compounds more than one mechanism of action involving both CAs IX and XII and other not yet identified target(s).

Multivalency has impressive effects on (bio)molecular recognition, through the simultaneous presentation of multiple copies of a ligand, which can change a weak millimolar binder into a potent nanomolar one. The implementation of multivalency in enzyme inhibition is rather recent, being exemplified by few serendipitous discoveries, and hitherto relying on the random exploration of new multivalent structures as potential enzyme inhibitors. Here, a straightforward and versatile method is reported that enables the construction of multivalent systems for the inhibition of carbonic anhydrases (CA), widespread enzymes that catalyze a fundamental biochemical reaction. Oxime and hydrazone click-type bioconjugation techniques were successfully used for the preparation of tetravalent peptide conjugates tethered with sulfonamide CA inhibitors. The enzyme inhibition assays show that multivalent effects were present with these novel compounds, but also reveal various structural effects provided by the scaffolds. The versatility of this approach may facilitate the exploration of structure-activity relationships for other types of enzyme inhibitors., (© 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

New phenolic mono and bis Mannich bases incorporating benzimidazole, such as 2-(aminomethyl)-4-(1H-benzimidazol-2-yl)phenol and 2,6-bis(aminomethyl)-4-(1H-benzimidazol-2-yl)phenol were synthesized starting from 4-(1H-benzimidazol-2-yl)phenol. Amines used for the synthesis included dimethylamine, pyrrolidine, piperidine, N-methylpiperazine and morpholine. The CA inhibitory properties of these compounds were tested on the human carbonic anhydrase (CA, EC 4.2.1.1) isoforms hCA I and hCA II. These new compounds, as many phenols show moderate CA inhibitory properties.

N-protected amino acids (Gly, Ala and Phe protected with Boc and Z groups) were reacted with sulfonamide derivatives, leading to the corresponding N-protected amino acid-sulfonamide conjugates. The carbonic anhydrase (CA, EC 4.2.1.1) inhibitory activity of the new compounds was assessed against four human (h) isoforms, hCA I, hCA II, hCA IV and hCA XII. Among them, hCA II, IV and XII are antiglaucoma drug targets, being involved in aqueous humor secretion within the eye. Low nanomolar inhibition was measured against all four isoforms with the 20 reported sulfonamides, but no selective inhibitory profiles, except for some CA XII-selective derivatives, were observed. hCA I, II and XII were generally better inhibited by sulfonamides incorporating longer scaffolds and Gly/Ala, whereas the best hCA IV inhibitors were homosulfanilamide derivatives, incorporating Phe moieties. The amino acid-sulfonamide conjugates show good water solubility and effective hCA II, IV and XII inhibition, and may be considered as interesting candidates for antiglaucoma studies.

Mannich bases of thymol were synthesized. The aminomethylation reaction was realised in the ortho position of the phenol for compounds 2 (dipropylamine), 3 (benzylamine), and 4 (dibenzylamine) while it was from para position for 1 (dimethylamine), 5 (piperidine), 6 (morpholine) and 7 (N-methylpiperazine). The carbonic anhydrase (CA, EC 4.2.1.1) inhibitory effects of the compounds were asssessed against hCA I and hCA II. All compounds moderately inhibited hCA I and hCA II. The cytotoxicity of the compounds against four human oral squamous cell carcinoma cell lines were compared those against three normal oral cells. Tumor specificity values were about 2 or slightly more for the compounds 2, 3, 4, 5 and 6. Compound 2 showed cytostatic activity against OSCC cell lines at 16 to 32-fold lower concentrations as compared with normal cells. This suggests that compound 2 can be considered as cytotoxicity enhancing drug candidate for further investigations.

N-protected amino acids were reacted with substituted benzothiazoles to give the corresponding N-protected amino acid-benzothiazole conjugates (60-89%). Their structures were confirmed by proton nuclear magnetic resonance ((1)H NMR), carbon-13 nuclear magnetic resonance ((13)C NMR), IR and elemental analysis. Their carbonic anhydrase (CA, EC 4.2.1.1) inhibitory activities were determined against two cytosolic human isoforms (hCA I and hCA II), one membrane-associated (hCA IV) and one transmembrane (hCA XII) enzyme by a stopped-flow CO2 hydrase assay method. The new compounds showed rather weak, micromolar inhibitory activity against most of these enzymes.

Phenolic mono Mannich bases [2-[4-hydroxy-3-(aminomethyl)benzylidene]-2,3-dihydro-1H-inden-1-one (8-15)] and bis Mannich bases [2-[4-hydroxy-3,5-bis(aminomethyl)benzylidene]-2, 3-dihydro-1H-inden-1-one (2-7)] were synthesized starting from 2-(4-hydroxybenzylidene)-2, 3-dihydro-inden-1-one (1). This study was designed in order to investigate the carbonic anhydrase (CA, EC 4.2.1.1) inhibitory properties of a library of compounds incorporating the phenol functional group. All prepared compounds showed a low inhibition percentages on both human (h) isoforms hCA I and hCA II compared to the reference sulfonamide acetazolamide. Mannich bases 2-15 had lower inhibition percentages than the compound 1 on hCA I and hCA II, except compound 14, which is a Mannich base derivative of dipropylamine, which had a similar inhibitory power as compound 1 on hCA II. All compounds synthesized 1-15 were 1.3-1.9 times more effective on hCA II comparing with the effectivenes of the compounds on hCA I.

N-Protected amino acids (Gly, Ala and Phe) were reacted with amino substituted coumarin and quinolinone derivatives, leading to the corresponding N-protected amino acid-coumarin/quinolinone conjugates. The carbonic anhydrase (CA, EC 4.2.1.1) inhibitory activity of the new compounds was assessed against various human (h) isoforms, such as hCA I, hCA II, hCA IV and hCA XII. The quinolinone conjugates were inactive as enzyme inhibitors, whereas the coumarins were ineffective hCA I/II inhibitors (KIs > 50 μM) but were submicromolar hCA IV and XII inhibitors, with inhibition constants ranging between 92 nM and 1.19 μM for hCA IV, and between 0.11 and 0.79 μM for hCA XII. These coumarin derivatives, as many others reported earlier, thus show an interesting selective inhibitory profile for the membrane-bound over the cytosolic CA isoforms.

N-substituted maleimides were synthesized from maleic anhydride and primary amines. 1,4-Dibromo-dibenzo[e,h]bicyclo-[2,2,2]octane-2,3-dicarboximide derivatives (4a-f) were prepared by the [4+2] cycloaddition reaction of dibromoanthracenes with the N-substituted maleimide derivatives. The carbonic anhydrase (CA, EC 4.2.1.1) inhibitory effects of the new derivatives were assayed against the human (h) isozymes hCA I, II, IX, and XII. All tested bicyclo dicarboximide derivatives exhibited excellent inhibitory effects in the nanomolar range, with Ki values in the range of 117.73-232.87 nM against hCA I and of 69.74-111.51 nM against hCA II, whereas they were low micromolar inhibitors against hCA IX and XII., (© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

A series of Mannich bases having piperidine moiety were reacted with 2-mercaptoethanol, leading to 1-aryl-3-piperidine-4-yl-1-propanone hydrochlorides. The cytotoxicity and carbonic anhydrase inhibitory activities of these new compounds were evaluated. Among the compounds, only one derivative, nitro substituent bearing EU9, showed an effective cytotoxicity, although weak tumor specificity against human oral malignant versus nonmalignant cells. The compound induced apoptosis in HSC-2 oral squamous cell carcinoma cells, but not in human gingival fibroblast. Chemical modifications of this lead are thus necessary to further investigate it as a drug candidate and to obtain compounds with a better activity profile.

New series of benzenesulfonamide derivatives incorporating pyrazole and isatin moieties were prepared using celecoxib as lead molecule. Biological evaluation of the target compounds was performed against the metalloenzyme carbonic anhydrase (CA, EC 4.2.1.1) and more precisely against the human isoforms hCA I, II (cytosolic), IX and XII (transmembrane, tumor-associated enzymes). Most of the tested compounds efficiently inhibited hCA I, II and IX, with KIs of 2.5-102 nM, being more effective than the reference drug acetazolamide. Compounds 11e, 11f, 16e and 16f were found to inhibit hCA XII with Ki of 3.7, 6.5, 5.4 and 7.2 nM, respectively. Compounds 11e and 16e, with 5-NO2 substitution on the isatin ring, were found to be selective inhibitors of hCA IX and hCA XII. Docking studies revealed that the NO2 group of both compounds participate in interactions with Asp132 within the hCA IX active site, and with residues Lys67 and Asp130 in hCA XII, respectively., (Copyright © 2015 Elsevier Masson SAS. All rights reserved.)

6-Substituted sulfocoumarins bearing the carboxamido, trimethylammonium as well as the cyano and methoxy moieties with interesting inhibitory activity/selectivity against the tumor associated carbonic anhydrase (CA, EC 4.2.1.1) isoforms hCA IX and XII are reported. Moieties leading to the best inhibition were tert-butylcarboxamido, phenylcarboxamido, and 4-pyridylcarboxamido, with K(I) values of 2.1-8.1 nM. No inhibition of the off-target hCA II and I was observed. A number of these compounds were evaluated against HT-29 colon cancer cell lines ex vivo. Compounds 9c and 9e revealed effective cytotoxic effects after 72 h of incubation in both normoxic and hypoxic conditions, unlike sulfonamide CA inhibitors that show such effects only in hypoxia. These results may be of particular importance for the choice of future drug candidates targeting hypoxic tumors and metastases, considering the fact that a sulfonamide CA IX inhibitor (SLC-0111) is presently in phase I clinical trials.

1-N-Alkylated-6-sulfamoyl saccharin derivatives were prepared and assayed as carbonic anhydrase inhibitors (CAIs). During X-ray crystallographic experiments an unexpected hydrolysis of the isothiazole ring was evidenced which allowed us to prepare highly potent enzyme inhibitors with selectivity for some isoforms with medical applications.

A series of 6-aryl-substituted 1,2-benzoxathiine 2,2-dioxides was obtained by reacting 6-iodo-sulfocoumarin with arylboronic acids in Suzuki cross-coupling conditions. The new sulfocoumarins incorporating various substituted phenyl moieties in position 6 of the heterocyclic ring were investigated for the inhibition of four human (h) carbonic anhydrase (CA, EC 4.2.1.1) isoforms with medicinal chemistry applications, the cytosolic hCA I and II, and the transmembrane, tumor-associated hCA IX and XII. The aryl-substituted sulfocoumarins did not inhibit the ubiquitous, off-target cytosolic isoforms hCA I and II (KIs>10μM) but showed effective inhibition against the two transmembrane CAs, with KIs ranging from 9.0 to 95.3nM against hCA IX, and between 3.5 and 14.2nM against hCA XII. As hCA IX and XII are validated anti-tumor targets, such sulfocoumarin, isoform-selective inhibitors may be useful for identifying suitable drug candidates for further clinical trials of this class of pharmacologic agents., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Cyclopropylcarboxylic acids and esters and cyclopropylmethanols incorporating bromophenol moieties were investigated as inhibitors of the carbonic anhydrase enzyme (CA; EC 4.2.1.1). The cis- and trans-esters 5 and 6 were obtained from the reaction of 4-allyl-1,2-dimethoxybenzene (4) with ethyl diazoacetate, which after bromination with Br2 gave two isomeric monobromides (11 and 15), four isomeric dibromides (12, 13, 16, and 17), and two isomeric tribromides (14 and 18). The carboxylic acids 7, 8, and 19-26 were thereafter obtained by hydrolysis of the synthesized esters. All these bromophenol derivatives were tested against human (h) CA isoenzymes I and II (cytosolic, ubiquitous isoforms) and hCA IX and XII (transmembrane, tumor-associated enzymes). All tested bromophenols exhibited excellent inhibitory effects, in the low nanomolar range, with Ki values in the range of 0.54-59 nM against hCA I and in the range of 0.97-12.14 nM against hCA II, whereas they were low micromolar inhibitors against hCA IX and XII. The best hCA I inhibition was observed in new bromophenol derivative 20 (Ki = 0.54 nM). On the other hand, new bromophenol derivative 12 showed a powerful inhibition effect against hCA II (Ki = 0.97 nM).

The α-carbonic anhydrases (CAs, EC 4.2.1.1) show catalytic versatility acting as esterases with carboxylic, sulfonic, and phosphate esters. Here we prove by kinetic, spectroscopic, and MS studies that they also possess thioesterase activity with a dithiocarbamate ester as a substrate (PhSO2NHCSSMe). Its CA-mediated hydrolysis leads to benzenesulfonamide, methyl mercaptan, and COS. The CA thioesterase activity may be useful for designing prodrug enzyme inhibitors, whereas some CA isoforms may use this activity for modulating physiologic/pathologic processes, which are possibly amenable to drug discovery of agents with multiple mechanisms of action.

A series of 6-substituted 2-benzoxathiine-2,2-dioxides were synthesized starting from 2,5-dihydroxybenzaldehyde, and then screened in vitro for their inhibition properties against five human carbonic anhydrase (hCA, EC 4.2.1.1) isoforms. All the compounds showed excellent selectivity against the mitochondrial (hCA VA) and the tumor associated (hCA IX and XII) enzymes.

A series of substituted pyrrolidines and piperidines were synthesized using superacid HF/SbF5 chemistry. Investigated as inhibitors of several human carbonic anhydrase (hCA, EC 4.2.1.1) isoforms, i.e. the cytosolic hCA I and II as well as the tumor-associated transmembrane isoforms hCA IX and XII, these compounds showed a never yet reported selectivity toward the human carbonic anhydrase hCA II. In the tertiary benzenesulfonamide family, this class of inhibitors points out a new mechanism of action for human carbonic anhydrase II inhibition.

Here we describe the screening via Dynamic Deconvolution of DCLs of inhibitors (CAIs) and activators (CAAs) of hCA I. The inhibitory effects dominate over the activating ones, while the CAAs may be identified in the absence of CAIs.

A series of 6-substituted sulfocoumarins incorporating substituted-1,2,3-triazol-4-yl-/5-yl moieties were synthesized by employing click chemistry. The new sulfocoumarins incorporated cycloalkyl, tert-butyl and substituted aryl moieties at the triazole ring, and were investigated for the inhibition of four human (h) carbonic anhydrase (hCA, EC 4.2.1.1) isoforms, the cytosolic hCA I and II; and the transmembrane, tumor-associated hCA IX and XII. The triazole-substituted sulfocoumarins did not inhibit the ubiquitous, off-target cytosolic isoforms hCA I and II (KIs >10 μM) but showed effective inhibition against the two transmembrane CAs, with KIs ranging from 7.2 to 10.5 nM against hCA IX, and between 5.5 and 17.7 nM against hCA XII. As hCA IX and XII are validated anti-tumor targets, such prodrug, isoform-selective inhibitors as the sulfocoumarins reported here, may be useful for identifying suitable drug candidates for clinical trials., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

A series of 6-substituted sulfocoumarins incorporating substituted-1,2,3,4-tetrazol-5-yl moieties were synthesized by reaction of 6-iodo-sulfocoumarin and the corresponding tetrazole via the CH activation reaction. The new sulfocoumarins incorporating alkyl and substituted aryl moieties at the 1-position of the tetrazole, were investigated for the inhibition of four human (h) carbonic anhydrase (hCA, EC 4.2.1.1) isoforms, the cytosolic hCA I and II; and the transmembrane, tumor-associated hCA IX and XII. The tetrazole-substituted sulfocoumarins did not inhibit the ubiquitous, off-target cytosolic isoforms (KIs >10 μM) but showed effective inhibition against the two transmembrane CAs, with KIs ranging from 6.5 to 68.6 nM against hCA IX, and between 4.3 and 59.8 nM against hCA XII. As hCA IX and XII are validated anti-tumor targets, such prodrug, isoform-selective inhibitors as the sulfocoumarins reported here, may be useful for identifying suitable drug candidates for clinical trials., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

We report here a series of 2-thiophene-sulfonamides incorporating 1-substituted aryl-1,2,3-triazolyl moieties, prepared by click chemistry from 5-ethynylthiophene-2-sulfonamide and substituted aryl azides. The new sulfonamides were investigated as inhibitors of the zinc metalloenzyme CA (EC 4.2.1.1), and more specifically against the human (h) cytosolic isoforms hCA I and II and the transmembrane, tumor-associated ones hCA IX and XII: The new compounds were medium-weak hCA I inhibitors (KIs in the range of 224-7544nM), but were compactly, highly effective, low nanomolar hCA II inhibitors (KIs of 2.2-7.7nM). The tumor-associated hCA IX was inhibited with KIs ranging between 5.4 and 811nM, whereas hCA XII with inhibition constants in the range of 3.4-239nM. The X-ray crystal structure of the adducts of two such compounds bound to hCA II (one incorporating 1-naphthyl, the other one 3-cyanophenyl moieties) evidenced the reasons of the high affinity for hCA II. Highly favorable, predominantly hydrophobic interactions between the sulfonamide scaffold and the hCA II active site were responsible for the binding, in addition to the coordination of the sulfamoyl moiety to the zinc ion. The tails of the two inhibitors adopted very diverse orientations when bound to the active site, with the naphthyltriazolyl moiety orientated towards the hydrophobic half of the active site, and the 3-cyanophenyl one pointing towards the hydrophilic half. These data may be used for the structure-based drug design of even more effective hCA II inhibitors, with potential use as antiglaucoma agents or as diuretics., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

A series of 7-substituted sulfocoumarins and 3,4-dihydrosulfocoumarins was obtained by cyclization of the methanesulfonate of 2,4-dihydroxy- or 2-hydroxy-4-methoxybenzaldehyde, followed by derivatization reactions. The new compounds incorporate a range of substituents in position 7 of the heterocyclic ring (hydroxyl, methoxy, carboxylic and alkylsulfonate ester). The compounds were tested for the inhibition of the zinc enzyme human (h) carbonic anhydrase (hCA, EC 4.2.1.1). Unlike the 6-substituted sulfocoumarins which were potent hCA IX and XII inhibitors and ineffective hCA I and II inhibitors, compounds from this series showed low nanomolar hCA II inhibitory properties, and inhibited the mitochondrial isoform hCA VA with KIs in the range of 91-9960 nM, but were ineffective as hCA I, IX and XII inhibitors. The structure activity relationship for this class of inhibitors was rather clear, with the nature of the 7-substituent strongly influencing hCA VA inhibition, whereas the nature of these groups were less relevant for hCA II inhibition (all reported compounds were highly effective hCA II inhibitors, with KIs in the range of 1.5-8.4 nM). Since both hCA II and hCA VA are important drug targets (hCA II for antiglaucoma agents; hCA VA for antiobesity drugs), these isoform-selective inhibitors reported here may be considered of interest for various biomedical applications., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

A series of novel sulfamides incorporating the dopamine scaffold were synthesized. Reaction of amines and tert-butyl-alcohol/benzyl alcohol in the presence of chlorosulfonyl isocyanate (CSI) afforded sulfamoyl carbamates, which were converted to the title compounds by treatment with trifluoroacetic acid or by palladium-catalyzed hydrogenolysis. Inhibition of six α-carbonic anhydrases (CAs, EC 4.2.1.1), that is, CA I, CA II, CA VA, CA IX, CA XII and CA XIV, and two β-CAs from Candida glabrata (CgCA) and Mycobacterium tuberculosis (Rv3588) with these sulfamides was investigated. All CA isozymes were inhibited in the low micromolar to nanomolar range by the dopamine sulfamide analogues. K(i)s were in the range of 0.061-1.822 μM for CA I, 1.47-2.94 nM for CA II, 2.25-3.34 μM for CA VA, 0.041-0.37 μM for CA IX, 0.021-1.52 μM for CA XII, 0.007-0.219 μM for CA XIV, 0.35-5.31 μM for CgCA and 0.465-4.29 μM for Rv3588. The synthesized sulfamides may lead to inhibitors targeting medicinally relevant CA isoforms with potential applications as antiepileptic, antiobesity antitumor agents or anti-infective., (Copyright © 2013 Elsevier Ltd. All rights reserved.)