Carbonic anhydrase inhibitors: inhibition of the transmembrane isozyme XIV with sulfonamides

The inhibition of the last human carbonic anhydrase (CA, EC 4.2.1.1) isozyme (hCA XIV) discovered has been investigated with a series of sulfonamides, including some clinically used derivatives (acetazolamide, methazolamide, ethoxzolamide, dichlorophenamide, dorzolamide, brinzolamide, benzolamide, and zonisamide), as well as the sulfamate antiepileptic drug topiramate. The full-length hCA XIV is an enzyme showing a medium-low catalytic activity, quite similar to that of hCA XII, with the following kinetic parameters at 20 degrees C and pH 7.5, for the CO2 hydration reaction: k(cat) = 3.12 x 10(5) s(-1) and k(cat)/K(M) = 3.9 x 10(7) M(-1) s(-1). All types of activities have been detected for the investigated compounds, with several micromolar inhibitors, including zonisamide, topiramate, and simple sulfanilamide derivatives (K(I)-s in the range of 1.46-6.50 microM). In addition, topiramate and zonisamide were observed to behave as weak hCA XII inhibitors, while zonisamide was an effective hCA IX inhibitor (K(I) of 5.1 nM). Some benzene-1,3-disulfonamide derivatives or simple five-membered heteroaromatic sulfonamides showed K(I)-s in the range of 180-680 nM against hCA XIV, whereas the most effective of such inhibitors, including 3-chloro-/bromo-sulfanilamide, benzolamide-like, ethoxzolamide-like, and acetazolamide/methazolamide-like derivatives, showed inhibition constant in the range of 13-48 nM. The best hCA XIV inhibitor was aminobenzolamide (K(I) of 13 nM), but no CA XIV-selective derivatives were evidenced. There are important differences of affinity of these sulfonamides/sulfamates for the three transmembrane CA isozymes, with CA XII showing the highest affinity, followed by CA IX, whereas CA XIV usually showed the lowest affinity for these inhibitors.     

QSAR studies for the inhibition of the transmembrane isozymes XII and XIV of human carbonic anhydrase with a series of sulfonamides

A diverse series of aromatic/heterocyclic sulfonamides possessing inhibitory action against the human transmembrane isoforms XII (cancer-associated) and XIV of the metalloenzyme carbonic anhydrase (CA, EC 4.2.1.1) has been used to develop QSAR models. Including all the 55 investigated sulfonamides in the calibration set, the predictive qualities of the QSAR equations were weak (r(2)=0.1771, F=5.70) for CA XII and good for CA XIV inhibition (r(2)=0.8222, F=57.04 before eliminating the outliers, and r(2)=0.8911, F=67.07 after eliminating them). The obtained models suggest a slightly different inhibition mechanism for the two isoforms. 3-Halogeno-4-amino-benzenesulfonamides were outliers for scaffold hopping for the inhibition of CA XIV. CA XIV inhibitory activity was proportional to the degree of molecular surface rugosity. For compounds of the type X-Ar-SO(2)NH(2) and Ar'-Ar-SO(2)NH(2) type, best inhibitors were detected when Ar/Ar' incorporates a heterocyclic moiety. These studies may be helpful for the design of more specific CA XII/XIV inhibitors, since this is the first QSAR model investigating them.     

Carbonic anhydrase inhibitors: inhibition of the human transmembrane isozyme XIV with a library of aromatic/heterocyclic sulfonamides

The first inhibition study of the transmembrane carbonic anhydrase (CA, EC 4.2.1.1) isozymes hCA XIV with a library of aromatic and heteroaromatic sulfonamides synthesized earlier is reported. Most of the inhibitors were sulfanilamide, homosulfanilamide and 4-aminoethyl-benzenesulfonamide derivatives, to which tails that would induce diverse physicochemical properties have been attached at the amino moiety. Several of these compounds were metanilamide, benzene-1,3-disulfonamide or the 1,3,4-thiadiazole/thiadiazoline-2-sulfonamide derivatives. The tails incorporated in these molecules were of the alkyl/aryl-carboxamido/ sulfonamido-, ureido- or thioureido-types. The sulfanilamides acylated at the 4-amino group with short aliphatic/aromatic moieties incorporating 2-6 carbon atoms showed modest hCA XIV inhibitory activity (K(I)-s in the range of 1.25-4.2 microM) which were anyhow better than that of sulfanilamide (K(I) of 5.4 microM). Better activity showed the homosulfanilamide and 4-aminoethyl-benzenesulfonamide derivatives bearing arylsulfonamido/ureido and thioureido moieties, with K(I)'s in the range of 203-935 nM. The best activity was observed for the heteroaromatic compounds incorporating 1,3,4-thiadiazole/thiadiazoline-2-sulfonamide and 5-arylcarboxamido/sulfonamido moieties, with K(I)'s in the range of 10-85 nM. All these compounds were generally also much better inhibitors of the other two transmembrane CA isozyme, hCA IX and XII. Thus, highly potent hCA XIV inhibitors were detected, but isozyme-specific inhibitors were not discovered for the moment.     

QSAR study on para-substituted aromatic sulfonamides as carbonic anhydrase II inhibitors using topological information indices

A linear quantitative structure-activity relationship has been developed for a series of para-substituted aromatic sulfonamides by using topological index methodologies. The compounds were studied for their carbonic anhydrase II (CAII) inhibitory activity. A large series of topological indices were calculated and the stepwise regression method was used to derive the most significant model. Very good results were obtained using multi-parametric regressions and showed that the information approach used in the present work is quite useful for modeling carbonic anhydrase inhibition.     

Carbonic anhydrase inhibitors: cloning, characterization, and inhibition studies of the cytosolic isozyme III with sulfonamides

The cytosolic human carbonic anhydrase (hCA, EC 4.2.1.1) isozyme III (hCA III) has been cloned and purified by the GST-fusion protein method. Recombinant pure hCA III had the following kinetic parameters for the CO(2) hydration reaction at 20 degrees C and pH 7.5: k(cat) of 1.3 x 10(4) s(-1) and k(cat)/K(M) of 2.5 x 10(5) M(-1) s(-1), being a slower catalyst for the physiological reaction as compared to the genetically related cytosolic isoforms hCA I and II. An inhibition study with a library of sulfonamides and one sulfamate, some which are clinically used compounds, is reported. hCA III is less prone to be inhibited by these compounds as compared to hCA I and II for which many low nanomolar inhibitors were detected earlier. The best hCA III inhibitors were prontosil, sulpiride, indisulam, benzolamide, aminobenzolamide, and 4-amino-6-chloro-benzene-1,3-disulfonamide which showed K(I)s in the range of 2.3-18.1 microM. Clinically used compounds such as acetazolamide, methazolamide, ethoxzolamide, dorzolamide, brinzolamide, topiramate, zonisamide, celecoxib, and valdecoxib were less effective hCA III inhibitors, with affinities in the range of 154-2200 microM. This is the first study in which low micromolar hCA III inhibitors are reported.     

Carbonic anhydrase inhibitors: inhibition of the tumor-associated isozyme IX with aromatic and heterocyclic sulfonamides

The inhibition of the tumor-associated transmembrane carbonic anhydrase IX (CA IX) isozyme has been investigated with a series of aromatic and heterocyclic sulfonamides, including the six clinically used derivatives acetazolamide, methazolamide, ethoxzolamide, dichlorophenamide, dorzolamide and brinzolamide. Inhibition data for the physiologically relevant isozymes I and II (cytosolic forms) and IV (membrane-bound) were also provided for comparison. A very interesting and unusual inhibition profile against CA IX with these sulfonamides has been observed. Several nanomolar (K(I)-s in the range of 14-50 nM) CA IX inhibitors have been detected, both among the aromatic (such as orthanilamide, homosulfonilamide, 4-carboxy-benzenesulfonamide, 1-naphthalenesulfonamide and 1,3-benzenedisulfonamide derivatives) as well as the heterocylic (such as 1,3,4-thiadizole-2-sulfonamide, etc.) sulfonamides examined. Because CA IX is a highly active isozyme predominantly expressed in tumor tissues with poor prognosis of disease progression, this finding is very promising for the potential design of CA IX-specific inhibitors with applications as anti-tumor agents.     

QSAR studies on carbonic anhydrase inhibitors: a case of ureido and thioureido derivatives of aromatic/heterocyclic sulfonamides

QSAR studies on modelling of biological activity (hCAI) for a series of ureido and thioureido derivatives of aromatic/heterocyclic sulfonamides have been made using a pool of topological indices. Regression analysis of the data showed that excellent results were obtained in multiparametric correlations upon introduction of indicator parameters. The predictive abilities of the models are discussed using cross-validation parameters.     

Modelling of carbonic anhydrase inhibitory activity of sulfonamides using molecular negentropy

The present paper deals with the modelling of carbonic anhydrase inhibitory activity of sulfonamides using molecular negentropy (N). Excellent results are obtained in multiple regression analysis upon introduction of indicator parameters. The results are critically discussed on the basis of statistical data obtained from regression analysis.     

QSAR studies on the activation of the human carbonic anhydrase cytosolic isoforms I and II and secretory isozyme VI with amino acids and amines

The first QSAR study on the activation of the human secretory isoform of the metalloenzyme carbonic anhydrase (CA, EC 4.2.1.1), CA VI, with a series of amines and amino acids is reported. A large set of topological indices have been used to obtain several tri-/tetra-parametric models. We compared the CA VI activating QSAR models with those calculated for activation of the cytosolic human isozymes hCA I and hCA II. In addition, the effect of D- and L-amino acids as activators of hCA I, hCA II and of hCA VI as compared to those of structurally related biogenic amines was investigated for obtaining statistically significant and predictive QSAR equations. The obtained models are discussed using a variety of statistical parameters. The best models were obtained for hCA II activation, followed by hCA I, whereas the QSAR models for the activation of hCA VI were statistically weaker.     

Carbonic anhydrase inhibitors: purification and inhibition studies of pigeon (Columba livia var. domestica) red blood cell carbonic anhydrase with sulfonamides

A carbonic anhydrase (CA, EC 4.2.1.1) from red blood cells of pigeons (Columba livia var. domestica), clCA, was purified to homogeneity. Its kinetic parameters for the CO₂ hydration reaction were measured. With a k_cat/K_m of 1.1?×?10⁸ M^?1 s^?1, and a k_cat of 1.3?×?10⁶ s^?1, clCA has a high activity, similar to that of the human isoform hCA II. A group of 25 aromatic/heterocyclic sulfonamides incorporating the sulfanilamide, homosulfanilamide, benzene-1,3-disulfonamide, and acetazolamide scaffolds showed variable inhibitory activity against the pigeon enzyme, with K_Is in the range of 1.9–3460?nM. Red blood cells of pigeons, like those of ostriches, contain thus just one CA isoform, unlike the blood of mammals, which normally contain two isoforms, one of low (CA I-like) and one of very high activity (CA II-like). However, from the sulfonamide inhibition viewpoint, the pigeon enzyme was more similar to hCA II than to the ostrich enzyme.     

Carbonic anhydrase inhibitors: purification and inhibition studies of pigeon (Columba livia var. domestica) red blood cell carbonic anhydrase with sulfonamides

A carbonic anhydrase (CA, EC 4.2.1.1) from red blood cells of pigeons (Columba livia var. domestica), clCA, was purified to homogeneity. Its kinetic parameters for the CO(2) hydration reaction were measured. With a k(cat)/K(m) of 1.1?×?10(8) M(-1) s(-1), and a k(cat) of 1.3?×?10(6) s(-1), clCA has a high activity, similar to that of the human isoform hCA II. A group of 25 aromatic/heterocyclic sulfonamides incorporating the sulfanilamide, homosulfanilamide, benzene-1,3-disulfonamide, and acetazolamide scaffolds showed variable inhibitory activity against the pigeon enzyme, with K(I)s in the range of 1.9-3460?nM. Red blood cells of pigeons, like those of ostriches, contain thus just one CA isoform, unlike the blood of mammals, which normally contain two isoforms, one of low (CA I-like) and one of very high activity (CA II-like). However, from the sulfonamide inhibition viewpoint, the pigeon enzyme was more similar to hCA II than to the ostrich enzyme.     

Spirocyclic sulfonamides with carbonic anhydrase inhibitory and anti-neuropathic pain activity

A novel series of 4-oxo-spirochromane bearing primary sulfonamide group were synthetized as Carbonic Anhydrase inhibitors (CAIs) and tested for their management of neuropathic pain. Indeed, CAs have been recently validated as novel therapeutic targets in neuropathic pain. All compounds, here reported, showed strong activity against hCA II and hCA VII with K_I values in the low or sub-nanomolar range. Two compounds (6d and 6l) showed good neuropathic pain attenuating effects and longer duration than drug reference acetazolamide in an animal model of oxaliplatin induced neuropathy.     

QSAR studies on benzene sulfonamide carbonic anhydrase inhibitors: need of hydrophobic parameter for topological modeling of binding constants of sulfonamides to human CA-II

The binding constants (logK) of benzene sulfonamides to human CA-II have been modeled using a large series of distance-based topological indices. The need or otherwise of the hydrophobic parameter (logP) for such topological modeling of logK has been examined critically. In both the cases excellent results have been obtained. In multiparametric models involving indicator parameters we observed that introduction of hydrophobic parameter (logP) yields much improved statistics. The results are discussed on the basis of statistical parameters and also by using cross-validation method.     

Fluoroenesulphonamides: N-sulphonylurea isosteres showing nanomolar selective cancer-related transmembrane human carbonic anhydrase inhibition

After hydrofluorination of ynesulphonamides in superacid or in the presence of hydrofluoric acid/base reagents, a series of α-fluoroenamides has been synthesised and tested for the inhibition of carbonic anhydrase (CA, EC 4.2.1.1) isoforms. This study reveals a new, highly selective family of cancer-related transmembrane human (h) CA IX/XII inhibitors. These original fluorinated ureido isosters do not inhibit the widespread cytosolic isoforms hCA I and II and selectively inhibit the transmembrane cancer-related hCA IX and XII, offering interesting new leads for future studies.     

Proton magnetic resonance studies of carbonic anhydrase. III. Binding of sulfonamides.

Resonances of the histidine region of human carbonic anhydrase B have been studied by proton magnetic resonance spectroscopy in the presence of seven sulfonamide inhibitors. Results of difference spectroscopy and observation of the C-2 resonance of an additional titratable histidine in some of these spectra suggest a conformational change in the enzyme, while the large number of unaltered resonances indicates involvement of only a few residues. Inhibition of carbonic anhydrase by sulfonamides appears to involve: stabilization of an appropriately oriented initial complex by hydrophobic binding of the aromatic ring of the inhibitor to residues of the cavity forming the active site; ionization of the sulfonamido group, facilitated by its proximity to zinc; protonation and displacement of the high pH ligand to the metal controlling catalytic activity, thought here to be a histidine residue; and formation by the sulfonamido group of an ionic bond to zinc and a hydrogen bond to the hydroxyl group of serine or threonine. Diversity of spectra produced with various sulfonamides suggests that substituents on the ring and heteroatoms within the ring interact with additional groups at the active site. Increase in inhibitory potency appears to involve optimizing the number as well as the strength of these interactions. An upper limit for the dissociation rate of these complexes of 10 sec-1 was obtained.     

Carbonic anhydrase inhibitors. Inhibition of cytosolic isozymes I and II and transmembrane, cancer-associated isozyme IX with lipophilic sulfonamides

A series of new compounds was obtained by reaction of aromatic/heterocyclic sulfonamides incorporating amino groups with N,N-diphenylcarbamoyl chloride and diphenylacetyl chloride. These sulfonamides were assayed for the inhibition of three carbonic anhydrase (CA, EC 4.2.1.1) isozymes: the cytosolic CA I and CA II, and the transmembrane, cancer-associated isozyme CA IX. Good inhibitors against all these isoforms were detected, and the inhibition profile of the newly investigated isozyme IX was observed to be different from that of the cytosolic isozymes, I and II. This may lead to the development of novel anticancer therapies based on the selective inhibition of CA IX.     

Carbonic anhydrase inhibitors: the first QSAR study on inhibition of tumor-associated isoenzyme IX with aromatic and heterocyclic sulfonamides

QSAR study on the tumor-associated transmembrane carbonic anhydrase IX (CA IX) isoenzyme has been made using a large pool of distance-based topological indices : W, Sz, PI (0)chi, (1)chi, (2)chi,(0)chi(v), (1)chi(v), (2)chi(v). A combined set of 32 aromatic and heterocyclic compounds, including the six clinically used derivatives: acetazolamide, methazolamide, ethoxyzolamide, dichlorophenamide, dorzolamide, and brinzolamide are used for this purpose. The results have shown that the inhibition of the tumor-associated isoenzyme IX with aromatic and heterocyclic sulfonamides can be modeled excellently in multiparametric regression after introduction of indicator parameters. The predictive power of the models is discussed using probable error of correlation (PE), variance-inflation factor (VIF), and cross-validation parameters: PRESS, SSY, r(2) (cv) (S) PRESS, and PSE. This is the first report on QSAR study on inhibition of tumor-associated isoenzyme IX.