RESUMO
A series of congeners structurally related to pritelivir, N-[5-(aminosulfonyl)-4-methyl-1,3-thiazol-2-yl]-N-methyl-2-[4-(2-pyridinyl)phenyl]acetamide, a helicase-primase inhibitor for the treatment of herpes simplex virus infections, was prepared. The synthesized primary and secondary sulfonamides were investigated as inhibitors of six physiologically and pharmacologically relevant human (h) carbonic anhydrase (hCA, EC 4.2.1.1) isoforms, the cytosolic enzymes hCA I and II, the mitochondrial ones hCA VA and VB, and the transmembrane, tumor associated hCA IX and XII. Low nanomolar inhibition KI values were detected for all of them, with a very interesting and well-defined structure-activity relationship. As many CAs are involved in serious pathologies, among which are cancer, obesity, epilepsy, glaucoma, etc., sulfonamide inhibitors as those reported here may be of interest as drug candidates. Furthermore, pritelivir itself is an effective inhibitor of some CAs, also inhibiting whole blood enzymes from several mammalian species, which may be a favorable pharmacokinetic feature of the drug which can be transported throughout the body bound to blood CA I and II.
Assuntos
Inibidores da Anidrase Carbônica/química , Inibidores da Anidrase Carbônica/farmacologia , Sulfonamidas/química , Sulfonamidas/farmacologia , Tiazóis/química , Tiazóis/farmacologia , Anidrases Carbônicas/química , Anidrases Carbônicas/metabolismo , Desenho de Fármacos , Humanos , Simulação de Acoplamento Molecular , Isoformas de Proteínas/antagonistas & inibidores , Isoformas de Proteínas/química , Isoformas de Proteínas/metabolismoRESUMO
A series of phenolic acid esters incorporating caffeic, ferulic, and p-coumaric acid, and benzyl, m/p-hydroxyphenethyl- as well as p-hydroxy-phenethoxy-phenethyl moieties were investigated for their inhibitory effects against the metalloenzyme carbonic anhydrase (CA, EC 4.2.1.1). Many of the mammalian isozymes of human (h) or murine (m) origin, hCA I-hCA XII, mCA XIII and hCA XIV, were inhibited in the submicromolar range by these derivatives (with KIs of 0.31-1.03 µM against hCA VA, VB, VI, VII, IX and XIV). The off-target, highly abundant isoforms hCA I and II, as well as hCA III, IV and XII were poorly inhibited by many of these esters, although the original phenolic acids were micromolar inhibitors. These phenols, like others investigated earlier, possess a CA inhibition mechanism distinct of the sulfonamides/sulfamates, clinically used drugs for the treatment of a multitude of pathologies, but with severe side effects due to hCA I/II inhibition. Unlike the sulfonamides, which bind to the catalytic zinc ion, phenols are anchored at the Zn(II)-coordinated water molecule, binding more externally within the active site cavity, and making contacts with amino acid residues at the entrance of the active site. As this is the region with the highest variability between the many CA isozymes found in mammals, this class of compounds shows isoform-selective inhibitory profiles, which may be exploited for obtaining pharmacological agents with less side effects compared to other classes of inhibitors.
Assuntos
Inibidores da Anidrase Carbônica/farmacologia , Anidrases Carbônicas/química , Anidrases Carbônicas/metabolismo , Ésteres/farmacologia , Hidroxibenzoatos/química , Animais , Inibidores da Anidrase Carbônica/química , Ativação Enzimática/efeitos dos fármacos , Humanos , Isoenzimas/antagonistas & inibidores , Isoenzimas/metabolismo , Camundongos , Relação Estrutura-AtividadeRESUMO
Autotaxin is an extracellular, two zinc-centered enzyme that hydrolyzes lysophosphatidyl choline to lysophosphatidic acid, involved in various cancerous processes, e.g. migration, proliferation and tumor progression. We examined the autotaxin inhibitory properties of extended structure carbamoylphosphonates (CPOs) PhOC(6)H(4)SO(2)NH(CH(2))nNHCOPO(3)H(2), with increasing lengths of methylene chains, (CH(2))(n), n = 4-8. Carbamoylphosphonates having n = 6, 7, 8 inhibited autotaxin in vitro with IC(50) ≈ 1.5 µM. Using an imaging probe we demonstrated that compound n = 6 inhibits recombinant autotaxin activity in vitro and in vivo, following oral CPO administration. Additionally, daily oral administration of compound n = 7 inhibited over 90% of lung metastases in a murine melanoma metastasis model. Both the carbamoylphosphonates and the enzymes reside and interact in the extracellular space expecting minimal toxic side effects, and presenting a novel approach for inhibiting tumor proliferation and metastasis dissemination.
Assuntos
Antineoplásicos/farmacologia , Neoplasias Pulmonares/tratamento farmacológico , Melanoma Experimental/tratamento farmacológico , Organofosfonatos/farmacologia , Diester Fosfórico Hidrolases/metabolismo , Sulfonamidas/farmacologia , Administração Oral , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/química , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Humanos , Neoplasias Pulmonares/secundário , Camundongos , Camundongos Endogâmicos C57BL , Estrutura Molecular , Organofosfonatos/administração & dosagem , Organofosfonatos/química , Relação Estrutura-Atividade , Sulfonamidas/administração & dosagem , Sulfonamidas/químicaRESUMO
Using hydroxy-/dihydroxybenzoic acids as leads, a series of methyl, ethyl and iso-propyl esters of 4-hydroxy-benzoic acid, 2,4-, 2,5-, 2,6-, 3,4-, and 3,5-dihydroxybenzoic acids and of coumaric acid, were obtained and investigated for the inhibition of six mammalian carbonic anhydrase (CA, EC 4.2.1.1) isoforms, that is, the cytosolic CA I, II and VII, and the transmembrane CA IX, XII and XIV, many of which are established drug targets. Other compounds incorporating phenol/catechol moieties were obtained from dopamine by reaction with fluorescein isothiocyanate or with 2,4,6-trisubstituted pyrylium salts. Some aminophenols were also derivatized in a similar manner, by using pyrylium salts. Many of these compounds showed increased inhibitory action compared to the lead compounds from which they were obtained, with efficacy in the submicromolar range against most investigated CA isoforms. As phenols are a class of less investigated CA inhibitors (CAIs) compared to the sulfonamides, and their mechanism of inhibition is less well understood, compounds of the type designed here may be helpful in gaining more insights into these phenomena.
Assuntos
Inibidores da Anidrase Carbônica/química , Anidrases Carbônicas/química , Hidroxibenzoatos/química , Fenol/química , Compostos de Piridínio/química , Animais , Anidrase Carbônica I/antagonistas & inibidores , Anidrase Carbônica I/metabolismo , Anidrase Carbônica II/antagonistas & inibidores , Anidrase Carbônica II/metabolismo , Inibidores da Anidrase Carbônica/metabolismo , Anidrases Carbônicas/metabolismo , Ésteres , Hidroxibenzoatos/metabolismo , Isoenzimas/antagonistas & inibidores , Isoenzimas/metabolismo , Cinética , Ligação Proteica , Compostos de Piridínio/metabolismo , Relação Estrutura-AtividadeRESUMO
A series of new, halogen containing N-substituted 4-aminobenzenesulfonamides were synthesized by using superacid HF/SbF5 chemistry and investigated as inhibitors of several human carbonic anhydrase (hCA, EC 4.2.1.1) isoforms, that is, the cytosolic hCA I and II and, the tumor-associated transmembrane isoforms hCA IX and XII. Despite the substitution of the sulfonamide function, the presence of fluorine atom(s) in ß position of the sulfonamide function strongly favors hCA inhibition. A similar effect of the ß-fluorinated alkyl substitution on the amino function has been also observed. Among the tested compounds, several chlorinated derivatives have been identified as selective nanomolar, tumor-associated isoforms inhibitors. These non-primary sulfonamides probably bind in the coumarin-binding site, at the entrance of the cavity, and not to the metal ion as the primary sulfonamide inhibitors.
Assuntos
Antígenos de Neoplasias/química , Inibidores da Anidrase Carbônica/síntese química , Anidrases Carbônicas/química , Flúor/química , Sulfonamidas/química , Antígenos de Neoplasias/metabolismo , Sítios de Ligação , Anidrase Carbônica I/química , Anidrase Carbônica I/metabolismo , Anidrase Carbônica II/química , Anidrase Carbônica II/metabolismo , Anidrase Carbônica IX , Inibidores da Anidrase Carbônica/química , Inibidores da Anidrase Carbônica/metabolismo , Anidrases Carbônicas/metabolismo , Domínio Catalítico , Humanos , Simulação de Acoplamento Molecular , Neoplasias/enzimologia , Neoplasias/metabolismo , Neoplasias/patologia , Ligação Proteica , Relação Estrutura-Atividade , Sulfonamidas/síntese química , Sulfonamidas/metabolismoRESUMO
A series of halogenated sulfanilamides and halogenated benzolamide derivatives have been investigated as inhibitors of three ß-carbonic anhydrases (CAs, EC 4.2.1.1) from the bacterial pathogen Mycobacterium tuberculosis, mtCA 1 (Rv1284), mtCA 2 (Rv3588c) and mtCA 3 (Rv3273). All three enzymes were inhibited with efficacies between the submicromolar to the micromolar one, depending on the substitution pattern at the sulfanilamide moiety/fragment of the molecule. Best inhibitors were the halogenated benzolamides (K(I)s in the range of 0.12-0.45 µM) whereas the halogenated sulfanilamides were slightly less inhibitory (K(I)s in the range of 0.41-4.74 µM). This class of ß-CA inhibitors may have the potential for developing antimycobacterial agents with a diverse mechanism of action compared to the clinically used drugs for which many strains exhibit multi-drug/extensive multi-drug resistance.
Assuntos
Benzolamida/farmacologia , Inibidores da Anidrase Carbônica/farmacologia , Anidrases Carbônicas/metabolismo , Mycobacterium tuberculosis/enzimologia , Sulfanilamidas/farmacologia , Benzolamida/síntese química , Benzolamida/química , Inibidores da Anidrase Carbônica/síntese química , Inibidores da Anidrase Carbônica/química , Anidrases Carbônicas/isolamento & purificação , Relação Dose-Resposta a Droga , Estrutura Molecular , Isoformas de Proteínas/antagonistas & inibidores , Isoformas de Proteínas/isolamento & purificação , Isoformas de Proteínas/metabolismo , Relação Estrutura-Atividade , Sulfanilamidas/síntese química , Sulfanilamidas/químicaRESUMO
A series of N-mono- and N,N-disubstituted dithiocarbamates have been investigated as inhibitors of two ß-carbonic anhydrases (CAs, EC 4.2.1.1) from the bacterial pathogen Mycobacterium tuberculosis, mtCA 1 (Rv1284) and mtCA 3 (Rv3273). Both enzymes were inhibited with efficacies between the subnanomolar to the micromolar one, depending on the substitution pattern at the nitrogen atom from the dithiocarbamate zinc-binding group. Aryl, arylalkyl-, heterocyclic as well as aliphatic and amino acyl such moieties led to potent mtCA 1 and 3 inhibitors in both the N-mono- and N,N-disubstituted dithiocarbamate series. This new class of ß-CA inhibitors may have the potential for developing antimycobacterial agents with a diverse mechanism of action compared to the clinically used drugs for which many strains exhibit multi-drug/extensive multi-drug resistance.
Assuntos
Inibidores da Anidrase Carbônica/farmacologia , Anidrases Carbônicas/metabolismo , Mycobacterium tuberculosis/enzimologia , Tiocarbamatos/farmacologia , Dióxido de Carbono/química , Dióxido de Carbono/metabolismo , Inibidores da Anidrase Carbônica/síntese química , Inibidores da Anidrase Carbônica/química , Relação Dose-Resposta a Droga , Ativação Enzimática/efeitos dos fármacos , Estrutura Molecular , Isoformas de Proteínas/antagonistas & inibidores , Isoformas de Proteínas/metabolismo , Relação Estrutura-Atividade , Tiocarbamatos/síntese química , Tiocarbamatos/químicaRESUMO
Natural products (NPs) have proven to be an invaluable source of new chemotherapies yet very few have been explored to source small molecule carbonic anhydrase (CA) inhibitors. CA enzymes underpin physiological pH and are critical to the progression of several diseases including cancer. The present study is the first to more widely investigate NP coumarins for CA inhibition following the recent discovery of a NP coumarin CA inhibitor. We assembled a NP library comprising 24 plant coumarins (compounds 4-27) and three ascidian coumarins (compounds 28-30) that together provide a diverse collection of structures containing the coumarin pharmacophore. This library was then evaluated for inhibition of six human CA isozymes (CAs I, II, VII, IX, XII and XIII) and a broad range of inhibition and isozyme selectivity profiles were evident. Our findings provide a platform to support further evaluation of NPs for the discovery of new chemotypes that inhibit disease relevant CA enzymes.
Assuntos
Inibidores da Anidrase Carbônica/química , Anidrases Carbônicas/química , Cumarínicos/química , Produtos Biológicos/química , Produtos Biológicos/metabolismo , Anidrases Carbônicas/metabolismo , Humanos , Isoenzimas/antagonistas & inibidores , Isoenzimas/metabolismo , Cinética , Plantas/química , Ligação ProteicaRESUMO
A set of sulfamides and sulfamates were synthesized and tested against several isoforms of carbonic anhydrase: CA I, CA II, CA VII, CA XII and CA XIV. The biological assays showed a broad range of inhibitory activity, and interesting results were found for several compounds in terms of activity (Ki <1µm) and selectivity: some aromatic sulfamides are active against CA I, CA II and/or CA VII; while they are less active in CA XII and CA XIV. On the other hand, bulky sulfamides are selective to CA VII. To understand the origin of the different inhibitory activity against each isozyme we used molecular modeling techniques such as docking and molecular dynamic simulations.
Assuntos
Inibidores da Anidrase Carbônica/química , Anidrases Carbônicas/química , Sulfonamidas/química , Sítios de Ligação , Anidrase Carbônica I/antagonistas & inibidores , Anidrase Carbônica I/metabolismo , Anidrase Carbônica II/antagonistas & inibidores , Anidrase Carbônica II/metabolismo , Inibidores da Anidrase Carbônica/síntese química , Inibidores da Anidrase Carbônica/metabolismo , Anidrases Carbônicas/metabolismo , Domínio Catalítico , Humanos , Simulação de Acoplamento Molecular , Ligação Proteica , Sulfonamidas/síntese química , Sulfonamidas/metabolismoRESUMO
Carbonic anhydrase inhibitors of primary sulfonamide type, RSO(2)NH(2), have clinical applications as diuretics, antiglaucoma, antiepileptic, antiobesity and antitumor drugs. Here we investigated inhibition of two human cytosolic isozymes, hCA I and II, with a series of secondary/tertiary sulfonamides, incorporating tosyl moieties (CH(3)C(6)H(4)SO(2)NR1R2). Most compounds inhibited both isoforms in low micromolar range, with inhibition constants between 0.181-6.01 µM against hCA I, and 0.209-0.779 µM against hCA II, respectively. These findings point out that substituted benzenesulfonamides may be used as leads for generating interesting CAIs probably possessing a distinct mechanism of action compared to primary sulfonamides. Indeed, classical RSO(2)NH(2) inhibitors bind in deprotonated form to the Zn(II) ion from the CA active site and participate in many other favorable interactions with amino acid residues lining the cavity. The secondary/tertiary sulfonamides cannot bind to the zinc due to steric hindrance and probably are accommodated at the entrance of the active site, in coumarin binding-site.
Assuntos
Inibidores da Anidrase Carbônica/farmacologia , Anidrases Carbônicas/metabolismo , Sulfonamidas/farmacologia , Dióxido de Carbono/química , Dióxido de Carbono/metabolismo , Inibidores da Anidrase Carbônica/síntese química , Inibidores da Anidrase Carbônica/química , Anidrases Carbônicas/isolamento & purificação , Relação Dose-Resposta a Droga , Humanos , Estrutura Molecular , Isoformas de Proteínas/antagonistas & inibidores , Isoformas de Proteínas/isolamento & purificação , Isoformas de Proteínas/metabolismo , Relação Estrutura-Atividade , Sulfonamidas/síntese química , Sulfonamidas/químicaRESUMO
Reaction of 6-/7-hydroxycoumarin with metronidazole afforded conjugates which incorporate two interesting chemotypes which may inhibit carbonic anhydrases (CAs, EC 4.2.1.1) due to the presence of the coumarin moiety and possess radiosensitizing effects due to the presence of the nitroazole. Another dual action compound, which may act both as CA inhibitor as well as monocarboxylate transporter inhibitor, is 3-cyano-7-hydroxy-coumarin. These compounds have been investigated as inhibitors of 11 human CA isoforms. Submicromolar inhibition was observed against hCA VA, hCA VB, hCA VI, hCA VII, hCA IX, hCA XII and hCA XIV, whereas isoforms hCA I, II and XIII were not inhibited by these compounds. These coumarins thus act as isoform-selective CA inhibitors with the possibility to target isoforms involved in pathologies such as obesity (CA VA/VB) or cancer (CA IX and XII) without inhibiting the physiologically dominant, highly abundant hCA I and II.
Assuntos
Inibidores da Anidrase Carbônica/farmacologia , Anidrases Carbônicas/metabolismo , Cumarínicos/farmacologia , Metronidazol/farmacologia , Biocatálise , Dióxido de Carbono/química , Dióxido de Carbono/metabolismo , Inibidores da Anidrase Carbônica/síntese química , Inibidores da Anidrase Carbônica/química , Cumarínicos/síntese química , Cumarínicos/química , Relação Dose-Resposta a Droga , Ativação Enzimática/efeitos dos fármacos , Humanos , Metronidazol/síntese química , Metronidazol/química , Estrutura Molecular , Isoformas de Proteínas/antagonistas & inibidores , Isoformas de Proteínas/metabolismo , Relação Estrutura-AtividadeRESUMO
The gastric pathogen Helicobacter pylori encodes two carbonic anhydrases (CAs, EC 4.2.1.1), an α- and a ß-class one, hpαCA and hpßCA, crucial for its survival in the acidic environment from the stomach. Sulfonamides, strong inhibitors of these enzymes, block the growth of the pathogen, in vitro and in vivo. Here we report the inhibition of the two H. pylori CAs with inorganic and complex anions and other molecules interacting with zinc proteins. hpαCA was inhibited in the low micromolar range by diethyldithiocarbamate, sulfamide, sulfamic acid, phenylboronic acid, and in the submillimolar one by cyanide, cyanate, hydrogen sulfide, divanadate, tellurate, perruthenate, selenocyanide, trithiocarbonate, iminodisulfonate. hpßCA generally showed a stronger inhibition with most of these anions, with several low micromolar and many submillimolar inhibitors detected. These inhibitors may be used as leads for developing anti-H. pylori agents with a diverse mechanism of action compared to clinically used antibiotics.
Assuntos
Inibidores da Anidrase Carbônica/farmacologia , Anidrases Carbônicas/metabolismo , Helicobacter pylori/enzimologia , Sulfanilamidas/farmacologia , Ânions/química , Inibidores da Anidrase Carbônica/síntese química , Inibidores da Anidrase Carbônica/química , Anidrases Carbônicas/isolamento & purificação , Relação Dose-Resposta a Droga , Helicobacter pylori/química , Humanos , Estrutura Molecular , Isoformas de Proteínas/antagonistas & inibidores , Isoformas de Proteínas/isolamento & purificação , Isoformas de Proteínas/metabolismo , Relação Estrutura-Atividade , Sulfanilamidas/síntese química , Sulfanilamidas/químicaRESUMO
The growth of Mycobacterium tuberculosis is strongly inhibited by weak acids although the mechanism by which these compounds act is not completely understood. A series of substituted benzoic acids, nipecotic acid, ortho- and para-coumaric acid, caffeic acid and ferulic acid were investigated as inhibitors of three ß-class carbonic anhydrases (CAs, EC 4.2.1.1) from this pathogen, mtCA 1 (Rv1284), mtCA 2 (Rv3588c) and mtCA 3 (Rv3273). All three enzymes were inhibited with efficacies between the submicromolar to the micromolar one, depending on the scaffold present in the carboxylic acid. mtCA 3 was the isoform mostly inhibited by these compounds (K(I)s in the range of 0.11-0.97 µM); followed by mtCA 2 (K(I)s in the range of 0.59-8.10 µM), whereas against mtCA 1, these carboxylic acids showed inhibition constants in the range of 2.25-7.13 µM. This class of relatively underexplored ß-CA inhibitors warrant further in vivo studies, as they may have the potential for developing antimycobacterial agents with a diverse mechanism of action compared to the clinically used drugs for which many strains exhibit multi-drug or extensive multi-drug resistance.
Assuntos
Inibidores da Anidrase Carbônica/farmacologia , Anidrases Carbônicas/metabolismo , Ácidos Carboxílicos/farmacologia , Mycobacterium tuberculosis/enzimologia , Biocatálise , Dióxido de Carbono/química , Dióxido de Carbono/metabolismo , Inibidores da Anidrase Carbônica/síntese química , Inibidores da Anidrase Carbônica/química , Ácidos Carboxílicos/síntese química , Ácidos Carboxílicos/química , Relação Dose-Resposta a Droga , Ativação Enzimática/efeitos dos fármacos , Estrutura Molecular , Mycobacterium tuberculosis/crescimento & desenvolvimento , Isoformas de Proteínas/antagonistas & inibidores , Isoformas de Proteínas/metabolismo , Relação Estrutura-AtividadeRESUMO
Carbonic anhydrases (CAs, EC 4.2.1.1) are ubiquitous isozymes involved in crucial physiological and pathological events, representing the targets of inhibitors with several therapeutic applications. In this connection, we report a new class of carbonic anhydrase inhibitors, based on the thiopyrano-fused pyrazole scaffold to which a pendant 4-sulfamoylphenyl moiety was attached. The new sulfonamides 3a-e were designed as constrained analogues of celecoxib and valdecoxib. The most interesting feature of sulfonamides 3 was their predominantly strong inhibition of human (h) CA I and II, as well as those of the mycobacterial ß-class enzymes (Rv1284, Rv3273, and Rv3588c), whereas their inhibitory action against hCA III, IV, VA, VB, VI, VII, IX, XII, XIII, and XIV was found to be at least 2 orders of magnitude lower. X-ray crystallography and structural superposition studies made it possible to explain the very distinct inhibition profile of the tricyclic sulfonamides, different from those of celecoxib and valdecoxib.
Assuntos
Inibidores da Anidrase Carbônica/farmacologia , Anidrases Carbônicas/química , Pirazóis/química , Sulfonamidas/química , Inibidores da Anidrase Carbônica/química , Domínio Catalítico , Cristalografia por Raios X , Humanos , Isoenzimas , Modelos Moleculares , Estrutura Molecular , Conformação Proteica , Relação Estrutura-AtividadeRESUMO
Carbamoylphosphonates (CPOs) have been identified as inhibitors of matrix metalloproteinases (MMPs) and as orally active, bioavailable, and safe antimetastatic agents. In this article, we focus on the direct antitumor activity of the CPOs. We discovered that CPOs also inhibit carbonic anhydrases (CAs), especially the IX and XII isoforms identified as cancer promoting factors. Thus, CPOs can be regarded as novel nontoxic drug candidates for tumor microenvironment targeted chemotherapy acting by two synergistic mechanisms, namely, inhibiting CAs and MMPs simultaneously. We have also demonstrated that the ionized CPO acid is unable to cross the cell membrane and thus limited to interact with the extracellular domains of isozymes CAIX and CAXII. Finally, applying CPOs against cancer cells in hypoxic conditions resulted in the dose dependent release of lactate dehydrogenase, confirming the direct interaction of the CPOs with the cancer related isozymes CAIX and XII and thereby promoting cellular damage.
Assuntos
Antineoplásicos/farmacologia , Inibidores da Anidrase Carbônica/farmacologia , Anidrases Carbônicas/efeitos dos fármacos , Metaloproteinase 2 da Matriz/efeitos dos fármacos , Organofosfonatos/farmacologia , Inibidores de Proteases/farmacologia , Microambiente Tumoral , Linhagem Celular Tumoral , HumanosRESUMO
Hydroxamates (R-CONHOH) have been scarcely investigated as carbonic anhydrase (CA, EC 4.2.1.1) inhibitors (CAIs). An inhibition/structural study of PhCONHOH is reported against all human isoforms. Comparing aliphatic (R = Me and CF(3)) and aromatic (R = Ph) hydroxamates as CAIs, we prove that CONHOH is a versatile zinc binding group. Depending on the nature of the R moiety, it can adopt different coordination modes to the catalytic ion within the CA active site.
Assuntos
Inibidores da Anidrase Carbônica/química , Anidrases Carbônicas/química , Quelantes/química , Ácidos Hidroxâmicos/química , Zinco/química , Anidrases Carbônicas/metabolismo , Domínio Catalítico , Complexos de Coordenação , Cristalografia por Raios X , Humanos , Isoenzimas/química , Isoenzimas/metabolismo , Modelos Moleculares , Ligação Proteica , Relação Estrutura-Atividade , Sulfonamidas/químicaRESUMO
Pazopanib, a new, multi-targeted tyrosine kinase inhibitor (TKI) used clinically for the treatment of several types of tumors, incorporates a primary sulfonamide moiety normally associated with the inhibition of the metallo enzyme carbonic anhydrase (CA, EC 4.2.1.1). Here we show that pazopanib and related sulfonamides such as indisulam, acetazolamide or ureido-substituted peptidomimetic benzenesulfonamides are low nanomolar inhibitors of many of the fifteen human isoforms hCA I-XIV. These data indicate that in addition to the TK inhibitory action, pazopanib may exert antitumor/antimetastatic effects also due to the potent inhibition of the tumor-associated, hypoxia-inducible enzymes CA IX and XII.
Assuntos
Anidrases Carbônicas/metabolismo , Sistemas de Liberação de Medicamentos , Inibidores Enzimáticos/farmacologia , Pirimidinas/farmacologia , Receptores Proteína Tirosina Quinases/antagonistas & inibidores , Sulfonamidas/farmacologia , Animais , Antineoplásicos/química , Antineoplásicos/farmacologia , Ativação Enzimática/efeitos dos fármacos , Humanos , Indazóis , Modelos MolecularesRESUMO
An inhibition study of several carbonic anhydrase (CA, EC 4.2.1.1) isoforms with flavones and aminoflavones, compounds possessing a rather similar scaffold with the coumarins, recently discovered inhibitors of this enzyme, is reported. The natural product flavone and some of its hydroxylated derivatives did not show time-dependent inhibition of the CAs, sign that they are not hydrolyzed within the enzyme active site as the (thio)coumarins and lactones. These compounds were low micromolar inhibitors of hCA I, II, IX and XII, with K(I)s in the range of 1.88-9.07 µM. A series of substituted 2-amino-3-phenyl-4H-chromen-4-ones, incorporating chloro- and methoxy substituents in various positions of the heterocycle, were then prepared and assayed as hCA I and II inhibitors, showing activity in the micromolar range. Some of these derivatives, as well as cis+trans resveratrol, were then assayed for the inhibition of all catalytically active mammalian CA isoforms, hCA I, II, III, IV, VA, VB, VI, VII, IX, XII, XIII, XIV and mCA XV (h=human, m=murine enzyme). These derivatives inhibited these CAs in the submicromolar-low micromolar range. Flavones, although not as active as the coumarins, may be considered as interesting leads for the design of non-sulfonamide CA inhibitors.
Assuntos
Benzopiranos/farmacologia , Inibidores da Anidrase Carbônica/farmacologia , Cumarínicos/farmacologia , Flavonas/farmacologia , Benzopiranos/química , Flavonas/química , Humanos , Hidroxilação , Isoformas de Proteínas , Relação Estrutura-AtividadeRESUMO
Site-directed mutagenesis has been used to change three amino acid residues involved in the binding of inhibitors (Asn67Ile; Gln92Val and Leu204Ser) within the active site of human carbonic anhydrase (CA, EC 4.2.1.1) II (hCA II). Residues 67, 92 and 204 were changed from hydrophobic to hydrophilic ones, and vice versa. The Asn67Ile and Leu204Ser mutants showed similar k(cat)/K(M) values compared to the wild type (wt) enzyme, whereas the Gln92Val mutant was around 30% less active as a catalyst for CO(2) hydration to bicarbonate compared to the wt protein. Affinity for sulfonamides/sulfamates was decreased in all three mutants compared to wt hCA II. The effect was stronger for the Asn67Ile mutant (the closest residue to the zinc ion), followed by the Gln92Val mutant (residue situated in the middle of the active site) and weakest for the Leu204Ser mutant, an amino acid situated far away from the catalytic metal ion, at the entrance of the cavity. This study shows that small perturbations within the active site architecture have influences on the catalytic efficiency but dramatically change affinity for inhibitors among the CA enzymes, especially when the mutated amino acid residues are nearby the catalytic metal ion.