磺胺类碳酸酐酶抑制剂研究进展*

碳酸酐酶抑制剂(CAIs)作为药物具有广泛的生物活性,如抗青光眼、利尿、抗肿瘤、抗癫痫、抗风湿、降血糖等。近几年,新一代抗青光眼磺胺类CAIs已经取得很大的研究成果,抗肿瘤磺胺类CAIs的研究也日益活跃,发展极为迅速。现综述磺胺类CAIs在抗青光眼和抗肿瘤领域的研究进展。     

A novel class of carbonic anhydrase inhibitors: glycoconjugate benzene sulfonamides prepared by "click-tailing"

Aryl and heteroaryl sulfonamides (ArSO(2)NH(2)) are therapeutically used to inhibit the catalytic activity of carbonic anhydrases (CAs). Using a "click-tail" approach a novel class of glycoconjugate benzene sulfonamides have been synthesized that contain diverse carbohydrate-triazole tails. These compounds were assessed for their ability to inhibit three human CA isozymes in vitro: cytosolic hCA I and hCA II and transmembrane, tumor-associated hCA IX. This isozyme has a minimal expression in normal tissue but is overexpressed in hypoxic tumors and its inhibition is a current approach toward new cancer therapies. The qualitative structure-activity for all derivatives demonstrated that the stereochemical diversity present within the carbohydrate tails effectively interrogated the CA active site topology, to generate several inhibitors that were potent and selective toward hCA IX, an important outcome in the quest for potential cancer therapy applications based on CA inhibition.     

Antiobesity carbonic anhydrase inhibitors

Few pharmacological approaches for the treatment of obesity exist at this time, and most of them are unsatisfactory, whereas this disease is widespread both in the developed and developing world. Novel effective approaches are needed for the development of antiobesity agents possessing different mechanisms of action. A possible new approach for the treatment and prophylaxis of obesity is based on the inhibition of carbonic anhydrases (CAs, EC 4.2.1.1), enzymes involved in several steps of de novo lipogenesis, both in the mitochondria and the cytosol of cells. Topiramate and zonisamide are two antiepileptic drugs that were shown to induce persistent weight loss in obese patients, but their mechanism of action is largely unknown. We demonstrated strong CA inhibitory properties for these two drugs, by means of kinetic studies in solution and X-ray crystallography, against several physiologically relevant isoforms, such as CA II, VA and VB. It has been proved that topiramate also inhibits lipogenesis in adipocytes, similarly to other sulfonamide CA inhibitors investigated earlier. A large number of new sulfonamides have been synthesized and assayed as possible inhibitors of CA isoforms involved in lipogenesis. This is the beginning of a very new and promising approach for the treatment of obesity, with the hope that new compounds showing this property will be soon developed and available for clinical use.     

选择性碳酸酐酶Ⅸ抑制剂的研究进展

碳酸酐酶(carbonic anhydrases,CAs)是一类广泛分布于生物体内的金属酶,催化生物体内二氧化碳和碳酸氢根离子的相互转化。CAⅨ在正常组织中几乎不表达而在许多实体瘤中过度表达,促进肿瘤细胞的存活及转移性扩散,因此被确定为肿瘤相关蛋白。自1992年CAⅨ的结构被确定以来,CAⅨ抑制剂发展为药物化学领域的研究热点之一,四大类经典的碳酸酐酶抑制剂(CAIs)以及一些新型选择性CAIs不断被发现,部分化合物进入了临床研究阶段。本文对近5年CAⅨ选择性抑制剂的研究进展进行综述。     

Thienothiopyran-2-sulfonamides: a novel class of water-soluble carbonic anhydrase inhibitors

An attempt to develop a water-soluble carbonic anhydrase inhibitor focused on exploring structure-activity relationships in the thienothiopyransulfonamide class. The strategy to influence water solubility while retaining carbonic anhydrase activity involved the introduction of a hydroxyl moiety and adjusting the oxidation state of the sulfur on the thiopyran portion of the molecule. Compounds 4 and 17 best fit the criteria of aqueous solubility and inhibitory potency vs. human carbonic anhydrase II and are candidates for evaluation as topically effective antiglaucoma agents.     

Fluorescence- and spin-labeled carbonic anhydrase inhibitors

Carbonic anhydrase IX (hCA IX) is a membrane-associated glycoprotein that is observed in many tumor tissues and is strongly overexpressed by hypoxia conditions. Hypoxia is a clinically important tumor parameter and this enzyme can play an important role as a potential marker of hypoxic tumor and as a therapeutic target too. In the last years, Carbonic Anhydrase IX Inhibitors which possess fluorescent probe were largely used for visualize hypoxic tumor cell lines and for understanding the biological roles of hCA IX in acidification of the external matrix. Here we resume the development pathways of such compounds from the design to the final biological evaluation. Furthermore, spin-labeled CAIs were included to have a complete overview of the potentiality of this enzyme as marker of hypoxic tumors.     

Direct measurements of the rate constants of sulfonamides with carbonic anhydrase.

Kinetic values for 14 sulfonamides and carbonic anhydrase (equivalent of isozyme II) were determined directly by measurement of association rate constants (kon) and equilibrium constants (KI), yielding dissociation rate constants, koff. Values for kon (in liter/mol sec-1) ranged from 0.003 to 31 x 10(6), whereas KI ranged from 0.7 to 17,000 x 10(-9) M. The koff range was very small, 0.01-0.05 sec-1. Thus, the activity, which is usually thought of as reflecting KI, is entirely a function of the association rate. This is not the common situation in enzyme-inhibitor reactions. The koff range is faster by several orders of magnitude than drug decay from plasma, so equilibrium is always achieved at the enzyme site in vivo, after parenteral administration of acetazolamide, methazolamide, or ethoxzolamide. For topical administration, as for MK-927 to reduce intraocular flow and pressure, the quantitative relation between free drug in tissue and that bound to enzyme in ciliary process is not so clear. Thus, koff might be an independent factor in pharmacological activity. The reaction of anions with carbonic anhydrase is entirely different, in that variations in KI are chiefly determined by koff.     

COX-2 selective inhibitors, carbonic anhydrase inhibition and anticancer properties of sulfonamides belonging to this class of pharmacological agents

The sulfonamides constitute an important class of drugs, with several types of pharmacological agents possessing antibacterial, anti-carbonic anhydrase, diuretic, hypoglycaemic, antithyroid, protease inhibitory and anticancer activity among others. A recently developed class of pharmacological agents incorporating primary sulfamoyl moieties in their molecule is constituted by the COX-2 selective inhibitors, with at least two clinically used drugs, celecoxib and valdecoxib. Another drug of this class, rofecoxib, does not contain sulfonamide moieties, but the isosteric and isoelectronic methylsulfone group. It was recently shown that the sulfonamide COX-2 selective inhibitors (but not the methylsulfone ones) also act as nanomolar inhibitors of several isozymes of the metallo-enzyme carbonic anhydrase (CA), some of which are strongly involved in tumourigenesis. In consequence, the potent anticancer effects of the sulfonamide COX-2 selective inhibitors and the much weaker such effects of rofecoxib, reported ultimately by many researchers, may be explained by the contribution of CA inhibition to such processes in addition to COX-2 inhibition.     

Malarial parasite carbonic anhydrase and its inhibitors

Plasmodium falciparum is responsible for the majority of life-threatening cases of human malaria. The global emergence of drug-resistant malarial parasites necessitates identification and characterization of novel drug targets. At present, carbonic anhydrase (CA) genes are identified in limited numbers of protozoa and helminthes parasites, however, they are demonstrated in at least 4 Plasmodium species. The CA gene of P. falciparum encodes an alpha-carbonic anhydrase enzyme possessing catalytic properties distinct of that of the human host enzymes. A small library of aromatic sulfonamides, most of which were Schiff's bases derived from sulfanilamide/homosulfanilamide/4-aminoethylbenzenesulfonamide and substituted-aromatic aldehydes, or ureido-substituted sulfonamides are good inhibitors of P. falciparum enzyme. The 4-(3,4-dichlorophenylureido-ethyl)-benzenesulfonamide is the most effective antimalarial activity against growth of P. falciparum in vitro. The nature of the groups substituting the aromatic-ureido- or aromatic-azomethine fragment of the molecule and the length of the parent sulfonamide (i.e., from sulfanilamide to 4-aminoethylbenzenesulfonamide) from which the Schiff's base obtained, are the critical parameters for the CA inhibitory activities of these aromatic sulfonamide derivatives, both against the malarial as well as human enzymes. Thus, the sulfonamide CA inhibitors may have the potential for the development of novel antimalarial drugs.     

Successful virtual screening for novel inhibitors of human carbonic anhydrase: strategy and experimental confirmation

Virtual screening of compound libraries is an alternative and complementary approach to high-throughput screening in the lead discovery process. A new strategy is described to search for possible leads of human carbonic anhydrase II, applying a protocol of several consecutive hierarchical filters involving a preselection based on functional group requirements and fast pharmacophore matching. A suitable pharmacophore is derived by a sophisticated "hot spot" analysis of the binding site to detect regions favorable for protein-ligand interactions. In subsequent steps, molecular similarity with known reference ligands is used to rerank the hits from the pharmacophore matching. Finally the best scored candidates are docked flexibly into the protein binding pocket. After examination of the affinity predictions, 13 compounds were selected for experimental testing. Of these 13, three could be shown to be subnanomolar, one is nanomolar, while a further seven are micromolar inhibitors. The binding mode of two hits could be confirmed by crystal structure analysis. The novelty of the discovered leads is best supported by the fact that a search in the patent literature showed the newly discovered subnanomolar compounds to comprise scaffolds not yet covered by existing patents.     

Carbonic anhydrase inhibitors. Design of fluorescent sulfonamides as probes of tumor-associated carbonic anhydrase IX that inhibit isozyme IX-mediated acidification of hypoxic tumors

Sulfonamides inhibit the catalytic activity of carbonic anhydrases (CAs, EC 4.2.1.1), enzymes participating in the regulation of acid-base balance and ion transport in many tissues. Carbonic anhydrase IX (CA IX), a transmembrane isoform with predominant association with tumors and limited distribution in normal tissues, is strongly overexpressed by hypoxia. Hypoxia increases the catalytic performance of CA IX contributing to microenvironmental acidosis, which influences cancer progression and treatment outcome. CA IX represents a target for detection and therapy of hypoxic tumors. Sulfonamide CA IX selective inhibitors accumulate only in hypoxic cells containing CA IX, reversing acidification mediated by this enzyme. The design of fluorescent sulfonamides that preferentially inhibit the activity of CA IX, showing reduced penetration through the plasma membranes and binding to hypoxic cells expressing CA IX, is reported here. These inhibitors represent promising candidates for developing anticancer therapies based on tumor-associated CA isozyme inhibition and offer interesting tools for imaging and further investigation of hypoxic tumors.     

Carbonic anhydrase inhibitors: synthesis and inhibition of cytosolic/membrane-associated carbonic anhydrase isozymes I, II, and IX with sulfonamides incorporating hydrazino moieties

Targeting proteins overexpressed in hypoxic tumors is as an important means of controlling cancer disease. One such protein is the carbonic anhydrase (CA) isoenzyme IX, which in some types of tumors is overexpressed 150-200-fold. We report here a series of sulfonamide derivatives, prepared from 2-carbohydrazido- and 4-carbohydrazido-benzenesulfonamides, which were further derivatized by reaction with aryl isocyanates or arylsulfonyl isocyanates. Several low nanomolar CA IX inhibitors were detected in this way. SAR is discussed for the diverse types of inhibitors and their affinity for different isozymes, with the aim of obtaining isozyme-specific CA IX inhibitors, with putative applications as antitumor drugs.     

Anticonvulsant/antiepileptic carbonic anhydrase inhibitors: a patent review

Introduction: An epileptic seizure is a transient occurrence of signs and/or symptoms due to abnormal excessive or synchronous neuronal activity in the brain. The International League Against Epilepsy classifies seizures in two broad categories: partial (localized to one cerebral hemisphere) and generalized (localized to both cerebral hemispheres). One indirect pathway for the treatment of epilepsy includes the inhibition of carbonic anhydrase (CA), thereby increasing CO₂ levels in the brain.

Areas covered: Carbonic anhydrases (EC 4.2.1.1) are ubiquitous metalloenzymes that catalyze the reversible hydration/dehydration of CO₂/HCO₃ ^-, respectively. CA inhibitors (CAIs) such as acetazolamide, methazolamide, topiramate, zonisamide, and sulthiame can reduce seizures through perturbation of the CO₂ equilibrium and/or the inhibition of ion channels. This review focuses on the mechanism of epilepsy, CA catalysis, and recent developments in the treatment of epilepsy using CAIs.

Expert opinion: Based on the observed active-site binding interactions of CAIs in crystal structures and their respective inhibition constants, structure–activity relationships can be mapped. Various CAIs along with novel techniques to administer them have been patented in the last four years. However, epilepsy continues to be a path less traveled when it comes to CAIs. A major area of research must focus toward the design of isoform-specific inhibitors using analogs of existing CAIs.     

Anti-infective carbonic anhydrase inhibitors: a patent and literature review

Introduction: All microorganisms that cause diseases or illnesses to their host are defined as pathogens. In this paper we will review the current state of the art for inhibiting parasite carbonic anhydrases (CAs, EC 4.2.1.1) with a goal of developing antibacterial, antifungal or antiprotozoan agents possessing a different mechanism of action compared with the clinically used drugs to which a considerable degree of drug resistance has been reported.

Areas covered: Cloning of the genomes of many pathogenic microorganisms offered the possibility of exploring alternative pathways for inhibiting virulence factors or proteins essential for their life cycle, and such an approach was applied systematically for CAs from prokaryotic and eukaryotic microorganisms. Investigations of CAs in bacteria, fungi and protozoa domains will undoubtedly reveal novel aspects of microbial virulence, these efforts being part of a more general approach of using metalloenzyme inhibitors for designing novel classes of anti-infectives.

Expert opinion: Covers an overview of the patent literature in the field of the anti-infective CA inhibitors. Most of the patents deal with sulfonamide CA inhibitors. Bacterial/fungal/protozoan CAs represent at this moment very promising targets for obtaining anti-infectives devoid of the resistance problems of the clinically used such agents but further studies are needed to validate these and other less investigated enzymes as novel drug targets.