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The fungal pathogen Candida glabrata encodes for a β-carbonic anhydrase (CA, EC 4.2.1.1), CgNce103, recently discovered. Only anions have been investigated as CgNce103 inhibitors up until now. Here we report the first sulfonamides inhibition study of this enzyme. Simple sulfonamides showed weak or moderate CgNce103 inhibitory properties, whereas acetazolamide, and a series of 4-substituted ureido-benzene-sulfonamides, sulfamates and sulfamides showed effective CgNce103 inhibitory properties, with K I s in the range of 4.1–115 nM, being also ineffective as human CA II inhibitors. As there is significant resistance of C. glabrata clinical isolates to many classical antifungal agents, inhibition of the β-CA from this organism may allow an interesting means of controlling the pathogen growth, eventually leading to antifungals with a novel mechanism of action.

A series of 2-(hydrazinocarbonyl)-3-substituted-phenyl-1H-indole-5-sulfonamides and 1-({[5-(aminosulfonyl)-3-phenyl-1H-indol-2-yl]carbonyl}amino)-2,4,6 trimethylpyridinium perchlorates possessing various 2-, 3- or 4-substituted phenyl groups with methyl-, halogeno- and methoxy-functionalities, as well as the perfluorophenyl moiety, have been evaluated as inhibitors of the beta-carbonic anhydrases (CAs, EC 4.2.1.1) from the pathogenic fungi Cryptococcus neoformans (Can2) and Candida albicans (CaNce103). Both enzymes were potently inhibited by these sulfonamides, K(I)s in the range of 4.4-118 nM against Can2, and of 5.1-128 against CaNce103, respectively. Minor structural changes in the 3-substituted phenyl moiety contribute significantly to the inhibitory activity. Some of the investigated sulfonamides showed promising selectivity ratios for inhibiting Can2 over the host, human enzymes CA I and II.

The introduction of highly active antiretroviral therapy (HAART) in 1996 dramatically changed the course of HIV infection. This therapy involves the use of at least three agents from two distinct classes of antivirals: a protease inhibitor (PI) in combination with two nucleoside/nucleotide reverse transcriptase inhibitors (N(t)RTIs), or a non-nucleoside reverse transcriptase inhibitor (NNRTI) in combination with NRTIs. Nine drugs containing PIs are clinically available: the first generation ones, saquinavir, ritonavir, indinavir, nelfinavir, and amprenavir, and the second generation ones, fosamprenavir (the amprenavir prodrug), lopinavir, atazanavir, and tipranavir. Many other compounds are in advanced clinical evaluation, such as among others TMC-114, whereas a lot of different other effective HIV protease inhibitors were reported, mainly by using amprenavir and TMC-114 as lead molecules. The main goals of research in this field are: (i) the design of better pharmacological agents, devoid of severe side effects, resistance problems and with simple administration schedules (preferably once daily), and (ii) achieving eradication of the virus, and possibly, a definitive cure of the disease. A review on the pharmacology and interactions of these agents with other drugs is presented here, with emphasis on how these pharmacological interferences may improve the clinical use of antivirals, or how side effects due to PI drugs may be managed better by taking them into account (such as for example ritonavir boosting of other PIs which reduces dosages and administration schedules of these drugs). Except for being highly effective in the treatment of HIV infection, recent reports showed this class of drugs to be effective as antitumor agents, as antibacterials (for example against Mycobacterium tuberculosis infection), antifungals (against Candida albicans), antimalarials, antiSARS and anti-influenza agents.

Sulthiame, a clinically used antiepileptic, was investigated for its interaction with 12 catalytically active mammalian carbonic anhydrase (CA, EC 4.2.1.1) isoforms. The drug is a potent inhibitor of CA II, VII, IX, and XII ( K I s of 6–56 nM), and a medium potency inhibitor against CA IV, VA, VB, and VI ( K I s of 81–134 nM). The high resolution crystal structure of the hCA II-sulthiame adduct revealed a large number of favorable interactions between the drug and the enzyme which explain its strong low nanomolar affinity for this isoform and may also be exploited for the design of effective inhibitors incorporating sultam moieties.

The activation of the metalloenzyme carbonic anhydrase (CA, EC 4.2.1.1) with l -adrenaline and histamine has been investigated by kinetic and X-ray crystallographic studies. l -Adrenaline behaves as a potent activator of isozyme CA I (activation constant of 90 nM), being a much weaker activator of isozyme CA II (activation constant of 96 μM). Isoforms CA IV, VA, VII, and XIV were activated by l -adrenaline with KAs in the range of 36–63 μM. The X-ray crystal structure of the CA II– l -adrenaline adduct revealed that the activator plugs the entrance of the active site cavity, obstructing it almost completely.

Considerable advances have been made in the last years in the design of derivatives acting as inhibitors of HIV entry and fusion. The discovery of chemokines focused the attention on cellular coreceptors used by the virus for entering within cells, and consequently the various steps of such processes have been characterized in detail. Intense research led to a wide range of effective compounds that are able to inhibit the initial steps of HIV life cycle. All steps in the process of HIV entry into the cell may be targeted by specific compounds that may be developed as novel types of antiretrovirals. Thus, several inhibitors of the gp120-CD4 interaction have been detected so far (zintevir, FP-21399 and BMS-378806 in clinical trials). Small molecule chemokine receptor antagonists acting as HIV entry inhibitors also were described in the last period, which interact both with the CXCR4 coreceptor (such as AMD3100; AMD3465; ALX40-4C; T22, T134 and T140), or which are antagonist of the CCR5 coreceptor (TAK-779, TAK-220, SCH-C, SCH-D, E913, AK-602, UK-427857 and NSC 651016 in clinical trials), together with new types of fusion inhibitors possessing the same mechanism of action as enfuvirtide (such as T1249). Recently, a third family of antivirals started to be used clinically (in addition to the reverse transcriptase and protease inhibitors), with the advent of enfuvirtide (T20), the first fusion inhibitor to be approved as an anti-HIV agent. Some of these compounds demonstrated in vitro synergism with other classes of antivirals, offering thus the rationale for their combination in therapies for HIV-infected individuals. Many HIV entry and fusion inhibitors are currently being investigated in controlled clinical trials, and a number of them is bioavailable as oral formulations. This is an essential feature for an extended use of these compounds with the purpose of ameliorating adherence of patients to these medications and preventing the development of drug resistance.

A series of positively charged sulfonamides were obtained by reaction of aminobenzolamide [5-(4-aminobenzenesulfonylamino)-1,3,4-thiadiazole-2-sulfonamide] with tri-/tetrasubstituted pyrilium salts possessing alkyl-, aryl- or combinations of alkyl and aryl groups at the pyridinium ring. The new compounds reported here were assayed for the inhibition of four physiologically relevant carbonic anhydrase (CA, EC 4.2.1.1) isozymes: the cytosolic hCA I and II, the membrane-anchored bCA IV, and the membrane-bound, tumor-associated isozyme hCA IX. They showed potent inhibitory activity against all investigated isozymes, although with different profiles. For CA I the new derivatives showed inhibition constants in the range of 3-12 nM, for CA II in the range of 0.20-5.96 nM, against CA IV in the range of 2.0-10.3 nM, and against CA IX in the range of 3-45 nM, respectively. These new compounds are membrane-impermeant due to their salt-like character. Some of these derivatives were also tested for their inhibitory activity against the Cl(-)/HCO(3)(-) anion exchanger AE1: two derivatives showed inhibitory activity in the low micromolar range, whereas one compound was inactive at these concentrations. The high affinity of these new derivatives for the tumor-associated isozyme CA IX and their membrane impermeability make this type of CA inhibitor interesting candidates for the selective inhibition of only the tumor-associated isozyme and not the cytosolic ones, for which they also show high potency. Furthermore, we prove here for the first time that the CA-AE metabolon can be inhibited by the same type of sulfonamide derivative.

The sulfonamides constitute an important class of drugs, with several types of pharmacological agents possessing antibacterial, anti- carbonic anhydrase, diuretic, hypoglycemic and antithyroid activity among others. A large number of structurally novel sulfonamide derivatives have ultimately been reported to show substantial antitumor activity in vitro and in vivo. Although they have a common chemical motif of aromatic/heterocyclic or amino acid sulfonamide, there are a variety of mechanisms of their antitumor action, such as carbonic anhydrase inhibition, cell cycle perturbation in the G1 phase, disruption of microtubule assembly, functional suppression of the transcriptional activator NF-Y, and angiogenesis (matrix metalloproteinase, MMP) inhibition among others. Some of these compounds selected via elaborate preclinical screenings or obtained through computer-based drug design, are currently being evaluated in clinical trials. The review summarizes recent classes of sulfonamides and related sulfonyl derivatives disclosed as effective tumor cell growth inhibitors, or for the treatment of different types of cancer. Another research line that progressed much in the last time regards different sulfonamides with remarkable antiviral activity. Thus, at least two clinically used HIV protease inhibitors possess sulfonamide moieties in their molecules, whereas a very large number of other derivatives are constantly being synthesized and evaluated in order to obtain compounds with less toxicity or activity against drug-resistant viruses. Several non nucleoside HIV reverse transcriptase or HIV integrase inhibitors containing sulfonamido groups were also reported. Another approach to inhibit the growth of retroviruses, including HIV, targets the ejection of zinc ions from critical zinc finger viral proteins, which has as a consequence the inhibition of viral replication in the absence of mutations leading to drug resistance phenotypes. Most compounds with antiviral activity possessing this mechanism of action incorporate in their molecules primary sulfonamide groups. Some small molecule chemokine antagonists acting as HIV entry inhibitors also possess sulfonamide functionalities in their scaffold.

A series of 9-sulfonylated/sulfenylated-6-mercaptopurines has been prepared by reaction of 6-mercaptopurine with sulfonyl/sulfenyl halides. These compounds constitute a new class of potent antimycobacterial agents, possessing MIC values against Mycobacterium tuberculosis H37Rv in the range of 0.39–3.39 μg/mL, as well as appreciable activity against Mycobacterium avium. Furthermore, a compound of this small series exhibited good activity (MIC under 1 μg/mL) against several drug resistant strains of M. tuberculosis.

Aryl/alkyl-sulfonyl-, aryl/alkylcarboxyl- and aryl(sulfonyl)carbamyl/thiocarbamyl-derivatives of dibenzo-1,4-dioxine-2-acetyloxime were prepared by reaction of the title compound with sulfonyl halides, sulfonic acid anhydrides, acyl chlorides/carboxylic acids, arylsulfonyl isocyanates, aryl/acyl isocyanates or isothiocyanates. Several of the newly synthesized compounds showed effective in vitro antifungal activity against Aspergillus and Candida spp., some of them showing activities comparable to ketoconazole (with minimum inhibitory concentrations in the range of 1.2-4 microg/mL) against the two Aspergillus strains, but possessing a lower activity as compared to ketoconazole against C. albicans. Of the three investigated strains, best activity was detected against A. flavus. The mechanism of action of these compounds probably involves inhibition of ergosterol biosynthesis by interaction with lanosterol-14-alpha-demethylase (CYP51A1), since reduced amounts of ergosterol were found by means of HPLC, in cultures of the sensitive strain A. flavus treated with some of these inhibitors. Thus, the compounds reported here might possess a similar mechanism of action at molecular level with that of the widely used azole antifungals.

A β-carbonic anhydrase (CA, EC 4.2.1.1) from the fungal pathogen Malassezia globosa has been cloned, characterized, and studied for its inhibition with sulfonamides. This enzyme, designated MG-CA, has significant catalytic activity in the CO(2) hydration reaction and was inhibited by sulfonamides, sulfamates, and sulfamides with K(I) in the nanomolar to micromolar range. Several sulfonamides have also been investigated for the inhibition of growth of M. globosa, M. dermatis, M. pachydermatic, and M. furfur in cultures, whereas a mouse model of dandruff showed that treatment with sulfonamides led to fragmented fungal hyphae, as for the treatment with ketoconazole, a clinically used antifungal agent. These data prompt us to propose MG-CA as a new antidandruff drug target.

A beta-carbonic anhydrase (CA, EC 4.2.1.1), the protein encoded by the NCE103 gene of Candida glabrata which also present in Candida albicans and Saccharomycescerevisiae, was cloned, purified, characterized kinetically and investigated for its inhibition by a series simple, inorganic anions such as halogenides, pseudohalogenides, bicarbonate, carbonate, nitrate, nitrite, hydrogen sulfide, bisulfite, perchlorate, sulfate and some isosteric species. The enzyme showed significant CO(2) hydrase activity, with a k(cat) of 3.8 x 10(5)s(-1) and k(cat)/K(M) of 4.8 x 10(7)M(-1)s(-1). The Ca glabrata CA (CgCA) was moderately inhibited by metal poisons (cyanide, azide, cyanate, thiocyanate, K(I)s of 0.60-1.12 mM) but strongly inhibited by bicarbonate, nitrate, nitrite and phenylarsonic acid (K(I)s of 86-98 microM). The other anions investigated showed inhibition constants in the low millimolar range, with the exception of bromide and iodide (K(I)s of 27-42 mM).

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.

A detailed inhibition study of five carbonic anhydrase (CA, EC 4.2.1.1) isozymes with inorganic phosphates, carbamoyl phosphate, the antiviral phosphonate foscarnet as well as formate is reported. The cytosolic isozyme hCA I was weakly inhibited by neutral phosphate, strongly inhibited by carbamoyl phosphate (K(I) of 9.4 microM), and activated by hydrogen- and dihydrogenphosphate, foscarnet and formate (best activator foscarnet, K(A)=12 microM). The cytosolic isozyme hCA II was weakly inhibited by all the investigated anions, with carbamoyl phosphate showing a K(I) of 0.31 mM. The membrane-associated isozyme hCA IV was the most sensitive to inhibition by phosphates/phosphonates, showing a K(I) of 84 nM for PO(4)(3-), of 9.8 microM for HPO(4)(2-), and of 9.9 microM for carbamoyl phosphate. Foscarnet was the best inhibitor of this isozyme (K(I) of 0.82 mM) highly abundant in the kidneys, which may explain some of the renal side effects of the drug. The mitochondrial isozyme hCA V was weakly inhibited by all phosphates/phosphonates, except carbamoyl phosphate, which showed a K(I) of 8.5 microM. Thus, CA V cannot be the isozyme involved in the carbamoyl phosphate synthetase I biosynthetic reaction, as hypothesized earlier. Furthermore, the relative resistance of CA V to inhibition by inorganic phosphates suggests an evolutionary adaptation of this mitochondrial isozyme to the presence of high concentrations of such anions in these energy-converting organelles, where high amounts of ATP are produced by ATP synthetase, from ADP and inorganic phosphates. The transmembrane, tumor-associated isozyme hCA IX was on the other hand slightly inhibited by all these anions.

Reaction of sodium N,N-dimethyldithiocarbamate or N,N-diethyldithiocarbamate with arylsulfonyl halides afforded a series of arylsulfonyl-N,N-dialkyl-dithiocarbamates. The reactivity of these new derivatives with cysteine and glutathione has been investigated in order to identify derivatives that might label a cysteine residue of the heterodimeric protein tubulin which plays a critical physiological function in cell division and also possesses enzymatic activity as a GTP-ase. Since many antitumor drugs exert their action by binding to tubulin, inhibiting in this way microtubule association and provoking cell death, some of the most reactive compounds against the thiol reagents found in this work have been assayed for their antitumor activity. Indeed strong tumor cell growth inhibitory properties against several leukemia, non-small cell lung, ovarian, melanoma, colon, CNS, renal, prostate and breast cancer has been found in vitro for some of the 4-halogeno-, 4-methyl- or 4-carboxyphenyl-substituted arylsulfonyl-N,N-dialkyl-dithiocarbamates. Furthermore, some of these derivative were shown to act as in vitro tubulin polymerization inhibitors using a turbidimetric assay.

A series of alkyl/arylsulfonyl-N,N-diethyl-dithiocarbamates has been prepared by reaction of sodium N,N-diethyldithiocarbamate with alkyl/arylsulfonyl halides. The reactivity of these new derivatives against cysteine and glutathione has been investigated in order to identify derivatives that might label a critical cysteine residue of tubulin (Cys 239 of human β2 tubulin chain). Some of the most reactive compounds showed moderate to powerful tumor growth inhibitory properties against several leukemia, non-small cell lung, ovarian, melanoma, colon, CNS, renal, prostate and breast cancer cell lines in vitro.

Aminobenzolamide (5-sulfanilylamido-1,3,4-thiadiazole-2-sulfonamide) is a potent inhibitor of the zinc enzyme carbonic anhydrase (CA, EC 4.2.1.1), being at the same time structurally similar to the antimicrobial sulfonamides. Here we report that the reaction of aminobenzolamide with arylsulfonyl isocyanates affords a series of new arylsulfonylureido derivatives which were subsequently used as ligands (in the form of conjugate bases, as sulfonamide anions) for the preparation of metal complexes containing Ag(I) and Zn(II). All the new compounds proved to be very potent inhibitors of CA (isozymes I, II and IV). The newly synthesized complexes, unlike the free ligands, also act as effective antifungal agents against several Aspergillus and Candida spp., some of them showing activities comparable to ketoconazole, with minimum inhibitory concentrations in the range of 1.8-5 microg/mL. The mechanism of antifungal action of these complexes seem to be unconnected with inhibition of lanosterol-14-alpha-demethylase, since the levels of sterols assessed in the fungi cultures were equal in the absence or in the presence of the tested compounds. Probably the new complexes act as inhibitors of phosphomannose isomerase, a key enzyme in the biosynthesis of yeast cell walls.