Synthesis and biological evaluation of histamine Schiff bases as carbonic anhydrase I,II, IV,VII, and IX activators

A series of 20 histamine Schiff base was synthesised by reaction of histamine, a well known carbonic anhydrase (CA, E.C 4.2.2.1.) activator pharmacophore, with substituted aldehydes. The obtained histamine Schiff bases were assayed as activators of five selected human (h) CA isozymes, the cytosolic hCA I, hCA II, and hCA VII, the membrane-anchored hCA IV and transmembrane hCA IX. Some of these compounds showed efficient activity (in the nanomolar range) against the cytosolic isoform hCA VII, which is a key CA enzyme involved in brain metabolism. Moderate activity was observed against hCA I and hCA IV (in the nanomolar to low micromolar range). The structure–activity relationship for activation of these isoforms with the new histamine Schiff bases is discussed in detail based on the nature of the aliphatic, aromatic, or heterocyclic moiety present in the aldehyde fragment of the molecule, which may participate in diverse interactions with amino acid residues at the entrance of the active site, where activators bind, and which is the most variable part among the different CA isoforms.     

Carbonic anhydrase activators: amino acyl/dipeptidyl histamine derivatives bind with high affinity to isozymes I, II and IV and act as efficient activators

Reaction of histamine (Hst) with tetrabromophthalic anhydride and protection of its imidazole moiety with tritylsulfenyl chloride, followed by hydrazinolysis, afforded N-1-tritylsulfenyl-histamine, a key intermediate which was further derivatized at its aminoethyl moiety. Reaction of the key intermediate with N-Boc-amino acids/dipeptides (Boc-AA) in the presence of carbodiimides afforded, after deprotection of the imidazolic and amino moieties, a series of compounds with the general formula AA-Hst (AA=amino acyl; dipeptidyl). The new derivatives were assayed as activators of three carbonic anhydrase (CA) isozymes, hCA I, hCA II (cytosolic forms) and bCA IV (membrane-bound form). Efficient activation was observed against all three isozymes, but especially against hCA I and bCA IV, with affinities in the nanomolar range for the best compounds. hCA II was, on the other hand, activatable with affinities around 10–20 nM. This new class of CA activators might lead to the development of drugs/diagnostic agents for the CA deficiency syndrome, a genetic disease of bone, brain and kidneys.     

Carbonic anhydrase activators: synthesis of high affinity isozymes I, II and IV activators, derivatives of 4-(4-tosylureido-amino acyl)ethyl-1H-imidazole (histamine derivatives)

Reaction of histamine (Hst) with tetrabromophthalic anhydride and protection of its imidazole moiety with tritylsulfenyl chloride, followed by hydrazinolysis, afforded N-1-tritylsulfenyl histamine, a key intermediate which was further derivatized at its aminoethyl moiety. Reaction of the key intermediate with 4-tosylureido amino acids/dipeptides (ts-AA) in the presence of carbodiimides, afforded after deprotection of the imidazole moiety, a series of compounds with the general formula ts-AA-Hst (ts=4-MeC(6) H(4) SO(2) NHCO). Some structurally related dipeptide derivatives with the general formula ts-AA1-AA2-Hst, were also prepared, by in a similar way to the amino acyl compounds mentioned above. The new derivatives were examined as activators of three carbonic anhydrase (CA) isozymes, hCA I, hCA II (cytosolic forms) and bCA IV (membrane-bound form). Efficient activation was observed against all three isozymes, but especially against hCA I and bCA IV, with affinities in the 1-10 nanomolar range for the best compounds. hCA II was on the other hand activatable with affinities around 20-50 nM. This new class of CA activators might lead to the development of drugs/diagnostic agents for the CA deficiency syndrome, a genetic disease of bone, brain and kidneys.     

Carbonic anhydrase activators: human isozyme II is strongly activated by oligopeptides incorporating the carboxyterminal sequence of the bicarbonate anion exchanger AE1

Di-/tri- and especially tetrapeptides incorporating the sequence DADD present in the carboxyterminal region of the bicarbonate/chloride anion exchanger AE1 strongly activate human carbonic anhydrase (CA) isozyme II, whereas they act as more inefficient activators of isozymes I and IV. This discovery suggests that in the metabolon hCA II-AE1, the last protein plays a role both as a CA activator as well as a bicarbonate transporter. A synthesis of the tripeptide DAD and the tetrapeptide DADD is also presented together with the possible explanation why such highly acidic oligopeptides efficiently bind to hCA II but not to the closely related isozymes I and IV.     

Carbonic anhydrase activators: synthesis of high affinity isozymes I, II and IV activators, derivatives of 4-(arylsulfonylureido-amino acyl)ethyl-1H-imidazole

Based on the X-ray crystallographic structure of the adduct of human carbonic anhydrase II (hCA II) with the weak activator histamine (Briganti, F., Mangani, S., Orioli, P., Scozzafava, A., Vernaglione, G. and Supuran, C.T. (1997) Biochemistry, 36, 10,384-10,392), a novel class of tight-binding CA activators was designed by using histamine (Hst) as lead molecule. Thus, N-1-tritylsulfenyl Hst was synthesized by reaction of Hst with tetrabromophthalic anhydride followed by protection of its imidazole moiety with tritylsulfenyl chloride. After hydrazinolysis, it afforded a key intermediate which was derivatized at the aliphatic amino group. Reaction of the key intermediate with 4-fluorophenylsulfonylureido amino acids (fpu-AA) or 2-toluenesulfonylureido amino acids (ots-AA) in the presence of carbodiimides, afforded after deprotection, a series of compounds with the general formula fpu/ots-AA-Hst (fpu = 4-FC6H4SO2NHCO; ots = 2-MeC6H4SO2NHCO). Some structurally related dipeptides with the general formula fpu/ots-AA1-AA2-Hst (AA, AA1 and AA2 represent amino acyl moieties), were also prepared, by a strategy similar to that used for the simple amino acyl compounds above. The new derivatives proved to be efficient in vitro activators of three CA isozymes. Best activity was shown against hCA I and bCA IV, for which some of the new compounds (such as the Lys, Arg, His or the dipeptide derivatives) showed affinities in the 2-12 nm range (h = human; b = bovine isozymes). hCA II was on the other hand somehow less prone to activation by the new derivatives, which possessed affinities around 30-60 nM for this isozyme. Ex vivo experiments showed some of the new activators to strongly enhance red cell CA activity (180-230%) after incubation with human erythrocytes. This new class of CA activators might lead to the development of drugs/diagnostic tools for the CA deficiency syndrome, a genetic disease of bone, brain and kidneys.     

Carbonic anhydrase activators: X-ray crystal structure of the adduct of human isozyme II with L-histidine as a platform for the design of stronger activators

Activation of the carbonic anhydrase (CA, EC 4.2.1.1) isoforms hCA I, II, and IV with l-histidine and some of its derivatives has been investigated by kinetic and X-ray crystallographic methods. l-His was a potent activator of isozymes I and IV (activation constants in the range of 4-33microM), and a moderate hCA II activator (activation constant of 113microM). Both carboxy- as well as amino-substituted l-His derivatives, such as the methyl ester or the dipeptide carnosine (beta-Ala-His), acted as more efficient activators as compared to l-His. The X-ray crystallographic structure of the hCA II-l-His adduct showed the activator to be anchored at the entrance of the active site cavity, participating in an extended network of hydrogen bonds with the amino acid residues His64, Asn67, and Gln92 and, with three water molecules connecting it to the zinc-bound water. Although the binding site of l-His is similar to that of histamine, the first CA activator for which the X-ray crystal structure has been reported in complex with hCA II (Briganti, F.; Mangani, S.; Orioli, P.; Scozzafava, A.; Vernaglione, G.; Supuran, C. T. Biochemistry1997, 36, 10384) there are important differences of binding between the two structurally related activators, since histamine interacts among others with Asn67 and Gln92 (similarly to l-His), but also with Asn62 and not His64, whereas the number of water molecules connecting them to the zinc-bound water is different (two for histamine, three for l-His). Furthermore, the imidazole moieties of the two activators adopt different conformations when bound to the enzyme active site. Since neither the amino- nor carboxy moieties of l-His participate in interactions with amino acid moieties of the active site, they can be derivatized for obtaining more potent activators, with pharmacological applications for the enhancement of synaptic efficacy. This may constitute a novel approach for the treatment of Alzheimer's disease, aging, and other conditions in need of achieving spatial learning and memory therapy.     

Carbonic anhydrase activators: Activation of human isozymes I,II and IX with phenylsulfonylhydrazido l-histidine derivatives

Activation of the human carbonic anhydrase (CA, EC 4.2.1.1) isozymes I, II (cytosolic) and IX (transmembrane, tumor-associated isoform) with a series of arylsulfonylhydrazido-l-histidines incorporating 4-substituted-phenyl, pentafluorophenyl- and β-naphthyl moieties was investigated. The compounds showed a weak hCA I activation profile, but were more efficient as hCA II and IX activators. The 4-iodophenyl-substituted derivative behaved as a strong and isozyme selective hCA II activator, with an activation constant of 0.21 μM. This is the first isoform-selective, potent CA activator reported to date.     

Carbonic anhydrase activators: activation of the human isoforms VII (cytosolic) and XIV (transmembrane) with amino acids and amines

An activation study of the human carbonic anhydrase (hCA, EC 4.2.1.1) isozymes VII and XIV using a small library of natural/non-natural amino acids and aromatic/heterocyclic amines is reported. hCA VII was efficiently activated by L-/D-His, dopamine and serotonin (K(A)s of 0.71-0.93 microM). The best hCA XIV activators were histamine (K(A) of 10 nM), L-Phe, L-/D-His and 4-amino-L-Phe (K(A)s of 0.24-2.90 microM). In view of the significant expression levels of CA VII and CA XIV in the brain, selective activation of these isoforms may be useful when developing pharmacologic agents for the management of major disorders such as epilepsy and Alzheimer's disease.     

Carbonic Anhydrase Activators: Synthesis of High Affinity Isozymes I,II and IV Activators,Derivatives of 4-(4-Tosylureido-Amino Acyl)Ethyl-1H-Imidazole (Histamine Derivatives)

Reaction of histamine (Hst) with tetrabromophthalic anhydride and protection of its imidazole moiety with tritylsulfenyl chloride, followed by hydrazinolysis, afforded N-1-tritylsulfenyl histamine, a key intermediate which was further derivatized at its aminoethyl moiety. Reaction of the key intermediate with 4-tosylureido amino acids/dipeptides (ts-AA) in the presence of car-bodiimides, afforded after deprotection of the imidazole moiety, a series of compounds with the general formula ts-AA-Hst (ts = 4-MeC₆H₄SO₂NHCO). Some structurally related dipeptide derivatives with the general formula ts-AA l-AA2-Hst, were also prepared, by in a similar way to the amino acyl compounds mentioned above. The new derivatives were examined as activators of three carbonic anhydrase (CA) isozymes, hCA I, hCA II (cytosolic forms) and bCA IV (membrane-bound form). Efficient activation was observed against all three isozymes, but especially against hCA I and bCA IV, with affinities in the 1-10 nanomolar range for the best compounds. hCA II was on the other hand activatable with affinities around 20-50 nM. This new class of CA activators might lead to the development of drugs/diagnostic agents for the CA deficiency syndrome, a genetic disease of bone, brain and kidneys.     

Carbonic anhydrase activators: an activation study of the human mitochondrial isoforms VA and VB with amino acids and amines

The mitochondrial isozymes of human carbonic anhydrase (hCA, EC 4.2.1.1), hCA VA and hCA VB, were investigated for activation with a series of amino acids and amines. D-His, L-DOPA, histamine, dopamine, and 4-(2-aminoethyl)morpholine were excellent hCA VA activators, with KAs in the range of 10-130 nM. Good hCA VB activating effects were identified for L-His, D-Phe, D-DOPA, L-Trp, L-Tyr, serotonin, and 2-(2-aminoethyl)-pyridine, with KAs in the range of 44-110 nM. All these activators enhanced kcat, having no effect on KM, favoring thus the rate-determining step in the catalytic cycle, the proton transfer reactions between the active site and environment. The activation pattern of the two mitochondrial isoforms is very different from each other and as compared to those of the cytosolic isoforms hCA I and II.     

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.     

Investigation of piperazines as human carbonic anhydrase I,II, IV and VII activators

Four human (h) carbonic anhydrase isoforms (CA, EC 4.2.1.1), hCA I, II, IV, and VII, were investigated for their activation profile with piperazines belonging to various classes, such as N-aryl-, N-alkyl-, N-acyl-piperazines as well as 2,4-disubstituted derivatives. As the activation mechanism involves participation of the activator in the proton shuttling between the zinc-coordinated water molecule and the external milieu, these derivatives possessing diverse basicity and different scaffolds were appropriate for being investigated as CA activators (CAAs). Most of these derivatives showed CA activating properties against hCA I, II, and VII (cytosolic isoforms) but were devoid of activity against the membrane-associated hCA IV. For hCA I, the K_As were in the range of 32.6–131?µM; for hCA II of 16.2–116?µM, and for hCA VII of 17.1–131?µM. The structure-activity relationship was intricate and not easy to rationalize, but the most effective activators were 1-(2-piperidinyl)-piperazine (K_A of 16.2?µM for hCA II), 2-benzyl-piperazine (K_A of 17.1?µM for hCA VII), and 1-(3-benzylpiperazin-1-yl)propan-1-one (K_A of 32.6?µM for hCA I). As CAAs may have interesting pharmacologic applications in cognition and for artificial tissue engineering, investigation of new classes of activators may be crucial for this relatively new research field.     

Carbonic anhydrase activators: the first activation study of the human secretory isoform VI with amino acids and amines

The secretory isozyme of human carbonic anhydrase (hCA, EC 4.2.1.1), hCA VI, has been cloned, expressed, and purified in a bacterial expression system. The kinetic parameters for the CO(2) hydration reaction proved hCA VI to possess a k(cat) of 3.4 x 10(5)s(-1) and k(cat)/K(M) of 4.9 x 10(7)M(-1)s(-1) (at pH 7.5 and 20 degrees C). hCA VI has a significant catalytic activity for the physiological reaction, of the same order of magnitude as the ubiquitous isoform CA I or the transmembrane, tumor-associated isozyme CA IX. A series of amino acids and amines were shown to act as CA VI activators, with variable efficacies. l-His, l-Trp, and dopamine showed weak CA VI activating effects (K(A)s in the range of 21-42 microM), whereas d-His, d-Phe, l-DOPA, l-Trp, serotonin, and some pyridyl-alkylamines were better activators, with K(A)s in the range of 13-19 microM. The best CA VI activators were l-Phe, d-DOPA, l-Tyr, 4-amino-l-Phe, and histamine, with K(A)s in the range of 1.23-9.31 microM. All these activators enhance k(cat), having no effect on K(M), participating thus in the rate determining step in the catalytic cycle, the proton transfer reactions between the enzyme active site and the environment.     

Carbonic anhydrase inhibitors. Interaction of isozymes I, II, IV, V, and IX with organic phosphates and phosphonates

The interaction of five human carbonic anhydrase (hCA, EC 4.2.1.1) isozymes, that is, hCA I, II, IV, V, and IX with a small library of phosphonic acids/organic phosphates, including methylphosphonic acid, MPA; phenylphosphonic acid, PPA; N-(phosphonoacetyl)-L-aspartic acid, PALA, methylene diphosphonic acid MDPA, the O-phosphates of serine (Ser-OP) and threonine (Thr-OP) as well as the antiviral phosphonate foscarnet has been studied. hCA I was activated by all these compounds, with the best activators being MPA and PPA (K(A)s of 0.10-1.20 microM). MPA and PPA were on the other hand nanomolar inhibitors of hCA II (K(I)s of 98-99 nM). PALA showed an affinity of 7.8 microM, whereas the other compounds were weak, millimolar inhibitors of this isozyme. The best hCA IV inhibitors were PALA (79 nM) and PPA (5.4 microM), whereas the other compounds showed K(I)s in the range of 0.31-5.34 mM. The mitochondrial isozyme was weakly inhibited by all these compounds (K(I)s in the range of 0.09-41.7 mM), similarly to the transmembrane, tumor-associated isozyme (K(I)s in the range of 0.86-2.25 mM). Thus, phosphonates may lead to CA inhibitors with selectivity against two physiologically relevant isozymes, the cytosolic hCA II or the membrane-bound hCA IV.     

The determination of the carbonic anhydrases activators in vitro effect of mixed donor crown ethers

Carbonic anhydrases (CAs) play an important function in various physiological and pathological processes. Therefore, many researchers work in this field in order to design and synthesize new drugs. Both inhibitors and activators of CAs, which are associated with the diagnosis and treatment of many diseases, are very important. The emergence of the use of CA activators in the treatment of Alzheimer has led many scholars to work on this issue. In this study, CA activators and inhibitors are determined. The crown ethers compounds ( 1 , 2 , 3 , 6 , 7 , 8 , and 9 ) were found to cause activation on enzyme activities of hCA I and II. The AC₅₀ values on hCA I and II of the compounds are in the range of 4.6565–374.979 μM. The 4 (IC₅₀; 1.301 and 3.215 μM for hCA I and II) and 5 (IC₅₀; 73.96 and 378.5 μM for hCA I and II) compounds were found to cause inhibition on enzyme activities of hCA I and II.     

A class of carbonic anhydrase I – selective activators

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.     

Carbonic anhydrase inhibitors. Inhibition of the cytosolic and tumor-associated carbonic anhydrase isozymes I, II, and IX with a series of 1,3,4-thiadiazole- and 1,2,4-triazole-thiols

A series of heterocyclic mercaptans incorporating 1,3,4-thiadiazole- and 1,2,4-triazole rings have been prepared and assayed for the inhibition of three physiologically relevant carbonic anhydrase (CA, EC 4.2.1.1) isozymes, the cytosolic human isozymes I and II, and the transmembrane, tumor-associated hCA IX. Against hCA I the investigated thiols showed inhibition constants in the range of 97 nM to 548 microM, against hCA II in the range of 7.9-618 microM, and against hCA IX in the range of 9.3-772 microM. Thiadiazoles were generally more active than triazoles against all investigated isozymes. Generally, the best inhibitors were the simple derivative 5-amino-1,3,4-thiadiazole-2-thiol and its N-acetylated derivative, which were anyhow at least two orders of magnitude less effective inhibitors when compared to the corresponding sulfonamides, acetazolamide, and its deacetylated derivative. An exception was constituted by 5-(2-pyridylcarboxamido)-1,3,4-thiadiazole-2-thiol, which is the first hCA I-selective inhibitor ever reported, possessing an inhibition constant of 97 nM against isozyme I, and being a 105 times less effective hCA II inhibitor, and 3154 times less effective hCA IX inhibitor. Thus, the thiol moiety may lead to effective CA inhibitors targeting isozyme I, whereas it is a less effective zinc-binding function for the design of CA II and CA IX inhibitors over the sulfonamide group.     

Effects of dopaminergic compounds on carbonic anhydrase isozymes I, II, and VI

Studies on carbonic anhydrase (CA, EC 4.2.1.1) inhibitors have increased due to several therapeutic applications while there are few investigations on activators. Here we investigated CA inhibitory and activatory capacities of a series of dopaminergic compounds on human carbonic anhydrase (hCA) isozymes I, II, and VI. 2-Amino-1,2,3,4-tetrahydronaphthalene-6,7-diol hydrobromide and 2-amino-1,2,3,4-tetrahydronaphthalene-5,6-diol hydrobromide were found to show effective inhibitory action on hCA I and II whereas 2-amino-5,6-dibromoindan hydrobromide and 2-amino-5-bromoindan hydrobromide exhibited only moderate inhibition against both isoforms, being more effective inhibitors of hCA VI. K(i) values of the molecules 3-6 were in the range of 41.12-363 μM against hCA I, of 0.381-470 μM against hCA II and of 0.578-1.152 μM against hCA VI, respectively. Compound 7 behaved as a CA activator with K(A) values of 27.3 μM against hCA I, of 18.4 μM against hCA II and of 8.73 μM against hCA VI, respectively.     

Carbonic Anhydrase Activators: Synthesis of High Affinity Isozymes I,II and IV Activators,Derivatives of 4-(Arylsulfonylureido-Amino Acyl)Ethyl-1H-Imidazole

Abstract

Based on the X-ray crystallographic structure of the adduct of human carbonic anhydrase II (hCA II) with the weak activator histamine (Briganti, F., Mangani, S., Orioli, P., Scozzafava, A., Vernaglione, G. and Supuran, C.T. (1997) Biochemistry, 36, 10 384–10 392), a novel class of tight-binding CA activators was designed by using histamine (Hst) as lead molecule. Thus, N-1-tritylsulfenyl Hst was synthesized by reaction of Hst with tetrabromophthalic anhydride followed by protection of its imidazole moiety with tritylsulfenyl chloride. After hydrazinolysis, it afforded a key intermediate which was derivatized at the aliphatic amino group. Reaction of the key intermediate with 4-fluorophenylsulfonylureido amino acids (fpu-AA) or 2-toluenesul-fonylureido amino acids (ots-AA) in the presence of carbodiimides, afforded after deprotection, a series of compounds with the general formula fpu/ots-AA-Hst (fpu = 4-FC₆H₄SO₂NHCO; ots = 2-MeC₆H₄SO₂NHCO). Some structurally related dipeptides with the general formula fpu/ ots-AA1-AA2-Hst (AA, AA1 and AA2 represent amino acyl moieties), were also prepared, by a strategy similar to that used for the simple amino acyl compounds above. The new derivatives proved to be efficient in vitro activators of three CA isozymes. Best activity was shown against hCA I and bCA IV, for which some of the new compounds (such as the Lys, Arg. His or the dipeptide derivatives) showed affinities in the 2–12 nm range (h = human; b = bovine isozymes). hCA II was on the other hand somehow less prone to activation by the new derivatives, which possessed affinities around 30–60 nM for this isozyme. Ex vivo experiments showed some of the new activators to strongly enhance red cell CA activity (180–230%) after incubation with human erythrocytes. This new class of CA activators might lead to the development of drugs/ diagnostic tools for the CA deficiency syndrome, a genetic disease of bone, brain and kidneys.     

α-Carbonic anhydrases are strongly activated by spinaceamine derivatives

A series of 4-substituted-spinaceamine (4,5,6,7-tetrahydro-imidazolo[4,5-c]pyridine) were prepared from histamine and aromatic aldehydes Schiff bases, and investigated as activators of four human (h) carbonic anhydrase (CA, EC 4.2.1.1) isoforms, the cytosolic hCA I, II and VII, and the membrane-associated hCA IV. All isoforms were effectively activated by the new derivatives, and the nature of the moiety in position 4 of the bicyclic system was the factor influencing activation properties against all isoforms. For hCA I, these compounds showed K_As in the range of 2.52–21.5?µM, the most effective activator being 4-(2-hydroxyphenyl)-spinaceamine. For hCA II the activation constants ranged between 0.60 and 17.2?µM, with 4-(2,3,5,6-tetrafluorophenyl)- spinaceamine the best activator. Affinity for hCA IV was in the range of 0.52–63.8?µM, and the same compound as for hCA II was the most effective activator. The most sensitive isoform for activation was the brain-associated hCA VII, for which K_As in the range of 82?nM–4.26?µM were observed. Effective hCA VII activators were the (2-bromophenyl)-, 2,3,5,6-tetrafluorophenyl- and furyl-substituted spineaceamines (K_As of 82–95?nM). As CA activators may have pharmacologic applications in various fields, this work provides interesting derivatives for further studies.